Sample records for adipose-derived stromal vascular

  1. Analysis for apoptosis and necrosis on adipocytes, stromal vascular fraction, and adipose-derived stem cells in human lipoaspirates after liposuction.

    PubMed

    Wang, Wei Z; Fang, Xin-Hua; Williams, Shelley J; Stephenson, Linda L; Baynosa, Richard C; Wong, Nancy; Khiabani, Kayvan T; Zamboni, William A

    2013-01-01

    Adipose-derived stem cells have become the most studied adult stem cells. The authors examined the apoptosis and necrosis rates for adipocyte, stromal vascular fraction, and adipose-derived stem cells in fresh human lipoaspirates. Human lipoaspirate (n = 8) was harvested using a standard liposuction technique. Stromal vascular fraction cells were separated from adipocytes and cultured to obtain purified adipose-derived stem cells. A panel of stem cell markers was used to identify the surface phenotypes of cultured adipose-derived stem cells. Three distinct stem cell subpopulations (CD90/CD45, CD105/CD45, and CD34/CD31) were selected from the stromal vascular fraction. Apoptosis and necrosis were determined by annexin V/propidium iodide assay and analyzed by flow cytometry. The cultured adipose-derived stem cells demonstrated long-term proliferation and differentiation evidenced by cell doubling time and positive staining with oil red O and alkaline phosphatase. Isolated from lipoaspirates, adipocytes exhibited 19.7 ± 3.7 percent apoptosis and 1.1 ± 0.3 percent necrosis; stromal vascular fraction cells revealed 22.0 ± 6.3 percent of apoptosis and 11.2 ± 1.9 percent of necrosis; stromal vascular fraction cells had a higher rate of necrosis than adipocytes (p < 0.05). Among the stromal vascular fraction cells, 51.1 ± 3.7 percent expressed CD90/CD45, 7.5 ± 1.0 percent expressed CD105/CD45, and 26.4 ± 3.8 percent expressed CD34/CD31. CD34/CD31 adipose-derived stem cells had lower rates of apoptosis and necrosis compared with CD105/CD45 adipose-derived stem cells (p < 0.05). Adipose-derived stem cells had a higher rate of apoptosis and necrosis than adipocytes. However, the extent of apoptosis and necrosis was significantly different among adipose-derived stem cell subpopulations.

  2. Comparative characterization of stromal vascular cells derived from three types of vascular wall and adipose tissue.

    PubMed

    Yang, Santsun; Eto, Hitomi; Kato, Harunosuke; Doi, Kentaro; Kuno, Shinichiro; Kinoshita, Kahori; Ma, Hsu; Tsai, Chi-Han; Chou, Wan-Ting; Yoshimura, Kotaro

    2013-12-01

    Multipotent stem/progenitor cells localize perivascularly in many organs and vessel walls. These tissue-resident stem/progenitor cells differentiate into vascular endothelial cells, pericytes, and other mesenchymal lineages, and participate in physiological maintenance and repair of vasculatures. In this study, we characterized stromal vascular cells obtained through the explant culture method from three different vessel walls in humans: arterial wall (ART; >500 μm in diameter), venous wall (VN; >500 μm in diameter), and small vessels in adipose tissue (SV; arterioles and venules, <100 μm in diameter). These were examined for functionality and compared with adipose-derived stem/stromal cells (ASCs). All stromal vascular cells of different origins presented fibroblast-like morphology and we could not visually discriminate one population from another. Flow cytometry showed that the cultured population heterogeneously expressed a variety of surface antigens associated with stem/progenitor cells, but CD105 was expressed by most cells in all groups, suggesting that the cells generally shared the characteristics of mesenchymal stem cells. Our histological and flow cytometric data suggested that the main population of vessel wall-derived stromal vascular cells were CD34(+)/CD31(-) and came from the tunica adventitia and areola tissue surrounding the adventitia. CD271 (p75NTR) was expressed by the vasa vasorum in the VN adventitia and by a limited population in the adventitia of SV. All three populations differentiated into multiple lineages as did ASCs. ART cells induced the largest quantity of calcium formation in the osteogenic medium, whereas ASCs showed the greatest adipogenic differentiation. SV and VN stromal cells had greater potency for network formation than did ART stromal cells. In conclusion, the three stromal vascular populations exhibited differential functional properties. Our results have clinical implications for vascular diseases such as

  3. Uncultivated stromal vascular fraction is equivalent to adipose-derived stem and stromal cells on porous polyurethrane scaffolds forming adipose tissue in vivo.

    PubMed

    Griessl, Michael; Buchberger, Anna-Maria; Regn, Sybille; Kreutzer, Kilian; Storck, Katharina

    2018-06-01

    To find an alternative approach to contemporary techniques in tissue augmentation and reconstruction, tissue engineering strategies aim to involve adipose-derived stem and stromal cells (ASCs) harboring a strong differentiation potential into various tissue types such as bone, cartilage, and fat. Animal research. The stromal vascular fraction (SVF) was used directly as a cell source to provide a potential alternative to contemporary ASC-based adipose tissue engineering. Seeded in TissuCol fibrin, we applied ASCs or SVF cells to porous, degradable polyurethane (PU) scaffolds. We successfully demonstrated the in vivo generation of volume-stable, well-vascularized PU-based constructs containing host-derived mature fat pads. Seeded human stem cells served as modulators of host-cell migration rather than differentiating themselves. We further demonstrated that preliminary culture of SVF cells was not necessary. Our results bring adipose tissue engineering, together with automated processing devices, closer to clinical applicability. The time-consuming and cost-intensive culture and induction of the ASCs is not necessary. NA. Laryngoscope, 128:E206-E213, 2018. © 2018 The American Laryngological, Rhinological and Otological Society, Inc.

  4. Adipose-Derived Stromal Vascular Fraction Differentially Expands Breast Progenitors in Tissue Adjacent to Tumors Compared to Healthy Breast Tissue

    PubMed Central

    Chatterjee, Sumanta; Laliberte, Mike; Blelloch, Sarah; Ratanshi, Imran; Safneck, Janice; Buchel, Ed

    2015-01-01

    Background: Autologous fat grafts supplemented with adipose-derived stromal vascular fraction are used in reconstructive and cosmetic breast procedures. Stromal vascular fraction contains adipose-derived stem cells that are thought to encourage wound healing, tissue regeneration, and graft retention. Although use of stromal vascular fraction has provided exciting perspectives for aesthetic procedures, no studies have yet been conducted to determine whether its cells contribute to breast tissue regeneration. The authors examined the effect of these cells on the expansion of human breast epithelial progenitors. Methods: From patients undergoing reconstructive breast surgery following mastectomies, abdominal fat, matching tissue adjacent to breast tumors, and the contralateral non–tumor-containing breast tissue were obtained. Ex vivo co-cultures using breast epithelial cells and the stromal vascular fraction cells were used to study the expansion potential of breast progenitors. Breast reduction samples were collected as a source of healthy breast cells. Results: The authors observed that progenitors present in healthy breast tissue or contralateral non–tumor-containing breast tissue showed significant and robust expansion in the presence of stromal vascular fraction (5.2- and 4.8-fold, respectively). Whereas the healthy progenitors expanded up to 3-fold without the stromal vascular fraction cells, the expansion of tissue adjacent to breast tumor progenitors required the presence of stromal vascular fraction cells, leading to a 7-fold expansion, which was significantly higher than the expansion of healthy progenitors with stromal vascular fraction. Conclusions: The use of stromal vascular fraction might be more beneficial to reconstructive operations following mastectomies compared with cosmetic corrections of the healthy breast. Future studies are required to examine the potential risk factors associated with its use. CLINICAL QUESTION/LEVEL OF EVIDENCE

  5. Stromal cells from the adipose tissue-derived stromal vascular fraction and culture expanded adipose tissue-derived stromal/stem cells: a joint statement of the International Federation for Adipose Therapeutics and Science (IFATS) and the International Society for Cellular Therapy (ISCT).

    PubMed

    Bourin, Philippe; Bunnell, Bruce A; Casteilla, Louis; Dominici, Massimo; Katz, Adam J; March, Keith L; Redl, Heinz; Rubin, J Peter; Yoshimura, Kotaro; Gimble, Jeffrey M

    2013-06-01

    Adipose tissue is a rich and very convenient source of cells for regenerative medicine therapeutic approaches. However, a characterization of the population of adipose-derived stromal and stem cells (ASCs) with the greatest therapeutic potential remains unclear. Under the authority of International Federation of Adipose Therapeutics and International Society for Cellular Therapy, this paper sets out to establish minimal definitions of stromal cells both as uncultured stromal vascular fraction (SVF) and as an adherent stromal/stem cells population. Phenotypic and functional criteria for the identification of adipose-derived cells were drawn from the literature. In the SVF, cells are identified phenotypically by the following markers: CD45-CD235a-CD31-CD34+. Added value may be provided by both a viability marker and the following surface antigens: CD13, CD73, CD90 and CD105. The fibroblastoid colony-forming unit assay permits the evaluation of progenitor frequency in the SVF population. In culture, ASCs retain markers in common with other mesenchymal stromal/stem cells (MSCs), including CD90, CD73, CD105, and CD44 and remain negative for CD45 and CD31. They can be distinguished from bone-marrow-derived MSCs by their positivity for CD36 and negativity for CD106. The CFU-F assay is recommended to calculate population doublings capacity of ASCs. The adipocytic, chondroblastic and osteoblastic differentiation assays serve to complete the cell identification and potency assessment in conjunction with a quantitative evaluation of the differentiation either biochemically or by reverse transcription polymerase chain reaction. The goal of this paper is to provide initial guidance for the scientific community working with adipose-derived cells and to facilitate development of international standards based on reproducible parameters. Copyright © 2013 International Society for Cellular Therapy. All rights reserved.

  6. Non-canonical Wnt Signaling Enhances differentiation of Sca1+/c-kit+ Adipose-derived Murine Stromal Vascular Cells into Spontaneously Beating Cardiac Myocytes

    PubMed Central

    Palpant, Nathan J.; Yasuda, So-ichiro; MacDougald, Ormond; Metzger, Joseph M.

    2007-01-01

    Recent reports have described a stem cell population termed stromal vascular cells (SVCs) derived from the stromal vascular fraction of adipose tissue, which are capable of intrinsic differentiation into spontaneously beating cardiomyocytes in vitro. The objective of this study was to further define the cardiac lineage differentiation potential of SVCs in vitro and to derive methods for enriching SVC-derived beating cardiac myocytes. SVCs were isolated from the stromal vascular fraction of murine adipose tissue. Cells were cultured in methylcellulose-based murine stem cell media. Analysis of SVC-derived beating myocytes included Western blot, and calcium imaging. Enrichment of acutely isolated SVCs was carried out using antibody tagged magnetic nanoparticles, and pharmacologic manipulation of Wnt and cytokine signaling. Under initial media conditions, spontaneously beating SVCs expressed both cardiac developmental and adult protein isoforms. Functionally, this specialized population can spontaneously contract and pace under field stimulation, and shows the presence of coordinated calcium transients. Importantly, this study provides evidence for two independent mechanisms of enriching the cardiac differentiation of SVCs. First, this study shows that differentiation of SVCs into cardiac myocytes is augmented by non-canonical Wnt agonists, canonical Wnt antagonists, and cytokines. Second, SVCs capable of cardiac lineage differentiation can be enriched by selection for stem cell-specific membrane markers Sca1 and c-kit. Adipose-derived SVCs are a unique population of stem cells that show evidence of cardiac lineage development making them a potential source for stem cell-based cardiac regeneration studies. PMID:17706246

  7. Non-canonical Wnt signaling enhances differentiation of Sca1+/c-kit+ adipose-derived murine stromal vascular cells into spontaneously beating cardiac myocytes.

    PubMed

    Palpant, Nathan J; Yasuda, So-ichiro; MacDougald, Ormond; Metzger, Joseph M

    2007-09-01

    Recent reports have described a stem cell population termed stromal vascular cells (SVCs) derived from the stromal vascular fraction of adipose tissue, which are capable of intrinsic differentiation into spontaneously beating cardiomyocytes in vitro. The objective of this study was to further define the cardiac lineage differentiation potential of SVCs in vitro and to establish methods for enriching SVC-derived beating cardiac myocytes. SVCs were isolated from the stromal vascular fraction of murine adipose tissue. Cells were cultured in methylcellulose-based murine stem cell media. Analysis of SVC-derived beating myocytes included Western blot and calcium imaging. Enrichment of acutely isolated SVCs was carried out using antibody-tagged magnetic nanoparticles, and pharmacologic manipulation of Wnt and cytokine signaling. Under initial media conditions, spontaneously beating SVCs expressed both cardiac developmental and adult protein isoforms. Functionally, this specialized population can spontaneously contract and pace under field stimulation and shows the presence of coordinated calcium transients. Importantly, this study provides evidence for two independent mechanisms of enriching the cardiac differentiation of SVCs. First, this study shows that differentiation of SVCs into cardiac myocytes is augmented by non-canonical Wnt agonists, canonical Wnt antagonists, and cytokines. Second, SVCs capable of cardiac lineage differentiation can be enriched by selection for stem cell-specific membrane markers Sca1 and c-kit. Adipose-derived SVCs are a unique population of stem cells that show evidence of cardiac lineage development making them a potential source for stem cell-based cardiac regeneration studies.

  8. Flow cytometry on the stromal-vascular fraction of white adipose tissue

    USDA-ARS?s Scientific Manuscript database

    Adipose tissue contains cell types other than adipocytes that may contribute to complications linked to obesity. For example, macrophages have been shown to infiltrate adipose tissue in response to a high-fat diet. Isolation of the stromal-vascular fraction of adipose tissue allows one to use flow c...

  9. Stromal vascular cells and adipogenesis: Cells within adipose depots regulate adipogenesis

    USDA-ARS?s Scientific Manuscript database

    A collection of investigations indicate the importance of adipose tissue stromal/stem cells to vasculogenesis and angiogenesis during adipogenesis. Early in development the stromal-vascular (S-V) elements control and dictate the extent of adipogenesis in a depot dependent manner. For instance, the...

  10. Successful management of an equine carpal chip fracture by intra-articularly injected adipose-derived stromal vascular fraction after arthroscopic removal.

    PubMed

    Tyrnenopoulou, P; Karayannopoulou, M; Angelopoulou, S; Pyrros, A; Mparous, E; Koliakos, G; Diakakis, N

    2016-01-01

    Carpal chip fractures are common causes of lameness in racehorses. Due to disadvantages in surgical management, adjuvant treatment modalities are usually necessary. Adipose-derived stem cells (ADSCs) have the potential to differentiate into other cell types including bone and cartilage cells. Adipose-derived stromal vascular fraction (SVF) is produced during ADSCs isolation from adipose tissue. The purpose of this report was to present the successful management of a grade III chip fracture in the right carpus of a 5-year-old Thoroughbred gelding by intra-articularly injected autologous SVF one month after the arthroscopic removal of the fracture. This treatment resulted in lameness improvement and short rehabilitation period to previous racing activities. High performance levels and no recurrent injuries were recorded during a twenty month follow-up period.

  11. Gender-dimorphic effects of adipose-derived stromal vascular fractions on HUVECs exposed to oxidative stress.

    PubMed

    Lim, Soyeon; Kim, Il-Kwon; Choi, Jung-Won; Seo, Hyang-Hee; Lim, Kyu Hee; Lee, Seahyoung; Lee, Hoon-Bum; Kim, Sang Woo; Hwang, Ki-Chul

    2017-01-01

    Stromal vascular fractions (SVFs) are a heterogeneous collection of cells within adipose tissue that are being studied for various clinical indications. In this study, we aimed to determine whether SVF transplantation into impaired tissues has differential effects on inflammatory and angiogenetic properties with regard to gender. As reactive oxygen species have been implicated in cardiovascular disease development, we investigated differences in gene and protein expression related to inflammation and angiogenesis in HUVECs co-cultured with adipose-derived SVFs from male (M group) and female (F group) individuals under oxidative stress conditions. The expression of several inflammatory (interleukin (IL)-33) and angiogenetic (platelet-derived growth factor (PDGF)) factors differed dramatically between male and female donors. Anti-inflammatory and pro-angiogenetic responses were observed in HUVECs co-cultured with SVFs under oxidative stress conditions, and these characteristics may exhibit partially differential effects according to gender. Using network analysis, we showed that co-culturing HUVECs with SVFs ameliorated pyroptosis/apoptosis via an increase in oxidative stress. Activation of caspase-1 and IL-1B was significantly altered in HUVECs co-cultured with SVFs from female donors. These findings regarding gender-dimorphic regulation of adipose-derived SVFs provide valuable information that can be used for evidence-based gender-specific clinical treatment of SVF transplantation for understanding of cardiovascular disease, allowing for the development of additional treatment.

  12. Flow cytometry on the stromal-vascular fraction of white adipose tissue.

    PubMed

    Brake, Danett K; Smith, C Wayne

    2008-01-01

    Adipose tissue contains cell types other than adipocytes that may contribute to complications linked to obesity. For example, macrophages have been shown to infiltrate adipose tissue in response to a high-fat diet. Isolation of the stromal-vascular fraction of adipose tissue allows one to use flow cytometry to analyze cell surface markers on leukocytes. Here, we present a technical approach to identify subsets of leukocytes that differentially express cell surface markers.

  13. Photobiomodulation of freshly isolated human adipose tissue-derived stromal vascular fraction cells by pulsed light-emitting diodes for direct clinical application.

    PubMed

    Priglinger, E; Maier, J; Chaudary, S; Lindner, C; Wurzer, C; Rieger, S; Redl, H; Wolbank, S; Dungel, P

    2018-06-01

    A highly interesting source for adult stem cells is adipose tissue, from which the stromal vascular fraction (SVF)-a heterogeneous cell population including the adipose-derived stromal/stem cells-can be obtained. To enhance the regenerative potential of freshly isolated SVF cells, low-level light therapy (LLLT) was used. The effects of pulsed blue (475 nm), green (516 nm), and red (635 nm) light from light-emitting diodes applied on freshly isolated SVF were analysed regarding cell phenotype, cell number, viability, adenosine triphosphate content, cytotoxicity, and proliferation but also osteogenic, adipogenic, and proangiogenic differentiation potential. The colony-forming unit fibroblast assay revealed a significantly increased colony size after LLLT with red light compared with untreated cells, whereas the frequency of colony-forming cells was not affected. LLLT with green and red light resulted in a stronger capacity to form vascular tubes by SVF when cultured within 3D fibrin matrices compared with untreated cells, which was corroborated by increased number and length of the single tubes and a significantly higher concentration of vascular endothelial growth factor. Our study showed beneficial effects after LLLT on the vascularization potential and proliferation capacity of SVF cells. Therefore, LLLT using pulsed light-emitting diode light might represent a new approach for activation of freshly isolated SVF cells for direct clinical application. Copyright © 2018 John Wiley & Sons, Ltd.

  14. Adipose Tissue-Derived Stromal Cells for Wound Healing.

    PubMed

    Goodarzi, Parisa; Alavi-Moghadam, Sepideh; Sarvari, Masoumeh; Tayanloo Beik, Akram; Falahzadeh, Khadijeh; Aghayan, Hamidreza; Payab, Moloud; Larijani, Bagher; Gilany, Kambiz; Rahim, Fakher; Adibi, Hossein; Arjmand, Babak

    2018-06-02

    Skin as the outer layer covers the body. Wounds can affect this vital organ negatively and disrupt its functions. Wound healing as a biological process is initiated immediately after an injury. This process consists of three stages: inflammation, proliferation, remodeling. Generally, these three stages occur continuously and timely. However, some factors such as infection, obesity and diabetes mellitus can interfere with these stages and impede the normal healing process which results in chronic wounds. Financial burden on both patients and health care systems, negative biologic effect on the patient's general health status and reduction in quality of life are a number of issues which make chronic wounds as a considerable challenge. During recent years, along with advances in the biomedical sciences, various surgical and non-surgical therapeutic methods have been suggested. All of these suggested treatments have their own advantages and disadvantages. Recently, cell-based therapies and regenerative medicine represent promising approaches to wound healing. Accordingly, several types of mesenchymal stem cells have been used in both preclinical and clinical settings for the treatment of wounds. Adipose-derived stromal cells are a cost-effective source of mesenchymal stem cells in wound management which can be easily harvest from adipose tissues through the less invasive processes with high yield rates. In addition, their ability to secrete multiple cytokines and growth factors, and differentiation into skin cells make them an ideal cell type to use in wound treatment. This is a concise overview on the application of adipose-derived stromal cells in wound healing and their role in the treatment of chronic wounds.

  15. Anti-apoptotic effects of adipose-derived adherent stromal cells in mesenchymal stem cells exposed to oxidative stress.

    PubMed

    Shin, Sunhye; Choi, Jung-Won; Lim, Soyeon; Lee, Seahyoung; Jun, Eun-Young; Sun, Hyun-Min; Kim, Il-Kwon; Lee, Hoon-Bum; Kim, Sang Woo; Hwang, Ki-Chul

    2018-06-19

    Adipose-derived stromal vascular fractions (SVFs) are a heterogeneous collection of cells, and their regenerative modality has been applied in various animal experiments and clinical trials. Despite the attractive advantages of SVFs in clinical interventions, the recent status of clinical studies involving the application of SVFs in many diseases has not been fully evaluated. Mesenchymal stem cells (MSCs) are multipotent stromal cells that can differentiate into a variety of cell types despite their low numbers in heart tissue. Here, we sought to determine if SVF implantation into impaired heart tissue affected endogenous MSCs in the heart. Therefore, we investigated the expression levels of proteins associated with oxidation, inflammation, and apoptosis in MSCs co-cultured with adipose-derived adherent stromal cells (ADASs) from 6 donors' SVFs under oxidative stress conditions for their roles in many physiological processes in the heart. Interestingly, p53 pathway proteins and mitogen-activated protein kinase (MAPK) signalling pathway components were up-regulated by H 2 O 2 but exhibited a downward trend in MSCs co-cultured with ADASs. These data suggest that ADASs may inhibit oxidative stress-induced apoptosis in MSCs via the p53 and MAPK pathways. Our findings also suggest that the positive effects of SVF implantation into damaged heart tissue may be attributed to the various responses of MSCs. This finding may provide new insights for the clinical application of adipose-derived SVF transplantation in cardiac diseases. We investigated the expression levels of proteins associated with oxidation, inflammation, and apoptosis in MSCs co-cultured with isolated ADASs from 6 donors' SVFs under oxidative stress conditions. Our results imply that isolated ADASs from SVFs may inhibit oxidative stress-induced cell cycle arrest and/or apoptosis in MSCs via a p53-dependent pathway. Furthermore, we identified an anti-apoptotic mechanism involving oxidative stress

  16. Cell sheet engineering using the stromal vascular fraction of adipose tissue as a vascularization strategy.

    PubMed

    Costa, Marina; Cerqueira, Mariana T; Santos, Tírcia C; Sampaio-Marques, Belém; Ludovico, Paula; Marques, Alexandra P; Pirraco, Rogério P; Reis, Rui L

    2017-06-01

    Current vascularization strategies for Tissue Engineering constructs, in particular cell sheet-based, are limited by time-consuming and expensive endothelial cell isolation and/or by the complexity of using extrinsic growth factors. Herein, we propose an alternative strategy using angiogenic cell sheets (CS) obtained from the stromal vascular fraction (SVF) of adipose tissue that can be incorporated into more complex constructs. Cells from the SVF were cultured in normoxic and hypoxic conditions for up to 8days in the absence of extrinsic growth factors. Immunocytochemistry against CD31 and CD146 revealed spontaneous organization in capillary-like structures, more complex after hypoxic conditioning. Inhibition of HIF-1α pathway hindered capillary-like structure formation in SVF cells cultured in hypoxia, suggesting a role of HIF-1α. Moreover, hypoxic SVF cells showed a trend for increased secretion of angiogenic factors, which was reflected in increased network formation by endothelial cells cultured on matrigel using that conditioned medium. In vivo implantation of SVF CS in a mouse hind limb ischemia model revealed that hypoxia-conditioned CS led to improved restoration of blood flow. Both in vitro and in vivo data suggest that SVF CS can be used as simple and cost-efficient tools to promote functional vascularization of TE constructs. Neovascularization after implantation is a major obstacle for producing clinically viable cell sheet-based tissue engineered constructs. Strategies using endothelial cells and extrinsic angiogenic growth factors are expensive and time consuming and may raise concerns of tumorigenicity. In this manuscript, we describe a simplified approach using angiogenic cell sheets fabricated from the stromal vascular fraction of adipose tissue. The strong angiogenic behavior of these cell sheets, achieved without the use of external growth factors, was further stimulated by low oxygen culture. When implanted in an in vivo model of hind limb

  17. Adipose Derived Stromal Cell (ADSC) Injections for Pain Management of Osteoarthritis in the Human Knee Joint.

    PubMed

    Fodor, Peter B; Paulseth, Stephen G

    2016-02-01

    This safety and feasibility study used autologous adipose-derived stromal vascular cells (the stromal vascular fraction [SVF] of adipose tissue), to treat 8 osteoarthritic knees in 6 patients of grade I to III (K-L scale) with initial pain of 4 or greater on a 10-point Visual Analog Scale (VAS). The primary objective of the study was evaluation of the safety of intra-articular injection of SVF. The secondary objective was to assess initial feasibility for reduction of pain in osteoarthritic knees. Adipose-derived SVF cells were obtained through enzymatic disaggregation of lipoaspirate, resuspension in 3 mL of Lactated Ringer's Solution, and injection directly into the intra-articular space of the knee, with a mean of 14.1 million viable, nucleated SVF cells per knee. Metrics included monitoring of adverse events and preoperative to postoperative changes in the Western Ontario and McMaster Universities Arthritis Index (WOMAC), the VAS pain scale, range of motion (ROM), timed up-and-go (TUG), and MRI. No infections, acute pain flares, or other adverse events were reported. At 3-months postoperative, there was a statistically significant improvement in WOMAC and VAS scores (P < .02 and P < .001, respectively), which was maintained at 1 year. Physical therapy measurements for ROM and TUG both improved from preoperative to 3-months postoperative. Standard MRI assessment from preoperative to 3-months postoperative showed no detectable structural differences. All patients attained full activity with decreased knee pain. Autologous SVF was shown to be safe and to present a new potential therapy for reduction of pain for osteoarthritis of the knee. LEVEL OF EVIDENCE 4: Therapeutic. © 2015 The American Society for Aesthetic Plastic Surgery, Inc. Reprints and permission: journals.permissions@oup.com.

  18. Adipose-derived stromal cells for the reconstruction of a human vesical equivalent.

    PubMed

    Rousseau, Alexandre; Fradette, Julie; Bernard, Geneviève; Gauvin, Robert; Laterreur, Véronique; Bolduc, Stéphane

    2015-11-01

    Despite a wide panel of tissue-engineering models available for vesical reconstruction, the lack of a differentiated urothelium remains their main common limitation. For the first time to our knowledge, an entirely human vesical equivalent, free of exogenous matrix, has been reconstructed using the self-assembly method. Moreover, we tested the contribution of adipose-derived stromal cells, an easily available source of mesenchymal cells featuring many potential advantages, by reconstructing three types of equivalent, named fibroblast vesical equivalent, adipose-derived stromal cell vesical equivalent and hybrid vesical equivalent--the latter containing both adipose-derived stromal cells and fibroblasts. The new substitutes have been compared and characterized for matrix composition and organization, functionality and mechanical behaviour. Although all three vesical equivalents displayed adequate collagen type I and III expression, only two of them, fibroblast vesical equivalent and hybrid vesical equivalent, sustained the development of a differentiated and functional urothelium. The presence of uroplakins Ib, II and III and the tight junction marker ZO-1 was detected and correlated with impermeability. The mechanical resistance of these tissues was sufficient for use by surgeons. We present here in vitro tissue-engineered vesical equivalents, built without the use of any exogenous matrix, able to sustain mechanical stress and to support the formation of a functional urothelium, i.e. able to display a barrier function similar to that of native tissue. Copyright © 2013 John Wiley & Sons, Ltd.

  19. Safety of intrathecal autologous adipose-derived mesenchymal stromal cells in patients with ALS

    PubMed Central

    Madigan, Nicolas N.; Morris, Jonathan; Jentoft, Mark; Sorenson, Eric J.; Butler, Greg; Gastineau, Dennis; Dietz, Allan; Windebank, Anthony J.

    2016-01-01

    Objective: To determine the safety of intrathecal autologous adipose-derived mesenchymal stromal cell treatment for amyotrophic lateral sclerosis (ALS). Methods: Participants with ALS were enrolled and treated in this phase I dose-escalation safety trial, ranging from 1 × 107 (single dose) to 1 × 108 cells (2 monthly doses). After intrathecal treatments, participants underwent standardized follow-up, which included clinical examinations, revised ALS Functional Rating Scale (ALSFRS-R) questionnaire, blood and CSF sampling, and MRI of the neuroaxis. Results: Twenty-seven patients with ALS were enrolled and treated in this study. The safety profile was positive, with the most common side effects reported being temporary low back and radicular leg pain at the highest dose level. These clinical findings were associated with elevated CSF protein and nucleated cells with MRI of thickened lumbosacral nerve roots. Autopsies from 4 treated patients did not show evidence of tumor formation. Longitudinal ALSFRS-R questionnaires confirmed continued progression of disease in all treated patients. Conclusions: Intrathecal treatment of autologous adipose-derived mesenchymal stromal cells appears safe at the tested doses in ALS. These results warrant further exploration of efficacy in phase II trials. Classification of evidence: This phase I study provides Class IV evidence that in patient with ALS, intrathecal autologous adipose-derived mesenchymal stromal cell therapy is safe. PMID:27784774

  20. Adipose-derived cells.

    PubMed

    Meliga, Emanuele; Strem, Brian M; Duckers, H J; Serruys, Patrick W

    2007-01-01

    Heart failure is by far the most common cause of hospitalization in Western countries, with onerous economic consequences. Cell therapy holds great promise for use in tissue regeneration and is increasingly used in an effort to improve outcomes in cardiac disease. Recently it has been shown that adipose tissue, in addition to committed adipogenic, endothelial progenitor cells and pluripotent vascular progenitor cells, also contains multipotent cell types (adipose-derived stem cells, ADSCs) that, in cell culture conditions, have shown to have an impressive developmental plasticity including the ability to undergo multilineage differentiation and self-renewal. ADSCs express multiple CD marker antigens similar to those observed on MSCs and are also capable of secreting a large number of angiogenesis-related cytokines, including vascular endothelial growth factor, granulocyte/macrophage colony stimulating factor, stromal-derived factor-1alpha, and hepatocyte growth factor. Adipose tissue can be harvested in large quantities with minimal morbidity in several regions of the body and, on average, 100 ml of human adipose tissue yields about 1 x 10(6) stem cells. Studies conducted in porcine AMI models have shown a significant LV functional improvement, with no report of any potentially fatal arrhythmias. The APOLLO trial, a prospective, double blind, randomized, placebo-controlled trial currently in the recruiting phase, is a "first-in-man" study that explores the safety and feasibility of ADSC transplantation in patients with acute MI.

  1. IFATS Collection: Adipose-Derived Stromal Cells Improve the Foreign Body Response

    PubMed Central

    Prichard, Heather L.; Reichert, William; Klitzman, Bruce

    2015-01-01

    Many implanted devices fail due to the formation of an avascular capsule surrounding the device. Additionally, fat has long been known to promote healing and vascularization. The goals of this study were to identify potential mechanisms of the provascular actions of adipose-derived stromal cells (ASCs) and to improve implant biocompatibility. First, adult ASCs and fibroblasts from rats were attached to polyurethane and polystyrene in vitro and their cytokine secretion profile was analyzed. Secretion of vascular endothelial growth factor (VEGF) from ASCs was 10 –70 times higher than fibroblasts after 3 and 6 days. Next, polyurethane, bare and with cellular coatings, was implanted subcutaneously in rats. The fibrous capsule surrounding bare polyurethane implants was 17%–32% thicker and the amount of collagen was 27% greater than the capsule surrounding ASC-coated implants. Finally, the microvessel density adjacent to ASC-coated polyurethane was approximately 50%–80% higher than bare polyurethane. In summary, ASCs attached to polyurethane have a dramatically increased VEGF production compared with fibroblasts in vitro, and these cells also produce an increased microvessel density in the surrounding tissue when implanted subcutaneously in rats. PMID:18436858

  2. Identification of active and quiescent adipose vascular stromal cells.

    PubMed

    Lin, Guiting; Xin, Zhongcheng; Zhang, Haiyang; Banie, Lia; Wang, Guifang; Qiu, Xuefeng; Ning, Hongxiu; Lue, Tom F; Lin, Ching-Shwun

    2012-02-01

    Recent studies have demonstrated the existence of both active and quiescent stem cells in bone marrow, hair follicle and intestine. We attempted to identify active and quiescent vascular stromal cells (VSC) in adipose tissue. For identification of active VSC, adult rats were injected intraperitoneally with thymidine analog 5-ethynyl-2-deoxyuridine (EdU) and their subcutaneous tissue harvested 3 days later. For identification of quiescent VSC, newborn rats were injected intraperitoneally with EdU and their subcutaneous tissue harvested 9 weeks later. The harvested adipose tissues were examined for the co-localization of EdU with VSC marker CD34, smooth muscle marker SMA, endothelial marker RECA and pericyte marker CD140b. In adult rat adipose tissues harvested 3 days after EdU injection, there were 28.80 ± 8.70 (mean ± SD) EdU+ cells/100 × microscopic field, and approximately 6.2% of cell nuclei were labeled with EdU. The percentages of EdU+ cells expressing the following markers were approximately: 84 for CD34, 5.6 for RECA (rat endothelial marker), 3.7 for SMA and 14.8 for CD140b. In the adipose tissues of newborn rats that were harvested 9 weeks after EdU injection, the percentages of EdU+ cells expressing the following markers were approximately: 76 for CD34, 1.8 for RECA, 0 for SMA and 12.9 for CD140b. In both the short-term (active) and long-term (quiescent) EdU-labeled adipose tissues, the EdU label was consistently co-localized with CD34 and in the proximity of CD140b stain or in the adventitia. Both active and quiescent VSC expressed CD34 and localized to capillaries and the adventitia of larger blood vessels.

  3. The effect of adipose derived stromal vascular fraction on stasis zone in an experimental burn model.

    PubMed

    Eyuboglu, Atilla Adnan; Uysal, Cagri A; Ozgun, Gonca; Coskun, Erhan; Markal Ertas, Nilgun; Haberal, Mehmet

    2018-03-01

    Stasis zone is the surrounding area of the coagulation zone which is an important part determining the extent of the necrosis in burn patients. In our study we aim to salvage the stasis zone by injecting adipose derived stromal vascular fraction (ADSVF). Thermal injury was applied on dorsum of Sprague-Dawley rats (n=20) by the "comb burn" model as described previously. When the burn injury was established on Sprague-Dawley rats (30min); rat dorsum was separated into 2 equal parts consisting of 4 burn zones (3 stasis zone) on each pair. ADSVF cells harvested from inguinal fat pads of Sprague-Dawley rats (n=5) were injected on the right side while same amount of phosphate buffered saline (PBS) injected on the left side of the same animal. One week later, average vital tissue on the statis zone was determined by macroscopy, angiography and microscopy. Vascular density, inflammatory cell density, gradient of fibrosis and epithelial thickness were determined via immunohistochemical assay. Macroscopic stasis zone tissue viability (32±3.28%, 57±4.28%) (p<0.01), average number of vessels (10.28±1.28, 19.43±1.72) (p<0.01), capillary count (15.67±1.97, 25.35±2.15) (p<0.01) vascular density (1.55±0.38, 2.14±0.45) (p<0.01) epithelial thickness (0.014±0.009mm, 0.024±0.0011mm) were higher on ADSVF side. Fibrosis gradient (1.87±0.51, 1.50±0.43) (p<0.01) and inflammatory cell density (1.33±0.40, 1.20±0.32) (p<0.01) were higher on the PBS side. Macroscopic and microscopic findings determined that ADSVF has a statistically significant benefit for salvaging stasis zone on acute burn injuries. Copyright © 2017 Elsevier Ltd and ISBI. All rights reserved.

  4. Adipose Stromal Vascular Fraction Isolation: A Head-to-Head Comparison of 4 Cell Separation Systems #2.

    PubMed

    Aronowitz, Joel A; Lockhart, Ryan A; Hakakian, Cloe S; Birnbaum, Zoe E

    2016-09-01

    With stromal vascular fraction (SVF) cell and adipose-derived stem cell-based technologies translating into the clinical setting, numerous isolation systems have been developed for the point of care isolation of SVF cells from adipose tissue. A relative lack of performance data on these systems can make objective assessment difficult for prospective clinicians. This study compared the performance of 4 SVF cell isolation systems. Four isolation systems were compared: the MultiStation by PNC International, the LipoKit by MediKhan, the GID SVF-2 platform by GID Europe Ltd, and the StemSource 900/MB system by Cytori Therapeutics, Inc. Identical lipoaspirate samples for 5 separate donors were used. Stromal vascular fraction output was compared in terms of nucleated cell yield, viability, residual collagenase activity, sterility of the output, colony-forming unit-fibroblast frequency, frequency of CD31-/CD34+/CD45- cells, and operating statistics. Mean process time ranged from 65.4 to 120.8 minutes. Mean nucleated cell yield per milliliter of tissue processed ranged from 1.01 × 10 cells/mL to 6.24 × 10 cells/mL. Mean cellular viability ranged from 50.3% to 84.02%. Residual collagenase activity was negligible across all systems. Observed colony-forming unit-fibroblast frequency ranged from 0.495% to 1.704%. No significant difference was observed in frequency of CD31-/CD34+/CD45- cells. Results of the anaerobic/aerobic cultures were mixed. There was considerable variability between the outputs of each system. The system used by a clinician should be tailored to the individual needs of the practice. There is a range of cost options available. This study may help clinicians make more educated decisions when choosing an isolation system to meet their clinical needs.

  5. Improvement of adipose tissue-derived cells by low-energy extracorporeal shock wave therapy.

    PubMed

    Priglinger, Eleni; Schuh, Christina M A P; Steffenhagen, Carolin; Wurzer, Christoph; Maier, Julia; Nuernberger, Sylvia; Holnthoner, Wolfgang; Fuchs, Christiane; Suessner, Susanne; Rünzler, Dominik; Redl, Heinz; Wolbank, Susanne

    2017-09-01

    Cell-based therapies with autologous adipose tissue-derived cells have shown great potential in several clinical studies in the last decades. The majority of these studies have been using the stromal vascular fraction (SVF), a heterogeneous mixture of fibroblasts, lymphocytes, monocytes/macrophages, endothelial cells, endothelial progenitor cells, pericytes and adipose-derived stromal/stem cells (ASC) among others. Although possible clinical applications of autologous adipose tissue-derived cells are manifold, they are limited by insufficient uniformity in cell identity and regenerative potency. In our experimental set-up, low-energy extracorporeal shock wave therapy (ESWT) was performed on freshly obtained human adipose tissue and isolated adipose tissue SVF cells aiming to equalize and enhance stem cell properties and functionality. After ESWT on adipose tissue we could achieve higher cellular adenosine triphosphate (ATP) levels compared with ESWT on the isolated SVF as well as the control. ESWT on adipose tissue resulted in a significantly higher expression of single mesenchymal and vascular marker compared with untreated control. Analysis of SVF protein secretome revealed a significant enhancement in insulin-like growth factor (IGF)-1 and placental growth factor (PLGF) after ESWT on adipose tissue. Summarizing we could show that ESWT on adipose tissue enhanced the cellular ATP content and modified the expression of single mesenchymal and vascular marker, and thus potentially provides a more regenerative cell population. Because the effectiveness of autologous cell therapy is dependent on the therapeutic potency of the patient's cells, this technology might raise the number of patients eligible for autologous cell transplantation. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  6. Ultrasound-assisted liposuction provides a source for functional adipose-derived stromal cells.

    PubMed

    Duscher, Dominik; Maan, Zeshaan N; Luan, Anna; Aitzetmüller, Matthias M; Brett, Elizabeth A; Atashroo, David; Whittam, Alexander J; Hu, Michael S; Walmsley, Graham G; Houschyar, Khosrow S; Schilling, Arndt F; Machens, Hans-Guenther; Gurtner, Geoffrey C; Longaker, Michael T; Wan, Derrick C

    2017-12-01

    Regenerative medicine employs human mesenchymal stromal cells (MSCs) for their multi-lineage plasticity and their pro-regenerative cytokine secretome. Adipose-derived mesenchymal stromal cells (ASCs) are concentrated in fat tissue, and the ease of harvest via liposuction makes them a particularly interesting cell source. However, there are various liposuction methods, and few have been assessed regarding their impact on ASC functionality. Here we study the impact of the two most popular ultrasound-assisted liposuction (UAL) devices currently in clinical use, VASER (Solta Medical) and Lysonix 3000 (Mentor) on ASCs. After lipoaspirate harvest and processing, we sorted for ASCs using fluorescent-assisted cell sorting based on an established surface marker profile (CD34 + CD31 - CD45 - ). ASC yield, viability, osteogenic and adipogenic differentiation capacity and in vivo regenerative performance were assessed. Both UAL samples demonstrated equivalent ASC yield and viability. VASER UAL ASCs showed higher osteogenic and adipogenic marker expression, but a comparable differentiation capacity was observed. Soft tissue healing and neovascularization were significantly enhanced via both UAL-derived ASCs in vivo, and there was no significant difference between the cell therapy groups. Taken together, our data suggest that UAL allows safe and efficient harvesting of the mesenchymal stromal cellular fraction of adipose tissue and that cells harvested via this approach are suitable for cell therapy and tissue engineering applications. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  7. Enhanced mitogenesis in stromal vascular cells derived from subcutaneous adipose tissue of Wagyu compared with those of Angus cattle.

    PubMed

    Wei, S; Fu, X; Liang, X; Zhu, M J; Jiang, Z; Parish, S M; Dodson, M V; Zan, L; Du, M

    2015-03-01

    Japanese Wagyu cattle are well known for their extremely high marbling and lower subcutaneous adipose tissue compared with Angus cattle. However, mechanisms for differences in adipose deposition are unknown. The objective of this paper was to evaluate breed differences in the structure of subcutaneous adipose tissue, adipogenesis, and mitogenesis of stromal vascular (SV) cells between Wagyu and Angus cattle. Subcutaneous biopsy samples were obtained from 5 Wagyu (BW = 302 ± 9 kg) and 5 Angus (BW = 398 ± 12 kg) heifers at 12 mo of age, and samples were divided into 3 pieces for histological examination, biochemical analysis, and harvest of SV cells. Adipogenesis of SV cells was assessed by the expression of adipogenic markers and Oil Red-O staining, while mitogenesis was evaluated by an MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium dromide) test, phosphorylation of extracellular signal-regulated kinase (ERK) and protein kinase B (PKB; AKT). Based on histological analysis, Wagyu had larger adipocytes compared with Angus. At the tissue level, protein expression of peroxisome proliferator-activated receptor γ (PPARG) in Wagyu was much lower compared with that of Angus. Similarly, a lower mRNA expression of PPARG was found in Wagyu SV cells. No significant difference was observed for the zinc finger protein 423 (ZNF423) expression between Wagyu and Angus. As assessed by Oil Red-O staining, Wagyu SV cells possessed a notable trend of lower adipogenic capability. Interestingly, higher mitogenic ability was discovered in Wagyu SV cells, which was associated with an elevated phosphorylation of ERK1/2. There was no difference in AKT phosphorylation of SV cells between Wagyu and Angus. Moreover, exogenous fibroblast growth factor 2 (FGF2) enhanced mitogenesis and ERK1/2 phosphorylation of SV cells to a greater degree in Angus compared with that in Wagyu. Expression of transforming growth factor β 3 (TGFB3) and bone morphogenetic protein 2 (BMP2) in Wagyu SV

  8. Canine adipose-derived stromal cell viability following exposure to synovial fluid from osteoarthritic joints.

    PubMed

    Kiefer, Kristina M; O'Brien, Timothy D; Pluhar, Elizabeth G; Conzemius, Michael

    2015-01-01

    Stem cell therapy used in clinical application of osteoarthritis in veterinary medicine typically involves intra-articular injection of the cells, however the effect of an osteoarthritic environment on the fate of the cells has not been investigated. Assess the viability of adipose derived stromal cells following exposure to osteoarthritic joint fluid. Adipose derived stromal cells (ASCs) were derived from falciform adipose tissue of five adult dogs, and osteoarthritic synovial fluid (SF) was obtained from ten patients undergoing surgical intervention on orthopedic diseases with secondary osteoarthritis. Normal synovial fluid was obtained from seven adult dogs from an unrelated study. ASCs were exposed to the following treatment conditions: culture medium, normal SF, osteoarthritic SF, or serial dilutions of 1:1 to 1:10 of osteoarthritic SF with media. Cells were then harvested and assessed for viability using trypan blue dye exclusion. There was no significant difference in the viability of cells in culture medium or normal SF. Significant differences were found between cells exposed to any concentration of osteoarthritic SF and normal SF and between cells exposed to undiluted osteoarthritic SF and all serial dilutions. Subsequent dilutions reduced cytotoxicity. Osteoarthritic synovial fluid in this ex vivo experiment is cytotoxic to ASCs, when compared with normal synovial fluid. Current practice of direct injection of ASCs into osteoarthritic joints should be re-evaluated to determine if alternative means of administration may be more effective.

  9. Bone marrow-derived stromal cells are more beneficial cell sources for tooth regeneration compared with adipose-derived stromal cells.

    PubMed

    Ye, Lanfeng; Chen, Lin; Feng, Fan; Cui, Junhui; Li, Kaide; Li, Zhiyong; Liu, Lei

    2015-10-01

    Tooth loss is presently a global epidemic and tooth regeneration is thought to be a feasible and ideal treatment approach. Choice of cell source is a primary concern in tooth regeneration. In this study, the odontogenic differentiation potential of two non-dental-derived stem cells, adipose-derived stromal cells (ADSCs) and bone marrow-derived stromal cells (BMSCs), were evaluated both in vitro and in vivo. ADSCs and BMSCs were induced in vitro in the presence of tooth germ cell-conditioned medium (TGC-CM) prior to implantation into the omentum majus of rats, in combination with inactivated dentin matrix (IDM). Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the mRNA expression levels of odontogenic-related genes. Immunofluorescence and immunohistochemical assays were used to detect the protein levels of odontogenic-specific genes, such as DSP and DMP-1 both in vitro and in vivo. The results suggest that both ADSCs and BMSCs have odontogenic differentiation potential. However, the odontogenic potential of BMSCs was greater compared with ADSCs, showing that BMSCs are a more appropriate cell source for tooth regeneration. © 2015 International Federation for Cell Biology.

  10. Bone regeneration by implantation of adipose-derived stromal cells expressing BMP-2

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Li Huiwu; Health and Science Center, SIBS CAS and SSMU, 225 South Chongqing Road, Shanghai 200025; Dai Kerong

    2007-05-18

    In this study, we reported that the adipose-derived stromal cells (ADSCs) genetically modified by bone morphogenetic protein 2 (BMP-2) healed critical-sized canine ulnar bone defects. First, the osteogenic and adipogenic differentiation potential of the ADSCs derived from canine adipose tissue were demonstrated. And then the cells were modified by the BMP-2 gene and the expression and bone-induction ability of BMP-2 were identified. Finally, the cells modified by BMP-2 gene were applied to a {beta}-tricalcium phosphate (TCP) carrier and implanted into ulnar bone defects in the canine model. After 16 weeks, radiographic, histological, and histomorphometry analysis showed that ADSCs modified bymore » BMP-2 gene produced a significant increase of newly formed bone area and healed or partly healed all of the bone defects. We conclude that ADSCs modified by the BMP-2 gene can enhance the repair of critical-sized bone defects in large animals.« less

  11. Enrichment of Adipose-Derived Stromal Cells for BMPR1A Facilitates Enhanced Adipogenesis

    PubMed Central

    Zielins, Elizabeth R.; Paik, Kevin; Ransom, Ryan C.; Brett, Elizabeth A.; Blackshear, Charles P.; Luan, Anna; Walmsley, Graham G.; Atashroo, David A.; Senarath-Yapa, Kshemendra; Momeni, Arash; Rennert, Robert; Sorkin, Michael; Seo, Eun Young; Chan, Charles K.; Gurtner, Geoffrey C.; Longaker, Michael T.

    2016-01-01

    Background: Reconstruction of soft tissue defects has traditionally relied on the use of grafts and flaps, which may be associated with variable resorption and/or significant donor site morbidity. Cell-based strategies employing adipose-derived stromal cells (ASCs), found within the stromal vascular fraction (SVF) of adipose tissue, may offer an alternative strategy for soft tissue reconstruction. In this study, we investigated the potential of a bone morphogenetic protein receptor type 1A (BMPR1A)(+) subpopulation of ASCs to enhance de novo adipogenesis. Methods: Human lipoaspirate was enzymatically digested to isolate SVF and magnetic-activated cell separation was utilized to obtain BMPR1A(+) and BMPR1A(−) cells. These cells, along with unenriched cells, were expanded in culture and evaluated for adipogenic gene expression and in vitro adipocyte formation. Cells from each group were also labeled with a green fluorescent protein (GFP) lentivirus and transplanted into the inguinal fat pads, an adipogenic niche, of immunocompromised mice to determine their potential for de novo adipogenesis. Confocal microscopy along with staining of lipid droplets and vasculature was performed to evaluate the formation of mature adipocytes by transplanted cells. Results: In comparison to BMPR1A(−) and unenriched ASCs, BMPR1A(+) cells demonstrated significantly enhanced adipogenesis when cultured in an adipogenic differentiation medium, as evidenced by increased staining with Oil Red O and increased expression of peroxisome proliferator-activating receptor gamma (PPAR-γ) and fatty acid-binding protein 4 (FABP4). BMPR1A(+) cells also formed significantly more adipocytes in vivo, as demonstrated by quantification of GFP+ adipocytes. Minimal formation of mature adipocytes was appreciated by BMPR1A(−) cells. Conclusions: BMPR1A(+) ASCs show an enhanced ability for adipogenesis in vitro, as shown by gene expression and histological staining. Furthermore, within an adipogenic niche

  12. Enrichment of Adipose-Derived Stromal Cells for BMPR1A Facilitates Enhanced Adipogenesis.

    PubMed

    Zielins, Elizabeth R; Paik, Kevin; Ransom, Ryan C; Brett, Elizabeth A; Blackshear, Charles P; Luan, Anna; Walmsley, Graham G; Atashroo, David A; Senarath-Yapa, Kshemendra; Momeni, Arash; Rennert, Robert; Sorkin, Michael; Seo, Eun Young; Chan, Charles K; Gurtner, Geoffrey C; Longaker, Michael T; Wan, Derrick C

    2016-02-01

    Reconstruction of soft tissue defects has traditionally relied on the use of grafts and flaps, which may be associated with variable resorption and/or significant donor site morbidity. Cell-based strategies employing adipose-derived stromal cells (ASCs), found within the stromal vascular fraction (SVF) of adipose tissue, may offer an alternative strategy for soft tissue reconstruction. In this study, we investigated the potential of a bone morphogenetic protein receptor type 1A (BMPR1A)(+) subpopulation of ASCs to enhance de novo adipogenesis. Human lipoaspirate was enzymatically digested to isolate SVF and magnetic-activated cell separation was utilized to obtain BMPR1A(+) and BMPR1A(-) cells. These cells, along with unenriched cells, were expanded in culture and evaluated for adipogenic gene expression and in vitro adipocyte formation. Cells from each group were also labeled with a green fluorescent protein (GFP) lentivirus and transplanted into the inguinal fat pads, an adipogenic niche, of immunocompromised mice to determine their potential for de novo adipogenesis. Confocal microscopy along with staining of lipid droplets and vasculature was performed to evaluate the formation of mature adipocytes by transplanted cells. In comparison to BMPR1A(-) and unenriched ASCs, BMPR1A(+) cells demonstrated significantly enhanced adipogenesis when cultured in an adipogenic differentiation medium, as evidenced by increased staining with Oil Red O and increased expression of peroxisome proliferator-activating receptor gamma (PPAR-γ) and fatty acid-binding protein 4 (FABP4). BMPR1A(+) cells also formed significantly more adipocytes in vivo, as demonstrated by quantification of GFP+ adipocytes. Minimal formation of mature adipocytes was appreciated by BMPR1A(-) cells. BMPR1A(+) ASCs show an enhanced ability for adipogenesis in vitro, as shown by gene expression and histological staining. Furthermore, within an adipogenic niche, BMPR1A(+) cells possessed an increased capacity

  13. Case Report: Industrial X-Ray Injury Treated With Non-Cultured Autologous Adipose-Derived Stromal Vascular Fraction (SVF).

    PubMed

    Iddins, C J; Cohen, S R; Goans, R E; Wanat, R; Jenkins, M; Christensen, D M; Dainiak, N

    2016-08-01

    Local cutaneous injuries induced by ionizing radiation (IR) are difficult to treat. Many have reported local injection of adipose-derived stromal vascular fraction (SVF), often with additional therapies, as an effective treatment of IR-induced injury even after other local therapies have failed. The authors report a case of a locally recurrent, IR-induced wound that was treated with autologous, non-cultured SVF without other concurrent therapy. A nondestructive testing technician was exposed to 130 kVp x rays to his non-dominant right thumb on 5 October 2011. The wound healed 4 mo after initial conservative therapy with oral/topical α-tocopherol, oral pentoxifylline, naproxen sodium, low-dose oral steroids, topical steroids, hyperbaric oxygen therapy (HBOT), oral antihistamines, and topical aloe vera. Remission lasted approximately 17 mo with one minor relapse in July 2012 after minimal trauma and subsequent healing. Aggressive wound breakdown during June 2013 required additional therapy with HBOT. An erythematous, annular papule developed over the following 12 mo (during which time the patient was not undergoing prescribed treatment). Electron paramagnetic resonance (EPR) done more than 2 mo after exposure to IR revealed dose estimates of 14 ± 3 Gy and 19 ± 6 Gy from two centers using different EPR techniques. The patient underwent debridement of the 0.5 cm papular area, followed by SVF injection into and around the wound bed and throughout the thumb without complication. Eleven months post SVF injection, the patient has been essentially asymptomatic with an intact integument. These results raise the possibility of prolonged benefit from SVF therapy without the use of cytokines. Since there is currently no consensus on the use of isolated SVF therapy in chronic, local IR-induced injury, assessment of this approach in an appropriately powered, controlled trial in experimental animals with local radiation injury appears to be indicated.

  14. [Adipose-derived stromal cells (ASC) - basics and therapeutic approaches in otorhinolaryngology].

    PubMed

    Frölich, K; Hagen, R; Kleinsasser, N

    2014-06-01

    Adipose-derived Stromal Cells (ASC) - Basics and Therapeutic Approaches in Otorhinolaryngology Mesenchymal stem cells from adipose tissue can be easily harvested with less discomfort, low donor-site morbidity and high amount compared to bone marrow-derived stem cells. Due to their multilineage differentiation potential in various cell types, immunmodulatory properties and their capability to enhance wound healing, ASC are a promising cell source for tissue engineering approaches and regenerative medicine. They are characterized by the expression of specific surface marker proteins and their differentiation potential into the mesenchymal lineages. Whereas only preclinical studies are published for otorhinolaryngology-related therapeutic options using ASC, various diseases, for instance graft-versus-host disease, have already been treated with ASC in single cases or clinical trials. Safety and genomic stability of ASC as well as the risk of spontaneous malignant transformation are still disputed. This review summarizes the current literature on characterization and anatomic localization of ASC. In addition, beside the presentation of preclinical studies concerning therapeutic approaches in otorhinolaryngology as well as of current clinical applications, the issue of safety of ASC in human stem cell therapy is discussed. © Georg Thieme Verlag KG Stuttgart · New York.

  15. Generation of a Bone Organ by Human Adipose-Derived Stromal Cells Through Endochondral Ossification.

    PubMed

    Osinga, Rik; Di Maggio, Nunzia; Todorov, Atanas; Allafi, Nima; Barbero, Andrea; Laurent, Frédéric; Schaefer, Dirk Johannes; Martin, Ivan; Scherberich, Arnaud

    2016-08-01

    : Recapitulation of endochondral ossification (ECO) (i.e., generation of marrow-containing ossicles through a cartilage intermediate) has relevance to develop human organotypic models for bone or hematopoietic cells and to engineer grafts for bone regeneration. Unlike bone marrow-derived stromal cells (also known as bone marrow-derived mesenchymal stromal/stem cells), adipose-derived stromal cells (ASC) have so far failed to form a bone organ by ECO. The goal of the present study was to assess whether priming human ASC to a defined stage of chondrogenesis in vitro allows their autonomous ECO upon ectopic implantation. ASC were cultured either as micromass pellets or into collagen sponges in chondrogenic medium containing transforming growth factor-β3 and bone morphogenetic protein-6 for 4 weeks (early hypertrophic templates) or for two additional weeks in medium supplemented with β-glycerophosphate, l-thyroxin, and interleukin1-β to induce hypertrophic maturation (late hypertrophic templates). Constructs were implanted in vivo and analyzed after 8 weeks. In vitro, ASC deposited cartilaginous matrix positive for glycosaminoglycans, type II collagen, and Indian hedgehog. Hypertrophic maturation induced upregulation of type X collagen, bone sialoprotein, and matrix metalloproteinase13 (MMP13). In vivo, both early and late hypertrophic templates underwent cartilage remodeling, as assessed by MMP13- and tartrate-resistant acid phosphatase-positive staining, and developed bone ossicles, including bone marrow elements, although to variable degrees of efficiency. In situ hybridization for human-specific sequences and staining with a human specific anti-CD146 antibody demonstrated the direct contribution of ASC to bone and stromal tissue formation. In conclusion, despite their debated skeletal progenitor nature, human ASC can generate bone organs through ECO when suitably primed in vitro. Recapitulation of endochondral ossification (ECO) (i.e., generation of marrow

  16. Prevention of skin flap necrosis by use of adipose-derived stromal cells with light-emitting diode phototherapy.

    PubMed

    Park, In-Su; Mondal, Arindam; Chung, Phil-Sang; Ahn, Jin Chul

    2015-03-01

    The aim of this study was to investigate the effects of low-level light therapy (LLLT) on transplanted human adipose-derived mesenchymal stromal cells (ASCs) in the skin flap of mice. LLLT, ASC transplantation and ASC transplantation with LLLT (ASC + LLLT) were applied to the skin flap. Immunostaining and Western blot analysis were performed to evaluate cell survival and differentiation and secretion of vascular endothelial growth factor and basic fibroblast growth factor by the ASCs. Vascular regeneration was assessed by means of immunostaining in addition to hematoxylin and eosin staining. In the ASC + LLLT group, the survival of ASCs was increased as the result of the decreased apoptosis of ASCs. The secretion of growth factors was higher in this group as compared with ASCs alone. ASCs contributed to tissue regeneration through vascular cell differentiation and secretion of angiogenic growth factors. The ASC + LLLT group displayed improved treatment efficacy including neovascularization and tissue regeneration compared with ASCs alone. Transplanting ASCs to ischemic skin flaps improved therapeutic efficacy for ischemia treatment as the result of enhanced cell survival and paracrine effects. These data suggest that LLLT is an effective biostimulator of ASCs in vascular regeneration, which enhances the survival of ASCs and stimulates the secretion of growth factors in skin flaps. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  17. Human adipose-derived stem cells promote vascularization of collagen-based scaffolds transplanted into nude mice

    PubMed Central

    Cherubino, Mario; Valdatta, Luigi; Balzaretti, Riccardo; Pellegatta, Igor; Rossi, Federica; Protasoni, Marina; Tedeschi, Alessandra; Accolla, Roberto S; Bernardini, Giovanni; Gornati, Rosalba

    2016-01-01

    Aim: After in vivo implantation of cell-loaded devices, only the cells close to the capillaries can obtain nutrients to maintain their functions. It is known that factors secreted by stem cells, rather than stem cells themselves, are fundamental to guarantee new vascularization in the area of implant. Materials & methods: To investigate this possibility, we have grafted mice with Bilayer and Flowable Integra® scaffolds, loaded or not with human adipose-derived stem cells. Results: Our results support the therapeutic potential of human adipose-derived stem cells to induce new vascular networks of engineered organs and tissues. Conclusion: This finding suggests that our approach can help to form new vascular networks that allow sufficient vascularization of engineered organs and tissues in cases of difficult wound healing due to ischemic conditions. PMID:26965659

  18. Extracellular matrix and α5β1 integrin signaling control the maintenance of bone formation capacity by human adipose-derived stromal cells

    PubMed Central

    Di Maggio, Nunzia; Martella, Elisa; Frismantiene, Agne; Resink, Therese J.; Schreiner, Simone; Lucarelli, Enrico; Jaquiery, Claude; Schaefer, Dirk J.; Martin, Ivan; Scherberich, Arnaud

    2017-01-01

    Stromal vascular fraction (SVF) cells of human adipose tissue have the capacity to generate osteogenic grafts with intrinsic vasculogenic properties. However, adipose-derived stromal/stem cells (ASC), even after minimal monolayer expansion, display poor osteogenic capacity in vivo. We investigated whether ASC bone-forming capacity may be maintained by culture within a self-produced extracellular matrix (ECM) that recapitulates the native environment. SVF cells expanded without passaging up to 28 days (Unpass-ASC) deposited a fibronectin-rich extracellular matrix and displayed greater clonogenicity and differentiation potential in vitro compared to ASC expanded only for 6 days (P0-ASC) or for 28 days with regular passaging (Pass-ASC). When implanted subcutaneously, Unpass-ASC produced bone tissue similarly to SVF cells, in contrast to P0- and Pass-ASC, which mainly formed fibrous tissue. Interestingly, clonogenic progenitors from native SVF and Unpass-ASC expressed low levels of the fibronectin receptor α5 integrin (CD49e), which was instead upregulated in P0- and Pass-ASC. Mechanistically, induced activation of α5β1 integrin in Unpass-ASC led to a significant loss of bone formation in vivo. This study shows that ECM and regulation of α5β1-integrin signaling preserve ASC progenitor properties, including bone tissue-forming capacity, during in vitro expansion. PMID:28290502

  19. Adipose Tissue-Derived Pericytes for Cartilage Tissue Engineering.

    PubMed

    Zhang, Jinxin; Du, Chunyan; Guo, Weimin; Li, Pan; Liu, Shuyun; Yuan, Zhiguo; Yang, Jianhua; Sun, Xun; Yin, Heyong; Guo, Quanyi; Zhou, Chenfu

    2017-01-01

    Mesenchymal stem cells (MSCs) represent a promising alternative source for cartilage tissue engineering. However, MSC culture is labor-intensive, so these cells cannot be applied immediately to regenerate cartilage for clinical purposes. Risks during the ex vivo expansion of MSCs, such as infection and immunogenicity, can be a bottleneck in their use in clinical tissue engineering. As a novel stem cell source, pericytes are generally considered to be the origin of MSCs. Pericytes do not have to undergo time-consuming ex vivo expansion because they are uncultured cells. Adipose tissue is another optimal stem cell reservoir. Because adipose tissue is well vascularized, a considerable number of pericytes are located around blood vessels in this accessible and dispensable tissue, and autologous pericytes can be applied immediately for cartilage regeneration. Thus, we suggest that adipose tissue-derived pericytes are promising seed cells for cartilage regeneration. Many studies have been performed to develop isolation methods for the adipose tissuederived stromal vascular fraction (AT-SVF) using lipoaspiration and sorting pericytes from AT-SVF. These methods are useful for sorting a large number of viable pericytes for clinical therapy after being combined with automatic isolation using an SVF device and automatic magnetic-activated cell sorting. These tools should help to develop one-step surgery for repairing cartilage damage. However, the use of adipose tissue-derived pericytes as a cell source for cartilage tissue engineering has not drawn sufficient attention and preclinical studies are needed to improve cell purity, to increase sorting efficiency, and to assess safety issues of clinical applications. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  20. Characterization of a Murine Pressure Ulcer Model to Assess Efficacy of Adipose-derived Stromal Cells

    PubMed Central

    Strong, Amy L.; Bowles, Annie C.; MacCrimmon, Connor P.; Lee, Stephen J.; Frazier, Trivia P.; Katz, Adam J.; Gawronska-Kozak, Barbara; Bunnell, Bruce A.

    2015-01-01

    Background: As the world’s population lives longer, the number of individuals at risk for pressure ulcers will increase considerably in the coming decades. In developed countries, up to 18% of nursing home residents suffer from pressure ulcers and the resulting hospital costs can account for up to 4% of a nation’s health care budget. Although full-thickness surgical skin wounds have been used as a model, preclinical rodent studies have demonstrated that repeated cycles of ischemia and reperfusion created by exposure to magnets most closely mimic the human pressure ulcer condition. Methods: This study uses in vivo and in vitro quantitative parameters to characterize the temporal kinetics and histology of pressure ulcers in young, female C57BL/6 mice exposed to 2 or 3 ischemia-reperfusion cycles. This pressure ulcer model was validated further in studies examining the efficacy of adipose-derived stromal/stem cell administration. Results: Optimal results were obtained with the 2-cycle model based on the wound size, histology, and gene expression profile of representative angiogenic and reparative messenger RNAs. When treated with adipose-derived stromal/stem cells, pressure ulcer wounds displayed a dose-dependent and significant acceleration in wound closure rates and improved tissue histology. Conclusion: These findings document the utility of this simplified preclinical model for the evaluation of novel tissue engineering and medical approaches to treat pressure ulcers in humans. PMID:25878945

  1. Immunomodulatory Effects of Adipose Stromal Vascular Fraction Cells Promote Alternative Activation Macrophages to Repair Tissue Damage.

    PubMed

    Bowles, Annie C; Wise, Rachel M; Gerstein, Brittany Y; Thomas, Robert C; Ogelman, Roberto; Febbo, Isabella; Bunnell, Bruce A

    2017-10-01

    The pathogenesis of many diseases is driven by the interactions between helper T (T H ) cells and macrophages. The phenotypes of these cells are functional dichotomies that are persuaded according to the surrounding milieu. In both multiple sclerosis and the experimental autoimmune encephalomyelitis (EAE) model, T H 1 and T H 17 cells propagate autoimmune signaling and inflammation in the peripheral lymphoid tissues. In turn, this proinflammatory repertoire promotes the classical activation, formerly the M1-type, macrophages. Together, these cells infiltrate into the central nervous system (CNS) tissues and generate inflammatory and demyelinating lesions. Our most recent report demonstrated the immunomodulatory and anti-inflammatory effects of adipose stromal vascular fraction (SVF) cells and adipose-derived stem cells (ASCs) that led to functional, immunological, and pathological improvements in the EAE model. Here, a deeper investigation revealed the induction of regulatory T cells and alternative activation, or M2-type, macrophages in the periphery followed by the presence of alternative activation macrophages, reduced cellular infiltrates, and attenuation of neuroinflammation in CNS tissues following intraperitoneal administration of these treatments. Spleens from treated EAE mice revealed diminished T H 1 and T H 17 cell activities and were markedly higher in the levels of anti-inflammatory cytokine interleukin-10. Interestingly, SVF cells were more effective than ASCs at mediating these beneficial changes, which were attributed to their localization to the spleens after administration. Together, SVF cells rapidly and robustly attenuated the propagation of autoimmune signaling in the periphery that provided a permissive milieu in the CNS for repair and possibly regeneration. Stem Cells 2017;35:2198-2207. © 2017 AlphaMed Press.

  2. Isolation of Human Adipose-Derived Stromal Cells Using Laser-Assisted Liposuction and Their Therapeutic Potential in Regenerative Medicine

    PubMed Central

    Chung, Michael T.; Zimmermann, Andrew S.; Paik, Kevin J.; Morrison, Shane D.; Hyun, Jeong S.; Lo, David D.; McArdle, Adrian; Montoro, Daniel T.; Walmsley, Graham G.; Senarath-Yapa, Kshemendra; Sorkin, Michael; Rennert, Robert; Chen, Hsin-Han; Chung, Andrew S.; Vistnes, Dean; Gurtner, Geoffrey C.; Longaker, Michael T.

    2013-01-01

    Harvesting adipose-derived stromal cells (ASCs) for tissue engineering is frequently done through liposuction. However, several different techniques exist. Although third-generation ultrasound-assisted liposuction has been shown to not have a negative effect on ASCs, the impact of laser-assisted liposuction on the quality and differentiation potential of ASCs has not been studied. Therefore, ASCs were harvested from laser-assisted lipoaspirate and suction-assisted lipoaspirate. Next, in vitro parameters of cell yield, cell viability and proliferation, surface marker phenotype, osteogenic differentiation, and adipogenic differentiation were performed. Finally, in vivo bone formation was assessed using a critical-sized cranial defect in athymic nude mice. Although ASCs isolated from suction-assisted lipoaspirate and laser-assisted lipoaspirate both successfully underwent osteogenic and adipogenic differentiation, the cell yield, viability, proliferation, and frequency of ASCs (CD34+CD31−CD45−) in the stromal vascular fraction were all significantly less with laser-assisted liposuction in vitro (p < .05). In vivo, quantification of osseous healing by micro-computed tomography revealed significantly more healing with ASCs isolated from suction-assisted lipoaspirate relative to laser-assisted lipoaspirate at the 4-, 6-, and 8-week time points (p < .05). Therefore, as laser-assisted liposuction appears to negatively impact the biology of ASCs, cell harvest using suction-assisted liposuction is preferable for tissue-engineering purposes. PMID:24018794

  3. Isolation of human adipose-derived stromal cells using laser-assisted liposuction and their therapeutic potential in regenerative medicine.

    PubMed

    Chung, Michael T; Zimmermann, Andrew S; Paik, Kevin J; Morrison, Shane D; Hyun, Jeong S; Lo, David D; McArdle, Adrian; Montoro, Daniel T; Walmsley, Graham G; Senarath-Yapa, Kshemendra; Sorkin, Michael; Rennert, Robert; Chen, Hsin-Han; Chung, Andrew S; Vistnes, Dean; Gurtner, Geoffrey C; Longaker, Michael T; Wan, Derrick C

    2013-10-01

    Harvesting adipose-derived stromal cells (ASCs) for tissue engineering is frequently done through liposuction. However, several different techniques exist. Although third-generation ultrasound-assisted liposuction has been shown to not have a negative effect on ASCs, the impact of laser-assisted liposuction on the quality and differentiation potential of ASCs has not been studied. Therefore, ASCs were harvested from laser-assisted lipoaspirate and suction-assisted lipoaspirate. Next, in vitro parameters of cell yield, cell viability and proliferation, surface marker phenotype, osteogenic differentiation, and adipogenic differentiation were performed. Finally, in vivo bone formation was assessed using a critical-sized cranial defect in athymic nude mice. Although ASCs isolated from suction-assisted lipoaspirate and laser-assisted lipoaspirate both successfully underwent osteogenic and adipogenic differentiation, the cell yield, viability, proliferation, and frequency of ASCs (CD34(+)CD31(-)CD45(-)) in the stromal vascular fraction were all significantly less with laser-assisted liposuction in vitro (p < .05). In vivo, quantification of osseous healing by micro-computed tomography revealed significantly more healing with ASCs isolated from suction-assisted lipoaspirate relative to laser-assisted lipoaspirate at the 4-, 6-, and 8-week time points (p < .05). Therefore, as laser-assisted liposuction appears to negatively impact the biology of ASCs, cell harvest using suction-assisted liposuction is preferable for tissue-engineering purposes.

  4. Adipose Extracellular Matrix/Stromal Vascular Fraction Gel Secretes Angiogenic Factors and Enhances Skin Wound Healing in a Murine Model.

    PubMed

    Sun, Mingliang; He, Yunfan; Zhou, Tao; Zhang, Pan; Gao, Jianhua; Lu, Feng

    2017-01-01

    Mesenchymal stem cells are an attractive cell type for cytotherapy in wound healing. The authors recently developed a novel, adipose-tissue-derived, injectable extracellular matrix/stromal vascular fraction gel (ECM/SVF-gel) for stem cell therapy. This study was designed to assess the therapeutic effects of ECM/SVF-gel on wound healing and potential mechanisms. ECM/SVF-gel was prepared for use in nude mouse excisional wound healing model. An SVF cell suspension and phosphate-buffered saline injection served as the control. The expression levels of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and monocyte chemotactic protein-1 (MCP-1) in ECM/SVF-gel were analyzed at different time points. Angiogenesis (tube formation) assays of ECM/SVF-gel extracts were evaluated, and vessels density in skin was determined. The ECM/SVF-gel extract promoted tube formation in vitro and increased the expression of the angiogenic factors VEGF and bFGF compared with those in the control. The expression of the inflammatory chemoattractant MCP-1 was high in ECM/SVF-gel at the early stage and decreased sharply during the late stage of wound healing. The potent angiogenic effects exerted by ECM/SVF-gel may contribute to the improvement of wound healing, and these effects could be related to the enhanced inflammatory response in ECM/SVF-gel during the early stage of wound healing.

  5. The Celution® System: Automated Processing of Adipose-Derived Regenerative Cells in a Functionally Closed System.

    PubMed

    Fraser, John K; Hicok, Kevin C; Shanahan, Rob; Zhu, Min; Miller, Scott; Arm, Douglas M

    2014-01-01

    Objective: To develop a closed, automated system that standardizes the processing of human adipose tissue to obtain and concentrate regenerative cells suitable for clinical treatment of thermal and radioactive burn wounds. Approach: A medical device was designed to automate processing of adipose tissue to obtain a clinical-grade cell output of stromal vascular cells that may be used immediately as a therapy for a number of conditions, including nonhealing wounds resulting from radiation damage. Results: The Celution ® System reliably and reproducibly generated adipose-derived regenerative cells (ADRCs) from tissue collected manually and from three commercial power-assisted liposuction devices. The entire process of introducing tissue into the system, tissue washing and proteolytic digestion, isolation and concentration of the nonadipocyte nucleated cell fraction, and return to the patient as a wound therapeutic, can be achieved in approximately 1.5 h. An alternative approach that applies ultrasound energy in place of enzymatic digestion demonstrates extremely poor efficiency cell extraction. Innovation: The Celution System is the first medical device validated and approved by multiple international regulatory authorities to generate autologous stromal vascular cells from adipose tissue that can be used in a real-time bedside manner. Conclusion: Initial preclinical and clinical studies using ADRCs obtained using the automated tissue processing Celution device described herein validate a safe and effective manner to obtain a promising novel cell-based treatment for wound healing.

  6. Expanded Stem Cells, Stromal-Vascular Fraction, and Platelet-Rich Plasma Enriched Fat: Comparing Results of Different Facial Rejuvenation Approaches in a Clinical Trial

    PubMed Central

    Rigotti, Gino; Charles-de-Sá, Luiz; Gontijo-de-Amorim, Natale Ferreira; Takiya, Christina Maeda; Amable, Paola Romina; Borojevic, Radovan; Benati, Donatella; Bernardi, Paolo; Sbarbati, Andrea

    2016-01-01

    Background In a previous study, the authors demonstrated that treatment with expanded adipose-derived stem cells or stromal vascular fraction (SVF)-enriched fat modify the pattern of the dermis in human beings, representing a skin rejuvenation effect. Considering that expanded stem cells require a cell factor, the authors wanted to assess similar results by replacing them with platelet-rich plasma (PRP), which is easier to obtain and for which an empirical regenerative effect has been already described. Objectives To determine if PRP injection could replace the cutaneous regenerative effect of adipose-derived stem cells. Methods This study was performed in 13 patients who were candidates for facelift. The patients underwent sampling of fat by liposuction from the abdomen and submitted to one of three protocols: injection of SVF-enriched fat or expanded adipose-derived stem cells or fat plus PRP in the preauricular areas. Fragments of skin were removed before and 3 months after treatment and analyzed by optical and electron microscopy. Results The use of fat plus PRP led to the presence of more pronounced inflammatory infiltrates and a greater vascular reactivity, increasing in vascular permeability and a certain reactivity of the nervous component. The addition of PRP did not improve the regenerative effect. Conclusion The use of PRP did not have significant advantages in skin rejuvenation over the use of expanded adipose-derived stem cells or SVF-enriched fat. The effect of increased vascular reactivity may be useful in pathological situations in which an intense angiogenesis is desirable, such as tissular ischemia. Level of Evidence: 4 Therapeutic PMID:26879294

  7. Transdifferentiation of mouse adipose-derived stromal cells into acinar cells of the submandibular gland using a co-culture system

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lee, Jingu; Park, Sangkyu; Roh, Sangho, E-mail: sangho@snu.ac.kr

    A loss of salivary gland function often occurs after radiation therapy in head and neck tumors, though secretion of saliva by the salivary glands is essential for the health and maintenance of the oral environment. Transplantation of salivary acinar cells (ACs), in part, may overcome the side effects of therapy. Here we directly differentiated mouse adipose-derived stromal cells (ADSCs) into ACs using a co-culture system. Multipotent ADSCs can be easily collected from stromal vascular fractions of adipose tissues. The isolated ADSCs showed positive expression of markers such as integrin beta-1 (CD29), cell surface glycoprotein (CD44), endoglin (CD105), and Nanog. Themore » cells were able to differentiate into adipocytes, osteoblasts, and neural-like cells after 14 days in culture. ADSCs at passage 2 were co-cultured with mouse ACs in AC culture medium using the double-chamber (co-culture system) to avoid mixing the cell types. The ADSCs in this co-culture system expressed markers of ACs, such as α-amylases and aquaporin5, in both mRNA and protein. ADSCs cultured in AC-conditioned medium also expressed AC markers. Cellular proliferation and senescence analyses demonstrated that cells in the co-culture group showed lower senescence and a higher proliferation rate than the AC-conditioned medium group at Days 14 and 21. The results above imply direct conversion of ADSCs into ACs under the co-culture system; therefore, ADSCs may be a stem cell source for the therapy for salivary gland damage. - Highlights: • ADSCs could transdifferentiate into acinar cells (ACs) using ACs co-culture (CCA). • Transdifferentiated ADSCs expressed ACs markers such as α-amylase and aquaporin5. • High proliferation and low senescence were presented in CCA at Day 14. • Transdifferentiation of ADSCs into ACs using CCA may be an appropriate method for cell-based therapy.« less

  8. Gold nanoparticles cellular toxicity and recovery: adipose Derived Stromal cells.

    PubMed

    Mironava, Tatsiana; Hadjiargyrou, Michael; Simon, Marcia; Rafailovich, Miriam H

    2014-03-01

    Gold nanoparticles (AuNPs) are currently used in numerous medical applications. Herein, we describe their in vitro impact on human adipose-derived stromal cells (ADSCs) using 13 nm and 45 nm citrate-coated AuNPs. In their non-differentiated state, ADSCs were penetrated by the AuNPs and stored in vacuoles. The presence of the AuNPs in ADSCs resulted in increased population doubling times, decreased cell motility and cell-mediated collagen contraction. The degree to which the cells were impacted was a function of particle concentration, where the smaller particles required a sevenfold higher concentration to have the same effect as the larger ones. Furthermore, AuNPs reduced adipogenesis as measured by lipid droplet accumulation and adiponectin secretion. These effects correlated with transient increases in DLK1 and with relative reductions in fibronectin. Upon removal of exogenous AuNPs, cellular NP levels decreased and normal ADSC functions were restored. As adiponectin helps regulate energy metabolism, local fluctuations triggered by AuNPs can lead to systemic changes. Hence, careful choice of size, concentration and clinical application duration of AuNPs is warranted.

  9. Therapeutic Potential of Human Adipose-Derived Stem/Stromal Cell Microspheroids Prepared by Three-Dimensional Culture in Non-Cross-Linked Hyaluronic Acid Gel.

    PubMed

    Mineda, Kazuhide; Feng, Jingwei; Ishimine, Hisako; Takada, Hitomi; Doi, Kentaro; Kuno, Shinichiro; Kinoshita, Kahori; Kanayama, Koji; Kato, Harunosuke; Mashiko, Takanobu; Hashimoto, Ichiro; Nakanishi, Hideki; Kurisaki, Akira; Yoshimura, Kotaro

    2015-12-01

    Three-dimensional culture of mesenchymal stem/stromal cells for spheroid formation is known to enhance their therapeutic potential for regenerative medicine. Spheroids were prepared by culturing human adipose-derived stem/stromal cells (hASCs) in a non-cross-linked hyaluronic acid (HA) gel and compared with dissociated hASCs and hASC spheroids prepared using a nonadherent dish. Preliminary experiments indicated that a 4% HA gel was the most appropriate for forming hASC spheroids with a relatively consistent size (20-50 µm) within 48 hours. Prepared spheroids were positive for pluripotency markers (NANOG, OCT3/4, and SOX-2), and 40% of the cells were SSEA-3-positive, a marker of the multilineage differentiating stress enduring or Muse cell. In contrast with dissociated ASCs, increased secretion of cytokines such as hepatocyte growth factor was detected in ASC spheroids cultured under hypoxia. On microarray ASC spheroids showed upregulation of some pluripotency markers and downregulation of genes related to the mitotic cell cycle. After ischemia-reperfusion injury to the fat pad in SCID mice, local injection of hASC spheroids promoted tissue repair and reduced the final atrophy (1.6%) compared with that of dissociated hASCs (14.3%) or phosphate-buffered saline (20.3%). Part of the administered hASCs differentiated into vascular endothelial cells. ASC spheroids prepared in a HA gel contain undifferentiated cells with therapeutic potential to promote angiogenesis and tissue regeneration after damage. This study shows the therapeutic value of human adipose-derived stem cell spheroids prepared in hyarulonic acid gel. The spheroids have various benefits as an injectable cellular product and show therapeutic potential to the stem cell-depleted conditions such as diabetic chronic skin ulcer. ©AlphaMed Press.

  10. Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Takahara, Kiyoshi; Ii, Masaaki, E-mail: masaii@art.osaka-med.ac.jp; Inamoto, Teruo

    2014-04-18

    Highlights: • AdSC transplantation exhibits inhibitory effect on tumor progressions of PCa cells. • AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway. • High expression of the TGF-β1 gene in AdSCs. - Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferationmore » of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa.« less

  11. Effect of TGF-β1 Stimulation on the Secretome of Human Adipose-Derived Mesenchymal Stromal Cells.

    PubMed

    Rodríguez, Tania M; Saldías, Alejandro; Irigo, Marcelo; Zamora, Jorge Velasco; Perone, Marcelo J; Dewey, Ricardo A

    2015-08-01

    Adipose tissue is an attractive source of mesenchymal stromal cells (MSCs) owing to the relative ease of obtaining large volumes with more MSC abundance compared with other sources. Increasing evidence supports the fact that trophic factors secreted by MSCs play a pivotal therapeutic role. Several strategies in regenerative medicine use MSCs, mainly exploiting their immunosuppressive effect and homing capacity to sites of damage. Transforming growth factor-β1 (TGF-β1) is a pleiotropic cytokine that, depending on the cell niche, can display either anti-inflammatory or proinflammatory effects. TGF-β1 expression increases in various tissues with damage, especially when accompanied by inflammation. Thus, we analyzed the effect of TGF-β1 on the secretion by adipose-derived mesenchymal stromal cells (ASCs) of a panel of 80 cytokines/chemokines using an antibody array. To avoid a possible effect of fetal bovine serum (FBS) on ASCs secretion, we performed our analysis by culturing cells in FBS-free conditions, only supplemented with 0.1% of bovine serum albumin. We report the cytokine profile secreted by ASCs. We also found that TGF-β1 exposure modulates 8 chemokines and 18 cytokines, including TGF-β1 and -β2, and other important cytokines involved in immunosuppression, allergic responses, and bone resorption. ©AlphaMed Press.

  12. Nonexpanded Adipose Stromal Vascular Fraction Local Therapy on Peripheral Nerve Regeneration Using Allografts.

    PubMed

    Mohammadi, Rahim; Mehrtash, Moein; Mehrtash, Moeid; Sajjadi, Seyedeh-Sepideh

    2016-06-01

    Adipose tissue possesses a population of multi-potent stem cells which can be differentiated to a Schwann cell phenotype and may be of benefit for treatment of peripheral nerve injuries. Effects of local therapy of nonexpanded adipose stromal vascular fraction (SVF) on peripheral nerve regeneration was studied using allografts in a rat sciatic nerve model. Thirty male white Wistar rats were divided into three experimental groups (n = 10), randomly: Sham-operated group (SHAM), allograft group (ALLO), SVF-treated group (ALLO/SVF). In SHAM group left sciatic nerve was exposed through a gluteal muscle incision and after homeostasis muscle was sutured. In the ALLO group the left sciatic nerve was exposed through a gluteal muscle incision and transected proximal to the tibio-peroneal bifurcation where a 10 mm segment was excised. The same procedure was performed in the ALLO/SVF group. The harvested nerves of the rats of ALLO group were served as allograft for ALLO/SVF group and vice versa. The SHAM and ALLO groups received 100 μL phosphate buffered saline and the ALLO/SVF group received 100 μL SVF (2.25 ± 0.45 × 10(7) cells) locally where the grafting was performed. Behavioral, functional, biomechanical, and gastrocnemius muscle mass showed earlier regeneration of axons in ALLO/SVF than in ALLO group (p < .05). Histomorphometic and immunohistochemical studies also showed earlier regeneration of axons in ALLO/SVF than in ALLO group (p < .05). Administration of nonexpanded SVF could accelerate functional recovery after nerve allografting in sciatic nerve. It may have clinical implications for the surgical management of patients after nerve transection.

  13. Osteochondral tissue formation through adipose-derived stromal cell differentiation on biomimetic polycaprolactone nanofibrous scaffolds with graded insulin and Beta-glycerophosphate concentrations.

    PubMed

    Erisken, Cevat; Kalyon, Dilhan M; Wang, Hongjun; Ornek-Ballanco, Ceren; Xu, Jiahua

    2011-05-01

    The ability to fabricate tissue engineering scaffolds containing systematic gradients in the distributions of stimulators provides additional means for the mimicking of the important gradients observed in native tissues. Here the concentration distributions of two bioactive agents were varied concomitantly for the first time (one increasing, whereas the other decreasing monotonically) in between the two sides of a nanofibrous scaffold. This was achieved via the application of a new processing method, that is, the twin-screw extrusion and electrospinning method, to generate gradients of insulin, a stimulator of chondrogenic differentiation, and β-glycerophosphate (β-GP), for mineralization. The graded poly(ɛ-caprolactone) mesh was seeded with human adipose-derived stromal cells and cultured over 8 weeks. The resulting tissue constructs were analyzed for and revealed indications of selective differentiation of human adipose-derived stromal cells toward chondrogenic lineage and mineralization as functions of position as a result of the corresponding concentrations of insulin and β-GP. Chondrogenic differentiation of the stem cells increased at insulin-rich locations and mineralization increased at β-GP-rich locations.

  14. The Celution® System: Automated Processing of Adipose-Derived Regenerative Cells in a Functionally Closed System

    PubMed Central

    Fraser, John K.; Hicok, Kevin C.; Shanahan, Rob; Zhu, Min; Miller, Scott; Arm, Douglas M.

    2014-01-01

    Objective: To develop a closed, automated system that standardizes the processing of human adipose tissue to obtain and concentrate regenerative cells suitable for clinical treatment of thermal and radioactive burn wounds. Approach: A medical device was designed to automate processing of adipose tissue to obtain a clinical-grade cell output of stromal vascular cells that may be used immediately as a therapy for a number of conditions, including nonhealing wounds resulting from radiation damage. Results: The Celution® System reliably and reproducibly generated adipose-derived regenerative cells (ADRCs) from tissue collected manually and from three commercial power-assisted liposuction devices. The entire process of introducing tissue into the system, tissue washing and proteolytic digestion, isolation and concentration of the nonadipocyte nucleated cell fraction, and return to the patient as a wound therapeutic, can be achieved in approximately 1.5 h. An alternative approach that applies ultrasound energy in place of enzymatic digestion demonstrates extremely poor efficiency cell extraction. Innovation: The Celution System is the first medical device validated and approved by multiple international regulatory authorities to generate autologous stromal vascular cells from adipose tissue that can be used in a real-time bedside manner. Conclusion: Initial preclinical and clinical studies using ADRCs obtained using the automated tissue processing Celution device described herein validate a safe and effective manner to obtain a promising novel cell-based treatment for wound healing. PMID:24761343

  15. Evaluation of Human Adipose Tissue Stromal Heterogeneity in Metabolic Disease Using Single Cell RNA-Seq

    DTIC Science & Technology

    2017-09-01

    1) define functional roles for individual genes and cell types in development of obesity and insulin resistance and 2) examine novel targets against...which we can design therapies to target specific pathogenic or or health-promoting cell types. 15. SUBJECT TERMS Obesity , Type 2 Diabetes Mellitus...compromised with chronic overnutrition ( obesity ). 4 KEYWORDS: Obesity , Diabetes, Insulin Resistance, Adipose, Adipocytes, Stromal Vascular Fraction, Single

  16. Evaluation of Human Adipose Tissue Stromal Heterogeneity in Metabolic Disease Using Single Cell RNA-Seq

    DTIC Science & Technology

    2016-09-01

    results are hypothesis-generating and provide the foundation for future studies that will 1) validate the role for newly identified mediators of obesity ...and insulin resistance in animal models and 2) examine novel targets against which we can design therapies to combat obesity and its related...complications. 15. SUBJECT TERMS Obesity , Type 2 Diabetes Mellitus, Insulin resistance, Adipose, Stromal vascular fraction 16. SECURITY CLASSIFICATION OF

  17. Decellularized adipose tissue microcarriers as a dynamic culture platform for human adipose-derived stem/stromal cell expansion.

    PubMed

    Yu, Claire; Kornmuller, Anna; Brown, Cody; Hoare, Todd; Flynn, Lauren E

    2017-03-01

    With the goal of designing a clinically-relevant expansion strategy for human adipose-derived stem/stromal cells (ASCs), methods were developed to synthesize porous microcarriers derived purely from human decellularized adipose tissue (DAT). An electrospraying approach was applied to generate spherical DAT microcarriers with an average diameter of 428 ± 41 μm, which were soft, compliant, and stable in long-term culture without chemical crosslinking. Human ASCs demonstrated enhanced proliferation on the DAT microcarriers relative to commercially-sourced Cultispher-S microcarriers within a spinner culture system over 1 month. ASC immunophenotype was maintained post expansion, with a trend for reduced expression of the cell adhesion receptors CD73, CD105, and CD29 under dynamic conditions. Upregulation of the early lineage-specific genes PPARγ, LPL, and COMP was observed in the ASCs expanded on the DAT microcarriers, but the cells retained their multilineage differentiation capacity. Comparison of adipogenic and osteogenic differentiation in 2-D cultures prepared with ASCs pre-expanded on the DAT microcarriers or Cultispher-S microcarriers revealed similar adipogenic and enhanced osteogenic marker expression in the DAT microcarrier group, which had undergone a higher population fold change. Further, histological staining results suggested a more homogeneous differentiation response in the ASCs expanded on the DAT microcarriers as compared to either Cultispher-S microcarriers or tissue culture polystyrene. A pilot chondrogenesis study revealed higher levels of chondrogenic gene and protein expression in the ASCs expanded on the DAT microcarriers relative to all other groups, including the baseline controls. Overall, this study demonstrates the promise of applying dynamic culture with tissue-specific DAT microcarriers as a means of deriving regenerative cell populations. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Effect of TGF-β1 Stimulation on the Secretome of Human Adipose-Derived Mesenchymal Stromal Cells

    PubMed Central

    Rodríguez, Tania M.; Saldías, Alejandro; Irigo, Marcelo; Zamora, Jorge Velasco; Perone, Marcelo J.

    2015-01-01

    Adipose tissue is an attractive source of mesenchymal stromal cells (MSCs) owing to the relative ease of obtaining large volumes with more MSC abundance compared with other sources. Increasing evidence supports the fact that trophic factors secreted by MSCs play a pivotal therapeutic role. Several strategies in regenerative medicine use MSCs, mainly exploiting their immunosuppressive effect and homing capacity to sites of damage. Transforming growth factor-β1 (TGF-β1) is a pleiotropic cytokine that, depending on the cell niche, can display either anti-inflammatory or proinflammatory effects. TGF-β1 expression increases in various tissues with damage, especially when accompanied by inflammation. Thus, we analyzed the effect of TGF-β1 on the secretion by adipose-derived mesenchymal stromal cells (ASCs) of a panel of 80 cytokines/chemokines using an antibody array. To avoid a possible effect of fetal bovine serum (FBS) on ASCs secretion, we performed our analysis by culturing cells in FBS-free conditions, only supplemented with 0.1% of bovine serum albumin. We report the cytokine profile secreted by ASCs. We also found that TGF-β1 exposure modulates 8 chemokines and 18 cytokines, including TGF-β1 and -β2, and other important cytokines involved in immunosuppression, allergic responses, and bone resorption. Significance Mesenchymal stromal cells (MSCs) secrete a broad spectrum of bioactive macromolecules that are both immunoregulatory and serve to structure regenerative microenvironments in fields of tissue injury. Increases or decreases in the production of TGF-β1 have been linked to numerous disease states, including autoimmune diseases and cancer. The secretome of MSCs stimulated with TGF-β1 is largely unknown. Thus, the present study makes an important contribution toward a better understanding of how MSCs could be affected by a cytokine normally upregulated in various diseases. PMID:26025982

  19. Conditioned medium of adipose-derived stromal cell culture in three-dimensional bioreactors for enhanced wound healing.

    PubMed

    Kwon, Sun Hyun; Bhang, Suk Ho; Jang, Hyeon-Ki; Rhim, Taiyoun; Kim, Byung-Soo

    2015-03-01

    It was previously shown that human adipose-derived stromal cell (hADSC)-conditioned medium (CM) promotes wound healing. An essential part of the wound healing process is neovascularization in the wound bed. We hypothesized that CM prepared from hADSCs cultured as spheroids in three-dimensional suspension bioreactors (spheroid CM) would contain much higher concentrations of angiogenic growth factors secreted by hADSCs, induce a higher extent of neovascularization in the wound bed, and improve wound healing as compared with CM prepared by conventional monolayer culture (monolayer CM). The concentrations of angiogenic growth factors (i.e., vascular endothelial growth factor, basic fibroblast growth factor, and hepatocyte growth factor) in spheroid CM were 20- to 145-fold higher than those in monolayer CM. Either fresh medium, monolayer CM, or spheroid CM was administered to full-thickness wounds created on the dorsal aspects of athymic mice. The monolayer CM promoted wound healing as compared with fresh medium or no treatment. Importantly, wound closure was faster, and dermal and epidermal regeneration was improved in the spheroid CM-treated mice compared with that in the monolayer CM-treated mice. The improved wound healing by spheroid CM may be attributed, at least in part, to enhanced neovascularization in the wound beds. The spheroid-based CM approach showed potential as a therapy for skin wound repair. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Depot-specific characteristics of adipose tissue-derived stromal cells in thyroid-associated orbitopathy.

    PubMed

    Wong, Janice Siu Chong; Chu, Wai Kit; Li, Benjamin Fuk-Loi; Pang, Chi-Pui; Chong, Kelvin Kam-Lung

    2018-04-17

    Thyroid-associated orbitopathy (TAO) causes inflammatory fibroproliferation of periocular connective tissues. We compared adipose tissue-derived stem/stromal cells (ADSCs) from three adipose depots of each patient with TAO on mesenchymal, myofibrogenic, adipogenic properties and associated hyaluronan (HA) synthesis. ADSCs were generated from periocular (eyelid, orbital) and subcutaneous (abdominal) adipose tissues of three patients with TAO. Mesenchymal markers were characterised by reverse transcription-PCR and immunofluorescent staining. A 3-week adipogenic induction was evaluated by Nile red staining and quantitative PCR (qPCR) of peroxisome proliferator-activated receptor (PPARγ), adiponectin and hyaluronan synthase (HAS)-2. A 7-day myofibrogenic induction was assayed by immunofluorescent staining and qPCR of α-smooth muscle actin (α-SMA). ADSCs from all depots expressed similar levels of mesenchymal markers CD44, CD90 and CD105 (p=0.288, p=0.43 and p=0.837, respectively). After adipogenic induction, intracellular lipid increased for more than 32% and PPARγ mRNA showed more than twofold increase from all three depots. However, adiponectin and HAS-2 mRNA levels were significantly higher in the eyelid and orbital ADSCs than those from the subcutaneous ADSCs after induction (2.4×10 7 , 3.9×10 6  folds vs below detection limit; 63.3-fold, 26.1-fold, vs 33% reduction, respectively; all p=0.002). Significantly more myofibroblasts and higher mRNA level of α-SMA were obtained from the orbital and eyelid compared with the subcutaneous ADSCs during myofibrogenic induction (80.2%, 70.6% vs 29.3%; 30.2-fold, 24.2-fold vs 1.7-fold, respectively; all p=0.002). ADSCs from different adipose depots of the same donors exhibited similar mesenchymal phenotypes but differed significantly in adipogenic, myofibrogenic potentials and associated HA synthesis. These depot-specific characteristics of ADSCs may contribute to site-specific adipose tissue involvement in TAO.

  1. Mechanically robust cryogels with injectability and bioprinting supportability for adipose tissue engineering.

    PubMed

    Qi, Dianjun; Wu, Shaohua; Kuss, Mitchell A; Shi, Wen; Chung, Soonkyu; Deegan, Paul T; Kamenskiy, Alexey; He, Yini; Duan, Bin

    2018-05-26

    Bioengineered adipose tissues have gained increased interest as a promising alternative to autologous tissue flaps and synthetic adipose fillers for soft tissue augmentation and defect reconstruction in clinic. Although many scaffolding materials and biofabrication methods have been investigated for adipose tissue engineering in the last decades, there are still challenges to recapitulate the appropriate adipose tissue microenvironment, maintain volume stability, and induce vascularization to achieve long-term function and integration. In the present research, we fabricated cryogels consisting of methacrylated gelatin, methacrylated hyaluronic acid, and 4arm poly(ethylene glycol) acrylate (PEG-4A) by using cryopolymerization. The cryogels were repeatedly injectable and stretchable, and the addition of PEG-4A improved the robustness and mechanical properties. The cryogels supported human adipose progenitor cell (HWA) and adipose derived mesenchymal stromal cell adhesion, proliferation, and adipogenic differentiation and maturation, regardless of the addition of PEG-4A. The HWA laden cryogels facilitated the co-culture of human umbilical vein endothelial cells (HUVEC) and capillary-like network formation, which in return also promoted adipogenesis. We further combined cryogels with 3D bioprinting to generate handleable adipose constructs with clinically relevant size. 3D bioprinting enabled the deposition of multiple bioinks onto the cryogels. The bioprinted flap-like constructs had an integrated structure without delamination and supported vascularization. Adipose tissue engineering is promising for reconstruction of soft tissue defects, and also challenging for restoring and maintaining soft tissue volume and shape, and achieving vascularization and integration. In this study, we fabricated cryogels with mechanical robustness, injectability, and stretchability by using cryopolymerization. The cryogels promoted cell adhesion, proliferation, and adipogenic

  2. Culture expansion of adipose derived stromal cells. A closed automated Quantum Cell Expansion System compared with manual flask-based culture.

    PubMed

    Haack-Sørensen, Mandana; Follin, Bjarke; Juhl, Morten; Brorsen, Sonja K; Søndergaard, Rebekka H; Kastrup, Jens; Ekblond, Annette

    2016-11-16

    Adipose derived stromal cells (ASCs) are a rich and convenient source of cells for clinical regenerative therapeutic approaches. However, applications of ASCs often require cell expansion to reach the needed dose. In this study, cultivation of ASCs from stromal vascular fraction (SVF) over two passages in the automated and functionally closed Quantum Cell Expansion System (Quantum system) is compared with traditional manual cultivation. Stromal vascular fraction was isolated from abdominal fat, suspended in α-MEM supplemented with 10% Fetal Bovine Serum and seeded into either T75 flasks or a Quantum system that had been coated with cryoprecipitate. The cultivation of ASCs from SVF was performed in 3 ways: flask to flask; flask to Quantum system; and Quantum system to Quantum system. In all cases, quality controls were conducted for sterility, mycoplasmas, and endotoxins, in addition to the assessment of cell counts, viability, immunophenotype, and differentiation potential. The viability of ASCs passage 0 (P0) and P1 was above 96%, regardless of cultivation in flasks or Quantum system. Expression of surface markers and differentiation potential was consistent with ISCT/IFATS standards for the ASC phenotype. Sterility, mycoplasma, and endotoxin tests were consistently negative. An average of 8.0 × 10 7 SVF cells loaded into a Quantum system yielded 8.96 × 10 7 ASCs P0, while 4.5 × 10 6 SVF cells seeded per T75 flask yielded an average of 2.37 × 10 6 ASCs-less than the number of SVF cells seeded. ASCs P1 expanded in the Quantum system demonstrated a population doubling (PD) around 2.2 regardless of whether P0 was previously cultured in flasks or Quantum, while ASCs P1 in flasks only reached a PD of 1.0. Manufacturing of ASCs in a Quantum system enhances ASC expansion rate and yield significantly relative to manual processing in T-flasks, while maintaining the purity and quality essential to safe and robust cell production. Notably, the use of the Quantum

  3. Enhanced angiogenic effect of adipose-derived stromal cell spheroid with low-level light therapy in hindlimb ischemia mice

    NASA Astrophysics Data System (ADS)

    Park, In-Su; Ahn, Jin-Chul; Chung, Phil-Sang

    2014-02-01

    Adipose-derived stromal cells (ASCs) are attractive cell source for tissue engineering. However, one obstacle to this approach is that the transplanted ASC population can decline rapidly in the recipient tissue. The aim of this study was to investigate the effects of low-level laser therapy (LLLT) on transplanted human ASCs (hASCs) spheroid in a hindlimb ischemia animal model. LLLT, hASCs spheroid and hASCs spheroid transplantation with LLLT (spheroid + LLLT) were applied to the ischemic hindlimbs in athymic mice. The survival, differentiation and secretion of vascular endothelial growth (VEGF) of spheroid ASCs were evaluated by immunohistochemistry. The spheroid + LLLT group enhanced the tissue regeneration, including angiogenesis, compared with other groups. The spheroid contributed tissue regeneration via differentiation and secretion of growth factors. In the spheroid + LLLT group, the survival of spheroid hASCs was increased by the decreased apoptosis of spheroid hASCs in the ischemic hindlimb. The secretion of growth factors was stimulated in the spheroid + LLLT group compared with the ASCs group and spheroid group. These data suggest that LLLT is an effective biostimulator of spheroid hASCs in tissue regeneration that enhances the survival of ASCs and stimulates the secretion of growth factors in the ischemic hindlimb.

  4. Autologous adipose tissue-derived stromal cells for treatment of spinal cord injury.

    PubMed

    Kang, Soo-Kyung; Shin, Myung-Joo; Jung, Jin Sup; Kim, Yong Geun; Kim, Cheul-Hong

    2006-08-01

    Isolated rat adipose tissue-derived stromal cells (rATSCs) contain pluripotent cells that can be differentiated into a variety of cell lineages, including neural cells. Recent work has shown that ATSCs can make neurosphere-like clumps and differentiate into neuron-like cells expressing neuronal markers, but their therapeutic effect is unclear. Here we report that intravenous infusion of oligodendrocyte precursor cells (OPCs) derived from rATSC autograft cells sources improve motor function in rat models of spinal cord injury (SCI). After 4-5 weeks, transplanted rATSC-OPC cells survived and migrated into the injured region of SCI very efficiently (30-35%) and migrated cells were partially differentiated into neurons and oligodendrocyte. Also, we found some of the engrafted OPCs migrated and integrated in the kidney, brain, lung, and liver through the intravenous system. Behavioral analysis revealed the locomotor functions of OPC-autografted SCI rats were significantly restored. Efficient migration of intravenously engrafted rATSC-OPCs cells into SCI lesion suggests that SCI-induced chemotaxic factors facilitate migration of rATSC-OPCs. Here, we verified that engrafted rATSCs and SCI-induced chemotaxic factors indeed play an important role in proliferation, migration, and differentiation of endogeneous spinal cord-derived neural progenitor cells in the injured region. In transplantation paradigms, the interaction between engrafted rATSC-OPCs and endogeneous spinal cord-derived neuronal progenitor cells will be important in promoting healing through fate decisions, resulting in coordinated induction of cell migration and differentiation.

  5. Transplantation of Adipose Derived Stromal Cells into the Developing Mouse Eye

    PubMed Central

    Yu, Song-Hee; Jang, Yu-Jin; Lee, Eun-Shil; Hwang, Dong-Youn; Jeon, Chang-Jin

    2010-01-01

    Adipose derived stromal cells (ADSCs) were transplanted into a developing mouse eye to investigate the influence of a developing host micro environment on integration and differentiation. Green fluorescent protein-expressing ADSCs were transplanted by intraocular injections. The age of the mouse was in the range of 1 to 10 days postnatal (PN). Survival dates ranged from 7 to 28 post transplantation (DPT), at which time immunohistochemistry was performed. The transplanted ADSCs displayed some morphological differentiations in the host eye. Some cells expressed microtubule associated protein 2 (marker for mature neuron), or glial fibrillary acid protein (marker for glial cell). In addition, some cells integrated into the ganglion cell layer. The integration and differentiation of the transplanted ADSCs in the 5 and 10 PN 7 DPT were better than in the host eye the other age ranges. This study was aimed at demonstrating how the age of host micro environment would influence the differentiation and integration of the transplanted ADSCs. However, it was found that the integration and differentiation into the developing retina were very limited when compared with other stem cells, such as murine brain progenitor cell. PMID:21245978

  6. Osteogenic capability of autologous rabbit adipose-derived stromal cells in repairing calvarial defects.

    PubMed

    Cheng, Shao-Wen; Lin, Zhong-Qin; Wang, Wei; Zhang, Wei; Kou, Dong-Quan; Ying, Xiao-Zhou; Chen, Qing-Yu; Shen, Yue; Cheng, Xiao-Jie; Peng, Lei; Lv, Chuan-Zhu

    2011-01-01

    To evaluate the in vitro and in vivo osteogenic capability of adipose-derived stromal cells (ASCs). ASCs were isolated from New Zealand white rabbits and determined by alkaline phosphatase (ALP) staining, von Kossa staining and alizarin red staining. Some specific markers of osteogenic differentiation, including ALP, osteocalcin (OCN), osteopontin (OPN) were examined by reverse transcription-polymerase chain reaction (RT-PCR). In vivo, demineralized bone matrix (DBM)-ASCs composites were implanted into the rabbit calvarial defects created at each side of the longitudinal midline. After 6 weeks, histologic properties of the transplants were analyzed. ASCs were successfully induced into osteogenesis. ALP staining, von Kossa staining and alizarin red staining showed positive results. The expressions of ALP, OCN and OPN were detected in ASCs after cultivation in osteogenic medium. Extensive new bone was observed in the defects transplanted with DBM-ASCs composites. ASCs have the potential to differentiate into osteogenic lineage and DBM-ASCs constructs are a promising method for regeneration in bone defects.

  7. Mirna biogenesis pathway is differentially regulated during adipose derived stromal/stem cell differentiation.

    PubMed

    Martin, E C; Qureshi, A T; Llamas, C B; Burow, M E; King, A G; Lee, O C; Dasa, V; Freitas, M A; Forsberg, J A; Elster, E A; Davis, T A; Gimble, J M

    2018-02-07

    Stromal/stem cell differentiation is controlled by a vast array of regulatory mechanisms. Included within these are methods of mRNA gene regulation that occur at the level of epigenetic, transcriptional, and/or posttranscriptional modifications. Current studies that evaluate the posttranscriptional regulation of mRNA demonstrate microRNAs (miRNAs) as key mediators of stem cell differentiation through the inhibition of mRNA translation. miRNA expression is enhanced during both adipogenic and osteogenic differentiation; however, the mechanism by which miRNA expression is altered during stem cell differentiation is less understood. Here we demonstrate for the first time that adipose-derived stromal/stem cells (ASCs) induced to an adipogenic or osteogenic lineage have differences in strand preference (-3p and -5p) for miRNAs originating from the same primary transcript. Furthermore, evaluation of miRNA expression in ASCs demonstrates alterations in both miRNA strand preference and 5'seed site heterogeneity. Additionally, we show that during stem cell differentiation there are alterations in expression of genes associated with the miRNA biogenesis pathway. Quantitative RT-PCR demonstrated changes in the Argonautes (AGO1-4), Drosha, and Dicer at intervals of ASC adipogenic and osteogenic differentiation compared to untreated ASCs. Specifically, we demonstrated altered expression of the AGOs occurring during both adipogenesis and osteogenesis, with osteogenesis increasing AGO1-4 expression and adipogenesis decreasing AGO1 gene and protein expression. These data demonstrate changes to components of the miRNA biogenesis pathway during stromal/stem cell differentiation. Identifying regulatory mechanisms for miRNA processing during ASC differentiation may lead to novel mechanisms for the manipulation of lineage differentiation of the ASC through the global regulation of miRNA as opposed to singular regulatory mechanisms.

  8. Comparison of Adipose-Derived and Bone Marrow Mesenchymal Stromal Cells in a Murine Model of Crohn's Disease.

    PubMed

    Xie, Minghao; Qin, Huabo; Luo, Qianxin; He, Xiaosheng; He, Xiaowen; Lan, Ping; Lian, Lei

    2017-01-01

    Mesenchymal stromal cells (MSCs) have been used in the treatment of Crohn's disease (CD) because of the immunomodulatory ability. The aim of this study was to investigate the therapeutic effect of adipose-derived MSCs (AD-MSCs) and to compare the therapeutic effect of AD-MSCs with that of bone marrow MSCs (BM-MSCs) in a murine model of CD. Murine colitis model of CD was created by trinitrobenzene sulfonic acid (TNBS). Twelve hours after treatment with TNBS, the mouse model was injected with MSCs intraperitoneally. Real-time polymerase chain reaction and immunohistochemistry staining were used to measure the expression levels of inflammatory cytokines in colonic tissues to investigate the therapeutic effect of AD-MSCs. The ten-day survival was recorded after infusion of MSCs. Intraperitoneal injection of MSCs alleviated the clinical and histopathologic severity of intestinal inflammation, and improved the survival of the TNBS-induced mouse model of CD. AD-MSCs could effectively increase the expression of interleukin-10 and reduce the secretion of pro-inflammatory cytokines including tumor necrosis factor-α, interleukin-12, and vascular endothelial growth factor. The mucosal injury was repaired by AD-MSCs. These effects were comparable between AD-MSCs and BM-MSCs. The therapeutic effect appears similar between AD-MSCs and BM-MSCs in treating CD. AD-MSCs may be a potential alternative of cell-based therapy for CD.

  9. Human Adipose-Derived Mesenchymal Stem/Stromal Cells Handling Protocols. Lipid Droplets and Proteins Double-Staining

    PubMed Central

    Gojanovich, Aldana D.; Gimenez, María C.; Masone, Diego; Rodriguez, Tania M.; Dewey, Ricardo A.; Delgui, Laura R.; Bustos, Diego M.; Uhart, Marina

    2018-01-01

    Human Adipose-derived mesenchymal stem/stromal cells (hASCs) are of great interest because of their potential for therapeutic approaches. The method described here covers every single step necessary for hASCs isolation from subcutaneous abdominal adipose tissue, multicolor phenotyping by flow cytometry, and quantitative determination of adipogenic differentiation status by means of lipid droplets (LDs) accumulation, and Western blot analysis. Moreover, to simultaneously analyze both LDs accumulation and cellular proteins localization by fluorescence microscopy, we combined Oil Red O (ORO) staining with immunofluorescence detection. For LDs quantification we wrote a program for automatic ORO-stained digital image processing implemented in Octave, a freely available software package. Our method is based on the use of the traditional low cost neutral lipids dye ORO, which can be imaged both by bright-field and fluorescence microscopy. The utilization of ORO instead of other more expensive lipid-specific dyes, together with the fact that the whole method has been designed employing cost-effective culture reagents (standard culture medium and serum), makes it affordable for tight-budget research laboratories. These may be replaced, if necessary or desired, by defined xeno-free reagents for clinical research and applications. PMID:29670879

  10. Human Adipose-Derived Mesenchymal Stem/Stromal Cells Handling Protocols. Lipid Droplets and Proteins Double-Staining.

    PubMed

    Gojanovich, Aldana D; Gimenez, María C; Masone, Diego; Rodriguez, Tania M; Dewey, Ricardo A; Delgui, Laura R; Bustos, Diego M; Uhart, Marina

    2018-01-01

    Human Adipose-derived mesenchymal stem/stromal cells (hASCs) are of great interest because of their potential for therapeutic approaches. The method described here covers every single step necessary for hASCs isolation from subcutaneous abdominal adipose tissue, multicolor phenotyping by flow cytometry, and quantitative determination of adipogenic differentiation status by means of lipid droplets (LDs) accumulation, and Western blot analysis. Moreover, to simultaneously analyze both LDs accumulation and cellular proteins localization by fluorescence microscopy, we combined Oil Red O (ORO) staining with immunofluorescence detection. For LDs quantification we wrote a program for automatic ORO-stained digital image processing implemented in Octave, a freely available software package. Our method is based on the use of the traditional low cost neutral lipids dye ORO, which can be imaged both by bright-field and fluorescence microscopy. The utilization of ORO instead of other more expensive lipid-specific dyes, together with the fact that the whole method has been designed employing cost-effective culture reagents (standard culture medium and serum), makes it affordable for tight-budget research laboratories. These may be replaced, if necessary or desired, by defined xeno-free reagents for clinical research and applications.

  11. In vitro differentiation of neural cells from human adipose tissue derived stromal cells.

    PubMed

    Dave, Shruti D; Patel, Chetan N; Vanikar, Aruna V; Trivedi, Hargovind L

    2018-01-01

    Stem cells, including neural stem cells (NSCs), are endowed with self-renewal capability and hence hold great opportunity for the institution of replacement/protective therapy. We propose a method for in vitro generation of stromal cells from human adipose tissue and their differentiation into neural cells. Ten grams of donor adipose tissue was surgically resected from the abdominal wall of the human donor after the participants' informed consents. The resected adipose tissue was minced and incubated for 1 hour in the presence of an enzyme (collagenase-type I) at 37 0 C followed by its centrifugation. After centrifugation, the supernatant and pellets were separated and cultured in a medium for proliferation at 37 0 C with 5% CO2 for 9-10 days in separate tissue culture dishes for generation of mesenchymal stromal cells (MSC). At the end of the culture, MSC were harvested and analyzed. The harvested MSC were subjected for further culture for their differentiation into neural cells for 5-7 days using differentiation medium mainly comprising of neurobasal medium. At the end of the procedure, culture cells were isolated and studied for expression of transcriptional factor proteins: orthodenticle homolog-2 (OTX-2), beta-III-tubulin (β3-Tubulin), glial-fibrillary acid protein (GFAP) and synaptophysin-β2. In total, 50 neural cells-lines were generated. In vitro generated MSC differentiated neural cells' mean quantum was 5.4 ± 6.9 ml with the mean cell count being, 5.27 ± 2.65 × 10 3/ μl. All of them showed the presence of OTX-2, β3-Tubulin, GFAP, synaptophysin-β2. Neural cells can be differentiated in vitro from MSC safely and effectively. In vitro generated neural cells represent a potential therapy for recovery from spinal cord injuries and neurodegenerative disease.

  12. High Aldehyde Dehydrogenase Activity Identifies a Subset of Human Mesenchymal Stromal Cells with Vascular Regenerative Potential.

    PubMed

    Sherman, Stephen E; Kuljanin, Miljan; Cooper, Tyler T; Putman, David M; Lajoie, Gilles A; Hess, David A

    2017-06-01

    During culture expansion, multipotent mesenchymal stromal cells (MSCs) differentially express aldehyde dehydrogenase (ALDH), an intracellular detoxification enzyme that protects long-lived cells against oxidative stress. Thus, MSC selection based on ALDH-activity may be used to reduce heterogeneity and distinguish MSC subsets with improved regenerative potency. After expansion of human bone marrow-derived MSCs, cell progeny was purified based on low versus high ALDH-activity (ALDH hi ) by fluorescence-activated cell sorting, and each subset was compared for multipotent stromal and provascular regenerative functions. Both ALDH l ° and ALDH hi MSC subsets demonstrated similar expression of stromal cell (>95% CD73 + , CD90 + , CD105 + ) and pericyte (>95% CD146 + ) surface markers and showed multipotent differentiation into bone, cartilage, and adipose cells in vitro. Conditioned media (CDM) generated by ALDH hi MSCs demonstrated a potent proliferative and prosurvival effect on human microvascular endothelial cells (HMVECs) under serum-free conditions and augmented HMVEC tube-forming capacity in growth factor-reduced matrices. After subcutaneous transplantation within directed in vivo angiogenesis assay implants into immunodeficient mice, ALDH hi MSC or CDM produced by ALDH hi MSC significantly augmented murine vascular cell recruitment and perfused vessel infiltration compared with ALDH l ° MSC. Although both subsets demonstrated strikingly similar mRNA expression patterns, quantitative proteomic analyses performed on subset-specific CDM revealed the ALDH hi MSC subset uniquely secreted multiple proangiogenic cytokines (vascular endothelial growth factor beta, platelet derived growth factor alpha, and angiogenin) and actively produced multiple factors with chemoattractant (transforming growth factor-β, C-X-C motif chemokine ligand 1, 2, and 3 (GRO), C-C motif chemokine ligand 5 (RANTES), monocyte chemotactic protein 1 (MCP-1), interleukin [IL]-6, IL-8) and matrix

  13. "In vitro" and multicolor phenotypic characterization of cell subpopulations identified in fresh human adipose tissue stromal vascular fraction and in the derived mesenchymal stem cells.

    PubMed

    Astori, Giuseppe; Vignati, Francesca; Bardelli, Silvana; Tubio, Monica; Gola, Mauro; Albertini, Veronica; Bambi, Franco; Scali, Giancarlo; Castelli, Damiano; Rasini, Valeria; Soldati, Gianni; Moccetti, Tiziano

    2007-10-31

    The stromal vascular fraction (SVF) is a heterogeneous cell population derived from the adipose tissue. There is still a lack of information concerning the characterization of the cell subpopulations constituting the SVF as well as its mesenchymal and haematopoietic potential. Furthermore there are great variations in its phenotypical characterization. Composition of SVF was investigated by FACS analysis, cytological and "in vitro" assays. We studied CD34+ population by combining FACS with human CFC (colony-forming-cell haematopoietic assay). The endothelial fraction was investigated by quantifying the co-expression of specific markers (CD146, CD105, CD31 and UEA-1). Mesenchymal potential was assessed by CFU-F assay and cultured AT-MSC were characterized by a 5-color FACS analysis. The multipotent differentiation potential (osteogenic, adipogenic and chondrogenic) was investigated both at cellular and molecular level. We identified in the SVF two CD34+ populations with a marked difference in the intensity of antigen expression, the majority of the cells expressing CD34 at low intensity. Moreover, two CD146+ cell populations were clearly distinguishable in the SVF:a CD146 dim accounting for 9.9% of the total SVF cells and a CD146+ bright cell population accounting for about 39.3%. The frequency of CFC clones was comparable with the one reported for peripheral blood. Endothelial cells account for about 7.7% of the SVF cells. AT-MSC differenced in the osteogenic adipogenic and chondrogenic lineage. The SVF is not a homogeneous cell population, and its final composition could be influenced both by the flow cytometric technique analysis and the SVF extraction steps. The CFU-F frequency in the SVF was 1/4880, a value about seven times greater than the data reported for bone marrow. The antigenic profile of AT-MSC was comparable with bone-marrow derived MSC. AT-MSC were able to differentiate along the osteogenic adipogenic and chondrogenic lineages. The data here reported

  14. Cigarette Smoking Impairs Adipose Stromal Cell Vasculogenic Activity and Abrogates Potency to Ameliorate Ischemia.

    PubMed

    Barwinska, Daria; Traktuev, Dmitry O; Merfeld-Clauss, Stephanie; Cook, Todd G; Lu, Hongyan; Petrache, Irina; March, Keith L

    2018-06-01

    Cigarette smoking (CS) adversely affects the physiologic function of endothelial progenitor, hematopoietic stem and progenitor cells. However, the effect of CS on the ability of adipose stem/stromal cells (ASC) to promote vasculogenesis and rescue perfusion in the context of ischemia is unknown. To evaluate this, ASC from nonsmokers (nCS-ASC) and smokers (CS-ASC), and their activity to promote perfusion in hindlimb ischemia models, as well as endothelial cell (EC) survival and vascular morphogenesis in vitro were assessed. While nCS-ASC improved perfusion in ischemic limbs, CS-ASC completely lost this therapeutic effect. In vitro vasculogenesis assays revealed that human CS-ASC and ASC from CS-exposed mice showed compromised support of EC morphogenesis into vascular tubes, and the CS-ASC secretome was less potent in supporting EC survival/proliferation. Comparative secretome analysis revealed that CS-ASC produced lower amounts of hepatocyte growth factor (HGF) and stromal cell-derived growth factor 1 (SDF-1). Conversely, CS-ASC secreted the angiostatic/pro-inflammatory factor Activin A, which was not detected in nCS-ASC conditioned media (CM). Furthermore, higher Activin A levels were measured in EC/CS-ASC cocultures than in EC/nCS-ASC cocultures. CS-ASC also responded to inflammatory cytokines with 5.2-fold increase in Activin A secretion, whereas nCS-ASC showed minimal Activin A induction. Supplementation of EC/CS-ASC cocultures with nCS-ASC CM or with recombinant vascular endothelial growth factor, HGF, or SDF-1 did not rescue vasculogenesis, whereas inhibition of Activin A expression or activity improved network formation up to the level found in EC/nCS-ASC cocultures. In conclusion, ASC of CS individuals manifest compromised in vitro vasculogenic activity as well as in vivo therapeutic activity. Stem Cells 2018;36:856-867. © 2018 AlphaMed Press.

  15. In vitro effects of direct current electric fields on adipose-derived stromal cells.

    PubMed

    Hammerick, Kyle E; Longaker, Michael T; Prinz, Fritz B

    2010-06-18

    Endogenous electric fields play an important role in embryogenesis, regeneration, and wound repair and previous studies have shown that many populations of cells, leukocytes, fibroblasts, epithelial cells, and endothelial cells, exhibit directed migration in response to electric fields. As regenerative therapies continue to explore ways to control mesenchymal progenitor cells to recreate desirable tissues, it is increasingly necessary to characterize the vast nature of biological responses imposed by physical phenomena. Murine adipose-derived stromal cells (mASCs) migrated toward the cathode in direct current (DC) fields of physiologic strength and show a dose dependence of migration rate to stronger fields. Electric fields also caused mASCs to orient perpendicularly to the field vector and elicited a transient increase in cytosolic calcium. Additionally, their galvanotactic response appears to share classic chemotactic signaling pathways that are involved in the migration of other cell types. Galvanotaxis is one predominant result of electric fields on mASCs and it may be exploited to engineer adult stem cell concentrations and locations within implanted grafts or toward sites of wound repair. Copyright (c) 2010 Elsevier Inc. All rights reserved.

  16. Development of large-scale manufacturing of adipose-derived stromal cells for clinical applications using bioreactors and human platelet lysate.

    PubMed

    Haack-Sørensen, Mandana; Juhl, Morten; Follin, Bjarke; Harary Søndergaard, Rebekka; Kirchhoff, Maria; Kastrup, Jens; Ekblond, Annette

    2018-04-17

    In vitro expanded adipose-derived stromal cells (ASCs) are a useful resource for tissue regeneration. Translation of small-scale autologous cell production into a large-scale, allogeneic production process for clinical applications necessitates well-chosen raw materials and cell culture platform. We compare the use of clinical-grade human platelet lysate (hPL) and fetal bovine serum (FBS) as growth supplements for ASC expansion in the automated, closed hollow fibre quantum cell expansion system (bioreactor). Stromal vascular fractions were isolated from human subcutaneous abdominal fat. In average, 95 × 10 6 cells were suspended in 10% FBS or 5% hPL medium, and loaded into a bioreactor coated with cryoprecipitate. ASCs (P0) were harvested, and 30 × 10 6 ASCs were reloaded for continued expansion (P1). Feeding rate and time of harvest was guided by metabolic monitoring. Viability, sterility, purity, differentiation capacity, and genomic stability of ASCs P1 were determined. Cultivation of SVF in hPL medium for in average nine days, yielded 546 × 10 6 ASCs compared to 111 × 10 6 ASCs, after 17 days in FBS medium. ASCs P1 yields were in average 605 × 10 6 ASCs (PD [population doublings]: 4.65) after six days in hPL medium, compared to 119 × 10 6 ASCs (PD: 2.45) in FBS medium, after 21 days. ASCs fulfilled ISCT criteria and demonstrated genomic stability and sterility. The use of hPL as a growth supplement for ASCs expansion in the quantum cell expansion system provides an efficient expansion process compared to the use of FBS, while maintaining cell quality appropriate for clinical use. The described process is an obvious choice for manufacturing of large-scale allogeneic ASC products.

  17. Diabetes impairs adipose tissue-derived stem cell function and efficiency in promoting wound healing.

    PubMed

    Cianfarani, Francesca; Toietta, Gabriele; Di Rocco, Giuliana; Cesareo, Eleonora; Zambruno, Giovanna; Odorisio, Teresa

    2013-01-01

    Adipose tissue-derived stem cells (ASCs) are gaining increasing consideration in tissue repair therapeutic application. Recent evidence indicates that ASCs enhance skin repair in animal models of impaired wound healing. To assess the therapeutic activity of autologous vs. allogeneic ASCs in the treatment of diabetic ulcers, we functionally characterized diabetic ASCs and investigated their potential to promote wound healing with respect to nondiabetic ones. Adipose tissue-derived cells from streptozotocin-induced type 1 diabetic mice were analyzed either freshly isolated as stromal vascular fraction (SVF), or following a single passage of culture (ASCs). Diabetic ASCs showed decreased proliferative potential and migration. Expression of surface markers was altered in diabetic SVF and cultured ASCs, with a reduction in stem cell marker-positive cells. ASCs from diabetic mice released lower amounts of hepatocyte growth factor, vascular endothelial growth factor (VEGF)-A, and insulin-like growth factor-1, growth factors playing important roles in skin repair. Accordingly, the supernatant of diabetic ASCs manifested reduced capability to promote keratinocyte and fibroblast proliferation and migration. Therapeutic potential of diabetic SVF administered to wounds of diabetic mice was blunted as compared with cells isolated from nondiabetic mice. Our data indicate that diabetes alters ASC intrinsic properties and impairs their function, thus affecting therapeutic potential in the autologous treatment for diabetic ulcers. © 2013 by the Wound Healing Society.

  18. Stromal vascular fraction isolated from lipo-aspirates using an automated processing system: bench and bed analysis.

    PubMed

    Doi, Kentaro; Tanaka, Shinsuke; Iida, Hideo; Eto, Hitomi; Kato, Harunosuke; Aoi, Noriyuki; Kuno, Shinichiro; Hirohi, Toshitsugu; Yoshimura, Kotaro

    2013-11-01

    The heterogeneous stromal vascular fraction (SVF), containing adipose-derived stem/progenitor cells (ASCs), can be easily isolated through enzymatic digestion of aspirated adipose tissue. In clinical settings, however, strict control of technical procedures according to standard operating procedures and validation of cell-processing conditions are required. Therefore, we evaluated the efficiency and reliability of an automated system for SVF isolation from adipose tissue. SVF cells, freshly isolated using the automated procedure, showed comparable number and viability to those from manual isolation. Flow cytometric analysis confirmed an SVF cell composition profile similar to that after manual isolation. In addition, the ASC yield after 1 week in culture was also not significantly different between the two groups. Our clinical study, in which SVF cells isolated with the automated system were transplanted with aspirated fat tissue for soft tissue augmentation/reconstruction in 42 patients, showed satisfactory outcomes with no serious side-effects. Taken together, our results suggested that the automated isolation system is as reliable a method as manual isolation and may also be useful in clinical settings. Automated isolation is expected to enable cell-based clinical trials in small facilities with an aseptic room, without the necessity of a good manufacturing practice-level cell processing area. Copyright © 2012 John Wiley & Sons, Ltd.

  19. Comparison of Synthetic Media Designed for Expansion of Adipose-Derived Mesenchymal Stromal Cells.

    PubMed

    Lensch, Michelle; Muise, Angela; White, Lisa; Badowski, Michael; Harris, David

    2018-05-14

    Mesenchymal stromal cells (MSCs) are multipotent cells that can differentiate into various cell types, such as osteoblasts, myocytes, and adipocytes. This characteristic makes the cells a useful tool in developing new therapies for a number of common maladies and diseases. The utilization of animal-derived growth serum, such as fetal bovine serum (FBS), for the expansion of MSCs has traditionally been used for cell culture. However, in clinical applications, animal-derived products present limitations and safety concerns for the recipient, as exposure to animal (xeno-) antigens and infectious agents is possible. Multiple synthetic, xeno-free media have been developed to combat these limitations of animal-derived growth serum and have the potential to be used in ex vivo MSC expansion for clinical use. The goal of this study was to determine if xeno-free media are adequate to significantly and efficiently expand MSCs derived from adipose tissue. MSCs were cultured in both standard FBS-containing as well as xeno-free media. The media were compared for cell yield, viability, and phenotypic expression via flow cytometry and directed differentiation. The xeno-free media that were tested were StemMACS MSC Expansion Media (Miltenyi Biotec, Bergisch Gladbach, Germany), PLTMax Human Platelet Lysate (Sigma-Aldrich, St. Louis, MO, USA), and MesenCult-hPL media (Stemcell Technologies, Vancouver, BC, Canada). All xeno-free media showed promise as a feasible replacement for animal-derived growth serums. The xeno-free media expanded MSCs more quickly than the FBS-containing medium and also showed great similarity in cell viability and phenotypic expression. In fact, each xeno-free media produced a greater viable cell yield than the standard FBS-containing medium.

  20. Amniotic Mesenchymal Stromal Cells Exhibit Preferential Osteogenic and Chondrogenic Differentiation and Enhanced Matrix Production Compared With Adipose Mesenchymal Stromal Cells.

    PubMed

    Topoluk, Natasha; Hawkins, Richard; Tokish, John; Mercuri, Jeremy

    2017-09-01

    Therapeutic efficacy of various mesenchymal stromal cell (MSC) types for orthopaedic applications is currently being investigated. While the concept of MSC therapy is well grounded in the basic science of healing and regeneration, little is known about individual MSC populations in terms of their propensity to promote the repair and/or regeneration of specific musculoskeletal tissues. Two promising MSC sources, adipose and amnion, have each demonstrated differentiation and extracellular matrix (ECM) production in the setting of musculoskeletal tissue regeneration. However, no study to date has directly compared the differentiation potential of these 2 MSC populations. To compare the ability of human adipose- and amnion-derived MSCs to undergo osteogenic and chondrogenic differentiation. Controlled laboratory study. MSC populations from the human term amnion were quantified and characterized via cell counting, histologic assessment, and flow cytometry. Differentiation of these cells in comparison to commercially purchased human adipose-derived mesenchymal stromal cells (hADSCs) in the presence and absence of differentiation media was evaluated via reverse transcription polymerase chain reaction (PCR) for bone and cartilage gene transcript markers and histology/immunohistochemistry to examine ECM production. Analysis of variance and paired t tests were performed to compare results across all cell groups investigated. The authors confirmed that the human term amnion contains 2 primary cell types demonstrating MSC characteristics-(1) human amniotic epithelial cells (hAECs) and (2) human amniotic mesenchymal stromal cells (hAMSCs)-and each exhibited more than 90% staining for MSC surface markers (CD90, CD105, CD73). Average viable hAEC and hAMSC yields at harvest were 2.3 × 10 6 ± 3.7 × 10 5 and 1.6 × 10 6 ± 4.7 × 10 5 per milliliter of amnion, respectively. As well, hAECs and hAMSCs demonstrated significantly greater osteocalcin ( P = .025), aggrecan ( P < .0001

  1. Safety, tolerability and potential efficacy of injection of autologous adipose-derived stromal vascular fraction in the fingers of patients with systemic sclerosis: an open-label phase I trial.

    PubMed

    Granel, Brigitte; Daumas, Aurélie; Jouve, Elisabeth; Harlé, Jean-Robert; Nguyen, Pierre-Sébastien; Chabannon, Christian; Colavolpe, Nathalie; Reynier, Jean-Charles; Truillet, Romain; Mallet, Stéphanie; Baiada, Antoine; Casanova, Dominique; Giraudo, Laurent; Arnaud, Laurent; Veran, Julie; Sabatier, Florence; Magalon, Guy

    2015-12-01

    In patients with systemic sclerosis (scleroderma, SSc), impaired hand function greatly contributes to disability and reduced quality of life, and is insufficiently relieved by currently available therapies. Adipose tissue-derived stromal vascular fraction (SVF) is increasingly recognised as an easily accessible source of regenerative cells with therapeutic potential in ischaemic or autoimmune diseases. We aimed to measure for the first time the safety, tolerability and potential efficacy of autologous SVF cells local injections in patients with SSc with hand disability. We did an open-label, single arm, at one study site with 6-month follow-up among 12 female SSc patients with Cochin Hand Function Scale score >20/90. Autologous SVF was obtained from lipoaspirates, using an automated processing system, and subsequently injected into the subcutaneous tissue of each finger in contact with neurovascular pedicles. Primary outcome was the number and the severity of adverse events related to SVF-based therapy. Secondary endpoints were changes in hand disability and fibrosis, vascular manifestations, pain and quality of life from baseline to 2 and 6 months after cell therapy. All enrolled patients had surgery, and there were no dropouts or patients lost to follow-up. No severe adverse events occurred during the procedure and follow-up. Four minor adverse events were reported and resolved spontaneously. A significant improvement in hand disability and pain, Raynaud's phenomenon, finger oedema and quality of life was observed. This study outlines the safety of the autologous SVF cells injection in the hands of patients with SSc. Preliminary assessments at 6 months suggest potential efficacy needing confirmation in a randomised placebo-controlled trial on a larger population. GFRS (Groupe Francophone de Recherche sur la Sclérodermie). NCT01813279. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to

  2. Wound Healing and Angiogenesis through Combined Use of a Vascularized Tissue Flap and Adipose-Derived Stem Cells in a Rat Hindlimb Irradiated Ischemia Model.

    PubMed

    Yoshida, Shuhei; Yoshimoto, Hiroshi; Hirano, Akiyoshi; Akita, Sadanori

    2016-05-01

    Treatment of critical limb ischemia is sometimes difficult because of the patient's condition, and some novel approaches are needed. The hindlimbs of Sprague-Dawley rats, after 20-Gy x-ray irradiation and surgical occlusion, were divided into four groups: with a superficial fascial flap, 5.0 × 10 adipose-derived stromal/stem cells, and both combined. The rats were tested for laser tissue blood flow, immunohistologic blood vessel density, and foot paw punch hole wound healing. Green fluorescent protein-tagged Sprague-Dawley rats were used for further investigation by cell tracking for 2 weeks. Laser tissue blood flow demonstrated a significant increase in the combined treatment of flap and adipose-derived stem cells at both 1 and 2 weeks. There were no significant differences between the treatment groups treated with flaps alone and those treated with adipose-derived stem cells alone. Wound healing was significantly increased following combined treatment at 1 week, and there was no wound by 2 weeks except for the no-flap and no-adipose-derived stem cell group. The number of vessels depicted by von Willebrand factor showed a significant increase in the combined treatment group, at both 1 week and 2 weeks. In the cell tracking group, at 2 weeks, the green fluorescent protein-tagged adipose-derived stem cells were significantly more positive in the no-flap group than in the flap group. Adipose-derived stem cells may be a potent cell source in irradiated and occluded limbs by enhancing tissue blood flow and blood vessel density. Adipose-derived stem cells may play an important role in some difficult ischemic conditions in terms of wound healing.

  3. Stromal Vascular Fraction Transplantation as an Alternative Therapy for Ischemic Heart Failure: Anti-inflammatory Role

    PubMed Central

    2011-01-01

    Background The aims of this study were: (1) to show the feasibility of using adipose-derived stromal vascular fraction (SVF) as an alternative to bone marrow mono nuclear cell (BM-MNC) for cell transplantation into chronic ischemic myocardium; and (2) to explore underlying mechanisms with focus on anti-inflammation role of engrafted SVF and BM-MNC post chronic myocardial infarction (MI) against left ventricular (LV) remodelling and cardiac dysfunction. Methods Four weeks after left anterior descending coronary artery ligation, 32 Male Lewis rats with moderate MI were divided into 3 groups. SVF group (n = 12) had SVF cell transplantation (6 × 106 cells). BM-MNC group (n = 12) received BM-MNCs (6 × 106) and the control (n = 10) had culture medium. At 4 weeks, after the final echocardiography, histological sections were stained with Styrus red and immunohistochemical staining was performed for α-smooth muscle actin, von Willebrand factor, CD3, CD8 and CD20. Results At 4 weeks, in SVF and BM-MNC groups, LV diastolic dimension and LV systolic dimension were smaller and fractional shortening was increased in echocardiography, compared to control group. Histology revealed highest vascular density, CD3+ and CD20+ cells in SVF transplanted group. SVF transplantation decreased myocardial mRNA expression of inflammatory cytokines TNF-α, IL-6, MMP-1, TIMP-1 and inhibited collagen deposition. Conclusions Transplantation of adipose derived SVF cells might be a useful therapeutic option for angiogenesis in chronic ischemic heart disease. Anti-inflammation role for SVF and BM transplantation might partly benefit for the cardioprotective effect for chronic ischemic myocardium. PMID:21453457

  4. Adipose tissue-derived stem cells enhance bioprosthetic mesh repair of ventral hernias.

    PubMed

    Altman, Andrew M; Abdul Khalek, Feras J; Alt, Eckhard U; Butler, Charles E

    2010-09-01

    Bioprosthetic mesh used for ventral hernia repair becomes incorporated into the musculofascial edge by cellular infiltration and vascularization. Adipose tissue-derived stem cells promote tissue repair and vascularization and may increase the rate or degree of tissue incorporation. The authors hypothesized that introducing these cells into bioprosthetic mesh would result in adipose tissue-derived stem cell engraftment and proliferation and enhance incorporation of the bioprosthetic mesh. Adipose tissue-derived stem cells were isolated from the subcutaneous adipose tissue of syngeneic Brown Norway rats, expanded in vitro, and labeled with green fluorescent protein. Thirty-six additional rats underwent inlay ventral hernia repair with porcine acellular dermal matrix. Two 12-rat groups had the cells (1.0 x 10(6)) injected directly into the musculofascial/porcine acellular dermal matrix interface after repair or received porcine acellular dermal matrix on which the cells had been preseeded; the 12-rat control group received no stem cells. At 2 weeks, adipose tissue-derived stem cells in both stem cell groups engrafted, survived, migrated, and proliferated. Mean cellular infiltration into porcine acellular dermal matrix at the musculofascial/graft interface was significantly greater in the preseeded and injected stem cell groups than in the control group. Mean vascular infiltration of the porcine acellular dermal matrix was significantly greater in both stem cell groups than in the control group. Preseeded and injected adipose tissue-derived stem cells engraft, migrate, proliferate, and enhance the vascularity of porcine acellular dermal matrix grafts at the musculofascial/graft interface. These cells can thus enhance incorporation of porcine acellular dermal matrix into the abdominal wall after repair of ventral hernias.

  5. Enrichment isolation of adipose-derived stem/stromal cells from the liquid portion of liposuction aspirates with the use of an adherent column.

    PubMed

    Doi, Kentaro; Kuno, Shinichiro; Kobayashi, Akira; Hamabuchi, Takahisa; Kato, Harunosuke; Kinoshita, Kahori; Eto, Hitomi; Aoi, Noriyuki; Yoshimura, Kotaro

    2014-03-01

    Adipose-derived stem/progenitor cells (ASCs) are typically obtained from the lipoaspirates; however, a smaller number of ASCs can be isolated without enzymatic digestion from the infranatant liposuction aspirate fluid (LAF). We evaluated the effectiveness of an adherent column, currently used to isolate mesenchymal stromal cells from bone marrow, to isolate LAF cells. We applied peripheral blood (PB), PB mixed with cultured ASCs (PB-ASC), and LAF solution to the column and divided it into two fractions, the adherent (positive) and the non-adherent (negative) fractions. We compared this method with hypotonic hemolysis (lysis) for the red blood cell count, nucleated cells count and cell compositions as well as functional properties of isolated mesenchymal cells. The column effectively removed red blood cells, though the removal efficiency was slightly inferior to hemolysis. After column processing of PB-ASC, 60.5% of ASCs (53.2% by lysis) were selectively collected in the positive fraction, and the negative fraction contained almost no ASCs. After processing of LAF solution, nucleated cell yields were comparable between the column and hemolysis; however, subsequent adherent culture indicated that a higher average ASC yield was obtained from the column-positive samples than from the lysis samples, suggesting that the column method may be superior to hemolysis for obtaining viable ASCs. Mesenchymal differentiation and network formation assays showed no statistical differences in ASC functions between the lysis and column-positive samples. Our results suggest that a column with non-woven rayon and polyethylene fabrics is useful for isolating stromal vascular fraction cells from LAF solutions for clinical applications. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  6. Novel culture system of mesenchymal stromal cells from human subcutaneous adipose tissue.

    PubMed

    Iwashima, Shigejiro; Ozaki, Takenori; Maruyama, Shoichi; Saka, Yousuke; Kobori, Masato; Omae, Kaoru; Yamaguchi, Hirotake; Niimi, Tomoaki; Toriyama, Kazuhiro; Kamei, Yuzuru; Torii, Shuhei; Murohara, Toyoaki; Yuzawa, Yukio; Kitagawa, Yasuo; Matsuo, Seiichi

    2009-05-01

    Accumulating evidence suggests that the delivery of human adipose tissue-derived stromal cells (hASCs) has great potential as regenerative therapy. This was performed to develop a method for expanding hASCs by reducing the amount of serum required. We demonstrate that hASCs were able to expand efficiently in media containing 2% serum and fibroblast growth factor-2. These cells, or low serum cultured hASCs (hLASCs), expressed cell surface markers similar to those on bone marrow-derived mesenchymal stem cells, and could be differentiated into cells of mesenchymal lineage. Of interest, hLASCs secreted higher levels of vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF) than hASCs cultured in 20% serum (hHASCs). Moreover, hLASC-conditioned media significantly increased endothelial cell (EC) proliferation and decreased EC apoptosis compared to that obtained from hHASCs or control media only. Antibodies against VEGF and HGF virtually negated these effects. When hASCs were administered into the ischemic hindlimbs of nude rats, hLASCs improved blood flow, increased capillary density, and raised the levels of VEGF and HGF in the muscles as compared with hHASCs. In conclusion, we demonstrate a novel low serum culture system for hASCs, which may have great potential in regenerative cell therapy for damaged organs in the clinical setting.

  7. Adipose-derived mesenchymal stromal cells from aged patients with coronary artery disease keep mesenchymal stromal cell properties but exhibit characteristics of aging and have impaired angiogenic potential.

    PubMed

    Efimenko, Anastasia; Dzhoyashvili, Nina; Kalinina, Natalia; Kochegura, Tatiana; Akchurin, Renat; Tkachuk, Vsevolod; Parfyonova, Yelena

    2014-01-01

    Tissue regeneration is impaired in aged individuals. Adipose-derived mesenchymal stromal cells (ADSCs), a promising source for cell therapy, were shown to secrete various angiogenic factors and improve vascularization of ischemic tissues. We analyzed how patient age affected the angiogenic properties of ADSCs. ADSCs were isolated from subcutaneous fat tissue of patients with coronary artery disease (CAD; n = 64, 43-77 years old) and without CAD (n = 31, 2-82 years old). ADSC phenotype characterized by flow cytometry was CD90(+)/CD73(+)/CD105(+)/CD45(-)/CD31(-) for all samples, and these cells were capable of adipogenic and osteogenic differentiation. ADSCs from aged patients had shorter telomeres (quantitative reverse transcription polymerase chain reaction) and a tendency to attenuated telomerase activity. ADSC-conditioned media (ADSC-CM) stimulated capillary-like tube formation by endothelial cells (EA.hy926), and this effect significantly decreased with the age of patients both with and without CAD. Angiogenic factors (vascular endothelial growth factor, placental growth factor, hepatocyte growth factor, angiopoetin-1, and angiogenin) in ADSC-CM measured by enzyme-linked immunosorbent assay significantly decreased with patient age, whereas levels of antiangiogenic factors thrombospondin-1 and endostatin did not. Expression of angiogenic factors in ADSCs did not change with patient age (real-time polymerase chain reaction); however, gene expression of factors related to extracellular proteolysis (urokinase and its receptor, plasminogen activator inhibitor-1) and urokinase-type plasminogen activator receptor surface expression increased in ADSCs from aged patients with CAD. ADSCs from aged patients both with and without CAD acquire aging characteristics, and their angiogenic potential declines because of decreasing proangiogenic factor secretion. This could restrict the effectiveness of autologous cell therapy with ADSCs in aged patients.

  8. GMP-compliant human adipose tissue-derived mesenchymal stem cells for cellular therapy.

    PubMed

    Aghayan, Hamid-Reza; Goodarzi, Parisa; Arjmand, Babak

    2015-01-01

    Stem cells, which can be derived from different sources, demonstrate promising therapeutic evidences for cellular therapies. Among various types of stem cell, mesenchymal stem cells are one of the most common stem cells that are used in cellular therapy. Human subcutaneous adipose tissue provides an easy accessible source of mesenchymal stem cells with some considerable advantages. Accordingly, various preclinical and clinical investigations have shown enormous potential of adipose-derived stromal cells in regenerative medicine. Consequently, increasing clinical applications of these cells has elucidated the importance of safety concerns regarding clinical transplantation. Therefore, clinical-grade preparation of adipose-derived stromal cells in accordance with current good manufacturing practice guidelines is an essential part of their clinical applications to ensure the safety, quality, characteristics, and identity of cell products. Additionally, GMP-compliant cell manufacturing involves several issues to provide a quality assurance system during translation from the basic stem cell sciences into clinical investigations and applications. On the other hand, advanced cellular therapy requires extensive validation, process control, and documentation. It also evidently elucidates the critical importance of production methods and probable risks. Therefore, implementation of a quality management and assurance system in accordance with GMP guidelines can greatly reduce these risks particularly in the higher-risk category or "more than minimally manipulated" products.

  9. Hyaluronan preserves the proliferation and differentiation potentials of long-term cultured murine adipose-derived stromal cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chen, P.-Y.; Huang, Lynn L.H.; Hsieh, H.-J.

    2007-08-17

    For long-term culture, murine adipose-derived stromal cells (mADSCs) at latter passages demonstrated a marked decline in proliferative activity, exhibited senescent morphology and reduced differentiation potentials, particularly osteogenesis. To extend the lifespan of mADSCs, two culture conditions containing hyaluronan (HA) was compared in our study, one as a culture medium supplement (SHA), and the other where HA was pre-coated on culture surface (CHA). mADSCs cultivated with SHA exhibited a prolonged lifespan, reduced cellular senescence, and enhanced osteogenic potential compared to regular culture condition (control). Upon CHA treatment, mADSCs tended to form cell aggregates with gradual growth profiles, while their differentiation activitiesmore » remained similar to SHA groups. After transferring mADSCs from CHA to control surface, they were shown to have an extended lifespan and an increase of osteogenic potential. Our results suggested that HA can be useful for preserving the proliferation and differentiation potentials of long-term cultured mADSCs.« less

  10. Tumor Associated Mesenchymal Stromal Cells Show Higher Immunosuppressive and Angiogenic Properties Compared to Adipose Derived MSCs.

    PubMed

    Langroudi, Ladan; Hassan, Zuhair Muhammad; Soleimani, Masoud; Hashemi, Seyed Mahmoud

    2015-12-01

    Differentiation, migratory properties and availability of Mesenchymal Stromal Cells (MSC) have become an important part of biomedical research. However, the functional heterogeneity of cells derived from different tissues has hampered providing definitive phenotypic markers for these cells. To characterize and compare the phenotype and cytokines of adipose derived MSCs (AD-MSCs) and tumoral-MSCs (T-MSCs) isolated from mammary tumors of BALB/c mice. Immunophenotyping and in vitro differentiation tests were used for MSC characterization. Cytokine and enzyme profiles were assessed using ELISA and Real-time PCR, respectively. T-MSCs expressed significantly higher levels of HLA-DR (p=0.04). Higher levels of PGE2 and COX-2 enzyme were also observed in T-MSCs (p=0.07 and p=0.00, respectively). Additionally, T-MSCs expressed higher levels of iNOS and MMP9 (p=0.01 and p=0.01, respectively). T-MSCs were also able to induce higher levels of proliferation and migration of HUVEC endothelial cells in wound scratch assay compared to AD-MSCs (p=0.015). Functional differences showed by the surface markers of MSCs, cytokine and enzyme production indicate the effect of different microenvironments on MSCs phenotype and function.

  11. Engineering vascularized soft tissue flaps in an animal model using human adipose-derived stem cells and VEGF+PLGA/PEG microspheres on a collagen-chitosan scaffold with a flow-through vascular pedicle.

    PubMed

    Zhang, Qixu; Hubenak, Justin; Iyyanki, Tejaswi; Alred, Erik; Turza, Kristin C; Davis, Greg; Chang, Edward I; Branch-Brooks, Cynthia D; Beahm, Elisabeth K; Butler, Charles E

    2015-12-01

    Insufficient neovascularization is associated with high levels of resorption and necrosis in autologous and engineered fat grafts. We tested the hypothesis that incorporating angiogenic growth factor into a scaffold-stem cell construct and implanting this construct around a vascular pedicle improves neovascularization and adipogenesis for engineering soft tissue flaps. Poly(lactic-co-glycolic-acid/polyethylene glycol (PLGA/PEG) microspheres containing vascular endothelial growth factor (VEGF) were impregnated into collagen-chitosan scaffolds seeded with human adipose-derived stem cells (hASCs). This setup was analyzed in vitro and then implanted into isolated chambers around a discrete vascular pedicle in nude rats. Engineered tissue samples within the chambers were harvested and analyzed for differences in vascularization and adipose tissue growth. In vitro testing showed that the collagen-chitosan scaffold provided a supportive environment for hASC integration and proliferation. PLGA/PEG microspheres with slow-release VEGF had no negative effect on cell survival in collagen-chitosan scaffolds. In vivo, the system resulted in a statistically significant increase in neovascularization that in turn led to a significant increase in adipose tissue persistence after 8 weeks versus control constructs. These data indicate that our model-hASCs integrated with a collagen-chitosan scaffold incorporated with VEGF-containing PLGA/PEG microspheres supported by a predominant vascular vessel inside a chamber-provides a promising, clinically translatable platform for engineering vascularized soft tissue flap. The engineered adipose tissue with a vascular pedicle could conceivably be transferred as a vascularized soft tissue pedicle flap or free flap to a recipient site for the repair of soft-tissue defects. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. Efficient generation of smooth muscle cells from adipose-derived stromal cells by 3D mechanical stimulation can substitute the use of growth factors in vascular tissue engineering.

    PubMed

    Parvizi, Mojtaba; Bolhuis-Versteeg, Lydia A M; Poot, André A; Harmsen, Martin C

    2016-07-01

    Occluding artery disease causes a high demand for bioartificial replacement vessels. We investigated the combined use of biodegradable and creep-free poly (1,3-trimethylene carbonate) (PTMC) with smooth muscle cells (SMC) derived by biochemical or mechanical stimulation of adipose tissue-derived stromal cells (ASC) to engineer bioartificial arteries. Biochemical induction of cultured ASC to SMC was done with TGF-β1 for 7d. Phenotype and function were assessed by qRT-PCR, immunodetection and collagen contraction assays. The influence of mechanical stimulation on non-differentiated and pre-differentiated ASC, loaded in porous tubular PTMC scaffolds, was assessed after culturing under pulsatile flow for 14d. Assays included qRT-PCR, production of extracellular matrix and scanning electron microscopy. ASC adhesion and TGF-β1-driven differentiation to contractile SMC on PTMC did not differ from tissue culture polystyrene controls. Mesenchymal and SMC markers were increased compared to controls. Interestingly, pre-differentiated ASC had only marginal higher contractility than controls. Moreover, in 3D PTMC scaffolds, mechanical stimulation yielded well-aligned ASC-derived SMC which deposited ECM. Under the same conditions, pre-differentiated ASC-derived SMC maintained their SMC phenotype. Our results show that mechanical stimulation can replace TGF-β1 pre-stimulation to generate SMC from ASC and that pre-differentiated ASC keep their SMC phenotype with increased expression of SMC markers. Copyright © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Generation of Human Adult Mesenchymal Stromal/Stem Cells Expressing Defined Xenogenic Vascular Endothelial Growth Factor Levels by Optimized Transduction and Flow Cytometry Purification

    PubMed Central

    Helmrich, Uta; Marsano, Anna; Melly, Ludovic; Wolff, Thomas; Christ, Liliane; Heberer, Michael; Scherberich, Arnaud; Martin, Ivan

    2012-01-01

    Adult mesenchymal stromal/stem cells (MSCs) are a valuable source of multipotent progenitors for tissue engineering and regenerative medicine, but may require to be genetically modified to widen their efficacy in therapeutic applications. For example, overexpression of the angiogenic factor vascular endothelial growth factor (VEGF) at controlled levels is an attractive strategy to overcome the crucial bottleneck of graft vascularization and to avoid aberrant vascular growth. Since the regenerative potential of MSCs is rapidly lost during in vitro expansion, we sought to develop an optimized technique to achieve high-efficiency retroviral vector transduction of MSCs derived from both adipose tissue (adipose stromal cells, ASCs) or bone marrow (BMSCs) and rapidly select cells expressing desired levels of VEGF with minimal in vitro expansion. The proliferative peak of freshly isolated human ASCs and BMSCs was reached 4 and 6 days after plating, respectively. By performing retroviral vector transduction at this time point, >90% efficiency was routinely achieved before the first passage. MSCs were transduced with vectors expressing rat VEGF164 quantitatively linked to a syngenic cell surface marker (truncated rat CD8). Retroviral transduction and VEGF expression did not affect MSC phenotype nor impair their in vitro proliferation and differentiation potential. Transgene expression was also maintained during in vitro differentiation. Furthermore, three subpopulations of transduced BMSCs homogeneously producing specific low, medium, and high VEGF doses could be prospectively isolated by flow cytometry based on the intensity of their CD8 expression already at the first passage. In conclusion, this optimized platform allowed the generation of populations of genetically modified MSCs, expressing specific levels of a therapeutic transgene, already at the first passage, thereby minimizing in vitro expansion and loss of regenerative potential. PMID:22070632

  14. Proinflammatory interleukins' production by adipose tissue-derived mesenchymal stromal cells: the impact of cell culture conditions and cell-to-cell interaction.

    PubMed

    Andreeva, Elena; Andrianova, Irina; Rylova, Julia; Gornostaeva, Aleksandra; Bobyleva, Polina; Buravkova, Ludmila

    2015-08-01

    The impact of culture conditions and interaction with activated peripheral blood mononuclear cells on the interleukin (IL) gene expression profile and proinflammatory IL-6 and IL-8 production by adipose-derived stromal cells (ASCs) was investigated. A microarray analysis revealed a wide range of IL genes either under standard (20%) or hypoxic (5%) O2 concentrations, some highly up-regulated at hypoxia. IL-6 and IL-8 production was inversely dependent on cell culture density. In early (first-third) passages, IL-6 and IL-8 concentration was higher at 20% O2 and in late (8th-12th) passages under 5% O2. Interaction between ASCs and mononuclear cells in indirect setting was accompanied with a significant decrease of IL-6 and did not result in the elevation of IL-8 concentration. Thereby, the production of proinflammatory interleukins (IL-6 and IL-8) may be affected by the ASC intrinsic features (density in culture, and duration of expansion), as well as by microenvironmental factors, such as hypoxia and the presence of blood-borne cells. These data are important for elucidating ASC paracrine activity regulation in vitro. They would also be on demand for optimisation of the cell therapy protocols, based on the application of ASC biologically active substances. SIGNIFICANCE PARAGRAPH: Ex vivo expansion is widely used for increasing the number of adipose-derived stromal cells (ASCs) and improving of their quality. The present study was designed to elucidate the particular factors influencing the interleukin production in ASCs. The presented data specified the parameters (i.e. cell density, duration of cultivation, hypoxia, etc.) that should be taken in mind when ASCs are intended to be used in protocols implying their paracrine activity. These data would be of considerable interest for researchers and clinicians working in the biomedical science. Copyright © 2015 John Wiley & Sons, Ltd.

  15. Dendritic cells maintain dermal adipose–derived stromal cells in skin fibrosis

    PubMed Central

    Chia, Jennifer J.; Zhu, Tong; Chyou, Susan; Dasoveanu, Dragos C.; Carballo, Camila; Tian, Sha; Magro, Cynthia M.; Rodeo, Scott; Spiera, Robert F.; Ruddle, Nancy H.; McGraw, Timothy E.; Browning, Jeffrey L.; Lafyatis, Robert; Gordon, Jessica K.; Lu, Theresa T.

    2016-01-01

    Scleroderma is a group of skin-fibrosing diseases for which there are no effective treatments. A feature of the skin fibrosis typical of scleroderma is atrophy of the dermal white adipose tissue (DWAT). Adipose tissue contains adipose-derived mesenchymal stromal cells (ADSCs) that have regenerative and reparative functions; however, whether DWAT atrophy in fibrosis is accompanied by ADSC loss is poorly understood, as are the mechanisms that might maintain ADSC survival in fibrotic skin. Here, we have shown that DWAT ADSC numbers were reduced, likely because of cell death, in 2 murine models of scleroderma skin fibrosis. The remaining ADSCs showed a partial dependence on dendritic cells (DCs) for survival. Lymphotoxin β (LTβ) expression in DCs maintained ADSC survival in fibrotic skin by activating an LTβ receptor/β1 integrin (LTβR/β1 integrin) pathway on ADSCs. Stimulation of LTβR augmented the engraftment of therapeutically injected ADSCs, which was associated with reductions in skin fibrosis and improved skin function. These findings provide insight into the effects of skin fibrosis on DWAT ADSCs, identify a DC-ADSC survival axis in fibrotic skin, and suggest an approach for improving mesenchymal stromal cell therapy in scleroderma and other diseases. PMID:27721238

  16. Osteogenic potential of human adipose-tissue-derived mesenchymal stromal cells cultured on 3D-printed porous structured titanium.

    PubMed

    Lewallen, Eric A; Jones, Dakota L; Dudakovic, Amel; Thaler, Roman; Paradise, Christopher R; Kremers, Hilal M; Abdel, Matthew P; Kakar, Sanjeev; Dietz, Allan B; Cohen, Robert C; Lewallen, David G; van Wijnen, Andre J

    2016-05-01

    Integration of porous metal prosthetics, which restore form and function of irreversibly damaged joints, into remaining healthy bone is critical for implant success. We investigated the biological properties of adipose-tissue-derived mesenchymal stromal/stem cells (AMSCs) and addressed their potential to alter the in vitro microenvironment of implants. We employed human AMSCs as a practical source for musculoskeletal applications because these cells can be obtained in large quantities, are multipotent, and have trophic paracrine functions. AMSCs were cultured on surgical-grade porous titanium disks as a model for orthopedic implants. We monitored cell/substrate attachment, cell proliferation, multipotency, and differentiation phenotypes of AMSCs upon osteogenic induction. High-resolution scanning electron microscopy and histology revealed that AMSCs adhere to the porous metallic surface. Compared to standard tissue culture plastic, AMSCs grown in the porous titanium microenvironment showed differences in temporal expression for genes involved in cell cycle progression (CCNB2, HIST2H4), extracellular matrix production (COL1A1, COL3A1), mesenchymal lineage identity (ACTA2, CD248, CD44), osteoblastic transcription factors (DLX3, DLX5, ID3), and epigenetic regulators (EZH1, EZH2). We conclude that metal orthopedic implants can be effectively seeded with clinical-grade stem/stromal cells to create a pre-conditioned implant. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Combined therapy for critical limb ischaemia: Biomimetic PLGA microcarriers potentiates the pro-angiogenic effect of adipose tissue stromal vascular fraction cells.

    PubMed

    Hoareau, Laurence; Fouchet, Florian; Planesse, Cynthia; Mirbeau, Sophie; Sindji, Laurence; Delay, Emmanuel; Roche, Régis; Montero-Menei, Claudia N; Festy, Franck

    2018-06-01

    We propose a regenerative solution in the treatment of critical limb ischaemia (CLI). Poly-lactic/glycolic acid microcarriers were prepared and coated with laminin to be sterilized through γ-irradiation of 25 kGy at low temperature. Stromal vascular fraction (SVF) cells were extracted through enzymatic digestion of adipose tissue. Streptozotocin-induced diabetic mice underwent arteriotomy and received an administration of SVF cells combined or not with biomimetic microcarriers. Functional evaluation of the ischaemic limb was then reported, and tissue reperfusion was evaluated through fluorescence molecular tomography. Microcarriers were stable and functional after γ-irradiation until at least 12 months of storage. Mice that received an injection of SVF cells in the ischaemic limb have 22% of supplementary blood supply within this limb 7 days after surgery compared with vehicle, whereas no difference was observed at Day 14. With the combined therapy, the improvement of blood flow is significantly higher compared with vehicle, of about 31% at Day 7 and of about 11% at Day 14. Injection of SVF cells induces a significant 27% decrease of necrosis compared with vehicle. This effect is more important when SVF cells were mixed with biomimetic microcarriers: -37% compared with control. Although SVF cells injection leads to a non-significant 22% proprioception recovery, the combined therapy induces a significant recovery of about 27% compared with vehicle. We show that the combination of SVF cells from adipose tissue with laminin-coated poly-lactic/glycolic acid microcarriers is efficient for critical limb ischaemia therapy in a diabetic mouse model. Copyright © 2018 John Wiley & Sons, Ltd.

  18. Effects of transplantation of adipose tissue-derived stem cells on prostate tumor.

    PubMed

    Lin, Guiting; Yang, Rong; Banie, Lia; Wang, Guifang; Ning, Hongxiu; Li, Long-Cheng; Lue, Tom F; Lin, Ching-Shwun

    2010-07-01

    Obesity is a risk factor for prostate cancer development, but the underlying mechanism is unknown. The present study tested the hypothesis that stromal cells of the adipose tissue might be recruited by cancer cells to help tumor growth. PC3 prostate cancer cells were transplanted into the subcutaneous space of the right flank of athymic mice. One week later, adipose tissue-derived stromal or stem cells (ADSC) or phosphate-buffered saline (PBS, as control) was transplanted similarly to the left flank. Tumor size was monitored for the next 34 days; afterwards, the mice were sacrificed and their tumors harvested for histological examination. The ability of PC3 cells to attract ADSC was tested by migration assay. The involvement of the CXCL12/CXCR4 axis was tested by migration assay in the presence of a specific inhibitor AMD3100. Throughout the entire course, the average size of PC3 tumors in ADSC-treated mice was larger than in PBS-treated mice. ADSC were identified inside the tumors of ADSC-treated mice; CXCR4 expression was also detected. Migration assay indicated the involvement of the CXCL12/CXCR4 axis in the migration of ADSC toward PC3 cells. Capillary density was twice as high in the tumors of ADSC-treated mice than in the tumors of PBS-treated mice. VEGF expression was similar but FGF2 expression was significantly higher in tumors of ADSC-treated mice than in the tumors of PBS-tread mice. Prostate cancer cells recruited ADSC by the CXCL12/CXCR4 axis. ADSC helps tumor growth by increasing tumor vascularity, and which was mediated by FGF2.

  19. Concise Review: Fat and Furious: Harnessing the Full Potential of Adipose‐Derived Stromal Vascular Fraction

    PubMed Central

    Dykstra, Jordan A.; Facile, Tiffany; Patrick, Ryan J.; Francis, Kevin R.; Milanovich, Samuel; Weimer, Jill M.

    2017-01-01

    Abstract Due to their capacity to self‐renew, proliferate and generate multi‐lineage cells, adult‐derived stem cells offer great potential for use in regenerative therapies to stop and/or reverse degenerative diseases such as diabetes, heart failure, Alzheimer's disease and others. However, these subsets of cells can be isolated from different niches, each with differing potential for therapeutic applications. The stromal vascular fraction (SVF), a stem cell enriched and adipose‐derived cell population, has garnered interest as a therapeutic in regenerative medicine due to its ability to secrete paracrine factors that accelerate endogenous repair, ease of accessibility and lack of identified major adverse effects. Thus, one can easily understand the rush to employ adipose‐derived SVF to treat human disease. Perhaps faster than any other cell preparation, SVF is making its way to clinics worldwide, while critical preclinical research needed to establish SVF safety, efficacy and optimal, standardized clinical procedures are underway. Here, we will provide an overview of the current knowledge driving this phenomenon, its regulatory issues and existing studies, and propose potential unmapped applications. Stem Cells Translational Medicine 2017;6:1096–1108 PMID:28186685

  20. The role of perivascular adipose tissue in vascular smooth muscle cell growth

    PubMed Central

    Miao, Chao-Yu; Li, Zhi-Yong

    2012-01-01

    Adipose tissue is the largest endocrine organ, producing various adipokines and many other substances. Almost all blood vessels are surrounded by perivascular adipose tissue (PVAT), which has not received research attention until recently. This review will discuss the paracrine actions of PVAT on the growth of underlying vascular smooth muscle cells (VSMCs). PVAT can release growth factors and inhibitors. Visfatin is the first identified growth factor derived from PVAT. Decreased adiponectin and increased tumour necrosis factor-α in PVAT play a pathological role for neointimal hyperplasia after endovascular injury. PVAT-derived angiotensin II, angiotensin 1–7, reactive oxygen species, complement component 3, NO and H2S have a paracrine action on VSMC contraction, endothelial or fibroblast function; however, their paracrine actions on VSMC growth remain to be directly verified. Factors such as monocyte chemoattractant protein-1, interleukin-6, interleukin-8, leptin, resistin, plasminogen activator inhibitor type-1, adrenomedullin, free fatty acids, glucocorticoids and sex hormones can be released from adipose tissue and can regulate VSMC growth. Most of them have been verified for their secretion by PVAT; however, their paracrine functions are unknown. Obesity, vascular injury, aging and infection may affect PVAT, causing adipocyte abnormality and inflammatory cell infiltration, inducing imbalance of PVAT-derived growth factors and inhibitors, leading to VSMC growth and finally resulting in development of proliferative vascular disease, including atherosclerosis, restenosis and hypertension. In the future, using cell-specific gene interventions and local treatments may provide definitive evidence for identification of key factor(s) involved in PVAT dysfunction-induced vascular disease and thus may help to develop new therapies. LINKED ARTICLES This article is part of a themed section on Fat and Vascular Responsiveness. To view the other articles in this section

  1. Comparison of in vivo immune responses following transplantation of vascularized and non-vascularized human dermo-epidermal skin substitutes.

    PubMed

    Klar, Agnes S; Biedermann, Thomas; Simmen-Meuli, Claudia; Reichmann, Ernst; Meuli, Martin

    2017-03-01

    Autologous bio-engineered dermo-epidermal skin substitutes (DESS) represent an alternative therapeutic option for a definitive treatment of skin defects in human patients. Largely, the interaction of host immune cells with transplanted DESS is considered to be essential for the granulation tissue formation, graft take, and its functionality. The aim of this study was to compare the spatiotemporal distribution and density of host-derived monocytes/macrophages and granulocytes in vascularized (vascDESS) versus non-vascularized DESS (non-vascDESS) in a rat model. Keratinocytes and the stromal vascular fraction (SVF) were derived from human skin or human adipose tissue, respectively. Human SVF containing both endothelial and mesenchymal/stromal progenitors was used to develop a vascularized collagen type I-based dermal component in vitro. The donor-matched, monolayer-expanded adipose stromal cells lacking endothelial cells were used as a negative control. Subsequently, human keratinocytes were seeded on top of hydrogels to build dermo-epidermal skin grafts. After transplantation onto full-thickness skin wounds on the back of immuno-incompetent rats, grafts were excised and analyzed after 1 and 3 weeks. The expression of distinct inflammatory cell markers specific for host-derived monocytes/macrophages (CD11b, CD68) or granulocytes (HIS48) was analyzed by immunofluorescence microscopy. All skin grafts were infiltrated by host-derived monocytes/macrophages (CD11b + , CD68 + ) and granulocytes (HIS48 + ) between 1-3 week post-transplantation. When compared to non-vascDESS, the vascDESS showed an increased granulocyte infiltration at all time points analyzed with the majority of cells scattered throughout the whole dermal part. Whereas a moderate number of rat monocytes/macrophages (CD11b + , CD68 + ) were found in vascDESS at 1 week, only a few cells were detected in non-vascDESS. We observed a time-dependent decrease of monocytes/macrophages in all transplants at 3

  2. Cryopreserved Off-the-Shelf Allogeneic Adipose-Derived Stromal Cells for Therapy in Patients with Ischemic Heart Disease and Heart Failure-A Safety Study.

    PubMed

    Kastrup, Jens; Haack-Sørensen, Mandana; Juhl, Morten; Harary Søndergaard, Rebekka; Follin, Bjarke; Drozd Lund, Lisbeth; Mønsted Johansen, Ellen; Ali Qayyum, Abbas; Bruun Mathiasen, Anders; Jørgensen, Erik; Helqvist, Steffen; Jørgen Elberg, Jens; Bruunsgaard, Helle; Ekblond, Annette

    2017-11-01

    The present first-in-human clinical trial evaluated the safety and feasibility of a newly developed and cryopreserved Cardiology Stem Cell Centre adipose-derived stromal cell (CSCC_ASC) product from healthy donors for intramyocardial injection in ten patients with ischemic heart disease and ischemic heart failure (IHF). Batches of CSCC_ASC were isolated from three healthy donors by liposuction from abdominal adipose tissue. Adipose mesenchymal stromal cells were culture expanded in bioreactors without the use of animal constituents, cryopreserved, and stored in vials in nitrogen dry-storage containers until use. Direct injection of CSCC_ASC into the myocardium did not cause any complications or serious adverse events related to either treatment or cell administration in a 6-month follow-up period. Four out of ten heart failure patients developed donor-specific de novo human leukocyte antigen (HLA) class I antibodies, and two out of ten patients had donor-specific HLA antibodies already at baseline. There were no clinical symptoms or changes in inflammatory parameters in the follow-up period that indicated an ongoing immune response. There was a tendency toward improvement in cardiac function after CSCC_ASC treatment at 6-month follow-up: left ventricular end systolic volume decreased and left ventricular ejection fraction increased. In addition, exercise capacity increased. These changes were independent of the presence or absence of HLA antibodies. It is concluded that the newly developed cryopreserved product CSCC_ASC from healthy donors was a safe and feasible treatment. We observed a tendency toward efficacy in patients with IHF. These findings have to be confirmed in larger placebo controlled clinical trials. Stem Cells Translational Medicine 2017;6:1963-1971. © 2017 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

  3. White Adipose Tissue Cells Are Recruited by Experimental Tumors and Promote Cancer Progression in Mouse Models

    PubMed Central

    Zhang, Yan; Daquinag, Alexes; Traktuev, Dmitry O.; Amaya-Manzanares, Felipe; Simmons, Paul J.; March, Keith L.; Pasqualini, Renata; Arap, Wadih; Kolonin, Mikhail G.

    2010-01-01

    The connection between obesity and accelerated cancer progression has been established, but the mediating mechanisms are not well understood. We have shown that stromal cells from white adipose tissue (WAT) cooperate with the endothelium to promote blood vessel formation through the secretion of soluble trophic factors. Here, we hypothesize that WAT directly mediates cancer progression by serving as a source of cells that migrate to tumors and promote neovascularization. To test this hypothesis, we have evaluated the recruitment of WAT-derived cells by tumors and the effect of their engraftment on tumor growth by integrating a transgenic mouse strain engineered for expansion of traceable cells with established allograft and xenograft cancer models. Our studies show that entry of adipose stromal and endothelial cells into systemic circulation leads to their homing to and engraftment into tumor stroma and vasculature, respectively. We show that recruitment of adipose stromal cells by tumors is sufficient to promote tumor growth. Finally, we show that migration of stromal and vascular progenitor cells from WAT grafts to tumors is also associated with acceleration of cancer progression. These results provide a biological insight for the clinical association between obesity and cancer, thus outlining potential avenues for preventive and therapeutic strategies. PMID:19491274

  4. Human adipose tissue-derived tenomodulin positive subpopulation of stem cells: A promising source of tendon progenitor cells.

    PubMed

    Gonçalves, A I; Gershovich, P M; Rodrigues, M T; Reis, R L; Gomes, M E

    2018-03-01

    Cell-based therapies are of particular interest for tendon and ligament regeneration given the low regenerative potential of these tissues. Adipose tissue is an abundant source of stem cells, which may be employed for the healing of tendon lesions. However, human adult multipotent adipose-derived stem cells (hASCs) isolated from the stromal vascular fraction of adipose tissue originate highly heterogeneous cell populations that hinder their use in specific tissue-oriented applications. In this study, distinct subpopulations of hASCs were immunomagnetic separated and their tenogenic differentiation capacity evaluated in the presence of several growth factors (GFs), namely endothelial GF, basic-fibroblast GF, transforming GF-β1 and platelet-derived GF-BB, which are well-known regulators of tendon development, growth and healing. Among the screened hASCs subpopulations, tenomodulin-positive cells were shown to be more promising for tenogenic applications and therefore this subpopulation was further studied, assessing tendon-related markers (scleraxis, tenomodulin, tenascin C and decorin) both at gene and protein level. Additionally, the ability for depositing collagen type I and III forming extracellular matrix structures were weekly assessed up to 28 days. The results obtained indicated that tenomodulin-positive cells exhibit phenotypical features of tendon progenitor cells and can be biochemically induced towards tenogenic lineage, demonstrating that this subset of hASCs can provide a reliable source of progenitor cells for therapies targeting tendon regeneration. Copyright © 2017 John Wiley & Sons, Ltd.

  5. Storage effect on viability and biofunctionality of human adipose tissue-derived stromal cells.

    PubMed

    Falah, Mizied; Rayan, Anwar; Srouji, Samer

    2015-09-01

    In our recent studies, the transplantation of human adipose tissue-derived stromal cells (ASCs) has shown promise for treatment of diseases related to bone and joint disorders. For the current clinical applications, ASCs were formulated and suspended in PlasmaLyte A supplemented with heparin, glucose and human serum albumin, balanced to pH 7.4 with sodium bicarbonate. This cell solution constitutes 20% of the overall transplanted mixture and is supplemented with hyaluronic acid (60%) and OraGraft particles (20%). We intended to investigate the effect of this transplantation mixture on the viability and biofunctionality of ASCs in bone formation. Freshly harvested cells were resuspended and incubated in the indicated mixture for up to 48 h at 4°C. Cell viability was assessed using trypan blue and AlamarBlue, and cell functionality was determined by quantifying their adhesion rate in vitro and bone formation in an ectopic mouse model. More than 80% of the ASCs stored in the transplantation mixture were viable for up to 24 h. Cell viability beyond 24 h in storage decreased to approximately 50%. In addition, an equal degree of bone formation was observed between the cells transplanted following incubation in transplantation mixture for up to 24 h and zero-time non-incubated cells (control). The viability and functionality of ASCs stored in the presented formulation will make such cell therapy accessible to larger and more remote populations. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  6. Notch3 is involved in adipogenesis of human adipose-derived stromal/stem cells.

    PubMed

    Sandel, Demi A; Liu, Mengcheng; Ogbonnaya, Ngozi; Newman, Jamie J

    2018-07-01

    Human adipose-derived stromal/stem cells (hASCs) have tremendous therapeutic potential and the ability to offer insight into human development and disease. Here we subject human ASCs to siRNA-mediated knockdown of Notch3 cultured under both self-renewing and adipogenic differentiation conditions. Self-renewal was monitored by assessing viability and proliferation rates through staining and alamarBlue assays, respectively. Adipogenesis was measured through Oil-Red O staining, western blot, and quantitative real-time RT-PCR that determined expression levels of multipotency and adipogenic markers over time. Notch3 was expressed in self-renewing hASCs but knockdown, as validated by qRT-PCR and western blot, showed no impact on cell viability, as measured through live-dead staining, or cell proliferation rates, as measured through alamarBlue assays. However, although Notch3 expression was observed to increase during adipogenesis, in the absence of Notch3 there was a significant increase in hASC adipogenesis as demonstrated through an increased number of lipid vesicles, and increased expression of adipogenic markers ppar-γ, adiponectin, fabp4, and plin2. Although Notch3 is only one of four Notch receptors expressed on the surface of hASCs, this receptor appears important for proper regulation of adipogenic differentiation, possibly serving as a negative regulator to prevent inappropriate adipogenesis or promote other lineage commitments of ASCs. Copyright © 2018 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  7. Hypoxia induced VEGF synthesis in visceral adipose depots of obese diabetic patients.

    PubMed

    Fusaru, Ana Marina; Pisoschi, Cătălina Gabriela; Bold, Adriana; Taisescu, C; Stănescu, R; Hîncu, Mihaela; Crăiţoiu, Stefania; Baniţă, Ileana Monica

    2012-01-01

    VEGF is one the pro-inflammatory adipokines synthesized by the "adipose secretoma" of obese subjects as a response to hypoxic conditions; but the main function of VEGF is angiogenesis, being recognized as the most important factor increasing blood capillaries in the adipose tissue by stimulating endothelial cell growth. In this paper, we propose a comparative study of the vascular response to VEGF synthesis in the subcutaneous and central-peritoneal adipose depots in lean, obese and obese diabetic patients. We used CD31 to label the endothelial cells in order to evaluate the response of the vascular network to VEGF synthesis. Our results showed an increase of VEGF protein synthesis in obese and obese-diabetic patients compared to lean subjects where the protein was absent. The positivity for VEGF in obese diabetic samples was observed in numerous structures from the adipose depots, both in the stromal vascular fraction--blood vessels and stromal cells--as well as in the cytoplasm of adipocytes. Positivity in the vascular wall was observed more frequently in areas of perivascular and intralobular fibrosis. Obese and diabetic patients showed similar incidence of CD31 immunoreactivity with lean subjects in both subcutaneous and peritoneal depots. In conclusion, human adipose depots show a different incidence of VEGF positive cells in relation with their disposal and the metabolic status. VEGF synthesis in visceral adipose tissue is inefficient being not followed by angiogenesis to counterbalance tissue hypoxia. We suggest that may be a pathogenic link between the degrees of intralobular fibrosis in adipose depots and VEGF expression.

  8. CD54+ rabbit adipose-derived stem cells overexpressing HIF-1α facilitate vascularized fat flap regeneration

    PubMed Central

    Liang, Zhi-Jie; Huang, Min-Hong; Peng, Qi-Liu; Zou, Dong-Hua; Gu, Rong-He; Xu, Fang-Tian; Gao, Hui; Chen, Zhen-Dong; Chi, Guang-Yi; Wei, Zhong-Heng; Chen, Li; Li, Hong-Mian

    2017-01-01

    Fat flap transplantation is frequently performed in patients suffering from soft tissue defects resulting from disease or trauma. This study explored the feasibility of constructing vascularized fat flaps using rabbit adipose-derived stem cells (rASCs) and collagen scaffolds in a rabbit model. We evaluated rASCs proliferation, paracrine function, adipogenesis, vascularization, and CD54 expression, with or without HIF-1α transfection in vitro and in vivo. We observed that adipogenic differentiation potential was greater in rASCs with high CD54 expression (CD54+rASCs) than in those with low expression (CD54–rASCs), both in vitro and in vivo. HIF-1α overexpression not only augmented this effect, but also enhanced cell proliferation and paracrine function in vitro. We also demonstrated that HIF-1α-transfected CD54+rASCs showed enhanced paracrine function and adipogenic capacity, and that paracrine function increases expression of angiogenesis-related markers. Thus, CD54+rASCs overexpressing HIF-1α enhanced large volume vascularized fat flap regeneration in rabbits, suggesting CD54 may be an ideal candidate marker for ASCs adipogenic differentiation. PMID:28423354

  9. Analysis of the Material Properties of Early Chondrogenic Differentiated Adipose-Derived Stromal Cells (ASC) Using an in vitro Three-dimensional Micromass Culture System

    PubMed Central

    Xu, Yue; Balooch, Guive; Chiou, Michael; Bekerman, Elena; Ritchie, Robert O.; Longaker, Michael T.

    2009-01-01

    Cartilage is an avascular tissue with only a limited potential to heal and chondrocytes in vitro have poor proliferative capacity. Recently, adipose-derived stromal cells (ASC) have demonstrated a great potential for application to tissue engineering due to their ability to differentiate into cartilage, bone, and fat. In this study, we have utilized a high density three-dimensional (3D) micromass model system of early chondrogenesis with ASC. The material properties of these micromasses showed a significant increase in dynamic and static elastic modulus during the early chondrogenic differentiation process. These data suggest that the 3D micromass culture system represents an in vitro model of early chondrogenesis with dynamic cell signaling interactions associated with the mechanical properties of chondrocyte differentiation. PMID:17543281

  10. Analysis of the material properties of early chondrogenic differentiated adipose-derived stromal cells (ASC) using an in vitro three-dimensional micromass culture system

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Xu, Yue; Balooch, Guive; Chiou, Michael

    2007-07-27

    Cartilage is an avascular tissue with only a limited potential to heal and chondrocytes in vitro have poor proliferative capacity. Recently, adipose-derived stromal cells (ASC) have demonstrated a great potential for application to tissue engineering due to their ability to differentiate into cartilage, bone, and fat. In this study, we have utilized a high density three-dimensional (3D) micromass model system of early chondrogenesis with ASC. The material properties of these micromasses showed a significant increase in dynamic and static elastic modulus during the early chondrogenic differentiation process. These data suggest that the 3D micromass culture system represents an in vitromore » model of early chondrogenesis with dynamic cell signaling interactions associated with the mechanical properties of chondrocyte differentiation.« less

  11. Adipocyte-derived factors in age-related dementia and their contribution to vascular and Alzheimer pathology.

    PubMed

    Ishii, Makoto; Iadecola, Costantino

    2016-05-01

    Age-related dementia is increasingly recognized as having a mixed pathology, with contributions from both cerebrovascular factors and pathogenic factors associated with Alzheimer's disease (AD). Furthermore, there is accumulating evidence that vascular risk factors in midlife, e.g., obesity, diabetes, and hypertension, increase the risk of developing late-life dementia. Since obesity and changes in body weight/adiposity often drive diabetes and hypertension, understanding the relationship between adiposity and age-related dementia may reveal common underlying mechanisms. Here we offer a brief appraisal of how changes in body weight and adiposity are related to both AD and dementia on vascular basis, and examine the involvement of two key adipocyte-derived hormones: leptin and adiponectin. The evidence suggests that in midlife increased body weight/adiposity and subsequent changes in adipocyte-derived hormones may increase the long-term susceptibility to dementia. On the other hand, later in life, decreases in body weight/adiposity and related hormonal changes are early manifestations of disease that precede the onset of dementia and may promote AD and vascular pathology. Understanding the contribution of adiposity to age-related dementia may help identify the underlying pathological mechanisms common to both vascular dementia and AD, and provide new putative targets for early diagnosis and therapy. This article is part of a Special Issue entitled: Vascular Contributions to Cognitive Impairment and Dementia, edited by M. Paul Murphy, Roderick A. Corriveau and Donna M. Wilcock. Copyright © 2015 Elsevier B.V. All rights reserved.

  12. Comparative Efficacy of Autologous Stromal Vascular Fraction and Autologous Adipose-Derived Mesenchymal Stem Cells Combined With Hyaluronic Acid for the Treatment of Sheep Osteoarthritis.

    PubMed

    Lv, Xiaoteng; He, Jiyin; Zhang, Xue; Luo, Xuan; He, Na; Sun, Zhongwei; Xia, Huitang; Liu, Victor; Zhang, Li; Lin, Xiangming; Lin, Liping; Yin, Huabin; Jiang, Dong; Cao, Wei; Wang, Richard; Zhou, Guangdong; Wang, Wen

    2018-01-01

    The current study explored whether intra-articular (IA) injection of autologous adipose mesenchymal stem cells (ASCs) combined with hyaluronic acid (HA) achieved better therapeutic efficacy than autologous stromal vascular fraction (SVF) combined with HA to prevent osteoarthritis (OA) progression and determined how long autologous ASCs combined with HA must remain in the joint to observe efficacy. OA models were established by performing anterior cruciate ligament transection (ACLT) and medial meniscectomy (MM). Autologous SVF (1×10 7 mononuclear cells), autologous low-dose ASCs (1×10 7 ), and autologous high-dose ASCs (5×10 7 ) combined with HA, and HA alone, or saline alone were injected into the OA model animals at 12 and 15 weeks after surgery, respectively. Compared with SVF+HA treatment, low-dose ASC+HA treatment yielded better magnetic resonance imaging (MRI) scores and macroscopic results, while the cartilage thickness of the tibial plateau did not differ between low, high ASC+HA and SVF+HA treatments detected by micro-computed tomography (µCT). Immunohistochemistry revealed that high-dose ASC+HA treatment rescued hypertrophic chondrocytes expressing collagen X in the deep area of articular cartilage. Western blotting analysis indicated the high- and low-dose ASC+HA groups expressed more collagen X than did the SVF+HA group. Enzyme-linked immunosorbent assay showed treatment with both ASC+HA and SVF+HA resulted in differing anti-inflammatory and trophic effects. Moreover, superparamagnetic iron oxide particle (SPIO)-labeled autologous ASC signals were detected by MRI at 2 and 18 weeks post-injection and were found in the lateral meniscus at 2 weeks and in the marrow cavity of the femoral condyle at 18 weeks post-injection. Thus, IA injection of autologous ASC+HA may demonstrate better efficacy than autologous SVF+HA in blocking OA progression and promoting cartilage regeneration, and autologous ASCs (5×10 7 cells) combined with HA potentially survive

  13. Engineered, axially-vascularized osteogenic grafts from human adipose-derived cells to treat avascular necrosis of bone in a rat model.

    PubMed

    Ismail, Tarek; Osinga, Rik; Todorov, Atanas; Haumer, Alexander; Tchang, Laurent A; Epple, Christian; Allafi, Nima; Menzi, Nadia; Largo, René D; Kaempfen, Alexandre; Martin, Ivan; Schaefer, Dirk J; Scherberich, Arnaud

    2017-11-01

    Avascular necrosis of bone (AVN) leads to sclerosis and collapse of bone and joints. The standard of care, vascularized bone grafts, is limited by donor site morbidity and restricted availability. The aim of this study was to generate and test engineered, axially vascularized SVF cells-based bone substitutes in a rat model of AVN. SVF cells were isolated from lipoaspirates and cultured onto porous hydroxyapatite scaffolds within a perfusion-based bioreactor system for 5days. The resulting constructs were inserted into devitalized bone cylinders mimicking AVN-affected bone. A ligated vascular bundle was inserted upon subcutaneous implantation of constructs in nude rats. After 1 and 8weeks in vivo, bone formation and vascularization were analyzed. Newly-formed bone was found in 80% of SVF-seeded scaffolds after 8weeks but not in unseeded controls. Human ALU+cells in the bone structures evidenced a direct contribution of SVF cells to bone formation. A higher density of regenerative, M2 macrophages was observed in SVF-seeded constructs. In both experimental groups, devitalized bone was revitalized by vascularized tissue after 8 weeks. SVF cells-based osteogenic constructs revitalized fully necrotic bone in a challenging AVN rat model of clinically-relevant size. SVF cells contributed to accelerated initial vascularization, to bone formation and to recruitment of pro-regenerative endogenous cells. Avascular necrosis (AVN) of bone often requires surgical treatment with autologous bone grafts, which is surgically demanding and restricted by significant donor site morbidity and limited availability. This paper describes a de novo engineered axially-vascularized bone graft substitute and tests the potential to revitalize dead bone and provide efficient new bone formation in a rat model. The engineering of an osteogenic/vasculogenic construct of clinically-relevant size with stromal vascular fraction of human adipose, combined to an arteriovenous bundle is described. This

  14. Different wound healing properties of dermis, adipose, and gingiva mesenchymal stromal cells.

    PubMed

    Boink, Mireille A; van den Broek, Lenie J; Roffel, Sanne; Nazmi, Kamran; Bolscher, Jan G M; Gefen, Amit; Veerman, Enno C I; Gibbs, Susan

    2016-01-01

    Oral wounds heal faster and with better scar quality than skin wounds. Deep skin wounds where adipose tissue is exposed, have a greater risk of forming hypertrophic scars. Differences in wound healing and final scar quality might be related to differences in mesenchymal stromal cells (MSC) and their ability to respond to intrinsic (autocrine) and extrinsic signals, such as human salivary histatin, epidermal growth factor, and transforming growth factor beta1. Dermis-, adipose-, and gingiva-derived MSC were compared for their regenerative potential with regards to proliferation, migration, and matrix contraction. Proliferation was assessed by cell counting and migration using a scratch wound assay. Matrix contraction and alpha smooth muscle actin was assessed in MSC populated collagen gels, and also in skin and gingival full thickness tissue engineered equivalents (reconstructed epithelium on MSC populated matrix). Compared to skin-derived MSC, gingiva MSC showed greater proliferation and migration capacity, and less matrix contraction in full thickness tissue equivalents, which may partly explain the superior oral wound healing. Epidermal keratinocytes were required for enhanced adipose MSC matrix contraction and alpha smooth muscle actin expression, and may therefore contribute to adverse scarring in deep cutaneous wounds. Histatin enhanced migration without influencing proliferation or matrix contraction in all three MSC, indicating that salivary peptides may have a beneficial effect on wound closure in general. Transforming growth factor beta1 enhanced contraction and alpha smooth muscle actin expression in all three MSC types when incorporated into collagen gels. Understanding the mechanisms responsible for the superior oral wound healing will aid us to develop advanced strategies for optimal skin regeneration, wound healing and scar formation. © 2015 by the Wound Healing Society.

  15. Decellularized skin/adipose tissue flap matrix for engineering vascularized composite soft tissue flaps.

    PubMed

    Zhang, Qixu; Johnson, Joshua A; Dunne, Lina W; Chen, Youbai; Iyyanki, Tejaswi; Wu, Yewen; Chang, Edward I; Branch-Brooks, Cynthia D; Robb, Geoffrey L; Butler, Charles E

    2016-04-15

    Using a perfusion decellularization protocol, we developed a decellularized skin/adipose tissue flap (DSAF) comprising extracellular matrix (ECM) and intact vasculature. Our DSAF had a dominant vascular pedicle, microcirculatory vascularity, and a sensory nerve network and retained three-dimensional (3D) nanofibrous structures well. DSAF, which was composed of collagen and laminin with well-preserved growth factors (e.g., vascular endothelial growth factor, basic fibroblast growth factor), was successfully repopulated with human adipose-derived stem cells (hASCs) and human umbilical vein endothelial cells (HUVECs), which integrated with DSAF and formed 3D aggregates and vessel-like structures in vitro. We used microsurgery techniques to re-anastomose the recellularized DSAF into nude rats. In vivo, the engineered flap construct underwent neovascularization and constructive remodeling, which was characterized by the predominant infiltration of M2 macrophages and significant adipose tissue formation at 3months postoperatively. Our results indicate that DSAF co-cultured with hASCs and HUVECs is a promising platform for vascularized soft tissue flap engineering. This platform is not limited by the flap size, as the entire construct can be immediately perfused by the recellularized vascular network following simple re-integration into the host using conventional microsurgical techniques. Significant soft tissue loss resulting from traumatic injury or tumor resection often requires surgical reconstruction using autologous soft tissue flaps. However, the limited availability of qualitative autologous flaps as well as the donor site morbidity significantly limits this approach. Engineered soft tissue flap grafts may offer a clinically relevant alternative to the autologous flap tissue. In this study, we engineered vascularized soft tissue free flap by using skin/adipose flap extracellular matrix scaffold (DSAF) in combination with multiple types of human cells. Following

  16. Inactivation of adipose angiotensinogen reduces adipose tissue macrophages and increases metabolic activity.

    PubMed

    LeMieux, Monique J; Ramalingam, Latha; Mynatt, Randall L; Kalupahana, Nishan S; Kim, Jung Han; Moustaïd-Moussa, Naïma

    2016-02-01

    The adipose renin-angiotensin system (RAS) has been linked to obesity-induced inflammation, though mechanisms are not completely understood. In this study, adipose-specific angiotensinogen knockout mice (Agt-KO) were generated to determine whether Agt inactivation reduces inflammation and alters the metabolic profile of the Agt-KO mice compared to wild-type (WT) littermates. Adipose tissue-specific Agt-KO mice were created using the Cre-LoxP system with both Agt-KO and WT littermates fed either a low-fat or high-fat diet to assess metabolic changes. White adipose tissue was used for gene/protein expression analyses and WAT stromal vascular cells for metabolic extracellular flux assays. No significant differences were observed in body weight or fat mass between both genotypes on either diet. However, improved glucose clearance was observed in Agt-KO compared to WT littermates, consistent with higher expression of genes involved in insulin signaling, glucose transport, and fatty acid metabolism. Furthermore, Agt inactivation reduced total macrophage infiltration in Agt-KO mice fed both diets. Lastly, stroma vascular cells from Agt-KO mice revealed higher metabolic activity compared to WT mice. These findings indicate that adipose-specific Agt inactivation leads to reduced adipose inflammation and increased glucose tolerance mediated in part via increased metabolic activity of adipose cells. © 2015 The Obesity Society.

  17. Adipose-derived stromal cells enhance auditory neuron survival in an animal model of sensory hearing loss.

    PubMed

    Schendzielorz, Philipp; Vollmer, Maike; Rak, Kristen; Wiegner, Armin; Nada, Nashwa; Radeloff, Katrin; Hagen, Rudolf; Radeloff, Andreas

    2017-10-01

    A cochlear implant (CI) is an electronic prosthesis that can partially restore speech perception capabilities. Optimum information transfer from the cochlea to the central auditory system requires a proper functioning auditory nerve (AN) that is electrically stimulated by the device. In deafness, the lack of neurotrophic support, normally provided by the sensory cells of the inner ear, however, leads to gradual degeneration of auditory neurons with undesirable consequences for CI performance. We evaluated the potential of adipose-derived stromal cells (ASCs) that are known to produce neurotrophic factors to prevent neural degeneration in sensory hearing loss. For this, co-cultures of ASCs with auditory neurons have been studied, and autologous ASC transplantation has been performed in a guinea pig model of gentamicin-induced sensory hearing loss. In vitro ASCs were neuroprotective and considerably increased the neuritogenesis of auditory neurons. In vivo transplantation of ASCs into the scala tympani resulted in an enhanced survival of auditory neurons. Specifically, peripheral AN processes that are assumed to be the optimal activation site for CI stimulation and that are particularly vulnerable to hair cell loss showed a significantly higher survival rate in ASC-treated ears. ASC transplantation into the inner ear may restore neurotrophic support in sensory hearing loss and may help to improve CI performance by enhanced AN survival. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  18. Endocrine modulators of mouse subcutaneous adipose tissue beige adipocyte markers

    USDA-ARS?s Scientific Manuscript database

    The stromal vascular fraction (SVF) of subcutaneous adipose tissue contains precursors that can give rise to beige adipocytes. Beige adipocytes are characterized by the expression of specific markers, but it is not clear which markers best evaluate beige adipocyte differentiation. Both regulators of...

  19. Defining adipose tissue-derived stem cells in tissue and in culture.

    PubMed

    Lin, Ching-Shwun; Xin, Zhong-Cheng; Deng, Chun-Hua; Ning, Hongxiu; Lin, Guiting; Lue, Tom F

    2010-06-01

    Adipose tissue-derived stem cells (ADSC) are routinely isolated from the stromal vascular fraction (SVF) of homogenized adipose tissue. Similar to other types of mesenchymal stem cells (MSC), ADSC remain difficult to define due to the lack of definitive cellular markers. Still, many types of MSC, including ADSC, have been shown to reside in a perivascular location, and increasing evidence shows that both MSC and ADSC may in fact be vascular stem cells (VSC). Locally, these cells differentiate into smooth muscle and endothelial cells that are assembled into newly formed blood vessels during angiogenesis and neovasculogenesis. Additionally, MSC or ADSC can also differentiate into tissue cells such as adipocytes in the adipose tissue. Systematically, MSC or ADSC are recruited to injury sites where they participate in the repair/regeneration of the injured tissue. Due to the vasculature's dynamic capacity for growth and multipotential nature for diversification, VSC in tissue are individually at various stages and on different paths of differentiation. Therefore, when isolated and put in culture, these cells are expected to be heterogeneous in marker expression, renewal capacity, and differentiation potential. Although this heterogeneity of VSC does impose difficulties and cause confusions in basic science studies, its impact on the development of VSC as a therapeutic cell source has not been as apparent, as many preclinical and clinical trials have reported favorable outcomes. With this understanding, ADSC are generally defined as CD34+CD31- although loss of CD34 expression in culture is well documented. In adipose tissue, CD34 is localized to the intima and adventitia of blood vessels but not the media where cells expressing alpha-smooth muscle actin (SMA) exist. By excluding the intima, which contains the CD34+CD31+ endothelial cells, and the media, which contains the CD34-CD31- smooth muscle cells, it leaves the adventitia as the only possible location for the CD34

  20. Chitosan-assisted differentiation of porcine adipose tissue-derived stem cells into glucose-responsive insulin-secreting clusters

    PubMed Central

    Lin, Yuan-Yu; Chen, Yu-Jen; Liu, Bing-Hsien; Wong, Shiu-Chung; Wu, Cheng-Yu; Chang, Yun-Tsui; Chou, Han-Yi E.

    2017-01-01

    The unique advantage of easy access and abundance make the adipose-derived stem cells (ADSCs) a promising system of multipotent cells for transplantation and regenerative medicine. Among the available sources, porcine ADSCs (pADSCs) deserve especial attention due to the close resemblance of human and porcine physiology, as well as for the upcoming availability of humanized porcine models. Here, we report on the isolation and conversion of pADSCs into glucose-responsive insulin-secreting cells. We used the stromal-vascular fraction of the dorsal subcutaneous adipose from 9-day-old male piglets to isolate pADSCs, and subjected the cells to an induction scheme for differentiation on chitosan-coated plates. This one-step procedure promoted differentiation of pADSCs into pancreatic islet-like clusters (PILC) that are characterized by the expression of a repertoire of pancreatic proteins, including pancreatic and duodenal homeobox (Pdx-1), insulin gene enhancer protein (ISL-1) and insulin. Upon glucose challenge, these PILC secreted high amounts of insulin in a dose-dependent manner. Our data also suggest that chitosan plays roles not only to enhance the differentiation potential of pADSCs, but also to increase the glucose responsiveness of PILCs. Our novel approach is, therefore, of great potential for transplantation-based amelioration of type 1 diabetes. PMID:28253305

  1. Immunomodulatory Effects of Adipose-Derived Stem Cells: Fact or Fiction?

    PubMed Central

    Leto Barone, Angelo A.; Khalifian, Saami; Lee, W. P. Andrew; Brandacher, Gerald

    2013-01-01

    Adipose-derived stromal cells (ASCs) are often referred to as adipose-derived stem cells due to their potential to undergo multilineage differentiation. Their promising role in tissue engineering and ability to modulate the immune system are the focus of extensive research. A number of clinical trials using ASCs are currently underway to better understand the role of such cell niche in enhancing or suppressing the immune response. If governable, such immunoregulatory role would find application in several conditions in which an immune response is present (i.e., autoimmune conditions) or feared (i.e., solid organ or reconstructive transplantation). Although allogeneic ASCs have been shown to prevent acute GvHD in both preclinical and clinical studies, their potential warrants further investigation. Well-designed and standardized clinical trials are necessary to prove the role of ASCs in the treatment of immune disorders or prevention of tissue rejection. In this paper we analyze the current literature on the role of ASCs in immunomodulation in vitro and in vivo and discuss their potential in regulating the immune system in the context of transplantation. PMID:24106704

  2. Cell Therapy Applications for Retinal Vascular Diseases: Diabetic Retinopathy and Retinal Vein Occlusion.

    PubMed

    Park, Susanna S

    2016-04-01

    Retinal vascular conditions, such as diabetic retinopathy and retinal vein occlusion, remain leading causes of vision loss. No therapy exists to restore vision loss resulting from retinal ischemia and associated retinal degeneration. Tissue regeneration is possible with cell therapy. The goal would be to restore or replace the damaged retinal vasculature and the retinal neurons that are damaged and/or degenerating from the hypoxic insult. Currently, various adult cell therapies have been explored as potential treatment. They include mesenchymal stem cells, vascular precursor cells (i.e., CD34+ cells, hematopoietic cells or endothelial progenitor cells), and adipose stromal cells. Preclinical studies show that all these cells have a paracrine trophic effect on damaged ischemic tissue, leading to tissue preservation. Endothelial progenitor cells and adipose stromal cells integrate into the damaged retinal vascular wall in preclinical models of diabetic retinopathy and ischemia-reperfusion injury. Mesenchymal stem cells do not integrate as readily but appear to have a primary paracrine trophic effect. Early phase clinical trials have been initiated and ongoing using mesenchymal stem cells or autologous bone marrow CD34+ cells injected intravitreally as potential therapy for diabetic retinopathy or retinal vein occlusion. Adipose stromal cells or pluripotent stem cells differentiated into endothelial colony-forming cells have been explored in preclinical studies and show promise as possible therapies for retinal vascular disorders. The relative safety or efficacy of these various cell therapies for treating retinal vascular disorders have yet to be determined.

  3. Umbilical Cord Blood-Derived Mesenchymal Stem Cells Inhibit, But Adipose Tissue-Derived Mesenchymal Stem Cells Promote, Glioblastoma Multiforme Proliferation

    PubMed Central

    Akimoto, Keiko; Kimura, Kenichi; Nagano, Masumi; Takano, Shingo; To'a Salazar, Georgina; Yamashita, Toshiharu

    2013-01-01

    Mesenchymal stem cells (MSCs) possess self-renewal and multipotential differentiation abilities, and they are thought to be one of the most reliable stem cell sources for a variety of cell therapies. Recently, cell therapy using MSCs has been studied as a novel therapeutic approach for cancers that show refractory progress and poor prognosis. MSCs from different tissues have different properties. However, the effect of different MSC properties on their application in anticancer therapies has not been thoroughly investigated. In this study, to characterize the anticancer therapeutic application of MSCs from different sources, we established two different kinds of human MSCs: umbilical cord blood-derived MSCs (UCB-MSCs) and adipose-tissue-derived MSCs (AT-MSCs). We used these MSCs in a coculture assay with primary glioblastoma multiforme (GBM) cells to analyze how MSCs from different sources can inhibit GBM growth. We found that UCB-MSCs inhibited GBM growth and caused apoptosis, but AT-MSCs promoted GBM growth. Terminal deoxynucleotidyl transferase-mediated biotinylated UTP nick-end labeling assay clearly demonstrated that UCB-MSCs promoted apoptosis of GBM via tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). TRAIL was expressed more highly by UCB-MSCs than by AT-MSCs. Higher mRNA expression levels of angiogenic factors (vascular endothelial growth factor, angiopoietin 1, platelet-derived growth factor, and insulin-like growth factor) and stromal-derived factor-1 (SDF-1/CXCL12) were observed in AT-MSCs, and highly vascularized tumors were developed when AT-MSCs and GBM were cotransplanted. Importantly, CXCL12 inhibited TRAIL activation of the apoptotic pathway in GBM, suggesting that AT-MSCs may support GBM development in vivo by at least two distinct mechanisms—promoting angiogenesis and inhibiting apoptosis. The opposite effects of AT-MSCs and UCB-MSCs on GBM clearly demonstrate that differences must be considered when choosing a stem cell source

  4. Autologous cell therapy for cisplatin-induced acute kidney injury by using non-expanded adipose tissue-derived cells.

    PubMed

    Yasuda, Kaoru; Ozaki, Takenori; Saka, Yousuke; Yamamoto, Tokunori; Gotoh, Momokazu; Ito, Yasuhiko; Yuzawa, Yukio; Matsuo, Seiichi; Maruyama, Shoichi

    2012-10-01

    Recent studies have demonstrated that cultured mesenchymal stromal cells derived from adipose tissue are useful for regenerative cell therapy. The stromal vascular fraction (SVF) can be obtained readily without culturing and may be clinically applicable. We investigated the therapeutic effects of SVF and used it in the treatment of acute kidney injury (AKI). Liposuction aspirates were obtained from healthy donors who had provided written informed consent. We harvested the SVF and determined the growth factor secretion and anti-apoptotic ability with conditioned medium. To investigate the effect of SVF on AKI, cisplatin was injected into rats and SVF was administrated into the subcupsula of the kidney. Both human and rat SVF cells secreted vascular endothelial growth factor-A (VEGF) and hepatocyte growth factor (HGF). Human SVF-conditioned media had an anti-apoptotic effect, which was inhibited by anti-HGF antibody (Ab) but not by anti-VEGF Ab. In vivo, SVF significantly ameliorated renal function, attenuated tubular damage and increased the cortical blood flow speed. In the SVF-treated group, VEGF levels in the cortex and HGF levels in both the cortex and medulla, especially tubules in the medulla, were significantly higher. Immunostaining revealed that SVF cells expressing VEGF and HGF and remained in the subcapsule on day 14. The present study demonstrates that a subcapsular injection of non-expanded SVF cells ameliorates rat AKI, and that the mechanism probably involves secretion of renoprotective molecules. Administration of human SVF may be clinically applicable and useful as a novel autologous cell therapy against kidney diseases.

  5. Tumor-extrinsic discoidin domain receptor 1 promotes mammary tumor growth by regulating adipose stromal interleukin 6 production in mice.

    PubMed

    Sun, Xiujie; Gupta, Kshama; Wu, Bogang; Zhang, Deyi; Yuan, Bin; Zhang, Xiaowen; Chiang, Huai-Chin; Zhang, Chi; Curiel, Tyler J; Bendeck, Michelle P; Hursting, Stephen; Hu, Yanfen; Li, Rong

    2018-02-23

    Discoidin domain receptor 1 (DDR1) is a collagen receptor that mediates cell communication with the extracellular matrix (ECM). Aberrant expression and activity of DDR1 in tumor cells are known to promote tumor growth. Although elevated DDR1 levels in the stroma of breast tumors are associated with poor patient outcome, a causal role for tumor-extrinsic DDR1 in cancer promotion remains unclear. Here we report that murine mammary tumor cells transplanted to syngeneic recipient mice in which Ddr1 has been knocked out (KO) grow less robustly than in WT mice. We also found that the tumor-associated stroma in Ddr1- KO mice exhibits reduced collagen deposition compared with the WT controls, supporting a role for stromal DDR1 in ECM remodeling of the tumor microenvironment. Furthermore, the stromal-vascular fraction (SVF) of Ddr1 knockout adipose tissue, which contains committed adipose stem/progenitor cells and preadipocytes, was impaired in its ability to stimulate tumor cell migration and invasion. Cytokine array-based screening identified interleukin 6 (IL-6) as a cytokine secreted by the SVF in a DDR1-dependent manner. SVF-produced IL-6 is important for SVF-stimulated tumor cell invasion in vitro , and, using antibody-based neutralization, we show that tumor promotion by IL-6 in vivo requires DDR1. In conclusion, our work demonstrates a previously unrecognized function of DDR1 in promoting tumor growth. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

  6. Isoliquiritigenin Attenuates Adipose Tissue Inflammation in vitro and Adipose Tissue Fibrosis through Inhibition of Innate Immune Responses in Mice.

    PubMed

    Watanabe, Yasuharu; Nagai, Yoshinori; Honda, Hiroe; Okamoto, Naoki; Yamamoto, Seiji; Hamashima, Takeru; Ishii, Yoko; Tanaka, Miyako; Suganami, Takayoshi; Sasahara, Masakiyo; Miyake, Kensuke; Takatsu, Kiyoshi

    2016-03-15

    Isoliquiritigenin (ILG) is a flavonoid derived from Glycyrrhiza uralensis and potently suppresses NLRP3 inflammasome activation resulting in the improvement of diet-induced adipose tissue inflammation. However, whether ILG affects other pathways besides the inflammasome in adipose tissue inflammation is unknown. We here show that ILG suppresses adipose tissue inflammation by affecting the paracrine loop containing saturated fatty acids and TNF-α by using a co-culture composed of adipocytes and macrophages. ILG suppressed inflammatory changes induced by the co-culture through inhibition of NF-κB activation. This effect was independent of either inhibition of inflammasome activation or activation of peroxisome proliferator-activated receptor-γ. Moreover, ILG suppressed TNF-α-induced activation of adipocytes, coincident with inhibition of IκBα phosphorylation. Additionally, TNF-α-mediated inhibition of Akt phosphorylation under insulin signaling was alleviated by ILG in adipocytes. ILG suppressed palmitic acid-induced activation of macrophages, with decreasing the level of phosphorylated Jnk expression. Intriguingly, ILG improved high fat diet-induced fibrosis in adipose tissue in vivo. Finally, ILG inhibited TLR4- or Mincle-stimulated expression of fibrosis-related genes in stromal vascular fraction from obese adipose tissue and macrophages in vitro. Thus, ILG can suppress adipose tissue inflammation by both inflammasome-dependent and -independent manners and attenuate adipose tissue fibrosis by targeting innate immune sensors.

  7. The features of adipose-derived stem cells in patients with inflammatory bowel diseases.

    PubMed

    Mizushima, Tsunekazu; Fukata, Tadafumi; Takeyama, Hiroshi; Takahashi, Hidekazu; Haraguchi, Naotsugu; Nishimura, Junichi; Hata, Taishi; Matsuda, Chu; Yamamoto, Hirofumi; Doki, Yuichiro; Mori, Masaki

    2018-03-01

    The treatment outcomes for inflammatory bowel disease (IBD) have been improving, but the development of better therapies is needed. Stem cell therapy is promising, but little is known about the differences in adipose-derived stem cells (ADSCs) between IBD patients and healthy individuals. ADSCs were isolated from subcutaneous adipose tissue (SAT) in IBD (Crohn's disease, 3; ulcerative colitis, 2) and non-IBD (colorectal cancer, 5; breast cancer, 1) patients. We also analyzed the effects of tumor necrosis factor (TNF)-α on murine ADSCs. The numbers of stromal vascular fraction (SVF) cells per gram of SAT were 7.72 ± 3.03 × 10 5 in IBD and 8.51 ± 8.80 × 10 5 in non-IBD patients (p > 0.05). The proportions of ADSCs in SVF cells were 4.98 ± 2.61% in IBD and 1.02 ± 0.67% in non-IBD patients. The numbers of ADSCs per gram of SAT were 4.16 ± 2.96 × 10 4 in IBD and 0.88 ± 1.04 × 10 4 in non-IBD patients. The proportions and numbers of ADSCs were significantly higher in IBD patients than in non-IBD patients. TNF-α significantly facilitated the proliferation and motility of murine ADSCs. These results showed the potential advantage of freshly isolated autologous ADSCs in IBD patients.

  8. Purification of human adipose-derived stem cells from fat tissues using PLGA/silk screen hybrid membranes.

    PubMed

    Chen, Da-Chung; Chen, Li-Yu; Ling, Qing-Dong; Wu, Meng-Hsueh; Wang, Ching-Tang; Suresh Kumar, S; Chang, Yung; Munusamy, Murugan A; Alarfajj, Abdullah A; Wang, Han-Chow; Hsu, Shih-Tien; Higuchi, Akon

    2014-05-01

    The purification of human adipose-derived stem cells (hADSCs) from human adipose tissue cells (stromal vascular fraction) was investigated using membrane filtration through poly(lactide-co-glycolic acid)/silk screen hybrid membranes. Membrane filtration methods are attractive in regenerative medicine because they reduce the time required to purify hADSCs (i.e., less than 30 min) compared with conventional culture methods, which require 5-12 days. hADSCs expressing the mesenchymal stem cell markers CD44, CD73, and CD90 were concentrated in the permeation solution from the hybrid membranes. Expression of the surface markers CD44, CD73, and CD99 on the cells in the permeation solution from the hybrid membranes, which were obtained using 18 mL of feed solution containing 50 × 10⁴ cells, was statistically significantly higher than that of the primary adipose tissue cells, indicating that the hADSCs can be purified in the permeation solution by the membrane filtration method. Cells expressing the stem cell-associated marker CD34 could be successfully isolated in the permeation solution, whereas CD34⁺ cells could not be purified by the conventional culture method. The hADSCs in the permeation solution demonstrated a superior capacity for osteogenic differentiation based on their alkali phosphatase activity, their osterix gene expression, and the results of mineralization analysis by Alizarin Red S and von Kossa staining compared with the cells from the suspension of human adipose tissue. These results suggest that the hADSCs capable of osteogenic differentiation preferentially permeate through the hybrid membranes. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. Combined introduction of Bmi-1 and hTERT immortalizes human adipose tissue-derived stromal cells with low risk of transformation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Tatrai, Peter, E-mail: peter.tatrai@biomembrane.hu; Department of Biochemistry and Molecular Biology, Medical and Health Science Center, University of Debrecen, Egyetem ter 1, H-4032 Debrecen; Szepesi, Aron, E-mail: aron.szepesi@biomembrane.hu

    2012-05-25

    Highlights: Black-Right-Pointing-Pointer We immortalized human adipose stromal cells (ASCs) with hTERT, Bmi-1, and SV40T. Black-Right-Pointing-Pointer hTERT-only ASCs are prone to transformation, while Bmi-only ASCs become senescent. Black-Right-Pointing-Pointer SV40T introduced along with hTERT abrogates proliferation control and multipotency. Black-Right-Pointing-Pointer hTERT combined with Bmi-1 yields stable phenotype up to 140 population doublings. -- Abstract: Adipose tissue-derived stromal cells (ASCs) are increasingly being studied for their usefulness in regenerative medicine. However, limited life span and donor-dependent variation of primary cells such as ASCs present major hurdles to controlled and reproducible experiments. We therefore aimed to establish immortalized ASC cell lines that provide steadymore » supply of homogeneous cells for in vitro work while retain essential features of primary cells. To this end, combinations of human telomerase reverse transcriptase (hTERT), murine Bmi-1, and SV40 large T antigen (SV40T) were introduced by lentiviral transduction into ASCs. The resulting cell lines ASC{sup hTERT}, ASC{sup Bmi-1}, ASC{sup Bmi-1+hTERT} and ASC{sup SV40T+hTERT} were tested for transgene expression, telomerase activity, surface immunomarkers, proliferation, osteogenic and adipogenic differentiation, karyotype, tumorigenicity, and cellular senescence. All cell lines have maintained expression of characteristic surface immunomarkers, and none was tumorigenic. However, ASC{sup Bmi-1} had limited replicative potential, while the rapidly proliferating ASC{sup SV40T+hTERT} acquired chromosomal aberrations, departed from MSC phenotype, and lost differentiation capacity. ASC{sup hTERT} and ASC{sup hTERT+Bmi-1}, on the other hand, preserved all essential MSC features and did not senesce after 100 population doublings. Notably, a subpopulation of ASC{sup hTERT} also acquired aberrant karyotype and showed signs of transformation after long

  10. Adipose-Derived Stem Cell Delivery for Adipose Tissue Engineering: Current Status and Potential Applications in a Tissue Engineering Chamber Model.

    PubMed

    Zhan, Weiqing; Tan, Shaun S; Lu, Feng

    2016-08-01

    In reconstructive surgery, there is a clinical need for adequate implants to repair soft tissue defects caused by traumatic injury, tumor resection, or congenital abnormalities. Adipose tissue engineering may provide answers to this increasing demand. This study comprehensively reviews current approaches to adipose tissue engineering, detailing different cell carriers under investigation, with a special focus on the application of adipose-derived stem cells (ASCs). ASCs act as building blocks for new tissue growth and as modulators of the host response. Recent studies have also demonstrated that the implantation of a hollow protected chamber, combined with a vascular pedicle within the fat flaps provides blood supply and enables the growth of large-volume of engineered soft tissue. Conceptually, it would be of value to co-regulate this unique chamber model with adipose-derived stem cells to obtain a greater volume of soft tissue constructs for clinical use. Our review provides a cogent update on these advances and details the generation of possible fat substitutes.

  11. Cell-based cytotoxicity assays for engineered nanomaterials safety screening: exposure of adipose derived stromal cells to titanium dioxide nanoparticles.

    PubMed

    Xu, Yan; Hadjiargyrou, M; Rafailovich, Miriam; Mironava, Tatsiana

    2017-07-11

    Increasing production of nanomaterials requires fast and proper assessment of its potential toxicity. Therefore, there is a need to develop new assays that can be performed in vitro, be cost effective, and allow faster screening of engineered nanomaterials (ENMs). Herein, we report that titanium dioxide (TiO 2 ) nanoparticles (NPs) can induce damage to adipose derived stromal cells (ADSCs) at concentrations which are rated as safe by standard assays such as measuring proliferation, reactive oxygen species (ROS), and lactate dehydrogenase (LDH) levels. Specifically, we demonstrated that low concentrations of TiO 2 NPs, at which cellular LDH, ROS, or proliferation profiles were not affected, induced changes in the ADSCs secretory function and differentiation capability. These two functions are essential for ADSCs in wound healing, energy expenditure, and metabolism with serious health implications in vivo. We demonstrated that cytotoxicity assays based on specialized cell functions exhibit greater sensitivity and reveal damage induced by ENMs that was not otherwise detected by traditional ROS, LDH, and proliferation assays. For proper toxicological assessment of ENMs standard ROS, LDH, and proliferation assays should be combined with assays that investigate cellular functions relevant to the specific cell type.

  12. Neurotrophic and neuroprotective potential of human limbus-derived mesenchymal stromal cells.

    PubMed

    Liang, Chang-Min; Weng, Shao-Ju; Tsai, Tung-Han; Li, I-Hsun; Lu, Pin-Hui; Ma, Kuo-Hsing; Tai, Ming-Cheng; Chen, Jiann-Torng; Cheng, Cheng-Yi; Huang, Yuahn-Sieh

    2014-10-01

    The purpose of this study was to examine neurotrophic and neuroprotective effects of limbus stroma-derived mesenchymal stromal cells (L-MSCs) on cortical neurons in vitro and in vivo. Cultured L-MSCs were characterized by flow cytometry and immunofluorescence through the use of specific MSC marker antibodies. Conditioned media were collected from normoxia- and hypoxia-treated L-MSCs to assess neurotrophic effects. Neuroprotective potentials were evaluated through the use of in vitro hypoxic cortical neuron culture and in vivo rat focal cerebral ischemia models. Neuronal morphology was confirmed by immunofluorescence with the use of anti-MAP2 antibody. Post-ischemic infarct volume and motor behavior were assayed by means of triphenyltetrazolium chloride staining and open-field testing, respectively. Human growth antibody arrays and enzyme-linked immunoassays were used to analyze trophic/growth factors contained in conditioned media. Isolated human L-MSCs highly expressed CD29, CD90 and CD105 but not CD34 and CD45. Mesenchymal lineage cell surface expression pattern and differentiation capacity were identical to MSCs derived form human bone marrow and adipose tissue. The L-MSC normoxic and hypoxic conditioned media both promoted neurite outgrowth in cultured cortical neurons. Hypoxic conditioned medium showed superior neurotrophic function and neuroprotective potential with reduced ischemic brain injury and improved functional recovery in rat focal cerebral ischemia models. Human growth factor arrays and enzyme-linked immunoassays measurements showed neuroprotective and growth-associated cytokines (vascular endothelial growth factor [VEGF], VEGFR3, brain-derived neurotrophic factor, insulin-like growth factor -2 and hepatocyte growth factor) contained in conditioned media. Hypoxic exposure caused VEGF and brain-derived neurotrophic factor upregulation, possibly contributing to neurotrophic and neuroprotective effects. L-MSCs can secrete various neurotrophic factors

  13. Human adipose-derived mesenchymal stromal cell pigment epithelium-derived factor cytotherapy modifies genetic and epigenetic profiles of prostate cancer cells.

    PubMed

    Zolochevska, Olga; Shearer, Joseph; Ellis, Jayne; Fokina, Valentina; Shah, Forum; Gimble, Jeffrey M; Figueiredo, Marxa L

    2014-03-01

    Adipose-derived mesenchymal stromal cells (ASCs) are promising tools for delivery of cytotherapy against cancer. However, ASCs can exert profound effects on biological behavior of tumor cells. Our study aimed to examine the influence of ASCs on gene expression and epigenetic methylation profiles of prostate cancer cells as well as the impact of expressing a therapeutic gene on modifying the interaction between ASCs and prostate cancer cells. ASCs were modified by lentiviral transduction to express either green fluorescent protein as a control or pigment epithelium-derived factor (PEDF) as a therapeutic molecule. PC3 prostate cancer cells were cultured in the presence of ASC culture-conditioned media (CCM), and effects on PC3 or DU145. Ras cells were examined by means of real-time quantitative polymerase chain reaction, EpiTect methyl prostate cancer-focused real-time quantitative polymerase chain reaction arrays, and luciferase reporter assays. ASCs transduced with lentiviral vectors were able to mediate expression of several tumor-inhibitory genes, some of which correlated with epigenetic methylation changes on cocultured PC3 prostate cancer cells. When PC3 cells were cultured with ASC-PEDF CCM, we observed a shift in the balance of gene expression toward tumor inhibition, which suggests that PEDF reduces the potential tumor-promoting activity of unmodified ASCs. These results suggest that ASC-PEDF CCM can promote reprogramming of tumor cells in a paracrine manner. An improved understanding of genetic and epigenetic events in prostate cancer growth in response to PEDF paracrine therapy would enable a more effective use of ASC-PEDF, with the goal of achieving safer yet more potent anti-tumor effects. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  14. Fluorescent Immortalized Human Adipose Derived Stromal Cells (hASCs-TS/GFP+) for Studying Cell Drug Delivery Mediated by Microvesicles.

    PubMed

    Cocce, Valentina; Balducci, Luigi; Falchetti, Maria L; Pascucci, Luisa; Ciusani, Emilio; Brini, Anna T; Sisto, Francesca; Piovani, Giovanna; Alessandri, Giulio; Parati, Eugenio; Cabeza, Laura; Pessina, Augusto

    2017-11-24

    A new tool for the drug delivery is based on the use of Mesenchymal Stromal Cells (MSCs) loaded in vitro with anti-cancer drugs. Unfortunately, the restricted lifespan of MSCs represents a significant limitation to produce them in high amounts and for long time studies. Immortalized MSCs from adipose tissue (hASCs) have been generated as good source of cells with stable features. These cells could improve the development of standardized procedures for both in vitro and preclinical studies. Furthermore they facilitate procedures for preparing large amounts of secretome containing microvesicles (MVs). We used human adipose tissue derived MSCs immortalized with hTERT+SV40 (TS) genes and transfected with GFP (hASCs-TS/GFP+). This line was investigated for its ability to uptake and release anticancer drugs. Microvesicles associated to paclitaxel (MVs/PTX) were isolated, quantified, and tested on pancreatic cancer cells. The line hASCs-TS/GFP+ maintained the main mesenchymal characters and was able to uptake and release, in active form, both paclitaxel and gemcitabine. From paclitaxel loaded hASCs-TS/GFP+ cells were isolated microvesicles in sufficient amount to inhibit "in vitro" the proliferation of pancreatic tumor cells. Our study suggests that human immortalized MSCs could be used for a large scale production of cells for mediated drug delivery. Moreover, the secretion of drug-associated MVs could represent a new way for producing new drug formulation by "biogenesis". In the context of the "advanced cell therapy procedure", the MVs/PTX production would use less resource and time and it could possibly contribute to simplification of GMP procedures. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  15. Dura Mater Stimulates Human Adipose-Derived Stromal Cells to Undergo Bone Formation in Mouse Calvarial Defects

    PubMed Central

    Levi, Benjamin; Nelson, Emily R.; Li, Shuli; James, Aaron W.; Hyun, Jeong S.; Montoro, Daniel T.; Lee, Min; Glotzbach, Jason P.; Commons, George W.; Longaker, Michael T.

    2015-01-01

    Human adipose-derived stromal cells (hASCs) have a proven capacity to aid in osseous repair of calvarial defects. However, the bone defect microenvironment necessary for osseous healing is not fully understood. In this study, we postulated that the cell-cell interaction between engrafted ASCs and host dura mater (DM) cells is critical for the healing of calvarial defects. hASCs were engrafted into critical sized calvarial mouse defects. The DM-hASC interaction was manipulated surgically by DM removal or by insertion of a semipermeable or nonpermeable membrane between DM and hASCs. Radiographic, histologic, and gene expression analyses were performed. Next, the hASC-DM interaction is assessed by conditioned media (CM) and coculture assays. Finally, bone morphogenetic protein (BMP) signaling from DM was investigated in vivo using novel BMP-2 and anti-BMP-2/4 slow releasing scaffolds. With intact DM, osseous healing occurs both from host DM and engrafted hASCs. Interference with the DM-hASC interaction dramatically reduced calvarial healing with abrogated BMP-2–Smad-1/5 signaling. Using CM and coculture assays, mouse DM cells stimulated hASC osteogenesis via BMP signaling. Through in vivo manipulation of the BMP-2 pathway, we found that BMP-2 plays an important role in DM stimulation of hASC osteogenesis in the context of calvarial bone healing. BMP-2 supplementation to a defect with disrupted DM allowed for bone formation in a nonhealing defect. DM is an osteogenic cell type that both participates in and stimulates osseous healing in a hASC-engrafted calvarial defect. Furthermore, DM-derived BMP-2 paracrine stimulation appears to play a key role for hASC mediated repair. PMID:21656608

  16. In Situ Blood Vessel Regeneration Using SP (Substance P) and SDF (Stromal Cell-Derived Factor)-1α Peptide Eluting Vascular Grafts.

    PubMed

    Shafiq, Muhammad; Zhang, Qiuying; Zhi, Dengke; Wang, Kai; Kong, Deling; Kim, Dong-Hwee; Kim, Soo Hyun

    2018-05-31

    The objective of this study was to develop small-diameter vascular grafts capable of eluting SDF (stromal cell-derived factor)-1α-derived peptide and SP (substance P) for in situ vascular regeneration. Polycaprolactone (PCL)/collagen grafts containing SP or SDF-1α-derived peptide were fabricated by electrospinning. SP and SDF-1α peptide-loaded grafts recruited significantly higher mesenchymal stem cells than that of the control group. The in vivo potential of PCL/collagen, SDF-1, and SP grafts was assessed by implanting them in a rat abdominal aorta for up to 4 weeks. All grafts remained patent as observed using color Doppler and stereomicroscope. Host cells infiltrated into the graft wall and the neointima was formed in peptides-eluting grafts. The lumen of the SP grafts was covered by the endothelial cells with cobblestone-like morphology, which were elongated in the direction of the blood flow, as discerned using scanning electron microscopy. Moreover, SDF-1α and SP grafts led to the formation of a confluent endothelium as evaluated using immunofluorescence staining with von Willebrand factor antibody. SP and SDF-1α grafts also promoted smooth muscle cell regeneration, endogenous stem cell recruitment, and blood vessel formation, which was the most prominent in the SP grafts. Evaluation of inflammatory response showed that 3 groups did not significantly differ in terms of the numbers of proinflammatory macrophages, whereas SP grafts showed significantly higher numbers of proremodeling macrophages than that of the control and SDF-1α grafts. SDF-1α and SP grafts can be potential candidates for in situ vascular regeneration and are worthy for future investigations. © 2018 American Heart Association, Inc.

  17. Burn Eschar Stimulates Fibroblast and Adipose Mesenchymal Stromal Cell Proliferation and Migration but Inhibits Endothelial Cell Sprouting

    PubMed Central

    Monsuur, Hanneke N.; van den Broek, Lenie J.; Jhingoerie, Renushka L.; Vloemans, Adrianus F. P. M.

    2017-01-01

    The majority of full-thickness burn wounds heal with hypertrophic scar formation. Burn eschar most probably influences early burn wound healing, since granulation tissue only forms after escharotomy. In order to investigate the effect of burn eschar on delayed granulation tissue formation, burn wound extract (BWE) was isolated from the interface between non-viable eschar and viable tissue. The influence of BWE on the activity of endothelial cells derived from dermis and adipose tissue, dermal fibroblasts and adipose tissue-derived mesenchymal stromal cells (ASC) was determined. It was found that BWE stimulated endothelial cell inflammatory cytokine (CXCL8, IL-6 and CCL2) secretion and migration. However, BWE had no effect on endothelial cell proliferation or angiogenic sprouting. Indeed, BWE inhibited basic Fibroblast Growth Factor (bFGF) induced endothelial cell proliferation and sprouting. In contrast, BWE stimulated fibroblast and ASC proliferation and migration. No difference was observed between cells isolated from dermis or adipose tissue. The inhibitory effect of BWE on bFGF-induced endothelial proliferation and sprouting would explain why excessive granulation tissue formation is prevented in full-thickness burn wounds as long as the eschar is still present. Identifying the eschar factors responsible for this might give indications for therapeutic targets aimed at reducing hypertrophic scar formation which is initiated by excessive granulation tissue formation once eschar is removed. PMID:28820426

  18. A Bilayer Construct Controls Adipose-Derived Stem Cell Differentiation into Endothelial Cells and Pericytes without Growth Factor Stimulation

    DTIC Science & Technology

    2011-01-01

    A Bilayer Construct Controls Adipose-Derived Stem Cell Differentiation into Endothelial Cells and Pericytes Without Growth Factor Stimulation...Ph.D.3 This work describes the differentiation of adipose-derived mesenchymal stem cells (ASC) in a composite hy- drogel for use as a vascularized...tissue from a single population of ASC. This work underscores the importance of the extracellular matrix in controlling stem cell phenotype. It is our

  19. Short-term hypoxic preconditioning promotes prevascularization in 3D bioprinted bone constructs with stromal vascular fraction derived cells† †Electronic supplementary information (ESI) available: qPCR primers and Fig. S1. See DOI: 10.1039/c7ra04372d Click here for additional data file.

    PubMed Central

    Kuss, Mitchell A.; Harms, Robert; Wu, Shaohua; Wang, Ying; Untrauer, Jason B.; Carlson, Mark A.

    2017-01-01

    Reconstruction of complex, craniofacial bone defects often requires autogenous vascularized bone grafts, and still remains a challenge today. In order to address this issue, we isolated the stromal vascular fraction (SVF) from adipose tissues and maintained the phenotypes and the growth of endothelial lineage cells within SVF derived cells (SVFC) by incorporating an endothelial cell medium. We 3D bioprinted SVFC within our hydrogel bioinks and conditioned the constructs in either normoxia or hypoxia. We found that short-term hypoxic conditioning promoted vascularization-related gene expression, whereas long-term hypoxia impaired cell viability and vascularization. 3D bioprinted bone constructs composed of polycaprolactone/hydroxyapatite (PCL/HAp) and SVFC-laden hydrogel bioinks were then implanted into athymic mice, after conditioning in normoxic or short-term hypoxic environments, in order to determine the in vitro and in vivo vascularization and osteogenic differentiation of the constructs. Short-term hypoxic conditioning promoted microvessel formation in vitro and in vivo and promoted integration with existing host vasculature, but did not affect osteogenic differentiation of SVFC. These findings demonstrate the benefit of short-term hypoxia and the potential for utilization of SVFC and 3D bioprinting for generating prevascularized 3D bioprinted bone constructs. Furthermore, the ability to custom design complex anatomical shapes has promising applications for the regeneration of both large and small craniofacial bone defects. PMID:28670447

  20. Triactome: Neuro–Immune–Adipose Interactions. Implication in Vascular Biology

    PubMed Central

    Chaldakov, George Nikov; Fiore, Marco; Ghenev, Peter I.; Beltowski, Jerzy; Ranćić, Gorana; Tunçel, Neşe; Aloe, Luigi

    2014-01-01

    Understanding how the precise interactions of nerves, immune cells, and adipose tissue account for cardiovascular and metabolic biology is a central aim of biomedical research at present. A long standing paradigm holds that the vascular wall is composed of three concentric tissue coats (tunicae): intima, media, and adventitia. However, large- and medium-sized arteries, where usually atherosclerotic lesions develop, are consistently surrounded by periadventitial adipose tissue (PAAT), we recently designated tunica adiposa (in brief, adiposa like intima, media, and adventitia). Today, atherosclerosis is considered an immune-mediated inflammatory disease featured by endothelial dysfunction/intimal thickening, medial atrophy, and adventitial lesions associated with adipose dysfunction, whereas hypertension is characterized by hyperinnervation-associated medial thickening due to smooth muscle cell hypertrophy/hyperplasia. PAAT expansion is associated with increased infiltration of immune cells, both adipocytes and immunocytes secreting pro-inflammatory and anti-inflammatory (metabotrophic) signaling proteins collectively dubbed adipokines. However, the role of vascular nerves and their interactions with immune cells and paracrine adipose tissue is not yet evaluated in such an integrated way. The present review attempts to briefly highlight the findings in basic and translational sciences in this area focusing on neuro–immune–adipose interactions, herein referred to as triactome. Triactome-targeted pharmacology may provide a novel therapeutic approach in cardiovascular disease. PMID:24782857

  1. Triactome: neuro-immune-adipose interactions. Implication in vascular biology.

    PubMed

    Chaldakov, George Nikov; Fiore, Marco; Ghenev, Peter I; Beltowski, Jerzy; Ranćić, Gorana; Tunçel, Neşe; Aloe, Luigi

    2014-01-01

    Understanding how the precise interactions of nerves, immune cells, and adipose tissue account for cardiovascular and metabolic biology is a central aim of biomedical research at present. A long standing paradigm holds that the vascular wall is composed of three concentric tissue coats (tunicae): intima, media, and adventitia. However, large- and medium-sized arteries, where usually atherosclerotic lesions develop, are consistently surrounded by periadventitial adipose tissue (PAAT), we recently designated tunica adiposa (in brief, adiposa like intima, media, and adventitia). Today, atherosclerosis is considered an immune-mediated inflammatory disease featured by endothelial dysfunction/intimal thickening, medial atrophy, and adventitial lesions associated with adipose dysfunction, whereas hypertension is characterized by hyperinnervation-associated medial thickening due to smooth muscle cell hypertrophy/hyperplasia. PAAT expansion is associated with increased infiltration of immune cells, both adipocytes and immunocytes secreting pro-inflammatory and anti-inflammatory (metabotrophic) signaling proteins collectively dubbed adipokines. However, the role of vascular nerves and their interactions with immune cells and paracrine adipose tissue is not yet evaluated in such an integrated way. The present review attempts to briefly highlight the findings in basic and translational sciences in this area focusing on neuro-immune-adipose interactions, herein referred to as triactome. Triactome-targeted pharmacology may provide a novel therapeutic approach in cardiovascular disease.

  2. Adipose tissue as a stem cell source for musculo-skeletal regeneration

    PubMed Central

    Gimble, Jeffrey M.; Grayson, Warren; Guilak, Farshid; Lopez, Mandi J.; Vunjak-Novakovic, Gordana

    2013-01-01

    Adipose tissue is an abundant, easily accessible, and reproducible cell source for musculo-skeletal regenerative medicine applications. Initial derivation steps yield a heterogeneous population of cells collectively termed the stromal vascular fraction (SVF), which consist of endothelial cells, immune cells, pericytes, and pre-adipocytes. Subsequent selection of an adherent cell subset from the SVF results in a relatively homogeneous population of adipose-derived stromal/stem cells (ASCs). Mammalian ASCs exhibit the ability to selectively differentiate into chondrogenic, myogenic, and osteogenic lineages in response to inductive stimuli in vitro (when cultured on scaffolds in bioreactors) and in vivo (when implanted in pre-clinical animal models). Unlike hematopoietic cells, ASCs do not elicit a robust lymphocyte reaction and instead generate and release immunosuppressive factors, such as prostaglandin E2. These unique immunomodulatory features suggest that both allogeneic and autologous ASCs will engraft successfully following application for tissue regeneration purposes. The differentiation and expansion potential of ASCs can be modified by growth factors like bone morphogenetic protein 6, bio-inductive scaffolds, and bioreactors providing environmental control and biophysical stimulation. Gene therapy approaches using lentiviral transduction can also be used to direct differentiation of ASCs along particular lineage pathways. We discuss here the utility of ASCs for musculo-skeletal tissue repair and some of the technologies that can be implemented to unlock the full regenerative potential of these highly valuable cells. PMID:21196358

  3. Fluorine-19 Labeling of Stromal Vascular Fraction Cells for Clinical Imaging Applications

    PubMed Central

    Rose, Laura C.; Kadayakkara, Deepak K.; Wang, Guan; Bar-Shir, Amnon; Helfer, Brooke M.; O’Hanlon, Charles F.; Kraitchman, Dara L.; Rodriguez, Ricardo L.

    2015-01-01

    Stromal vascular fraction (SVF) cells are used clinically for various therapeutic targets. The location and persistence of engrafted SVF cells are important parameters for determining treatment failure versus success. We used the GID SVF-1 platform and a clinical protocol to harvest and label SVF cells with the fluorinated (19F) agent CS-1000 as part of a first-in-human phase I trial (clinicaltrials.gov identifier NCT02035085) to track SVF cells with magnetic resonance imaging during treatment of radiation-induced fibrosis in breast cancer patients. Flow cytometry revealed that SVF cells consisted of 25.0% ± 15.8% CD45+, 24.6% ± 12.5% CD34+, and 7.5% ± 3.3% CD31+ cells, with 2.1 ± 0.7 × 105 cells per cubic centimeter of adipose tissue obtained. Fluorescent CS-1000 (CS-ATM DM Green) labeled 87.0% ± 13.5% of CD34+ progenitor cells compared with 47.8% ± 18.5% of hematopoietic CD45+ cells, with an average of 2.8 ± 2.0 × 1012 19F atoms per cell, determined using nuclear magnetic resonance spectroscopy. The vast majority (92.7% ± 5.0%) of CD31+ cells were also labeled, although most coexpressed CD34. Only 16% ± 22.3% of CD45−/CD31−/CD34− (triple-negative) cells were labeled with CS-ATM DM Green. After induction of cell death by either apoptosis or necrosis, >95% of 19F was released from the cells, indicating that fluorine retention can be used as a surrogate marker for cell survival. Labeled-SVF cells engrafted in a silicone breast phantom could be visualized with a clinical 3-Tesla magnetic resonance imaging scanner at a sensitivity of approximately 2 × 106 cells at a depth of 5 mm. The current protocol can be used to image transplanted SVF cells at clinically relevant cell concentrations in patients. Significance Stromal vascular fraction (SVF) cells harvested from adipose tissue offer great promise in regenerative medicine, but methods to track such cell therapies are needed to ensure correct administration and monitor survival. A clinical protocol

  4. Migraine and tension-type headache treated with stromal vascular fraction: a case series

    PubMed Central

    2014-01-01

    Introduction Chronic migraines and tension-type headaches are debilitating diseases affecting 1.4 to 2.2% of the population with both quality of life and economic implications. To date, the pain associated with migraine and tension-type headaches has been controlled with a range of medications, with varying levels of success. In addition, the side-effect profile of these medications, as well as their potential for addiction, has been a cause for concern for both patients and physicians. Case presentations Four women with long histories of migraine or frequent tension-type headache that meet the International Classification of Headache Disorders criteria for Chronic Migraine or Tension-type Headaches were given a systemic treatment(s) of autologous stromal vascular fraction or autologous ‘StroMed’ isolated from lipoaspirate. StroMed is stromal vascular fraction cells prepared by ultrasonic cavitation. Two of the four patients, both of whom are Arab women aged 40 and 36 years, ceased having migraines after 1 month, for a period of 12 to 18 months. The third patient, a Slavic woman aged 43 years, had a significant decrease in the frequency and severity of migraines with only seven migraines over 18 months. The fourth patient, an Asian woman aged 44 years, obtained a temporary decrease for a period of a month and was retreated 18 months later and has been free of migraines to date for 1 month. Pain medication was typically reduced from prescribed opioid analgesia to non-steroidal anti-inflammatory drugs and paracetamol. Conclusions This case series is the first to provide evidence of the efficacy of autologous StroMed and stromal vascular fraction in the treatment of migraine and tension-type headache. The treatment of this disease by stromal vascular fraction adds a new dimension to its clinical applicability and suggests a relatively simple treatment that may help address the symptoms of the disease. Given what is known about the components of the stromal

  5. Migraine and tension-type headache treated with stromal vascular fraction: a case series.

    PubMed

    Bright, Ralph; Bright, Matthew; Bright, Pelin; Hayne, Shannon; Thomas, Wayne D

    2014-06-30

    Chronic migraines and tension-type headaches are debilitating diseases affecting 1.4 to 2.2% of the population with both quality of life and economic implications. To date, the pain associated with migraine and tension-type headaches has been controlled with a range of medications, with varying levels of success. In addition, the side-effect profile of these medications, as well as their potential for addiction, has been a cause for concern for both patients and physicians. Four women with long histories of migraine or frequent tension-type headache that meet the International Classification of Headache Disorders criteria for Chronic Migraine or Tension-type Headaches were given a systemic treatment(s) of autologous stromal vascular fraction or autologous 'StroMed' isolated from lipoaspirate. StroMed is stromal vascular fraction cells prepared by ultrasonic cavitation. Two of the four patients, both of whom are Arab women aged 40 and 36 years, ceased having migraines after 1 month, for a period of 12 to 18 months. The third patient, a Slavic woman aged 43 years, had a significant decrease in the frequency and severity of migraines with only seven migraines over 18 months. The fourth patient, an Asian woman aged 44 years, obtained a temporary decrease for a period of a month and was retreated 18 months later and has been free of migraines to date for 1 month. Pain medication was typically reduced from prescribed opioid analgesia to non-steroidal anti-inflammatory drugs and paracetamol. This case series is the first to provide evidence of the efficacy of autologous StroMed and stromal vascular fraction in the treatment of migraine and tension-type headache. The treatment of this disease by stromal vascular fraction adds a new dimension to its clinical applicability and suggests a relatively simple treatment that may help address the symptoms of the disease. Given what is known about the components of the stromal vascular fraction and how they act, the information

  6. Adipose-derived mesenchymal stem cells and regenerative medicine.

    PubMed

    Konno, Masamitsu; Hamabe, Atsushi; Hasegawa, Shinichiro; Ogawa, Hisataka; Fukusumi, Takahito; Nishikawa, Shimpei; Ohta, Katsuya; Kano, Yoshihiro; Ozaki, Miyuki; Noguchi, Yuko; Sakai, Daisuke; Kudoh, Toshihiro; Kawamoto, Koichi; Eguchi, Hidetoshi; Satoh, Taroh; Tanemura, Masahiro; Nagano, Hiroaki; Doki, Yuichiro; Mori, Masaki; Ishii, Hideshi

    2013-04-01

    Adipose tissue-derived mesenchymal stem cells (ADSCs) are multipotent and can differentiate into various cell types, including osteocytes, adipocytes, neural cells, vascular endothelial cells, cardiomyocytes, pancreatic β-cells, and hepatocytes. Compared with the extraction of other stem cells such as bone marrow-derived mesenchymal stem cells (BMSCs), that of ADSCs requires minimally invasive techniques. In the field of regenerative medicine, the use of autologous cells is preferable to embryonic stem cells or induced pluripotent stem cells. Therefore, ADSCs are a useful resource for drug screening and regenerative medicine. Here we present the methods and mechanisms underlying the induction of multilineage cells from ADSCs. © 2013 The Authors Development, Growth & Differentiation © 2013 Japanese Society of Developmental Biologists.

  7. Effect of adipose-derived mesenchymal stromal cells on tendon healing in aging and estrogen deficiency: an in vitro co-culture model.

    PubMed

    Veronesi, Francesca; Della Bella, Elena; Torricelli, Paola; Pagani, Stefania; Fini, Milena

    2015-11-01

    Aging and estrogen deficiency play a pivotal role in reducing tenocyte proliferation, collagen turnover and extracellular matrix remodeling. Mesenchymal stromal cells are being studied as an alternative for tendon regeneration, but little is known about the molecular events of adipose-derived mesenchymal stromal cells (ADSCs) on tenocytes in tendons compromised by aging and estrogen deficiency. The present in vitro study aims to compare the potential therapeutic effects of ADSCs, harvested from healthy young (sham) and aged estrogen-deficient (OVX) subjects, for tendon healing. An indirect co-culture system was set up with ADSCs, isolated from OVX or sham rats, and tenocytes from OVX rats. Cell proliferation, healing rate and gene expression were evaluated in both a standard culture condition and a microwound-healing model. It was observed that tenocyte proliferation, healing rate and collagen expression improved after the addition of sham ADSCs in both culture situations. OVX ADSCs also increased tenocyte proliferation and healing rate but less compared with sham ADSCs. Decorin and Tenascin C expression increased in the presence of OVX ADSCs. Findings suggest that ADSCs might be a promising treatment for tendon regeneration in advanced age and estrogen deficiency. However, some differences between allogenic and autologous cells were found and should be investigated in further in vivo studies. It appears that allogenic ADSCs improve tenocyte proliferation, collagen expression and the healing rate more than autologous cells. Autologous cells increase collagen expression only in the absence of an injury and increase Decorin and Tenascin C more than allogenic cells. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  8. Fibrin glue as the cell-delivery vehicle for mesenchymal stromal cells in regenerative medicine.

    PubMed

    Wu, Xiuwen; Ren, Jianan; Li, Jieshou

    2012-05-01

    The use of tissue-engineering techniques such as stem-cell therapy to renew injured tissues is a promising strategy in regenerative medicine. As a cell-delivery vehicle, fibrin glues (FG) facilitate cell attachment, growth and differentiation and, ultimately, tissue formation and organization by its three-dimensional structure. Numerous studies have provided evidence that stromal cells derived from bone marrow (bone marrow stromal cells; BMSC) and adipose tissue (adipose-derived stromal cells; ADSC) contain a population of adult multipotent mesenchymal stromal cells (MSC) and endothelial progenitor cells that can differentiate into several lineages. By combining MSC with FG, the implantation could take advantage of the mutual benefits. Researchers and physicians have pinned their hopes on stem cells for developing novel approaches in regenerative medicine. This review focuses on the therapeutic potential of MSC with FG in bone defect reconstruction, cartilage and tendon injury repair, ligament, heart and nerve regeneration, and, furthermore, wound healing.

  9. Cyclohexane-1,2-dicarboxylic acid diisononyl ester and metabolite effects on rat epididymal stromal vascular fraction differentiation of adipose tissue

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Campioli, Enrico; Department of Medicine, McGill University, Montréal, Québec; Duong, Tam B.

    Plastics are generally mixed with additives like plasticizers to enhance their flexibility, pliability, and elasticity proprieties. Plasticizers are easily released into the environment and are absorbed mainly through ingestion, dermal contact, and inhalation. One of the main classes of plasticizers, phthalates, has been associated with endocrine and reproductive diseases. In 2002, 1,2-cyclohexane dicarboxylic acid diisononyl ester (DINCH) was introduced in the market for use in plastic materials and articles intended to come into contact with food, and it received final approval from the European Food Safety Authority in 2006. At present, there is limited knowledge about the safety and potentialmore » metabolic and endocrine-disrupting properties of DINCH and its metabolites. The purpose of this study was to evaluate the biological effects of DINCH and its active metabolites, cyclohexane-1,2-dicarboxylic acid (CHDA) and cyclohexane-1,2-dicarboxylic acid mono isononyl ester (MINCH), on rat primary stromal vascular fraction (SVF) of adipose tissue. DINCH and its metabolite, CHDA, were not able to directly affect SVF differentiation. However, exposure of SVF to 50 μM and 100 μM concentrations of MINCH affected the expression of Cebpa and Fabp4, thus inducing SVF preadipocytes to accumulate lipids and fully differentiate into mature adipocytes. The effect of MINCH was blocked by the specific peroxisome proliferator-activated receptor (PPAR)-α antagonist, GW6471. Taken together, these results suggest that MINCH is a potent PPAR-α agonist and a metabolic disruptor, capable of inducing SVF preadipocyte differentiation, that may interfere with the endocrine system in mammals. - Highlights: • DINCH and CHDA did not affect the adipogenesis of the SVF. • MINCH affected the adipogenesis of the SVF. • MINCH effect was blocked by the specific PPAR-α antagonist GW6471. • MINCH exerted a similar effect as MEHP on SVF adipogenesis. • DINCH/MINCH are potential

  10. Augmented expression and secretion of adipose-derived pigment epithelium-derived factor does not alter local angiogenesis or contribute to the development of systemic metabolic derangements.

    PubMed

    Lakeland, Thomas V; Borg, Melissa L; Matzaris, Maria; Abdelkader, Amany; Evans, Roger G; Watt, Matthew J

    2014-06-15

    Impaired coupling of adipose tissue expansion and vascularization is proposed to lead to adipocyte hypoxia and inflammation, which in turn contributes to systemic metabolic derangements. Pigment epithelium-derived factor (PEDF) is a powerful antiangiogenic factor that is secreted by adipocytes, elevated in obesity, and implicated in the development of insulin resistance. We explored the angiogenic and metabolic role of adipose-derived PEDF through in vivo studies of mice with overexpression of PEDF in adipocytes (PEDF-aP2). PEDF expression in white adipocytes and PEDF secretion from adipose tissue was increased in transgenic mice, but circulating levels of PEDF were not increased. Overexpression of PEDF did not alter vascularization, the partial pressure of O2, cellular hypoxia, or gene expression of inflammatory markers in adipose tissue. Energy expenditure and metabolic substrate utilization, body mass, and adiposity were not altered in PEDF-aP2 mice. Whole body glycemic control was normal as assessed by glucose and insulin tolerance tests, and adipocyte-specific glucose uptake was unaffected by PEDF overexpression. Adipocyte lipolysis was increased in PEDF-aP2 mice and associated with increased adipose triglyceride lipase and decreased perilipin 1 expression. Experiments conducted in mice rendered obese by high-fat feeding showed no differences between PEDF-aP2 and wild-type mice for body mass, adiposity, whole body energy expenditure, glucose tolerance, or adipose tissue oxygenation. Together, these data indicate that adipocyte-generated PEDF enhances lipolysis but question the role of PEDF as a major antiangiogenic or proinflammatory mediator in adipose tissue in vivo. Copyright © 2014 the American Physiological Society.

  11. Primary adipose-derived stem cells enriched by growth factor treatment improves cell adaptability toward cardiovascular differentiation in a rodent model of acute myocardial infarction.

    PubMed

    Chang, Jui-Chih; Lee, Ping-Chun; Lin, Yu-Chun; Lee, Kung-Wei; Hsu, Shan-hui

    2011-01-01

    The heterogeneous cell population in primary adipose-derived adult stem cells (ADAS) and difficulty in keeping their primitive properties have posed certain limitations on using these cells for cell therapy. Therefore, our objective was to generate a population of cells enriched from the adipose stromal-vascular fraction (SVF) with greater differentiation potential than ADAS and to explore the mechanism behind the repair of the injured myocardium in vivo. The distinct population of adipose stromal cells was enriched by immediate treatment of the growth factor cocktail (EGF and PDGF-BB) to the freshly isolated SVF. These cells (ADAS-GFs) had distinct cell morphology from ADAS and in average had a smaller size. They presented co-expression of CD140a (pericytic markers) and CD34 (hematopoietic marker), more obvious mesenchymal (CD13, CD29, CD44, CD90 and CD117) markers, but rare KDR, and were negative for CD45 and CD31. ADAS-GFs not only spontaneously expressed endothelial cell markers and formed capillary-like tubes on Matrigel but also clearly expressed early cardiomyocyte marker genes when embedded in methylcellulose-based medium. In Sprague-Dawley (SD) rats with left anterior descending artery (LAD)-induced myocardial infarction (MI), the ADAS-GFs transplanted group had the left ventricular function significantly improved compared with the ADAS transplanted group or the control group at 12 weeks post transplantation. The immunofluorescence staining revealed that the transplanted ADAS-GFs expressed GATA4, betamyosin heavy chain and troponin T protein but not vWF. More capillaries were also observed around the infarcted zone in the ADAS-GFs transplanted group. These data suggested that ADAS-GFs with a higher proangiogenic potential may restore the cardiac function of infarcted myocardium via the direct cardiomyocyte differentiation as well as angiogenesis recruitment.

  12. Mitochondrial dysfunction of immortalized human adipose tissue-derived mesenchymal stromal cells from patients with Parkinson's disease.

    PubMed

    Moon, Hyo Eun; Yoon, Seung Hee; Hur, Yong Suk; Park, Hyung Woo; Ha, Ji Young; Kim, Kyung-Hee; Shim, Jung Hee; Yoo, Seung Hyun; Son, Jin H; Paek, Seung Leal; Kim, In Keyoung; Hwang, Jae Ha; Kim, Dong Gyu; Kim, Han-Joon; Jeon, Beom Seok; Park, Sung Sup; Paek, Sun Ha

    2013-12-01

    Mitochondrial dysfunction in dopaminergic neurons of patients with idiopathic and familial Parkinson's disease (PD) is well known although the underlying mechanism is not clear. We established a homogeneous population of human adipose tissue-derived mesenchymal stromal cells (hAD-MSCs) from human adult patients with early-onset hereditary familial Parkin-defect PD as well as late-onset idiopathic PD by immortalizing cells with the hTERT gene to better understand the underlying mechanism of PD. The hAD-MSCs from patients with idiopathic PD were designated as "PD", from patients with Parkin-defect PD as "Parkin" and from patients with pituitary adenomas as "non-PD" in short. The pGRN145 plasmid containing hTERT was introduced to establish telomerase immortalized cells. The established hTERT-immortalized cell lines showed chromosomal aneuploidy sustained stably over two-years. The morphological study of mitochondria in the primary and immortalized hAD-MSCs showed that the mitochondria of the non-PD were normal; however, those of the PD and Parkin were gradually damaged. A striking decrease in mitochondrial complex I, II, and IV activities was observed in the hTERT-immortalized cells from the patients with idiopathic and Parkin-defect PD. Comparative Western blot analyses were performed to investigate the expressions of PD specific marker proteins in the hTERT-immortalized cell lines. This study suggests that the hTERT-immortalized hAD-MSC cell lines established from patients with idiopathic and familial Parkin-defect PD could be good cellular models to evaluate mitochondrial dysfunction to better understand the pathogenesis of PD and to develop early diagnostic markers and effective therapy targets for the treatment of PD.

  13. Mitochondrial Dysfunction of Immortalized Human Adipose Tissue-Derived Mesenchymal Stromal Cells from Patients with Parkinson's Disease

    PubMed Central

    Moon, Hyo Eun; Yoon, Seung Hee; Hur, Yong Suk; Park, Hyung Woo; Ha, Ji Young; Kim, Kyung-Hee; Shim, Jung Hee; Yoo, Seung Hyun; Son, Jin H.; Paek, Seung Leal; Kim, In Keyoung; Hwang, Jae Ha; Kim, Dong Gyu; Kim, Han-Joon; Jeon, Beom Seok; Park, Sung Sup

    2013-01-01

    Mitochondrial dysfunction in dopaminergic neurons of patients with idiopathic and familial Parkinson's disease (PD) is well known although the underlying mechanism is not clear. We established a homogeneous population of human adipose tissue-derived mesenchymal stromal cells (hAD-MSCs) from human adult patients with early-onset hereditary familial Parkin-defect PD as well as late-onset idiopathic PD by immortalizing cells with the hTERT gene to better understand the underlying mechanism of PD. The hAD-MSCs from patients with idiopathic PD were designated as "PD", from patients with Parkin-defect PD as "Parkin" and from patients with pituitary adenomas as "non-PD" in short. The pGRN145 plasmid containing hTERT was introduced to establish telomerase immortalized cells. The established hTERT-immortalized cell lines showed chromosomal aneuploidy sustained stably over two-years. The morphological study of mitochondria in the primary and immortalized hAD-MSCs showed that the mitochondria of the non-PD were normal; however, those of the PD and Parkin were gradually damaged. A striking decrease in mitochondrial complex I, II, and IV activities was observed in the hTERT-immortalized cells from the patients with idiopathic and Parkin-defect PD. Comparative Western blot analyses were performed to investigate the expressions of PD specific marker proteins in the hTERT-immortalized cell lines. This study suggests that the hTERT-immortalized hAD-MSC cell lines established from patients with idiopathic and familial Parkin-defect PD could be good cellular models to evaluate mitochondrial dysfunction to better understand the pathogenesis of PD and to develop early diagnostic markers and effective therapy targets for the treatment of PD. PMID:24465144

  14. Transforming growth factor β-activated kinase 1 negatively regulates interleukin-1α-induced stromal-derived factor-1 expression in vascular smooth muscle cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yang, Bin; Li, Wei; Zheng, Qichang

    Stromal-derived Factor-1 (SDF-1) derived from vascular smooth muscle cells (VSMCs) contributes to vascular repair and remodeling in various vascular diseases. In this study, the mechanism underlying regulation of SDF-1 expression by interleukin-1α (IL-1α) was investigated in primary rat VSMCs. We found IL-1α promotes SDF-1 expression by up-regulating CCAAT-enhancer-binding protein β (C/EBPβ) in an IκB kinase β (IKKβ) signaling-dependent manner. Moreover, IL-1α-induced expression of C/EBPβ and SDF-1 was significantly potentiated by knockdown of transforming growth factor β-activated kinase 1 (TAK1), an upstream activator of IKKβ signaling. In addition, we also demonstrated that TAK1/p38 mitogen-activated protein kinase (p38 MAPK) signaling exerted negativemore » effect on IL-1α-induced expression of C/EBPβ and SDF-1 through counteracting ROS-dependent up-regulation of nuclear factor erythroid 2-related factor 2 (NRF2). In conclusion, TAK1 acts as an important regulator of IL-1α-induced SDF-1 expression in VSMCs, and modulating activity of TAK1 may serve as a potential strategy for modulating vascular repair and remodeling. - Highlights: • IL-1α induces IKKβ signaling-dependent SDF-1 expression by up-regulating C/EBPβ. • Activation of TAK1 by IL-1α negatively regulates C/EBPβ-dependent SDF-1 expression. • IL-1α-induced TAK1/p38 MAPK signaling counteracts ROS-dependent SDF-1 expression. • TAK1 counteracts IL-1α-induced SDF-1 expression by attenuating NRF2 up-regulation.« less

  15. Neural differentiation of adipose-derived stem cells isolated from GFP transgenic mice

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Fujimura, Juri; Department of Pediatrics, Nippon Medical School, Tokyo; E-mail: juri-f@nms.ac.jp

    2005-07-22

    Taking advantage of homogeneously marked cells from green fluorescent protein (GFP) transgenic mice, we have recently reported that adipose-derived stromal cells (ASCs) could differentiate into mesenchymal lineages in vitro. In this study, we performed neural induction using ASCs from GFP transgenic mice and were able to induce these ASCs into neuronal and glial cell lineages. Most of the neurally induced cells showed bipolar or multipolar appearance morphologically and expressed neuronal markers. Electron microscopy revealed their neuronal morphology. Some cells also showed glial phenotypes, as shown immunocytochemically. The present study clearly shows that ASCs derived from GFP transgenic mice differentiate intomore » neural lineages in vitro, suggesting that these cells might provide an ideal source for further neural stem cell research with possible therapeutic application for neurological disorders.« less

  16. In Vivo Exposure to Inorganic Arsenic Alters Differentiation-Specific Gene Expression of Adipose-Derived Mesenchymal Stem/Stromal Cells in C57BL/6J Mouse Model

    PubMed Central

    Shearer, Joseph J.; Figueiredo Neto, Manoel; Umbaugh, C. Samuel; Figueiredo, Marxa L.

    2017-01-01

    Abstract The number of mesenchymal stem cell (MSC) therapeutic modalities has grown in recent years. Adipose-derived mesenchymal stem/stromal cells (ASCs) can be isolated and expanded relatively easily as compared with their bone-marrow counterparts, making them a particularly promising source of MSCs. And although the biological mechanisms surrounding ASCs are actively being investigated, little is known about the effects that in vivo environmental exposures might have on their ability to properly differentiate. Therefore, we hypothesized that ASCs isolated from mice exposed to inorganic arsenic (iAs) would have an altered response towards adipogenic, osteogenic, and/or chondrogenic differentiation. To test this hypothesis, C57BL/6J male mice were provided drinking water containing 0, 300, or 1000 ppb iAs. ASCs were then isolated and differentiated, which was assessed by immunocytochemistry and real-time quantitative PCR (RT-qPCR). Our results showed that total urinary arsenic equilibrated within 1 week of exposure to iAs and was maintained throughout the study. ASCs isolated from each exposure group maintained differentiation capabilities for each lineage. The magnitude of differentiation-specific gene expression, however, appeared to be concentration dependent. For osteogenesis and chondrogenesis, differentiation-specific gene expression decreased, whereas adipogenesis showed a biphasic response with an initial decrease followed by an increase in adipogenic-related gene expression following iAs exposure. These results suggest that the level in which differentiation-specific genes are induced within these stromal cells might be sensitive to environmental contaminants. These findings highlight the need to take into account potential environmental exposures prior to selecting stromal cell donors, so ASCs can achieve optimal efficiency in regenerative therapy applications. PMID:28206643

  17. Characterization of Canine Adipose-Derived Mesenchymal Stromal/Stem Cells in Serum-Free Medium.

    PubMed

    Liu, Zhuoming; Screven, Rudell; Boxer, Lynne; Myers, Michael J; Devireddy, Lax R

    2018-06-20

    In this article, we report on the development of a defined serum-free medium capable of supporting the culture expansion of mesenchymal stromal/stem cells (MSCs) from canine adipose tissue (canine Ad-MSCs). The potential benefits of serum-free media can only be utilized if cells cultured in serum-free media maintain the same functional characteristics as cells cultured in serum-containing media. Therefore, we analyze the characteristics of canine Ad-MSCs cultured in this serum-free medium or in serum-containing medium through evaluation of growth kinetics, clonogenic capacity, senescence, and differentiation capacity. Both, serum-containing medium and our serum-free medium, supported efficient growth and colony formation of canine Ad-MSCs. In addition, canine Ad-MSCs cultured in both media demonstrated similar viability after freeze/thaw, similar cell surface marker expression, and were capable of trilineage differentiation. While canine Ad-MSCs cultured in both media were generally similar, under the conditions of our study, canine Ad-MSCs cultured in serum-free medium demonstrated a shorter lag phase and higher colony-forming capacity, accelerated population doubling, maintained multipotentiality at higher passage numbers, and underwent senescence at higher passage numbers compared to canine Ad-MSCs cultured in conventional serum-containing medium. These results suggest that canine Ad-MSCs cultured in serum-free medium retain the basic characteristics associated with canine Ad-MSCs cultured in serum-containing medium, although some differences in growth kinetics were observed.

  18. Depot-dependent effects of adipose tissue explants on co-cultured hepatocytes.

    PubMed

    Du, Zhen-Yu; Ma, Tao; Lock, Erik-Jan; Hao, Qin; Kristiansen, Karsten; Frøyland, Livar; Madsen, Lise

    2011-01-01

    We have developed an in vitro hepatocyte-adipose tissue explant (ATE) co-culture model enabling examination of the effect of visceral and subcutaneous adipose tissues on primary rat hepatocytes. Initial analyses of inflammatory marker genes were performed in fractionated epididymal or inguinal adipose tissues. Expressions of inflammation related genes (IL-6, TNF-α, COX-2) were higher in the inguinal than the epididymal ATE. Similarly, expressions of marker genes of macrophage and monocyte (MPEG-1, CD68, F4/80, CD64) were higher in the stromal vascular fraction (SVF) isolated from inguinal ATE than that from epididymal ATE. However, expressions of lipolysis related genes (ATGL, HSL, perilipin-1) were higher in the epididymal adipocytes than inguinal adipocytes. Moreover, secretion of IL-6 and PGE(2) was higher from inguinal ATEs than from epididymal ATEs. There was a trend that the total levels of IL-6, TNF-α and PGE(2) in the media from inguinal ATEs co-cultured with primary rat hepatocytes were higher than that in the media from epididymal ATEs co-cultured with hepatocytes, although the significant difference was only seen in PGE(2). Lipolysis, measured as glycerol release, was similar in the ATEs isolated from inguinal and epididymal adipose tissues when cultured alone, but the glycerol release was higher in the ATEs isolated from epididymal than from inguinal adipose tissue when co-cultured with hepatocytes. Compared to epididymal ATEs, the ATEs from inguinal adipose tissue elicited a stronger cytotoxic response and higher level of insulin resistance in the co-cultured hepatocytes. In conclusion, our results reveal depot-dependent effects of ATEs on co-cultured primary hepatocytes, which in part may be related to a more pronounced infiltration of stromal vascular cells (SVCs), particularly macrophages, in inguinal adipose tissue resulting in stronger responses in terms of hepatotoxicity and insulin-resistance.

  19. Effect of single intralesional treatment of surgically induced equine superficial digital flexor tendon core lesions with adipose-derived mesenchymal stromal cells: a controlled experimental trial.

    PubMed

    Geburek, Florian; Roggel, Florian; van Schie, Hans T M; Beineke, Andreas; Estrada, Roberto; Weber, Kathrin; Hellige, Maren; Rohn, Karl; Jagodzinski, Michael; Welke, Bastian; Hurschler, Christof; Conrad, Sabine; Skutella, Thomas; van de Lest, Chris; van Weeren, René; Stadler, Peter M

    2017-06-05

    Adipose tissue is a promising source of mesenchymal stromal cells (MSCs) for the treatment of tendon disease. The goal of this study was to assess the effect of a single intralesional implantation of adipose tissue-derived mesenchymal stromal cells (AT-MSCs) on artificial lesions in equine superficial digital flexor tendons (SDFTs). During this randomized, controlled, blinded experimental study, either autologous cultured AT-MSCs suspended in autologous inactivated serum (AT-MSC-serum) or autologous inactivated serum (serum) were injected intralesionally 2 weeks after surgical creation of centrally located SDFT lesions in both forelimbs of nine horses. Healing was assessed clinically and with ultrasound (standard B-mode and ultrasound tissue characterization) at regular intervals over 24 weeks. After euthanasia of the horses the SDFTs were examined histologically, biochemically and by means of biomechanical testing. AT-MSC implantation did not substantially influence clinical and ultrasonographic parameters. Histology, biochemical and biomechanical characteristics of the repair tissue did not differ significantly between treatment modalities after 24 weeks. Compared with macroscopically normal tendon tissue, the content of the mature collagen crosslink hydroxylysylpyridinoline did not differ after AT-MSC-serum treatment (p = 0.074) while it was significantly lower (p = 0.027) in lesions treated with serum alone. Stress at failure (p = 0.048) and the modulus of elasticity (p = 0.001) were significantly lower after AT-MSC-serum treatment than in normal tendon tissue. The effect of a single intralesional injection of cultured AT-MSCs suspended in autologous inactivated serum was not superior to treatment of surgically created SDFT lesions with autologous inactivated serum alone in a surgical model of tendinopathy over an observation period of 22 weeks. AT-MSC treatment might have a positive influence on collagen crosslinking of remodelling scar

  20. Functional Characterization of Preadipocytes Derived from Human Periaortic Adipose Tissue

    PubMed Central

    Camacho, Jaime; Duque, Juan; Carreño, Marisol; Acero, Edward; Pérez, Máximo; Ramirez, Sergio; Umaña, Juan; Obando, Carlos; Guerrero, Albert; Sandoval, Néstor; Rodríguez, Gina

    2017-01-01

    Adipose tissue can affect the metabolic control of the cardiovascular system, and its anatomic location can affect the vascular function differently. In this study, biochemical and phenotypical characteristics of adipose tissue from periaortic fat were evaluated. Periaortic and subcutaneous adipose tissues were obtained from areas surrounding the ascending aorta and sternotomy incision, respectively. Adipose tissues were collected from patients undergoing myocardial revascularization or mitral valve replacement surgery. Morphological studies with hematoxylin/eosin and immunohistochemical assay were performed in situ to quantify adipokine expression. To analyze adipogenic capacity, adipokine expression, and the levels of thermogenic proteins, adipocyte precursor cells were isolated from periaortic and subcutaneous adipose tissues and induced to differentiation. The precursors of adipocytes from the periaortic tissue accumulated less triglycerides than those from the subcutaneous tissue after differentiation and were smaller than those from subcutaneous adipose tissue. The levels of proteins involved in thermogenesis and energy expenditure increased significantly in periaortic adipose tissue. Additionally, the expression levels of adipokines that affect carbohydrate metabolism, such as FGF21, increased significantly in mature adipocytes induced from periaortic adipose tissue. These results demonstrate that precursors of periaortic adipose tissue in humans may affect cardiovascular events and might serve as a target for preventing vascular diseases. PMID:29209367

  1. Concise Review: Therapeutic Potential of Adipose Tissue-Derived Angiogenic Cells

    PubMed Central

    Brinchmann, Jan E.

    2012-01-01

    Inadequate blood supply to tissues is a leading cause of morbidity and mortality today. Ischemic symptoms caused by obstruction of arterioles and capillaries are currently not treatable by vessel replacement or dilatation procedures. Therapeutic angiogenesis, the treatment of tissue ischemia by promoting the proliferation of new blood vessels, has recently emerged as one of the most promising therapies. Neovascularization is most often attempted by introduction of angiogenic cells from different sources. Emerging evidence suggests that adipose tissue (AT) is an excellent reservoir of autologous cells with angiogenic potential. AT yields two cell populations of importance for neovascularization: AT-derived mesenchymal stromal cells, which likely act predominantly as pericytes, and AT-derived endothelial cells (ECs). In this concise review we discuss different physiological aspects of neovascularization, briefly present cells isolated from the blood and bone marrow with EC properties, and then discuss isolation and cell culture strategies, phenotype, functional capabilities, and possible therapeutic applications of angiogenic cells obtained from AT. PMID:23197872

  2. NANOG Plays a Hierarchical Role in the Transcription Network Regulating the Pluripotency and Plasticity of Adipose Tissue-Derived Stem Cells

    PubMed Central

    Pitrone, Maria; Pizzolanti, Giuseppe; Tomasello, Laura; Coppola, Antonina; Morini, Lorenzo; Pantuso, Gianni; Ficarella, Romina; Guarnotta, Valentina; Perrini, Sebastio; Giorgino, Francesco; Giordano, Carla

    2017-01-01

    The stromal vascular cell fraction (SVF) of visceral and subcutaneous adipose tissue (VAT and SAT) has increasingly come into focus in stem cell research, since these compartments represent a rich source of multipotent adipose-derived stem cells (ASCs). ASCs exhibit a self-renewal potential and differentiation capacity. Our aim was to study the different expression of the embryonic stem cell markers NANOG (homeobox protein NANOG), SOX2 (SRY (sex determining region Y)-box 2) and OCT4 (octamer-binding transcription factor 4) and to evaluate if there exists a hierarchal role in this network in ASCs derived from both SAT and VAT. ASCs were isolated from SAT and VAT biopsies of 72 consenting patients (23 men, 47 women; age 45 ± 10; BMI between 25 ± 5 and 30 ± 5 range) undergoing elective open-abdominal surgery. Sphere-forming capability was evaluated by plating cells in low adhesion plastic. Stem cell markers CD90, CD105, CD29, CD31, CD45 and CD146 were analyzed by flow cytometry, and the stem cell transcription factors NANOG, SOX2 and OCT4 were detected by immunoblotting and real-time PCR. NANOG, SOX2 and OCT4 interplay was explored by gene silencing. ASCs from VAT and SAT confirmed their mesenchymal stem cell (MSC) phenotype expressing the specific MSC markers CD90, CD105, NANOG, SOX2 and OCT4. NANOG silencing induced a significant OCT4 (70 ± 0.05%) and SOX2 (75 ± 0.03%) downregulation, whereas SOX2 silencing did not affect NANOG gene expression. Adipose tissue is an important source of MSC, and siRNA experiments endorse a hierarchical role of NANOG in the complex transcription network that regulates pluripotency. PMID:28545230

  3. Human platelet lysate as a fetal bovine serum substitute improves human adipose-derived stromal cell culture for future cardiac repair applications.

    PubMed

    Naaijkens, B A; Niessen, H W M; Prins, H-J; Krijnen, P A J; Kokhuis, T J A; de Jong, N; van Hinsbergh, V W M; Kamp, O; Helder, M N; Musters, R J P; van Dijk, A; Juffermans, L J M

    2012-04-01

    Adipose-derived stromal cells (ASC) are promising candidates for cell therapy, for example to treat myocardial infarction. Commonly, fetal bovine serum (FBS) is used in ASC culturing. However, FBS has several disadvantages. Its effects differ between batches and, when applied clinically, transmission of pathogens and antibody development against FBS are possible. In this study, we investigated whether FBS can be substituted by human platelet lysate (PL) in ASC culture, without affecting functional capacities particularly important for cardiac repair application of ASC. We found that PL-cultured ASC had a significant 3-fold increased proliferation rate and a significantly higher attachment to tissue culture plastic as well as to endothelial cells compared with FBS-cultured ASC. PL-cultured ASC remained a significant 25% smaller than FBS-cultured ASC. Both showed a comparable surface marker profile, with the exception of significantly higher levels of CD73, CD90, and CD166 on PL-cultured ASC. PL-cultured ASC showed a significantly higher migration rate compared with FBS-cultured ASC in a transwell assay. Finally, FBS- and PL-cultured ASC had a similar high capacity to differentiate towards cardiomyocytes. In conclusion, this study showed that culturing ASC is more favorable in PL-supplemented medium compared with FBS-supplemented medium.

  4. Implications of adipose-derived stromal cells in a 3D culture system for osteogenic differentiation: an in vitro and in vivo investigation.

    PubMed

    Shen, Francis H; Werner, Brian C; Liang, Haixiang; Shang, Hulan; Yang, Ning; Li, Xudong; Shimer, Adam L; Balian, Gary; Katz, Adam J

    2013-01-01

    Healthy mammalian cells in normal tissues are organized in complex three-dimensional (3D) networks that display nutrient and signaling gradients. Conventional techniques that grow cells in a 2D monolayer fail to reproduce the environment that is observed in vivo. In recent years, 3D culture systems have been used to mimic tumor microenvironments in cancer research and to emulate embryogenesis in stem cell cultures. However, there have been no studies exploring the ability for adipose-derived stromal (ADS) cells in a 3D culture system to undergo osteogenic differentiation. To characterize and investigate the in vitro and in vivo potential for human ADS cells in a novel 3D culture system to undergo osteogenic differentiation. Basic science and laboratory study. Human ADS cells were isolated and prepared as either a 2D monolayer or 3D multicellular aggregates (MAs). Multicellular aggregates were formed using the hanging droplet technique. Cells were treated in osteogenic medium in vitro, and cellular differentiation was investigated using gene expression, histology, and microCT at 1-, 2-, and 4-week time points. In vivo investigation involved creating a muscle pouch by developing the avascular muscular interval in the vastus lateralis of male athymic rats. Specimens were then pretreated with osteogenic medium and surgically implanted as (1) carrier (Matrigel) alone (control), (2) carrier with human ADS cells in monolayer, or (3) human ADS cells as MAs. In vivo evidence of osteogenic differentiation was evaluated with micro computed tomography and histologic sectioning at a 2-week time point. Human ADS cells cultured by the hanging droplet technique successfully formed MAs at the air-fluid interface. Adipose-derived stromal cells cultured in monolayer or as 3D MAs retain their ability to self-replicate and undergo multilineage differentiation as confirmed by increased runx2/Cbfa2, ALP, and OCN and increased matrix mineralization on histologic sectioning

  5. Progranulin, a Major Secreted Protein of Mouse Adipose-Derived Stem Cells, Inhibits Light-Induced Retinal Degeneration

    PubMed Central

    Tsuruma, Kazuhiro; Yamauchi, Mika; Sugitani, Sou; Otsuka, Tomohiro; Ohno, Yuta; Nagahara, Yuki; Ikegame, Yuka; Shimazawa, Masamitsu; Yoshimura, Shinichi; Iwama, Toru

    2014-01-01

    Adipose tissue stromal vascular fraction contains mesenchymal stem cells, which show protective effects when administered to damaged tissues, mainly through secreted trophic factors. We examined the protective effects of adipose-derived stem cells (ASCs) and ASC-conditioned medium (ASC-CM) against retinal damage and identified the neuroprotective factors in ASC-CM. ASCs and mature adipocytes were isolated from mouse subcutaneous tissue. ASCs were injected intravitreally in a mouse model of light-induced retinal damage, and ASC injection recovered retinal function as measured by electroretinogram and inhibited outer nuclear layer, thinning, without engraftment of ASCs. ASC-CM and mature adipocyte-conditioned medium were collected after 72 hours of culture. In vitro, H2O2- and light-induced cell death was reduced in a photoreceptor cell line with ASC-CM but not with mature adipocyte-conditioned medium. In vivo, light-induced photoreceptor damage was evaluated by measurement of outer nuclear layer thickness at 5 days after light exposure and by electroretinogram recording. ASC-CM significantly inhibited photoreceptor degeneration and retinal dysfunction after light exposure. Progranulin was identified as a major secreted protein of ASCs that showed protective effects against retinal damage in vitro and in vivo. Furthermore, progranulin phosphorylated extracellular signal-regulated kinase, cAMP response element binding protein, and hepatocyte growth factor receptor, and protein kinase C signaling pathways were involved in the protective effects of progranulin. These findings suggest that ASC-CM and progranulin have neuroprotective effects in the light-induced retinal-damage model. Progranulin may be a potential target for the treatment of the degenerative diseases of the retina. PMID:24233842

  6. Progranulin, a major secreted protein of mouse adipose-derived stem cells, inhibits light-induced retinal degeneration.

    PubMed

    Tsuruma, Kazuhiro; Yamauchi, Mika; Sugitani, Sou; Otsuka, Tomohiro; Ohno, Yuta; Nagahara, Yuki; Ikegame, Yuka; Shimazawa, Masamitsu; Yoshimura, Shinichi; Iwama, Toru; Hara, Hideaki

    2014-01-01

    Adipose tissue stromal vascular fraction contains mesenchymal stem cells, which show protective effects when administered to damaged tissues, mainly through secreted trophic factors. We examined the protective effects of adipose-derived stem cells (ASCs) and ASC-conditioned medium (ASC-CM) against retinal damage and identified the neuroprotective factors in ASC-CM. ASCs and mature adipocytes were isolated from mouse subcutaneous tissue. ASCs were injected intravitreally in a mouse model of light-induced retinal damage, and ASC injection recovered retinal function as measured by electroretinogram and inhibited outer nuclear layer, thinning, without engraftment of ASCs. ASC-CM and mature adipocyte-conditioned medium were collected after 72 hours of culture. In vitro, H2O2- and light-induced cell death was reduced in a photoreceptor cell line with ASC-CM but not with mature adipocyte-conditioned medium. In vivo, light-induced photoreceptor damage was evaluated by measurement of outer nuclear layer thickness at 5 days after light exposure and by electroretinogram recording. ASC-CM significantly inhibited photoreceptor degeneration and retinal dysfunction after light exposure. Progranulin was identified as a major secreted protein of ASCs that showed protective effects against retinal damage in vitro and in vivo. Furthermore, progranulin phosphorylated extracellular signal-regulated kinase, cAMP response element binding protein, and hepatocyte growth factor receptor, and protein kinase C signaling pathways were involved in the protective effects of progranulin. These findings suggest that ASC-CM and progranulin have neuroprotective effects in the light-induced retinal-damage model. Progranulin may be a potential target for the treatment of the degenerative diseases of the retina.

  7. Human Adipose Mesenchymal Cells Inhibit Melanocyte Differentiation and the Pigmentation of Human Skin via Increased Expression of TGF-β1.

    PubMed

    Klar, Agnes S; Biedermann, Thomas; Michalak, Katarzyna; Michalczyk, Teresa; Meuli-Simmen, Claudia; Scherberich, Arnaud; Meuli, Martin; Reichmann, Ernst

    2017-12-01

    There is accumulating evidence that interactions between epidermal melanocytes and stromal cells play an important role in the regulation of skin pigmentation. In this study we established a pigmented dermo-epidermal skin model, melDESS, of human origin to investigate the effects of distinct stromal cells on melanogenesis. melDESS is a complex, clinically relevant skin equivalent composed of an epidermis containing both melanocytes and keratinocytes. Its dermal compartment consists either of adipose tissue-derived stromal cells, dermal fibroblasts (Fbs), or a mixture of both cell types. These skin substitutes were transplanted for 5 weeks on the backs of immuno-incompetent rats and analyzed. Gene expression and Western blot analyses showed a significantly higher expression of transforming growth factor-β1 by adipose tissue-derived stromal cells compared with dermal Fbs. In addition, we showed that melanocytes responded to the increased levels of transforming growth factor-β1 by down-regulating the expression of key melanogenic enzymes such as tyrosinase. This caused decreased melanin synthesis and, consequently, greatly reduced pigmentation of melDESS. The conclusions are of utmost clinical relevance, namely that adipose tissue-derived stromal cells derived from the hypodermis fail to appropriately interact with epidermal melanocytes, thus preventing the sustainable restoration of the patient's native skin color in bioengineered skin grafts. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  8. Lipopolysaccharide induces proliferation and osteogenic differentiation of adipose-derived mesenchymal stromal cells in vitro via TLR4 activation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Herzmann, Nicole; Salamon, Achim; Fiedler, Tomas

    Multipotent mesenchymal stromal cells (MSC) are capable of multi-lineage differentiation and support regenerative processes. In bacterial infections, resident MSC can come intocontact with and need to react to bacterial components. Lipopolysaccharide (LPS), a typical structure of Gram-negative bacteria, increases the proliferation and osteogenic differentiation of MSC. LPS is usually recognized by the toll-like receptor (TLR) 4 and induces pro-inflammatory reactions in numerous cell types. In this study, we quantified the protein expression of TLR4 and CD14 on adipose-derived MSC (adMSC) in osteogenic differentiation and investigated the effect of TLR4 activation by LPS on NF-κB activation, proliferation and osteogenic differentiation ofmore » adMSC. We found that TLR4 is expressed on adMSC whereas CD14 is not, and that osteogenic differentiation induced an increase of the amount of TLR4 protein whereas LPS stimulation did not. Moreover, we could show that NF-κB activation via TLR4 occurs upon LPS treatment. Furthermore, we were able to show that competitive inhibition of TLR4 completely abolished the stimulatory effect of LPS on the proliferation and osteogenic differentiation of adMSC. In addition, the inhibition of TLR4 leads to the complete absence of osteogenic differentiation of adMSC, even when osteogenically stimulated. Thus, we conclude that LPS induces proliferation and osteogenic differentiation of adMSC in vitro through the activation of TLR4 and that the TLR4 receptor seems to play a role during osteogenic differentiation of adMSC.« less

  9. Molecular Validation of Chondrogenic Differentiation and Hypoxia Responsiveness of Platelet-Lysate Expanded Adipose Tissue-Derived Human Mesenchymal Stromal Cells.

    PubMed

    Galeano-Garces, Catalina; Camilleri, Emily T; Riester, Scott M; Dudakovic, Amel; Larson, Dirk R; Qu, Wenchun; Smith, Jay; Dietz, Allan B; Im, Hee-Jeong; Krych, Aaron J; Larson, A Noelle; Karperien, Marcel; van Wijnen, Andre J

    2017-07-01

    To determine the optimal environmental conditions for chondrogenic differentiation of human adipose tissue-derived mesenchymal stromal/stem cells (AMSCs). In this investigation we specifically investigate the role of oxygen tension and 3-dimensional (3D) culture systems. Both AMSCs and primary human chondrocytes were cultured for 21 days in chondrogenic media under normoxic (21% oxygen) or hypoxic (2% oxygen) conditions using 2 distinct 3D culture methods (high-density pellets and poly-ε-caprolactone [PCL] scaffolds). Histologic analysis of chondro-pellets and the expression of chondrocyte-related genes as measured by reverse transcriptase quantitative polymerase chain reaction were used to evaluate the efficiency of differentiation. AMSCs are capable of expressing established cartilage markers including COL2A1, ACAN, and DCN when grown in chondrogenic differentiation media as determined by gene expression and histologic analysis of cartilage markers. Expression of several cartilage-related genes was enhanced by low oxygen tension, including ACAN and HAPLN1. The pellet culture environment also promoted the expression of hypoxia-inducible cartilage markers compared with cells grown on 3D scaffolds. Cell type-specific effects of low oxygen and 3D environments indicate that mesenchymal cell fate and differentiation potential is remarkably sensitive to oxygen. Genetic programming of AMSCs to a chondrocytic phenotype is effective under hypoxic conditions as evidenced by increased expression of cartilage-related biomarkers and biosynthesis of a glycosaminoglycan-positive matrix. Lower local oxygen levels within cartilage pellets may be a significant driver of chondrogenic differentiation.

  10. Adipose stromal cells improve healing of vocal fold scar: Morphological and functional evidences.

    PubMed

    de Bonnecaze, Guillaume; Chaput, Benoit; Woisard, Virginie; Uro-Coste, Emmanuelle; Swider, Pascal; Vergez, Sebastien; Serrano, Elie; Casteilla, Louis; Planat-Benard, Valerie

    2016-08-01

    Adipose derived stromal cells (ASCs) are abundant and easy to prepare. Such cells may be useful for treating severe vocal disturbance caused by acute vocal fold scars. Prospective animal experiments with controls. Twenty New-Zealand white rabbits were used in the present study. We evaluated vocal fold healing, with or without injection of autologous ASCs, after acute scarring. A defined lesion was created and the ASCs were immediately injected. Vocal fold regeneration was evaluated histomorphometrically and via viscoelastic analysis using an electrodynamic shaker. Six weeks after ASC injection, vocal folds exhibited significantly less inflammation than control folds (P < 0.005). In addition, hypertrophy of the lamina propria and fibrosis were significantly reduced upon ASC injection (P < 0.02). The decrease in viscoelastic parameters was less important in the ASC injected group compared to the noninjected group (P = 0.08). Injection of autologous ASCs improved vocal fold healing in our preclinical model. Further studies are needed, but this method may be useful in humans. NA. Laryngoscope, 126:E278-E285, 2016. © 2016 The American Laryngological, Rhinological and Otological Society, Inc.

  11. Stromal Vascular Fraction from Lipoaspirate Infranatant: Comparison Between Suction-Assisted Liposuction and Nutational Infrasonic Liposuction.

    PubMed

    Bowen, Robert E

    2016-06-01

    Lipoaspirate has shown great promise as a source of progenitor cells for use in regenerative medicine. The stromal vascular fraction (SVF) can be isolated from lipoaspirate using enzyme digestion and centrifugation, but this approach may be limited by the labor-intensive nature of the technique as well as ambiguities in current governmental regulations. An alternative approach to obtain SVF from lipoaspirate was studied. Paired (collected from contralateral regions) lipoaspirate specimens were acquired from 30 consenting patients (age 24-62; 22 females, 8 males) by suction-assisted liposuction (SAL) and nutational infrasonic liposuction (NIL). The infranatant from 50 ml of adipose tissue (LAF) was centrifuged at 400g × 5 min and the resultant pellet was collected with a pipette. Time = 15-20 min. The respective SVFs cell populations were counted using an optical fluorescent cell counter (Nexcelom A2000) and the fluorescent stains-acridine orange (AO) and propidium iodide (PI). The number of nucleated, live cells from SAL infranatant was 97,345 ± 23,435 per ml of adipose tissue and from NIL infranatant was 335,621 ± 81,274 per ml of adipose tissue. The p value is <0.00001, n = 30. Regenerative cells can be isolated from the lipoaspirate infranatant from either SAL or NIL, although in lower quantities than from enzyme digestion. NIL acquisition yielded 3.5× the number of cells over that acquired from SAL. The time, skill, and cost of producing SVF from infranatant is less than using enzyme digestion, which potentially make these regenerative therapies accessible to more physicians and patients. This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

  12. A clinical scalable cryopreservation method of adipose tissue for reconstructive surgery assessed by stromal vascular fraction and mice studies.

    PubMed

    Chaput, Benoit; Orio, Julie; Garrido, Ignacio; De Bonnecaze, Guillaume; Espagnolle, Nicolas; Gadelorge, Melanie; Chavoin, Jean-Pierre; Grolleau-Raoux, Jean-Louis; Casteilla, Louis; Planat, Valérie; Bourin, Philippe

    2014-04-01

    Adipose tissue is widely used in plastic surgery. The main obstacle is that it can be used only immediately after liposuction, while reconstruction often requires several procedures to achieve optimal results. This study aimed to develop a cryopreservation protocol directly applicable to clinical situations, allowing repetitive procedures without multiple tissue harvests. The authors first tested scalable bags suitable for therapeutic uses. All subsequent experiments were performed in those bags. The authors evaluated in vitro, on the basis of cell viability, cell number, phenotype, and stromal cell proliferation, the efficacy of six cryopreservation media composed of an external cryoprotectant (human albumin or hydroxylethyl starch) with or without an internal cryoprotectant (dimethyl sulfoxide). Two storage temperatures (-196°C and -80°C) were tested in vitro and in vivo (subcutaneous graft in 30 nude mice) with the selected medium. The combination of 5% dimethyl sulfoxide and 95% hydroxylethyl yielded in vitro results that were good and the most consistent. With this cryoprotective solution, the authors observed no significant difference in vitro for a storage period of 7 days. When the storage was extended to 1 month, the cell viability was decreased by 10 percent for both storage temperatures. The in vivo experiments assessed the superiority of cryopreservation at -80°C with less graft resorption (60 percent and 70 percent, respectively, for -80°C and -196°C) and less fibrosis. The study's protocol with a chemically defined cryoprotective solution, specific scalable bags constrained in an aluminum holder, and a storage temperature of -80°C is promising for long-term adipose tissue cryopreservation.

  13. Short-term preconditioning enhances the therapeutic potential of adipose-derived stromal/stem cell-conditioned medium in cisplatin-induced acute kidney injury.

    PubMed

    Overath, Jürgen M; Gauer, Stefan; Obermüller, Nicholas; Schubert, Ralf; Schäfer, Richard; Geiger, Helmut; Baer, Patrick C

    2016-03-15

    The development of new strategies to preserve renal function after acute kidney injury (AKI) is necessary due to limited clinical intervention options. The organ-protective effects of mesenchymal stromal/stem cells (MSCs) and their conditioned medium (CM) have been investigated demonstrating that both separately promoted tubular recovery and ameliorated the outcome of AKI. Nevertheless, strategies to optimise the regenerative potential of both are highly needed. Here we investigated the effects of CM from adipose-derived MSCs (ASCs) preincubated in a hypoxic environment (Hyp). Protective factors were investigated by PCR analysis and a protein array in vitro. The expression of 64 of the 308 proteins assayed was found to be more than two-fold increased after Hyp. CM of Hyp-pretreated ASCs (pCM) was used to enhance regeneration in a mouse model of cisplatin-induced AKI (cisAKI). Renal function was assessed by measurements of markers for AKI and serum cytokine levels. The pCM significantly ameliorated serum creatinine and neutrophil gelatinase-associated lipocalin values, and also the levels of inflammatory cytokines IL-1β and IL-6 in the serum of mice with AKI. Our work clearly showed that a Hyp preconditioning significantly increases the release of protective factors in ASCs and enhances the therapeutic effects of CM in cisAKI in mice. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Effect of Adipose Tissue-Derived Osteogenic and Endothelial Cells on Bone Allograft Osteogenesis and Vascularization in Critical-Sized Calvarial Defects

    DTIC Science & Technology

    2012-05-10

    1% peni - cillin/streptomycin, and 50 ng/mL recombinant rat VEGF-C (Promocell, Heidelberg, Germany). The media were changed every other day for 8...various animal models that have demonstrated an enhanced osteogenic effect after treating bone allografts with adipose tissue or bone marrow-derived... enhanced 1560 CORNEJO ET AL. performance of bone allografts using osteogenic differentiated adipose derived mesenchymal stem cells. Biomaterials 32, 8880

  15. Senescence and quiescence in adipose-derived stromal cells: Effects of human platelet lysate, fetal bovine serum and hypoxia.

    PubMed

    Søndergaard, Rebekka Harary; Follin, Bjarke; Lund, Lisbeth Drozd; Juhl, Morten; Ekblond, Annette; Kastrup, Jens; Haack-Sørensen, Mandana

    2017-01-01

    Adipose-derived stromal cells (ASCs) are attractive sources for cell-based therapies. The hypoxic niche of ASCs in vivo implies that cells will benefit from hypoxia during in vitro expansion. Human platelet lysate (hPL) enhances ASC proliferation rates, compared with fetal bovine serum (FBS) at normoxia. However, the low proliferation rates of FBS-expanded ASCs could be signs of senescence or quiescence. We aimed to determine the effects of hypoxia and hPL on the expansion of ASCs and whether FBS-expanded ASCs are senescent or quiescent. ASCs expanded in FBS or hPL at normoxia or hypoxia until passage 7 (P7), or in FBS until P5 followed by culture in hPL until P7, were evaluated by proliferation rates, cell cycle analyses, gene expression and β-galactosidase activity. hPL at normoxia and hypoxia enhanced proliferation rates and expression of cyclins, and decreased G0/G1 fractions and expression of p21 and p27, compared with FBS. The shift from FBS to hPL enhanced cyclin levels, decreased p21 and p27 levels and tended to decrease G0/G1 fractions. Hypoxia does not add to the effect of hPL during ASC expansion with regard to proliferation, cell cycle regulation and expression of cyclins, p21 and p27. hPL rejuvenates FBS-expanded ASCs with regard to cell cycle regulation and expression of cyclins, p21 and p27. This indicates a reversible arrest. Therefore, we conclude that ASCs expanded until P7 are not senescent regardless of culture conditions. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  16. Comparison of human platelet lysate alternatives using expired and freshly isolated platelet concentrates for adipose-derived stromal cell expansion.

    PubMed

    Dessels, Carla; Durandt, Chrisna; Pepper, Michael S

    2018-03-19

    Pooled human platelet lysate (pHPL) has been used to expand adipose-derived stromal cells (ASCs) and can be formulated using fresh or expired buffy coats (BCs) which are then resuspended in either plasma or an additive solution. Not much is known about the effects that expired products and additive solutions have on ASC expansion, and the need for quality control and release criteria has been expressed. This pilot study compared proliferation, cell size, morphology and immunophenotype of ASCs expanded in the different pHPL alternatives versus foetal bovine serum (FBS). Quality control criteria were assessed prior to and during the manufacture of the pHPL alternatives. ASCs were then expanded in 1%, 2.5%, 5% or 10% of the different pHPL alternatives or in 10% FBS. Cell size, morphology, cell number and immunophenotype were measured using microscopy and flow cytometry. The majority of the pHPL alternatives were within the recommended ranges for the quality control criteria. ASCs expanded in the pHPL alternatives were smaller in size, displayed a tighter spindle-shaped morphology, increased cell growth and had a similar immunophenotype (with the exception of CD34 and CD36) when compared to ASCs expanded in FBS. Here we report on the effects that expired BC products and additive solutions have on ASC expansion. When taken together, our findings indicate that all of the pHPL alternatives can be considered to be suitable replacements for FBS for ASC expansion, and that expired BC products can be used as an alternative to fresh BC products.

  17. Human adipose derived mesenchymal stromal cells transduced with GFP lentiviral vectors: assessment of immunophenotype and differentiation capacity in vitro.

    PubMed

    van Vollenstee, Fiona A; Jackson, Carlo; Hoffmann, Danie; Potgieter, Marnie; Durandt, Chrisna; Pepper, Michael S

    2016-10-01

    Adipose derived mesenchymal stromal/stem cells (ASCs) are a heterogeneous population characterized by (a) their ability to adhere to plastic; (b) immunophenotypic expression of certain cell surface markers, while lacking others; and (c) the capacity to differentiate into lineages of mesodermal origin including osteocytes, chondrocytes and adipocytes. The long-term goal is to utilize these cells for clinical translation into cell-based therapies. However, preclinical safety and efficacy need to be demonstrated in animal models. ASCs can also be utilized as biological vehicles for vector-based gene delivery systems, since they are believed to home to sites of inflammation and infection in vivo. These factors motivated the development of a labelling system for ASCs using lentiviral vector-based green fluorescent protein (GFP) transduction. Human ASCs were transduced with GFP-expressing lentiviral vectors. A titration study determined the viral titer required to transduce the maximum number of ASCs. The effect of the transduced GFP lentiviral vector on ASC immunophenotypic expression of surface markers as well as their ability to differentiate into osteocytes and adipocytes were assessed in vitro. A transduction efficiency in ASC cultures of approximately 80 % was observed with an MOI of ~118. No significant immunophenotypic differences were observed between transduced and non-transduced cells and both cell types successfully differentiated into adipocytes and osteocytes in vitro. We obtained >80 % transduction of ASCs using GFP lentiviral vectors. Transduced ASCs maintained plastic adherence, demonstrated ASC immunophenotype and the ability to differentiate into cells of the mesodermal lineage. This GFP-ASC transduction technique offers a potential tracking system for future pre-clinical studies.

  18. Effect of Adipose-Derived Stromal Cells and BMP12 on Intrasynovial Tendon Repair: A Biomechanical, Biochemical, and Proteomics Study

    PubMed Central

    Gelberman, Richard H.; Shen, Hua; Kormpakis, Ioannis; Rothrauff, Benjamin; Yang, Guang; Tuan, Rocky S.; Xia, Younan; Sakiyama-Elbert, Shelly; Silva, Matthew J.; Thomopoulos, Stavros

    2016-01-01

    The outcomes of flexor tendon repair are highly variable. As recent efforts to improve healing have demonstrated promise for growth factor- and cell-based therapies, the objective of the current study was to enhance repair via application of autologous adipose derived stromal cells (ASCs) and the tenogenic growth factor bone morphogenetic protein (BMP) 12. Controlled delivery of cells and growth factor was achieved in a clinically relevant canine model using a nanofiber/fibrin-based scaffold. Control groups consisted of repair-only (no scaffold) and acellular scaffold. Repairs were evaluated after 28 days of healing using biomechanical, biochemical, and proteomics analyses. Range of motion was reduced in the groups that received scaffolds compared to normal. There was no effect of ASC+BMP12 treatment for range of motion or tensile properties outcomes versus repair-only. Biochemical assays demonstrated increased DNA, glycosaminoglycans, and crosslink concentration in all repair groups compared to normal, but no effect of ASC+BMP12. Total collagen was significantly decreased in the acellular scaffold group compared to normal and significantly increased in the ASC+BMP12 group compared to the acellular scaffold group. Proteomics analysis comparing healing tendons to uninjured tendons revealed significant increases in proteins associated with inflammation, stress response, and matrix degradation. Treatment with ASC+BMP12 amplified these unfavorable changes. In summary, the treatment approach used in this study induced a negative inflammatory reaction at the repair site leading to poor healing. Future approaches should consider cell and growth factor delivery methods that do not incite negative local reactions. PMID:26445383

  19. Engineering vascularized soft tissue flaps in an animal model using human adipose–derived stem cells and VEGF+PLGA/PEG microspheres on a collagen-chitosan scaffold with a flow-through vascular pedicle

    PubMed Central

    Zhang, Qixu; Hubenak, Justin; Iyyanki, Tejaswi; Alred, Erik; Turza, Kristin C.; Davis, Greg; Chang, Edward I.; Branch-Brooks, Cynthia D.; Beahm, Elisabeth K.; Butler, Charles E.

    2015-01-01

    Insufficient neovascularization is associated with high levels of resorption and necrosis in autologous and engineered fat grafts. We tested the hypothesis that incorporating angiogenic growth factor into a scaffold–stem cell construct and implanting this construct around a vascular pedicle improves neovascularization and adipogenesis for engineering soft tissue flaps. Poly(lactic-co-glycolic-acid/polyethylene glycol (PLGA/PEG) microspheres containing vascular endothelial growth factor (VEGF) were impregnated into collagen-chitosan scaffolds seeded with human adipose-derived stem cells (hASCs). This setup was analyzed in vitro and then implanted into isolated chambers around a discrete vascular pedicle in nude rats. Engineered tissue samples within the chambers were harvested and analyzed for differences in vascularization and adipose tissue growth. In vitro testing showed that the collagen-chitosan scaffold provided a supportive environment for hASC integration and proliferation. PLGA/PEG microspheres with slow-release VEGF had no negative effect on cell survival in collagen-chitosan scaffolds. In vivo, the system resulted in a statistically significant increase in neovascularization that in turn led to a significant increase in adipose tissue persistence after 8 weeks versus control constructs. These data indicate that our model—hASCs integrated with a collagen-chitosan scaffold incorporated with VEGF-containing PLGA/PEG microspheres supported by a predominant vascular vessel inside a chamber—provides a promising, clinically translatable platform for engineering vascularized soft tissue flap. The engineered adipose tissue with a vascular pedicle could conceivably be transferred as a vascularized soft tissue pedicle flap or free flap to a recipient site for the repair of soft-tissue defects. PMID:26410787

  20. Local delivery of allogeneic bone marrow and adipose tissue-derived mesenchymal stromal cells for cutaneous wound healing in a porcine model.

    PubMed

    Hanson, Summer E; Kleinbeck, Kyle R; Cantu, David; Kim, Jaeyhup; Bentz, Michael L; Faucher, Lee D; Kao, W John; Hematti, Peiman

    2016-02-01

    Wound healing remains a major challenge in modern medicine. Bone marrow- (BM) and adipose tissue- (AT) derived mesenchymal stromal/stem cells (MSCs) are of great interest for tissue reconstruction due to their unique immunological properties and regenerative potential. The purpose of this study was to characterize BM and AT-MSCs and evaluate their effect when administered in a porcine wound model. MSCs were derived from male Göttingen Minipigs and characterized according to established criteria. Allogeneic BM- or AT-MSCs were administered intradermally (1 x 10(6) cells) into partial-thickness wounds created on female animals, and covered with Vaseline® gauze or fibrin in a randomized pattern. Animals were euthanized at 7, 10, 14 and 21 days. Tissues were analyzed visually for healing and by microscopic examination for epidermal development and remodelling. Polymerase chain reaction (PCR) was used to detect the presence of male DNA in the specimens. All wounds were healed by 14 days. MSC-injected wounds were associated with improved appearance and faster re-epithelialization compared to saline controls. Evaluation of rete ridge depth and architecture showed that MSC treatment promoted a faster rate of epidermal maturation. Male DNA was detected in all samples at days 7 and 10, suggesting the presence of MSCs. We showed the safety, feasibility and potential efficacy of local injection of allogeneic BM- and AT-MSCs for treatment of wounds in a preclinical model. Our data in this large animal model support the potential use of BM- and AT-MSC for treatment of cutaneous wounds through modulation of healing and epithelialization. Copyright © 2013 John Wiley & Sons, Ltd.

  1. Different response to hypoxia of adipose-derived multipotent cells from obese subjects with and without metabolic syndrome

    PubMed Central

    Moreno-Indias, Isabel; Coín-Aragüez, Leticia; Lhamyani, Said; Alcaide Torres, Juan; Fernández-Veledo, Sonia; Vendrell, Joan; Camargo, Antonio; El Bekay, Rajaa; Tinahones, Francisco José

    2017-01-01

    Background/Objectives Multiple studies suggest that hypoxia, together with inflammation, could be one of the phenomena involved in the onset and progression of obesity-related insulin resistance. In addition, dysfunction of adipose tissue in obese subjects with metabolic syndrome is associated with decreased angiogenesis. However, some subjects with a high body mass index do not develop metabolic abnormalities associated with obesity. The aim of the current study was to examine the neovascular properties of visceral adipose tissue-derived multipotent mesenchymal cells subjected to hypoxia (hypox-visASCs) from normal-weight subjects (Nw) and obese patients with metabolic syndrome (MS) and without metabolic syndrome (NonMS). Methods This was a 2-year study to enroll subjects who underwent bariatric surgery or cholecystectomy. Eight patients who underwent either bariatric surgery or cholecystectomy (27 patients) participated in the study. Visceral adipose tissue samples from Nw, MS and NonMS subjects were processed by enzymatic digestion. VisASCs cultured under hypoxic conditions were characterized by tubule formation assay, ELISA, flow cytometry, migration rate, and qRT-PCR, and the effects of visASCs-conditioned medium on survival and endothelial cell tubule formation were evaluated. Results Hypox-visASCs from NonMS subjects showed a greater capacity for tubule formation than hypox-visASCs from Nw and MS subjects. The lower percentage of CD140b+/CD44+ and CD140b+/CD184+ cells observed in hypox-visASCs from NonMS subjects compared to MS subjects was accompanied not only by a lower migration rate from the chemotactic effects of stromal cell derived factor 1α, but also by lower levels of NOX5 mRNA expression. While the levels of monocyte chemoattractant protein 1 mRNA expressed by hypox-visASCs correlated positively with the body mass index and waist circumference of the subjects, the concentration of vascular endothelial growth factor present in hypox

  2. Magnetic Resonance Imaging of Human Tissue-Engineered Adipose Substitutes

    PubMed Central

    Proulx, Maryse; Aubin, Kim; Lagueux, Jean; Audet, Pierre; Auger, Michèle

    2015-01-01

    Adipose tissue (AT) substitutes are being developed to answer the strong demand in reconstructive surgery. To facilitate the validation of their functional performance in vivo, and to avoid resorting to excessive number of animals, it is crucial at this stage to develop biomedical imaging methodologies, enabling the follow-up of reconstructed AT substitutes. Until now, biomedical imaging of AT substitutes has scarcely been reported in the literature. Therefore, the optimal parameters enabling good resolution, appropriate contrast, and graft delineation, as well as blood perfusion validation, must be studied and reported. In this study, human adipose substitutes produced from adipose-derived stem/stromal cells using the self-assembly approach of tissue engineering were implanted into athymic mice. The fate of the reconstructed AT substitutes implanted in vivo was successfully followed by magnetic resonance imaging (MRI), which is the imaging modality of choice for visualizing soft ATs. T1-weighted images allowed clear delineation of the grafts, followed by volume integration. The magnetic resonance (MR) signal of reconstructed AT was studied in vitro by proton nuclear magnetic resonance (1H-NMR). This confirmed the presence of a strong triglyceride peak of short longitudinal proton relaxation time (T1) values (200±53 ms) in reconstructed AT substitutes (total T1=813±76 ms), which establishes a clear signal difference between adjacent muscle, connective tissue, and native fat (total T1 ∼300 ms). Graft volume retention was followed up to 6 weeks after implantation, revealing a gradual resorption rate averaging at 44% of initial substitute's volume. In addition, vascular perfusion measured by dynamic contrast-enhanced-MRI confirmed the graft's vascularization postimplantation (14 and 21 days after grafting). Histological analysis of the grafted tissues revealed the persistence of numerous adipocytes without evidence of cysts or tissue necrosis. This study

  3. Regulation of adipose-tissue-derived stromal cell orientation and motility in 2D- and 3D-cultures by direct-current electrical field.

    PubMed

    Yang, Gang; Long, Haiyan; Ren, Xiaomei; Ma, Kunlong; Xiao, Zhenghua; Wang, Ying; Guo, Yingqiang

    2017-02-01

    Cell alignment and motility play a critical role in a variety of cell behaviors, including cytoskeleton reorganization, membrane-protein relocation, nuclear gene expression, and extracellular matrix remodeling. Direct current electric field (EF) in vitro can direct many types of cells to align vertically to EF vector. In this work, we investigated the effects of EF stimulation on rat adipose-tissue-derived stromal cells (ADSCs) in 2D-culture on plastic culture dishes and in 3D-culture on various scaffold materials, including collagen hydrogels, chitosan hydrogels and poly(L-lactic acid)/gelatin electrospinning fibers. Rat ADSCs were exposed to various physiological-strength EFs in a homemade EF-bioreactor. Changes of morphology and movements of cells affected by applied EFs were evaluated by time-lapse microphotography, and cell survival rates and intracellular calcium oscillations were also detected. Results showed that EF facilitated ADSC morphological changes, under 6 V/cm EF strength, and that ADSCs in 2D-culture aligned vertically to EF vector and kept a good cell survival rate. In 3D-culture, cell galvanotaxis responses were subject to the synergistic effect of applied EF and scaffold materials. Fast cell movement and intracellular calcium activities were observed in the cells of 3D-culture. We believe our research will provide some experimental references for the future study in cell galvanotaxis behaviors. © 2017 Japanese Society of Developmental Biologists.

  4. Low-molecular weight heparin protamine complex augmented the potential of adipose-derived stromal cells to ameliorate limb ischemia.

    PubMed

    Kishimoto, Satoko; Inoue, Ken-Ichi; Nakamura, Shingo; Hattori, Hidemi; Ishihara, Masayuki; Sakuma, Masashi; Toyoda, Shigeru; Iwaguro, Hideki; Taguchi, Isao; Inoue, Teruo; Yoshida, Ken-Ichiro

    2016-06-01

    Heparin/protamine micro/nanoparticles (LH/P-MPs) were recently developed as low-molecular weight, biodegradable carriers for adipose-derived stromal cells (ADSCs). These particles can be used for a locally delivered stem cell therapy that promotes angiogenesis. LH/P-MPs bind to the cell surface of ADSCs and promote cell-to-cell interaction and aggregation of ADSCs. Cultured ADSC/LH/P-MP aggregates remain viable. Here, we examined the ability of these aggregates to rescue limb loss in a mouse model of hindlimb ischemia. Unilateral hindlimb ischemia was induced in adult male BALB/c mice by ligation of the iliac artery and hindlimb vein. For allotransplantation of ADSCs from the same inbred strain, we injected ADSC alone or ADSC/LH/P-MP aggregates or control medium (sham-treated) directly into the ischemic muscles. Ischemic limb blood perfusion, vessel density, and vessel area were recorded. The extent of ischemic limb necrosis or limb loss was assessed on postoperative days 2, 7, and 14. Compared with the sham-treatment control, treatment with ADSCs alone showed modest effects on blood perfusion recovery and increased the number of α-SMA-positive vessels. Response to ADSC/LH/P-MP aggregates was significantly greater than ADSCs alone for every endpoint. ADSC/LH/P-MP aggregates more effectively prevented the loss of ischemic hindlimbs than ADSCs alone or the sham-treatment. The LH/P-MPs augmented the effects of ADSCs on angiogenesis and reversal of limb ischemia. Use of ADSC/LH/P-MP aggregates offers a novel and convenient treatment method and potentially represents a promising new therapeutic approach to inducing angiogenesis in ischemic diseases. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  5. Enhanced angiogenic effect of adipose-derived stromal cell spheroid with low-level light therapy in hind limb ischemia mice.

    PubMed

    Park, In-Su; Chung, Phil-Sang; Ahn, Jin Chul

    2014-11-01

    The aim of this study was to investigate the effects of low-level laser therapy (LLLT) on transplanted human adipose-derived mesenchymal stem cells (hASCs) spheroid in a hind limb ischemia animal model. LLLT, hASCs spheroid and hASCs spheroid transplantation with LLLT (spheroid + LLLT) were applied to the ischemic hind limbs in athymic mice. The survival, differentiation and secretion of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (FGF), and hepatocyte growth factor (HGF) of the spheroid ASCs were evaluated by immunohistochemistry and western blots. Spheroid + LLLT group had enhanced the tissue regeneration, including angiogenesis, compared with the ASC group. The spheroid ASCs contributed to tissue regeneration via differentiation and secretion of growth factors. In the spheroid + LLLT group, the survival of spheroid hASCs increased with a concomitant decrease in apoptosis of spheroid hASCs in the ischemic hind limb. The secretion of growth factors was stimulated in the spheroid + LLLT group compared with the ASCs and spheroid group. These data suggested that LLLT is an effective biostimulator of spheroid hASCs in tissue regeneration that enhanced the survival of ASCs and stimulated the secretion of growth factors in the ischemic hind limb. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Microencapsulated VEGF gene-modified umbilical cord mesenchymal stromal cells promote the vascularization of tissue-engineered dermis: an experimental study.

    PubMed

    Han, Yanfu; Tao, Ran; Han, Yanqing; Sun, Tianjun; Chai, Jiake; Xu, Guang; Liu, Jing

    2014-02-01

    Tissue-engineered dermis (TED) is thought to be the best treatment for skin defect wounds; however, lack of vascular structures in these products can cause slow vascularization or even transplant failure. We assessed the therapeutic potential of microencapsulated human umbilical cord mesenchymal stromal cells (hUCMSCs) expressing vascular endothelial growth factor (VEGF) in vascularization of TED. hUCMSCs were isolated by means of enzymatic digestion and identified by means of testing biological characteristics. hUCMSCs were induced to differentiate into dermal fibroblasts in conditioned induction media. Collagen-chitosan laser drilling acellular dermal matrix (ADM) composite scaffold was prepared by means of the freeze dehydration and dehydrothermal cross-linking method. hUCMSC-derived fibroblasts were implanted on composite scaffolds to construct TED. TED with microencapsulated VEGF gene-modified hUCMSCs was then transplanted into skin defect wounds in pigs. The angiogenesis of TED at 1 week and status of wound healing at 3 weeks were observed. The collagen-chitosan laser ADM composite has a uniform microporous structure. This composite has been used to grow hUCMSC-derived fibroblasts in vitro and to successfully construct stem cell-derived TED. Microencapsulated VEGF gene-modified hUCMSCs were prepared with the use of a sodium alginate-barium chloride one-step encapsulation technology. Seven days after the transplantation of the stem cell-derived TED and microencapsulated VEGF gene-modified hUCMSCs into the skin defect wounds on the backs of miniature pigs, the VEGF expression increased and the TED had a higher degree of vascularization. Re-epithelialization of the wound was completed after 3 weeks. Microencapsulated VEGF gene-modified hUCMSCs can effectively improve the vascularization of TED and consequently the quality of wound healing. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  7. Adipose-derived stem cells were impaired in restricting CD4+T cell proliferation and polarization in type 2 diabetic ApoE-/- mouse.

    PubMed

    Liu, Ming-Hao; Li, Ya; Han, Lu; Zhang, Yao-Yuan; Wang, Di; Wang, Zhi-Hao; Zhou, Hui-Min; Song, Ming; Li, Yi-Hui; Tang, Meng-Xiong; Zhang, Wei; Zhong, Ming

    2017-07-01

    Atherosclerosis (AS) is the most common and serious complication of type 2 diabetes mellitus (T2DM) and is accelerated via chronic systemic inflammation rather than hyperglycemia. Adipose tissue is the major source of systemic inflammation in abnormal metabolic state. Pro-inflammatory CD4 + T cells play pivotal role in promoting adipose inflammation. Adipose-derived stem cells (ADSCs) for fat regeneration have potent ability of immunosuppression and restricting CD4 + T cells as well. Whether T2DM ADSCs are impaired in antagonizing CD4 + T cell proliferation and polarization remains unclear. We constructed type 2 diabetic ApoE -/- mouse models and tested infiltration and subgroups of CD4 + T cell in stromal-vascular fraction (SVF) in vivo. Normal/T2DM ADSCs and normal splenocytes with or without CD4 sorting were separated and co-cultured at different scales ex vivo. Immune phenotypes of pro- and anti-inflammation of ADSCs were also investigated. Flow cytometry (FCM) and ELISA were applied in the experiments above. CD4 + T cells performed a more pro-inflammatory phenotype in adipose tissue in T2DM ApoE -/- mice in vivo. Restriction to CD4 + T cell proliferation and polarization was manifested obviously weakened after co-cultured with T2DM ADSCs ex vivo. No obvious distinctions were found in morphology and growth type of both ADSCs. However, T2DM ADSCs acquired a pro-inflammatory immune phenotype, with secreting less PGE2 and expressing higher MHC-II and co-stimulatory molecules (CD40, CD80). Normal ADSCs could also obtain the phenotypic change after cultured with T2DM SVF supernatant. CD4 + T cell infiltration and pro-inflammatory polarization exist in adipose tissue in type 2 diabetic ApoE -/- mice. T2DM ADSCs had impaired function in restricting CD4 + T lymphocyte proliferation and pro-inflammatory polarization due to immune phenotypic changes. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Exosome-Like Vesicles Derived from Adipose Tissue Provide Biochemical Cues for Adipose Tissue Regeneration.

    PubMed

    Dai, Minjia; Yu, Mei; Zhang, Yan; Tian, Weidong

    2017-11-01

    There is an emerging need for soft tissue replacements in the field of reconstructive surgery for the treatment of congenital deformities, posttraumatic repair, and cancer rehabilitation. Previous studies have shown that the bioactive adipose tissue extract can induce adipogenesis without additional stem cells or growth factors. In this study, we innovatively investigated whether exosome-like vesicles derived from adipose tissue (ELV-AT) could direct stem cell differentiation and trigger adipose tissue regeneration. In vitro, ELV-AT can induce adipogenesis of adipose-derived stem cells and promote proliferation, migration, and angiogenic potential of the aorta endothelial cells. In vivo, ELV-AT were transplanted to a chamber on the back of nude mice and neoadipose tissues were formed. Our findings indicated that ELV-AT could be used as a cell-free therapeutic approach for adipose tissue regeneration.

  9. Identification of the Centrifuged Lipoaspirate Fractions Suitable for Postgrafting Survival.

    PubMed

    Qiu, Lihong; Su, Yingjun; Zhang, Dongliang; Song, Yajuan; Liu, Bei; Yu, Zhou; Guo, Shuzhong; Yi, Chenggang

    2016-01-01

    The Coleman centrifugation procedure generates fractions with different adipocyte and progenitor cell densities. This study aimed to identify all fractions that are feasible for implantation. Human lipoaspirates were processed by Coleman centrifugation. The centrifugates were divided arbitrarily into upper, middle, and lower layers. Adipocyte viability, morphology, numbers of stromal vascular fraction cells, and adipose-derived mesenchymal stem cells of each layer were determined. The 12-week volume retention of subcutaneously implanted 0.3-ml lipoasperate of each layer was investigated in an athymic mice model. Most damaged adipocytes were located in the upper layers, whereas the intact adipocytes were distributed in the middle and lower layers. A gradient of stromal vascular fraction cell density was formed in the centrifugates. The implant volume retentions of samples from the upper, middle, and lower layers were 33.44 ± 5.9, 55.11 ± 4.4, and 71.2 ± 5.8 percent, respectively. Furthermore, the middle and lower layers contained significantly more adipose-derived stem cells than did the upper layer. The lower layer contains more viable adipocytes and stromal vascular fraction cells leading to the highest implant volume retention, whereas the most impaired cells are distributed in the upper layer, leading to the least volume retention. Although with a lower stromal vascular fraction content, the middle layer has a substantial number of intact adipocytes that are capable of retaining partial adipose tissue volume after implantation, suggesting that the middle layer may be an alternative fat source when large volumes of fat grafts are needed for transplantation.

  10. Myocardial regeneration potential of adipose tissue-derived stem cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Bai, Xiaowen, E-mail: baixw01@yahoo.com; Alt, Eckhard, E-mail: ealt@mdanderson.org

    Research highlights: {yields} Various tissue resident stem cells are receiving tremendous attention from basic scientists and clinicians and hold great promise for myocardial regeneration. {yields} For practical reasons, human adipose tissue-derived stem cells are attractive stem cells for future clinical application in repairing damaged myocardium. {yields} This review summarizes the characteristics of cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential and the, underlying mechanisms, and safety issues. -- Abstract: Various tissue resident stem cells are receiving attention from basic scientists and clinicians as they hold promise for myocardial regeneration. For practical reasons, adipose tissue-derivedmore » stem cells (ASCs) are attractive cells for clinical application in repairing damaged myocardium based on the following advantages: abundant adipose tissue in most patients and easy accessibility with minimally invasive lipoaspiration procedure. Several recent studies have demonstrated that both cultured and freshly isolated ASCs could improve cardiac function in animal model of myocardial infarction. The mechanisms underlying the beneficial effect of ASCs on myocardial regeneration are not fully understood. Growing evidence indicates that transplantation of ASCs improve cardiac function via the differentiation into cardiomyocytes and vascular cells, and through paracrine pathways. Paracrine factors secreted by injected ASCs enhance angiogenesis, reduce cell apoptosis rates, and promote neuron sprouts in damaged myocardium. In addition, Injection of ASCs increases electrical stability of the injured heart. Furthermore, there are no reported cases of arrhythmia or tumorigenesis in any studies regarding myocardial regeneration with ASCs. This review summarizes the characteristics of both cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential, and

  11. Paracrine Engineering of Human Explant-Derived Cardiac Stem Cells to Over-Express Stromal-Cell Derived Factor 1α Enhances Myocardial Repair.

    PubMed

    Tilokee, Everad L; Latham, Nicholas; Jackson, Robyn; Mayfield, Audrey E; Ye, Bin; Mount, Seth; Lam, Buu-Khanh; Suuronen, Erik J; Ruel, Marc; Stewart, Duncan J; Davis, Darryl R

    2016-07-01

    First generation cardiac stem cell products provide indirect cardiac repair but variably produce key cardioprotective cytokines, such as stromal-cell derived factor 1α, which opens the prospect of maximizing up-front paracrine-mediated repair. The mesenchymal subpopulation within explant derived human cardiac stem cells underwent lentiviral mediated gene transfer of stromal-cell derived factor 1α. Unlike previous unsuccessful attempts to increase efficacy by boosting the paracrine signature of cardiac stem cells, cytokine profiling revealed that stromal-cell derived factor 1α over-expression prevented lv-mediated "loss of cytokines" through autocrine stimulation of CXCR4+ cardiac stem cells. Stromal-cell derived factor 1α enhanced angiogenesis and stem cell recruitment while priming cardiac stem cells to readily adopt a cardiac identity. As compared to injection with unmodified cardiac stem cells, transplant of stromal-cell derived factor 1α enhanced cells into immunodeficient mice improved myocardial function and angiogenesis while reducing scarring. Increases in myocardial stromal-cell derived factor 1α content paralleled reductions in myocyte apoptosis but did not influence long-term engraftment or the fate of transplanted cells. Transplantation of stromal-cell derived factor 1α transduced cardiac stem cells increased the generation of new myocytes, recruitment of bone marrow cells, new myocyte/vessel formation and the salvage of reversibly damaged myocardium to enhance cardiac repair after experimental infarction. Stem Cells 2016;34:1826-1835. © 2016 AlphaMed Press.

  12. H2O2 generated from mitochondrial electron transport chain in thoracic perivascular adipose tissue is crucial for modulation of vascular smooth muscle contraction.

    PubMed

    Costa, Rafael M; Filgueira, Fernando P; Tostes, Rita C; Carvalho, Maria Helena C; Akamine, Eliana H; Lobato, Nubia S

    2016-09-01

    The perivascular adipose tissue (PVAT) releases a variety of factors that affect vascular function. PVAT in the thoracic aorta shares characteristics with the brown adipose tissue, including a large amount of mitochondria. PVAT-derived factors influence both endothelial and smooth muscle function via several signaling mechanisms including the release/generation of reactive nitrogen and oxygen species. Considering the importance of reactive oxygen species (ROS) on vascular function and that mitochondria are an important source of ROS, we hypothesized that mitochondria-derived ROS in the PVAT modulates vascular reactivity. Vascular reactivity to norephinephrine (NE) was evaluated in thoracic aortic rings, with or without endothelium and/or PVAT, from male Wistar rats. Mitochondrial uncoupling, as well as hydrogen peroxide (H2O2) removal, increased the contraction in vessels surrounded by PVAT. PVAT stimulated with NE exhibited increased protein expression, determined by Western blot analysis, of manganese superoxide dismutase (Mn-SOD) and decreased protein expression of catalase. Ultimately, NE increased superoxide anion (O2(-)) generation in PVAT via increases in intracellular calcium. These results clearly demonstrate that mitochondrial electron transport chain (mETC) in PVAT contributes to modulation of aortic muscle contraction by generating higher amounts of O2(-) that is, in turn, dismutated to hydrogen peroxide, which then acts as a pivotal signaling molecule regulating vascular smooth muscle contraction. Copyright © 2015 Elsevier Inc. All rights reserved.

  13. Application of adipose-derived stem cells on scleral contact lens carrier in an animal model of severe acute alkaline burn.

    PubMed

    Espandar, Ladan; Caldwell, Delmar; Watson, Richard; Blanco-Mezquita, Tomas; Zhang, Shijia; Bunnell, Bruce

    2014-07-01

    To evaluate the therapeutic effect of human adipose-derived stem cells (hASCs) overlaid on a scleral contact lens (SCL) carrier in a rabbit model of ocular alkaline burn. After inducing alkaline burn in 11 New Zealand white rabbits, hASCs cultured on SCLs were placed on the right eye of 5 rabbits, SCLs without cells were used in 5, and no treatment was applied in 1 eye. Each eye was examined and photographed for corneal vascularization, opacities, and epithelial defect in week 1, 2, and 4 after surgery. After 1 month, rabbits were killed and the corneas were removed and cut in half for electron and light microscopy examination. Human adipose-derived stem cells were attached to SCL surface and confluent easily. Human adipose-derived stem cells on SCL eyes showed smaller epithelial defect, less corneal opacity, corneal neovascularization relative to SCL eyes. Both groups showed no symblepharon. However, the cornea in the untreated eye was melted in 2 weeks and developed severe symblepharon. Human adipose-derived stem cells on SCL can reduce inflammation and corneal haziness in severe ocular alkaline burn injury in rabbits.

  14. Application of Adipose-Derived Stem Cells on Scleral Contact Lens Carrier in an Animal Model of Severe Acute Alkaline Burn

    PubMed Central

    Espandar, Ladan; Caldwell, Delmar; Watson, Richard; Blanco-Mezquita, Tomas; Zhang, Shijia; Bunnell, Bruce

    2015-01-01

    Purpose To evaluate the therapeutic effect of human adipose-derived stem cells (hASCs) overlaid on a scleral contact lens (SCL) carrier in a rabbit model of ocular alkaline burn. Materials and Methods After inducing alkaline burn in 11 New Zealand white rabbits, hASCs cultured on SCLs were placed on the right eye of 5 rabbits, SCLs without cells were used in 5, and no treatment was applied in 1 eye. Each eye was examined and photographed for corneal vascularization, opacities, and epithelial defect in week 1, 2, and 4 after surgery. After 1 month, rabbits were killed and the corneas were removed and cut in half for electron and light microscopy examination. Results Human adipose-derived stem cells were attached to SCL surface and confluent easily. Human adipose-derived stem cells on SCL eyes showed smaller epithelial defect, less corneal opacity, corneal neovascularization relative to SCL eyes. Both groups showed no symblepharon. However, the cornea in the untreated eye was melted in 2 weeks and developed severe symblepharon. Conclusion Human adipose-derived stem cells on SCL can reduce inflammation and corneal haziness in severe ocular alkaline burn injury in rabbits. PMID:24901976

  15. Complete resolution of avascular necrosis of the human femoral head treated with adipose tissue-derived stem cells and platelet-rich plasma.

    PubMed

    Pak, Jaewoo; Lee, Jung Hun; Jeon, Jeong Ho; Lee, Sang Hee

    2014-12-01

    We report a case of a 43-year-old man with early stage (stage 1) avascular necrosis (AVN) of the femoral head treated with adipose tissue-derived stem cells (ASCs) and platelet-rich plasma (PRP). ASC-containing stromal vascular fraction was mixed with PRP and hyaluronic acid. This mixture was then injected into the diseased hip under ultrasound guidance. The affected hip was reinjected weekly with additional PRP for 4 weeks. The patient was followed-up with sequential magnetic resonance imaging (MRI) scans at 3, 18, and 21 months after treatment, together with Visual Analogue Scale (VAS) Walking Index, Functional Rating Index, Harris Hip Score, and Range of Motion (ROM) assessments. The patient's severe hip pain was considerably improved at 3 months after treatment, with pain scores, ROM and MRI showing near complete resolution of AVN. Pain scores, ROM and MRI at 18 and 21 months after treatment indicated complete resolution of AVN. This case represents the first evidence of complete resolution of early stage AVN of the hip following treatment with ASCs/PRP. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

  16. Adipose Tissue-Derived Mesenchymal Stem Cells Attenuate Staphylococcal Enterotoxin A-Induced Toxic Shock

    PubMed Central

    Asano, Krisana; Yoshimura, Sayuri

    2015-01-01

    Adipose tissue-derived stem cells (ASCs), which are mesenchymal stromal cells isolated from adipose tissues, exhibit immunomodulatory effects that are promising for several applications, including the therapeutics of inflammatory diseases. In the present study, the effect of ASCs on bacterial toxin-induced inflammation was investigated. Intraperitoneal administration of ASCs rescued mice from lethal shock induced by staphylococcal enterotoxin A (SEA) potentiated with lipopolysaccharide. In the sera and/or spleens of mice administered ASCs, the production of proinflammatory cytokines, including interferon gamma, tumor necrosis factor alpha, interleukin-6 (IL-6), and IL-2 was reduced. By quantitative real-time PCR, the expression of Foxp3 in the mice administered ASCs was not altered. On the other hand, the expression of IL-12 receptor and STAT4 was decreased with ASC administration. These results imply that the effect of ASCs is not involved in the lineage of regulatory T cells but that these cells may modulate TH1 differentiation. This information provides evidence that ASCs have properties that are effective to attenuate SEA-induced toxic shock and should prompt further exploration on other inflammatory diseases caused by bacterial toxins or bacterial infections. PMID:26099581

  17. Spirulina platensis Improves Mitochondrial Function Impaired by Elevated Oxidative Stress in Adipose-Derived Mesenchymal Stromal Cells (ASCs) and Intestinal Epithelial Cells (IECs), and Enhances Insulin Sensitivity in Equine Metabolic Syndrome (EMS) Horses.

    PubMed

    Nawrocka, Daria; Kornicka, Katarzyna; Śmieszek, Agnieszka; Marycz, Krzysztof

    2017-08-03

    Equine Metabolic Syndrome (EMS) is a steadily growing life-threatening endocrine disorder linked to insulin resistance, oxidative stress, and systemic inflammation. Inflammatory microenvironment of adipose tissue constitutes the direct tissue milieu for various cell populations, including adipose-derived mesenchymal stromal cells (ASCs), widely considered as a potential therapeutic cell source in the course of the treatment of metabolic disorders. Moreover, elevated oxidative stress induces inflammation in intestinal epithelial cells (IECs)-the first-line cells exposed to dietary compounds. In the conducted research, we showed that in vitro application of Spirulina platensis contributes to the restoration of ASCs' and IECs' morphology and function through the reduction of cellular oxidative stress and inflammation. Enhanced viability, suppressed senescence, and improved proliferation of ASCs and IECs isolated from metabolic syndrome-affected individuals were evident following exposition to Spirulina. A protective effect of the investigated extract against mitochondrial dysfunction and degeneration was also observed. Moreover, our data demonstrate that Spirulina extract effectively suppressed LPS-induced inflammatory responses in macrophages. In vivo studies showed that horses fed with a diet based on Spirulina platensis supplementation lost weight and their insulin sensitivity improved. Thus, our results indicate the engagement of Spirulina platensis nourishing as an interesting alternative approach for supporting the conventional treatment of equine metabolic syndrome.

  18. Spirulina platensis Improves Mitochondrial Function Impaired by Elevated Oxidative Stress in Adipose-Derived Mesenchymal Stromal Cells (ASCs) and Intestinal Epithelial Cells (IECs), and Enhances Insulin Sensitivity in Equine Metabolic Syndrome (EMS) Horses

    PubMed Central

    Nawrocka, Daria; Kornicka, Katarzyna; Śmieszek, Agnieszka

    2017-01-01

    Equine Metabolic Syndrome (EMS) is a steadily growing life-threatening endocrine disorder linked to insulin resistance, oxidative stress, and systemic inflammation. Inflammatory microenvironment of adipose tissue constitutes the direct tissue milieu for various cell populations, including adipose-derived mesenchymal stromal cells (ASCs), widely considered as a potential therapeutic cell source in the course of the treatment of metabolic disorders. Moreover, elevated oxidative stress induces inflammation in intestinal epithelial cells (IECs)—the first-line cells exposed to dietary compounds. In the conducted research, we showed that in vitro application of Spirulina platensis contributes to the restoration of ASCs’ and IECs’ morphology and function through the reduction of cellular oxidative stress and inflammation. Enhanced viability, suppressed senescence, and improved proliferation of ASCs and IECs isolated from metabolic syndrome-affected individuals were evident following exposition to Spirulina. A protective effect of the investigated extract against mitochondrial dysfunction and degeneration was also observed. Moreover, our data demonstrate that Spirulina extract effectively suppressed LPS-induced inflammatory responses in macrophages. In vivo studies showed that horses fed with a diet based on Spirulina platensis supplementation lost weight and their insulin sensitivity improved. Thus, our results indicate the engagement of Spirulina platensis nourishing as an interesting alternative approach for supporting the conventional treatment of equine metabolic syndrome. PMID:28771165

  19. CD90 (Thy-1)-positive selection enhances osteogenic capacity of human adipose-derived stromal cells.

    PubMed

    Chung, Michael T; Liu, Chunjun; Hyun, Jeong S; Lo, David D; Montoro, Daniel T; Hasegawa, Masakazu; Li, Shuli; Sorkin, Michael; Rennert, Robert; Keeney, Michael; Yang, Fan; Quarto, Natalina; Longaker, Michael T; Wan, Derrick C

    2013-04-01

    Stem cell-based bone tissue engineering with adipose-derived stromal cells (ASCs) has shown great promise for revolutionizing treatment of large bone deficits. However, there is still a lack of consensus on cell surface markers identifying osteoprogenitors. Fluorescence-activated cell sorting has identified a subpopulation of CD105(low) cells with enhanced osteogenic differentiation. The purpose of the present study was to compare the ability of CD90 (Thy-1) to identify osteoprogenitors relative to CD(105). Unsorted cells, CD90(+), CD90(-), CD105(high), and CD105(low) cells were treated with an osteogenic differentiation medium. For evaluation of in vitro osteogenesis, alkaline phosphatase (ALP) staining and alizarin red staining were performed at 7 days and 14 days, respectively. RNA was harvested after 7 and 14 days of differentiation, and osteogenic gene expression was examined by quantitative real-time polymerase chain reaction. For evaluation of in vivo osteogenesis, critical-sized (4-mm) calvarial defects in nude mice were treated with the hydroxyapatite-poly(lactic-co-glycolic acid) scaffold seeded with the above-mentioned subpopulations. Healing was followed using micro-CT scans for 8 weeks. Calvaria were harvested at 8 weeks postoperatively, and sections were stained with Movat's Pentachrome. Transcriptional analysis revealed that the CD90(+) subpopulation was enriched for a more osteogenic subtype relative to the CD105(low) subpopulation. Staining at day 7 for ALP was greatest in the CD90(+) cells, followed by the CD105(low) cells. Staining at day 14 for alizarin red demonstrated the greatest amount of mineralized extracellular matrix in the CD90(+) cells, again followed by the CD105(low) cells. Quantification of in vivo healing at 2, 4, 6, and 8weeks postoperatively demonstrated increased bone formation in defects treated with CD90(+) ASCs relative to all other groups. On Movat's Pentachrome-stained sections, defects treated with CD90(+) cells showed the

  20. Development of a Vascularized Skin Construct Using Adipose-Derived Stem Cells from Debrided Burned Skin

    DTIC Science & Technology

    2012-01-01

    additional morbidity from subcutaneous liposuction . Because tangential debridement of skin often leads to debridement of some viable tissue, we have shown...32, 33]; however, in cases of extensive burn injury, liposuction may increase infectious complications of an already traumatized and immunologically...J. Foster et al., “Yield of human adipose-derived adult stem cells from liposuction aspirates,” Cytotherapy, vol. 6, no. 1, pp. 7–14, 2004. [33] B

  1. Adipose-Derived Stem-Cell-Seeded Non-Cross-Linked Porcine Acellular Dermal Matrix Increases Cellular Infiltration, Vascular Infiltration, and Mechanical Strength of Ventral Hernia Repairs

    PubMed Central

    Iyyanki, Tejaswi S.; Dunne, Lina W.; Zhang, Qixu; Hubenak, Justin; Turza, Kristin C.

    2015-01-01

    Adipose-derived stem cells (ASCs) facilitate wound healing by improving cellular and vascular recruitment to the wound site. Therefore, we investigated whether ASCs would augment a clinically relevant bioprosthetic mesh—non-cross-linked porcine acellular dermal matrix (ncl-PADM)—used for ventral hernia repairs in a syngeneic animal model. ASCs were isolated from the subcutaneous adipose tissue of Brown Norway rats, expanded, and labeled with green fluorescent protein. ASCs were seeded (2.5×104 cells/cm2) onto ncl-PADM for 24 h before surgery. In vitro ASC adhesion to ncl-PADM was assessed at 0.5, 1, and 2 h after seeding, and cell morphology on ncl-PADM was visualized by scanning electron microscopy. Ventral hernia defects (2×4 cm) were created and repaired with ASC-seeded (n=31) and control (n=32) ncl-PADM. Explants were harvested at 1, 2, and 4 weeks after surgery. Explant remodeling outcomes were evaluated using gross evaluation (bowel adhesions, surface area, and grade), histological analysis (hematoxylin and eosin and Masson's trichrome staining), immunohistochemical analysis (von Willebrand factor VIII), fluorescent microscopy, and mechanical strength measurement at the tissue-bioprosthetic mesh interface. Stem cell markers CD29, CD90, CD44, and P4HB were highly expressed in cultured ASCs, whereas endothelial and hematopoietic cell markers, such as CD31, CD90, and CD45 had low expression. Approximately 85% of seeded ASCs adhered to ncl-PADM within 2 h after seeding, which was further confirmed by scanning electron microcopy examination. Gross evaluation of the hernia repairs revealed weak omental adhesion in all groups. Ultimate tensile strength was not significantly different in control and treatment groups. Conversely, elastic modulus was significantly greater at 4 weeks postsurgery in the ASC-seeded group (p<0.001). Cellular infiltration was significantly higher in the ASC-seeded group at all time points (p<0.05). Vascular infiltration was

  2. Adipose-derived stem-cell-seeded non-cross-linked porcine acellular dermal matrix increases cellular infiltration, vascular infiltration, and mechanical strength of ventral hernia repairs.

    PubMed

    Iyyanki, Tejaswi S; Dunne, Lina W; Zhang, Qixu; Hubenak, Justin; Turza, Kristin C; Butler, Charles E

    2015-02-01

    Adipose-derived stem cells (ASCs) facilitate wound healing by improving cellular and vascular recruitment to the wound site. Therefore, we investigated whether ASCs would augment a clinically relevant bioprosthetic mesh-non-cross-linked porcine acellular dermal matrix (ncl-PADM)-used for ventral hernia repairs in a syngeneic animal model. ASCs were isolated from the subcutaneous adipose tissue of Brown Norway rats, expanded, and labeled with green fluorescent protein. ASCs were seeded (2.5×10(4) cells/cm(2)) onto ncl-PADM for 24 h before surgery. In vitro ASC adhesion to ncl-PADM was assessed at 0.5, 1, and 2 h after seeding, and cell morphology on ncl-PADM was visualized by scanning electron microscopy. Ventral hernia defects (2×4 cm) were created and repaired with ASC-seeded (n=31) and control (n=32) ncl-PADM. Explants were harvested at 1, 2, and 4 weeks after surgery. Explant remodeling outcomes were evaluated using gross evaluation (bowel adhesions, surface area, and grade), histological analysis (hematoxylin and eosin and Masson's trichrome staining), immunohistochemical analysis (von Willebrand factor VIII), fluorescent microscopy, and mechanical strength measurement at the tissue-bioprosthetic mesh interface. Stem cell markers CD29, CD90, CD44, and P4HB were highly expressed in cultured ASCs, whereas endothelial and hematopoietic cell markers, such as CD31, CD90, and CD45 had low expression. Approximately 85% of seeded ASCs adhered to ncl-PADM within 2 h after seeding, which was further confirmed by scanning electron microcopy examination. Gross evaluation of the hernia repairs revealed weak omental adhesion in all groups. Ultimate tensile strength was not significantly different in control and treatment groups. Conversely, elastic modulus was significantly greater at 4 weeks postsurgery in the ASC-seeded group (p<0.001). Cellular infiltration was significantly higher in the ASC-seeded group at all time points (p<0.05). Vascular infiltration was

  3. Yield and proliferation rate of adipose-derived stromal cells as a function of age, body mass index and harvest site-increasing the yield by use of adherent and supernatant fractions?

    PubMed

    Buschmann, Johanna; Gao, Shuping; Härter, Luc; Hemmi, Sonja; Welti, Manfred; Werner, Clement M L; Calcagni, Maurizio; Cinelli, Paolo; Wanner, Guido A

    2013-09-01

    Adipose-derived stem cells are easily accessed and have a relatively high density compared with other mesenchymal stromal cells. Isolation protocols of adipose-derived stem cells (ASC) rely on the cell's ability to adhere to tissue culture plastic overnight. It was evaluated whether the floating ASC fractions are also of interest for cell-based therapies. In addition, the impact of age, body mass index (BMI) and harvest site was assessed. The surface protein profile with the use of flow cytometry, the cell yield and the doubling time of passages 4, 5 and 6 of ASC from 30 donors were determined. Adherent and supernatant fractions were compared. The impact of age, BMI and harvest site on cell yield and doubling times was determined. Both adherent and supernatant fractions showed high mean fluorescence intensities for CD13, CD29, CD44, CD73, CD90 and CD105 and comparatively low mean fluorescence intensities for CD11b, CD62L, intracellular adhesion molecule-1 and CD34. Doubling times of adherent and supernatant fractions did not differ significantly. Whereas the old age group had a significantly lower cell yield compared with the middle aged group, BMI and harvest site had no impact on cell yield. Finally, doubling times for passages 4, 5 and 6 were not influenced by the age and BMI of the donors, nor the tissue-harvesting site. The floating ASC fraction is an equivalent second cell source just like the adherent ASC fraction. Donor age, BMI and harvest site do not influence cell yield and proliferation rate. Copyright © 2013 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  4. Enrichment of IFN-γ producing cells in different murine adipose tissue depots upon infection with an apicomplexan parasite

    PubMed Central

    Teixeira, Luzia; Marques, Raquel M.; Ferreirinha, Pedro; Bezerra, Filipa; Melo, Joana; Moreira, João; Pinto, Ana; Correia, Alexandra; Ferreira, Paula G.; Vilanova, Manuel

    2016-01-01

    Here we report that lean mice infected with the intracellular parasite Neospora caninum show a fast but sustained increase in the frequency of IFN-γ-producing cells noticeable in distinct adipose tissue depots. Moreover, IFN-γ-mediated immune memory could be evoked in vitro in parasite antigen-stimulated adipose tissue stromal vascular fraction cells collected from mice infected one year before. Innate or innate-like cells such as NK, NK T and TCRγδ+ cells, but also CD4+ and CD8+ TCRβ+ lymphocytes contributed to the IFN-γ production observed since day one of infection. This early cytokine production was largely abrogated in IL-12/IL23 p40-deficient mice. Moreover, production of IFN-γ by stromal vascular fraction cells isolated from these mice was markedly lower than that of wild-type counterparts upon stimulation with parasite antigen. In wild-type mice the increased IFN-γ production was concomitant with up-regulated expression of genes encoding interferon-inducible GTPases and nitric oxide synthase, which are important effector molecules in controlling intracellular parasite growth. This increased gene expression was markedly impaired in the p40-deficient mice. Overall, these results show that NK cells but also diverse T cell populations mediate a prompt and widespread production of IFN-γ in the adipose tissue of N. caninum infected mice. PMID:27001522

  5. Comparison of clinical grade human platelet lysates for cultivation of mesenchymal stromal cells from bone marrow and adipose tissue.

    PubMed

    Juhl, Morten; Tratwal, Josefine; Follin, Bjarke; Søndergaard, Rebekka H; Kirchhoff, Maria; Ekblond, Annette; Kastrup, Jens; Haack-Sørensen, Mandana

    2016-01-01

    The utility of mesenchymal stromal cells (MSCs) in therapeutic applications for regenerative medicine has gained much attention. Clinical translation of MSC-based approaches requires in vitro culture-expansion to achieve a sufficient number of cells. The ideal cell culture medium should be devoid of any animal derived components. We have evaluated whether human Platelet Lysate (hPL) could be an attractive alternative to animal supplements. MSCs from bone marrow (BMSCs) and adipose tissue-derived stromal cells (ASCs) obtained from three donors were culture expanded in three different commercially available hPL fulfilling good manufacturing practice criteria for clinical use. BMSCs and ASCs cultured in Minimum Essential Medium Eagle-alpha supplemented with 5% PLT-Max (Mill Creek), Stemulate™ PL-S and Stemulate™ PL-SP (COOK General Biotechnology) were compared to standard culture conditions with 10% fetal bovine serum (FBS). Cell morphology, proliferation, phenotype, genomic stability, and differentiation potential were analyzed. Regardless of manufacturer, BMSCs and ASCs cultured in hPL media showed a significant increase in proliferation capacity compared to FBS medium. In general, the immunophenotype of both BMSCs and ASCs fulfilled International Society for Cellular Therapy (ISCT) criteria after hPL media expansion. Comparative genomic hybridization measurements demonstrated no unbalanced chromosomal rearrangements for BMSCs or ASCs cultured in hPL media or FBS medium. The BMSCs and ASCs could differentiate into osteogenic, adipogenic, or chondrogenic lineages in all four culture conditions. All three clinically approved commercial human platelet lysates accelerated proliferation of BMSCs and ASCs and the cells meet the ISCT mesenchymal phenotypic requirements without exhibiting chromosomal aberrations.

  6. Morphological changes in paraurethral area after introduction of tissue engineering construct on the basis of adipose tissue stromal cells.

    PubMed

    Makarov, A V; Arutyunyan, I V; Bol'shakova, G B; Volkov, A V; Gol'dshtein, D V

    2009-10-01

    We studied morphological changes in the paraurethral area of Wistar rats after introduction of tissue engineering constructs on the basis of multipotent mesenchymal stem cells and gelatin sponge. The tissue engineering construct containing autologous culture of the stromal fraction of the adipose tissue was most effective. After introduction of this construct we observed more rapid degradation of the construct matrix and more intensive formation of collagen fibers.

  7. Characterization of the Tissue and Stromal Cell Components of Micro-Superficial Enhanced Fluid Fat Injection (Micro-SEFFI) for Facial Aging Treatment.

    PubMed

    Rossi, Martina; Roda, Barbara; Zia, Silvia; Vigliotta, Ilaria; Zannini, Chiara; Alviano, Francesco; Bonsi, Laura; Zattoni, Andrea; Reschiglian, Pierluigi; Gennai, Alessandro

    2018-06-14

    New microfat preparations provide material suitable for use as a regenerative filler for different facial areas. To support the development of new robust techniques for regenerative purposes, the cellular content of the sample should be considered. To evaluate the stromal vascular fraction (SVF) cell components of micro-superficial enhanced fluid fat injection (SEFFI) samples via a technique to harvest re-injectable tissue with minimum manipulation. The results were compared to those obtained from SEFFI samples. Microscopy analysis was performed to visualize the tissue structure. Micro-SEFFI samples were also fractionated using Celector ®, an innovative non-invasive separation technique, to provide an initial evaluation of sample fluidity and composition. SVFs obtained from SEFFI and micro-SEFFI were studied. Adipose stromal cells (ASCs) were isolated and characterized by proliferation and differentiation capacity assays. Microscopic and quality analyses of micro-SEFFI samples by Celector® confirmed the high fluidity and sample cellular composition in terms of red blood cell contamination, the presence of cell aggregates and extracellular matrix fragments. ASCs were isolated from adipose tissue harvested using SEFFI and micro-SEFFI systems. These cells were demonstrated to have a good proliferation rate and differentiation potential towards mesenchymal lineages. Despite the small sizes and low cellularity observed in micro-SEFFI-derived tissue, we were able to isolate stem cells. This result partially explains the regenerative potential of autologous micro-SEFFI tissue grafts. In addition, using this novel Celector® technology, tissues used for aging treatment were characterized analytically, and the adipose tissue composition was evaluated with no need for extra sample processing.

  8. Adipose-derived stem cells and periodontal tissue engineering.

    PubMed

    Tobita, Morikuni; Mizuno, Hiroshi

    2013-01-01

    Innovative developments in the multidisciplinary field of tissue engineering have yielded various implementation strategies and the possibility of functional tissue regeneration. Technologic advances in the combination of stem cells, biomaterials, and growth factors have created unique opportunities to fabricate tissues in vivo and in vitro. The therapeutic potential of human multipotent mesenchymal stem cells (MSCs), which are harvested from bone marrow and adipose tissue, has generated increasing interest in a wide variety of biomedical disciplines. These cells can differentiate into a variety of tissue types, including bone, cartilage, fat, and nerve tissue. Adipose-derived stem cells have some advantages compared with other sources of stem cells, most notably that a large number of cells can be easily and quickly isolated from adipose tissue. In current clinical therapy for periodontal tissue regeneration, several methods have been developed and applied either alone or in combination, such as enamel matrix proteins, guided tissue regeneration, autologous/allogeneic/xenogeneic bone grafts, and growth factors. However, there are various limitations and shortcomings for periodontal tissue regeneration using current methods. Recently, periodontal tissue regeneration using MSCs has been examined in some animal models. This method has potential in the regeneration of functional periodontal tissues because the various secreted growth factors from MSCs might not only promote the regeneration of periodontal tissue but also encourage neovascularization of the damaged tissues. Adipose-derived stem cells are especially effective for neovascularization compared with other MSC sources. In this review, the possibility and potential of adipose-derived stem cells for regenerative medicine are introduced. Of particular interest, periodontal tissue regeneration with adipose-derived stem cells is discussed.

  9. Effect of N-Acetylcysteine on Adipose-Derived Stem Cell and Autologous Fat Graft Survival in a Mouse Model.

    PubMed

    Gillis, Joshua; Gebremeskel, Simon; Phipps, Kyle D; MacNeil, Lori A; Sinal, Christopher J; Johnston, Brent; Hong, Paul; Bezuhly, Michael

    2015-08-01

    Autologous fat grafting is a popular reconstructive technique, but is limited by inconsistent graft retention. The authors examined whether a widely available, clinically safe antioxidant, N-acetylcysteine, could improve adipose-derived stem cell survival and graft take when added to tumescent solution during fat harvest. Inguinal fat pads were harvested from C57BL/6 mice using tumescent solution with or without N-acetylcysteine. Flow cytometric, proliferation, and differentiation assays were performed on isolated primary adipose-derived stem cells and 3T3-L1 preadipocytes treated with or without hydrogen peroxide and/or N-acetylcysteine. N-Acetylcysteine-treated or control grafts were injected under recipient mouse scalps and assessed by serial micro-computed tomographic volumetric analysis. Explanted grafts underwent immunohistochemical analysis. In culture, N-acetylcysteine protected adipose-derived stem cells from oxidative stress and improved cell survival following hydrogen peroxide treatment. Combined exposure to both N-acetylcysteine and hydrogen peroxide led to a 200-fold increase in adipose-derived stem cell proliferation, significantly higher than with either agent alone. N-Acetylcysteine decreased differentiation of adipose-derived stem cells into mature adipocytes, as evidenced by decreased transcription of adipocyte differentiation markers and reduced Oil Red-O staining. In vivo, N-acetylcysteine treatment resulted in improved graft retention at 3 months compared with control (46 versus 17 percent; p = 0.027). N-Acetylcysteine-treated grafts demonstrated less fibrosis and inflammation, and a 33 percent increase in adipocyte density compared with controls (p < 0.001) that was not associated with increased vascularity. These findings provide proof of principle for the addition of N-acetylcysteine to tumescent harvest solution in the clinical setting to optimize fat graft yields.

  10. Impaired autophagy activity is linked to elevated ER-stress and inflammation in aging adipose tissue.

    PubMed

    Ghosh, Amiya Kumar; Mau, Theresa; O'Brien, Martin; Garg, Sanjay; Yung, Raymond

    2016-10-24

    Adipose tissue dysfunction in aging is associated with inflammation, metabolic syndrome and other diseases. We propose that impaired protein homeostasis due to compromised lysosomal degradation (micro-autophagy) might promote aberrant ER stress response and inflammation in aging adipose tissue. Using C57BL/6 mouse model, we demonstrate that adipose tissue-derived stromal vascular fraction (SVF) cells from old (18-20 months) mice have reduced expression of autophagy markers as compared to the younger (4-6 months) cohort. Elevated expressions of ER-stress marker CHOP and autophagy substrate SQSTM1/p62 are observed in old SVFs compared to young, when treated with either vehicle or with thapsigargin (Tg), an ER stress inducer. Treatment with bafilomycin A1 (Baf), a vacuolar-type H (+)-ATPase, or Tg elevated expressions of CHOP, and SQSTM1/p62 and LC-3-II, in 3T3-L1-preadipocytes. We also demonstrate impaired autophagy activity in old SVFs by analyzing increased accumulation of autophagy substrates LC3-II and p62. Compromised autophagy activity in old SVFs is correlated with enhanced release of pro-inflammatory cytokines IL-6 and MCP-1. Finally, SVFs from calorie restricted old mice (CR-O) have shown enhanced autophagy activity compared to ad libitum fed old mice (AL-O). Our results support the notion that diminished autophagy activity with aging contributes to increased adipose tissue ER stress and inflammation.

  11. Flow cytometric single cell analysis reveals heterogeneity between adipose depots

    PubMed Central

    Boumelhem, Badwi B.; Assinder, Stephen J.; Bell-Anderson, Kim S.; Fraser, Stuart T.

    2017-01-01

    ABSTRACT Understanding adipose tissue heterogeneity is hindered by the paucity of methods to analyze mature adipocytes at the single cell level. Here, we report a system for analyzing live adipocytes from different adipose depots in the adult mouse. Single cell suspensions of buoyant adipocytes were separated from the stromal vascular fraction and analyzed by flow cytometry. Compared to other lipophilic dyes, Nile Red uptake effectively distinguished adipocyte populations. Nile Red fluorescence increased with adipocyte size and granularity and could be combined with MitoTracker® Deep Red or fluorescent antibody labeling to further dissect adipose populations. Epicardial adipocytes exhibited the least mitochondrial membrane depolarization and highest fatty-acid translocase CD36 surface expression. In contrast, brown adipocytes showed low surface CD36 expression. Pregnancy resulted in reduced mitochondrial membrane depolarisation and increased CD36 surface expression in brown and epicardial adipocyte populations respectively. Our protocol revealed unreported heterogeneity between adipose depots and highlights the utility of flow cytometry for screening adipocytes at the single cell level. PMID:28453382

  12. Insulin-secreting adipose-derived mesenchymal stromal cells with bone marrow-derived hematopoietic stem cells from autologous and allogenic sources for type 1 diabetes mellitus.

    PubMed

    Thakkar, Umang G; Trivedi, Hargovind L; Vanikar, Aruna V; Dave, Shruti D

    2015-07-01

    Stem cell therapy (SCT) is now the up-coming therapeutic modality for treatment of type 1 diabetes mellitus (T1DM). Our study was a prospective, open-labeled, two-armed trial for 10 T1DM patients in each arm of allogenic and autologous adipose-derived insulin-secreting mesenchymal stromal cells (IS-AD-MSC)+bone marrow-derived hematopoietic stem cell (BM-HSC) infusion. Group 1 received autologous SCT: nine male patients and one female patient; mean age, 20.2 years, disease duration 8.1 years; group 2 received allogenic SCT: six male patients and four female patients, mean age, 19.7 years and disease duration, 7.9 years. Glycosylated hemoglobin (HbA1c) was 10.99%; serum (S.) C-peptide, 0.22 ng/mL and insulin requirement, 63.9 IU/day in group 1; HbA1c was 11.93%, S.C-peptide, 0.028 ng/mL and insulin requirement, 57.55 IU/day in group 2. SCs were infused into the portal+thymic circulation and subcutaneous tissue under non-myelo-ablative conditioning. Patients were monitored for blood sugar, S.C-peptide, glutamic acid decarboxylase antibodies and HbA1c at 3-month intervals. Group 1 received mean SCs 103.14 mL with 2.65 ± 0.8 × 10(4) ISCs/kg body wt, CD34+ 0.81% and CD45-/90+/73+, 81.55%. Group 2 received mean SCs 95.33 mL with 2.07 ± 0.67 × 10(4) ISCs/kg body wt, CD34+ 0.32% and CD45-/90+/73+ 54.04%. No untoward effect was observed with sustained improvement in HbA1c and S.C-peptide in both groups with a decrease in glutamic acid decarboxylase antibodies and reduction in mean insulin requirement. SCT is a safe and viable treatment option for T1DM. Autologous IS-AD-MSC+ BM-HSC co-infusion offers better long-term control of hyperglycemia as compared with allogenic SCT. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  13. Osteogenic Differentiation of Mesenchymal Stromal Cells: A Comparative Analysis Between Human Subcutaneous Adipose Tissue and Dental Pulp.

    PubMed

    D'Alimonte, Iolanda; Mastrangelo, Filiberto; Giuliani, Patricia; Pierdomenico, Laura; Marchisio, Marco; Zuccarini, Mariachiara; Di Iorio, Patrizia; Quaresima, Raimondo; Caciagli, Francesco; Ciccarelli, Renata

    2017-06-01

    White adipose tissue is a source of mesenchymal stromal/stem cells (MSCs) that are actively studied for their possible therapeutic use in bone tissue repair/remodeling. To better appreciate the osteogenic potential of these cells, we compared some properties of MSCs from human subcutaneous adipose tissue [subcutaneous-adipose stromal cells (S-ASCs)] and dental pulp stem cell (DPSCs) of third-impacted molars, the latter representing a well-established MSC source. Both undifferentiated cell types showed similar fibroblast-like morphology and mesenchymal marker expression. However, undifferentiated S-ASCs displayed a faster doubling time coupled to greater proliferation and colony-forming ability than DPSCs. Also, the osteogenic differentiation of S-ASCs was greater than that of DPSCs, as evaluated by the higher levels of expression of early osteogenic markers Runt-related transcription factor-2 (RUNX2) and alkaline phosphatase at days 3-14 and of extracellular matrix mineralization at days 14-21. Moreover, S-ASCs showed a better colonization of the titanium scaffold. In addition, we investigated whether S-ASC osteogenic commitment was enhanced by adenosine A1 receptor (A1R) stimulation, as previously shown for DPSCs. Although A1R expression was constant during DPSC differentiation, it increased in S-ASC at day 21 from osteogenesis induction. Accordingly, A1R stimulation by the agonist 2-chloro-N 6 -cyclopentyl-adenosine, added to the cultures at each medium change, stimulated proliferation only in differentiating DPSC and enhanced the osteogenic differentiation earlier in DPSCs than in S-ASCs. These effects were counteracted by cell pretreatment with a selective A1R antagonist. Thus, our findings suggest that S-ASCs could be advantageously used in regenerative orthopedics/dentistry, and locally released or exogenously added purines may play a role in bone repair/remodeling, even though this aspect should be more thoroughly evaluated.

  14. Contribution of Adipose Tissue to Development of Cancer

    PubMed Central

    Cozzo, Alyssa J.; Fuller, Ashley M.; Makowski, Liza

    2018-01-01

    Solid tumor growth and metastasis require the interaction of tumor cells with the surrounding tissue, leading to a view of tumors as tissue-level phenomena rather than exclusively cell-intrinsic anomalies. Due to the ubiquitous nature of adipose tissue, many types of solid tumors grow in proximate or direct contact with adipocytes and adipose-associated stromal and vascular components, such as fibroblasts and other connective tissue cells, stem and progenitor cells, endothelial cells, innate and adaptive immune cells, and extracellular signaling and matrix components. Excess adiposity in obesity both increases risk of cancer development and negatively influences prognosis in several cancer types, in part due to interaction with adipose tissue cell populations. Herein, we review the cellular and noncellular constituents of the adipose “organ,” and discuss the mechanisms by which these varied microenvironmental components contribute to tumor development, with special emphasis on obesity. Due to the prevalence of breast and prostate cancers in the United States, their close anatomical proximity to adipose tissue depots, and their complex epidemiologic associations with obesity, we particularly highlight research addressing the contribution of adipose tissue to the initiation and progression of these cancer types. Obesity dramatically modifies the adipose tissue microenvironment in numerous ways, including induction of fibrosis and angiogenesis, increased stem cell abundance, and expansion of proinflammatory immune cells. As many of these changes also resemble shifts observed within the tumor microenvironment, proximity to adipose tissue may present a hospitable environment to developing tumors, providing a critical link between adiposity and tumorigenesis. PMID:29357128

  15. Adipose Tissue in HIV Infection.

    PubMed

    Koethe, John R

    2017-09-12

    HIV infection and antiretroviral therapy (ART) treatment exert diverse effects on adipocytes and stromal-vascular fraction cells, leading to changes in adipose tissue quantity, distribution, and energy storage. A HIV-associated lipodystrophic condition was recognized early in the epidemic, characterized by clinically apparent changes in subcutaneous, visceral, and dorsocervical adipose depots. Underlying these changes is altered adipose tissue morphology and expression of genes central to adipocyte maturation, regulation, metabolism, and cytokine signaling. HIV viral proteins persist in circulation and locally within adipose tissue despite suppression of plasma viremia on ART, and exposure to these proteins impairs preadipocyte maturation and reduces adipocyte expression of peroxisome proliferator-activated receptor gamma (PPAR-γ) and other genes involved in cell regulation. Several early nucleoside reverse transcriptase inhibitor and protease inhibitor antiretroviral drugs demonstrated substantial adipocyte toxicity, including reduced mitochondrial DNA content and respiratory chain enzymes, reduced PPAR-γ and other regulatory gene expression, and increased proinflammatory cytokine production. Newer-generation agents, such as integrase inhibitors, appear to have fewer adverse effects. HIV infection also alters the balance of CD4+ and CD8+ T cells in adipose tissue, with effects on macrophage activation and local inflammation, while the presence of latently infected CD4+ T cells in adipose tissue may constitute a protected viral reservoir. This review provides a synthesis of the literature on how HIV virus, ART treatment, and host characteristics interact to affect adipose tissue distribution, immunology, and contribution to metabolic health, and adipocyte maturation, cellular regulation, and energy storage. © 2017 American Physiological Society. Compr Physiol 7:1339-1357, 2017. Copyright © 2017 John Wiley & Sons, Inc.

  16. Acute Hypoxic Stress Affects Migration Machinery of Tissue O2-Adapted Adipose Stromal Cells.

    PubMed

    Udartseva, Olga O; Lobanova, Margarita V; Andreeva, Elena R; Buravkov, Sergey V; Ogneva, Irina V; Buravkova, Ludmila B

    2016-01-01

    The ability of mesenchymal stromal (stem) cells (MSCs) to be mobilised from their local depot towards sites of injury and to participate in tissue repair makes these cells promising candidates for cell therapy. Physiological O 2 tension in an MSC niche in vivo is about 4-7%. However, most in vitro studies of MSC functional activity are performed at 20% O 2 . Therefore, this study focused on the effects of short-term hypoxic stress (0.1% O 2 , 24 h) on adipose tissue-derived MSC motility at tissue-related O 2 level. No significant changes in integrin expression were detected after short-term hypoxic stress. However, O 2 deprivation provoked vimentin disassembly and actin polymerisation and increased cell stiffness. In addition, hypoxic stress induced the downregulation of ACTR3, DSTN, MACF1, MID1, MYPT1, NCK1, ROCK1, TIAM1 , and WASF1 expression, the products of which are known to be involved in leading edge formation and cell translocation. These changes were accompanied by the attenuation of targeted and nontargeted migration of MSCs after short-term hypoxic exposure, as demonstrated in scratch and transwell migration assays. These results indicate that acute hypoxic stress can modulate MSC function in their native milieu, preventing their mobilisation from sites of injury.

  17. Acute Hypoxic Stress Affects Migration Machinery of Tissue O2-Adapted Adipose Stromal Cells

    PubMed Central

    Lobanova, Margarita V.; Andreeva, Elena R.

    2016-01-01

    The ability of mesenchymal stromal (stem) cells (MSCs) to be mobilised from their local depot towards sites of injury and to participate in tissue repair makes these cells promising candidates for cell therapy. Physiological O2 tension in an MSC niche in vivo is about 4–7%. However, most in vitro studies of MSC functional activity are performed at 20% O2. Therefore, this study focused on the effects of short-term hypoxic stress (0.1% O2, 24 h) on adipose tissue-derived MSC motility at tissue-related O2 level. No significant changes in integrin expression were detected after short-term hypoxic stress. However, O2 deprivation provoked vimentin disassembly and actin polymerisation and increased cell stiffness. In addition, hypoxic stress induced the downregulation of ACTR3, DSTN, MACF1, MID1, MYPT1, NCK1, ROCK1, TIAM1, and WASF1 expression, the products of which are known to be involved in leading edge formation and cell translocation. These changes were accompanied by the attenuation of targeted and nontargeted migration of MSCs after short-term hypoxic exposure, as demonstrated in scratch and transwell migration assays. These results indicate that acute hypoxic stress can modulate MSC function in their native milieu, preventing their mobilisation from sites of injury. PMID:28115943

  18. Cavernous nerve repair with allogenic adipose matrix and autologous adipose-derived stem cells.

    PubMed

    Lin, Guiting; Albersen, Maarten; Harraz, Ahmed M; Fandel, Thomas M; Garcia, Maurice; McGrath, Mary H; Konety, Badrinath R; Lue, Tom F; Lin, Ching-Shwun

    2011-06-01

    To investigate whether adipose-derived matrix seeded with adipose-derived stem cells (ADSC) can facilitate the repair of injured cavernous nerves (CNs). Human and rat adipose tissues were decellularized and fabricated into various forms, including adipose tissue-derived acellular matrix thread (ADMT). ADMT seeded with ADSC were transplanted into subcutaneous space and examined for signs of inflammation. ADSC-seeded ADMTs were then used to repair CN injury in rats, followed by assessment of histology and erectile function. Adipose tissue can be fabricated into acellular matrices of various shapes and sizes, including threads and sheets. Seeding of ADMT occurred rapidly: within 24 hours, 55% of the surface was covered with ADSC and within 1 week, 90% was covered. Transplantation of the seeded ADMT into the subcutaneous space of an allogenic host showed no signs of inflammatory reaction. At 3 months after grafting into CN injury rats, approximately twice as many cells were found on seeded ADMT as on unseeded ADMT. The seeded ADMT also had various degrees of S100 and neuronal nitric oxide synthase expression, suggesting CN axonal ingrowth. Rats grafted with seeded ADMT overall had the best erectile function recovery when compared with those grafted with unseeded ADMT and those ungrafted. However, as a result of large variations, the differences did not reach statistic significance (P = .07). Grafting of ADSC-seeded matrix resulted in a substantial recovery of erectile function and improvement of histology. However, further refinement of the matrix architecture is needed to improve the success rate. Copyright © 2011 Elsevier Inc. All rights reserved.

  19. CD90 (Thy-1)-Positive Selection Enhances Osteogenic Capacity of Human Adipose-Derived Stromal Cells

    PubMed Central

    Chung, Michael T.; Liu, Chunjun; Hyun, Jeong S.; Lo, David D.; Montoro, Daniel T.; Hasegawa, Masakazu; Li, Shuli; Sorkin, Michael; Rennert, Robert; Keeney, Michael; Yang, Fan; Quarto, Natalina; Longaker, Michael T.

    2013-01-01

    Background Stem cell-based bone tissue engineering with adipose-derived stromal cells (ASCs) has shown great promise for revolutionizing treatment of large bone deficits. However, there is still a lack of consensus on cell surface markers identifying osteoprogenitors. Fluorescence-activated cell sorting has identified a subpopulation of CD105low cells with enhanced osteogenic differentiation. The purpose of the present study was to compare the ability of CD90 (Thy-1) to identify osteoprogenitors relative to CD105. Methods Unsorted cells, CD90+, CD90−, CD105high, and CD105low cells were treated with an osteogenic differentiation medium. For evaluation of in vitro osteogenesis, alkaline phosphatase (ALP) staining and alizarin red staining were performed at 7 days and 14 days, respectively. RNA was harvested after 7 and 14 days of differentiation, and osteogenic gene expression was examined by quantitative real-time polymerase chain reaction. For evaluation of in vivo osteogenesis, critical-sized (4-mm) calvarial defects in nude mice were treated with the hydroxyapatite-poly(lactic-co-glycolic acid) scaffold seeded with the above-mentioned subpopulations. Healing was followed using micro-CT scans for 8 weeks. Calvaria were harvested at 8 weeks postoperatively, and sections were stained with Movat's Pentachrome. Results Transcriptional analysis revealed that the CD90+ subpopulation was enriched for a more osteogenic subtype relative to the CD105low subpopulation. Staining at day 7 for ALP was greatest in the CD90+ cells, followed by the CD105low cells. Staining at day 14 for alizarin red demonstrated the greatest amount of mineralized extracellular matrix in the CD90+ cells, again followed by the CD105low cells. Quantification of in vivo healing at 2, 4, 6, and 8weeks postoperatively demonstrated increased bone formation in defects treated with CD90+ ASCs relative to all other groups. On Movat's Pentachrome-stained sections, defects treated with CD90+ cells showed

  20. Scaffold preferences of mesenchymal stromal cells and adipose-derived stem cells from green fluorescent protein transgenic mice influence the tissue engineering of bone.

    PubMed

    Wittenburg, Gretel; Flade, Viktoria; Garbe, Annette I; Lauer, Günter; Labudde, Dirk

    2014-05-01

    We have analysed the growth and differentiation of mesenchymal stromal cells (MSC) from bone marrow, and of adipose derived stem cells (ASC) from murine abdominal fat tissue, of green fluorescent protein (GFP) transgenic animals grown directly on two types of hydroxyapatite ceramic bone substitutes. BONITmatrix® and NanoBone® have specific mechanical and physiochemical properties such as porosity and an inner surface that influence cellular growth. Both MSC and ASC were separately seeded on 200mg of each biomaterial and cultured for 3 weeks under osteogenic differentiation conditions. The degree of mineralisation was assessed by alizarin red dye and the specific alkaline phosphatase activity of the differentiated cells. The morphology of the cells was examined by scanning electron microscopy and confocal microscopy. The osteoblastic phenotype of the cells was confirmed by analysing the expression of bone-specific genes (Runx2, osteocalcin, osteopontin, and osteonectin) by semiquantitative reverse transcriptase polymerase chain reaction (PCR). Comparison of BONITmatrix® and NanoBone® showed cell type-specific preferences in terms of osteogenic differentiation. MSC-derived osteoblast-like cells spread optimally on the surface of NanoBone® but not BONITmatrix® granules. In contrast BONITmatrix® granules conditioned the growth of osteoblast-like cells derived from ASC. The osteoblastic phenotype of the cultured cells on all matrices was confirmed by specific gene expression. Our results show that the in vitro growth and osteogenic differentiation of murine MSC or ASC of GFP transgenic mice are distinctly influenced by the ceramic substratum. While NanoBone® granules support the proliferation and differentiation of murine MSC isolated from bone marrow, the growth of murine ASC is supported by BONITmatrix® granules. NanoBone® is therefore recommended for use as scaffold in tissue engineering that requires MSC, whereas ASC can be combined with BONITmatrix® for

  1. Substance P enhances proliferation and paracrine potential of adipose-derived stem cells in vitro.

    PubMed

    Kim, Suna; Piao, Jiyuan; Son, Youngsook; Hong, Hyun Sook

    2017-03-25

    Stem cells have tremendous promise to treat intractable diseases. Notably, adipose-derived stem cells (ADSCs) are actively being investigated because of ease of sampling and high repopulation capacity in vitro. ADSCs can exert a therapeutic effect through differentiation and paracrine potential, and these actions have been proven in many diseases, including cutaneous and inflammatory diseases. Transplantation of ADSCs necessitates therapeutic quantities and thus, long term ex vivo culture of ADSCs. However, this procedure can impair the activity of ADSCs and provoke cellular senescence, leading to low efficacy in vivo. Accordingly, strategies to restore cellular activity and inhibit senescence of stem cells during ex vivo culture are needed for stem cell-based therapies. This study evaluated a potential supplementary role of Substance P (SP) in ADSC ex vivo culture. After confirming that the ADSC cell cycle was damaged by passage 6 (p6), ADSCs at p6 were cultured with SP, and their proliferation rates, cumulative cell numbers, cytokine profiles, and impact on T/endothelial cells were assessed. Long-term culture weakened proliferation ability and secretion of the cytokines, transforming growth factor-beta 1 (TGF-beta1), vascular endothelial growth factor (VEGF), and stromal cell derived factor-1 alpha (SDF-1alpha) in ADSCs. However, SP treatment reduced the population doubling time (PDT), enabling gain of a sufficient number of ADSCs at early passages. In addition, SP restored cytokine secretion, enhancing the ADSC-mediated paracrine effect on T cell and human umbilical vein endothelial cells (HUVECs). Taken together, these results suggest that SP can retain the therapeutic effect of ADSCs by elevating their proliferative and paracrine potential in ex vivo culture. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Role of adipose tissue-derived stem cells in the progression of renal disease.

    PubMed

    Donizetti-Oliveira, Cassiano; Semedo, Patricia; Burgos-Silva, Marina; Cenedeze, Marco Antonio; Malheiros, Denise Maria Avancini Costa; Reis, Marlene Antônia Dos; Pacheco-Silva, Alvaro; Câmara, Niels Olsen Saraiva

    2011-03-01

    To analyze the role of adipose tissue-derived stem cells in reducing the progression of renal fibrosis. adipose tissue-derived stem cells were isolated from C57Bl/6 mice and characterized by cytometry and differentiation. Renal fibrosis was established after unilateral clamping of the renal pedicle for 1 hour. Four hours after reperfusion, 2.105 adipose tissue-derived stem cells were administered intraperitoneally and the animals were followed for 24 hours during 6 weeks. In another experimental group, 2.105adipose tissue-derived stem cells were administered only after 6 weeks of reperfusion, and they were euthanized and studied 4 weeks later. Twenty-four hours after reperfusion, the animals treated with adipose tissue-derived stem cells displayed reduced renal and tubular dysfunction and an increase of the regenerative process. Renal expression of IL-6 and TNF mRNA were decreased in the animals treated with adipose tissue-derived stem cells, while the levels of IL-4, IL-10, and HO-1 were increased, despite the fact that adipose tissue-derived stem cells were not observed in the kidneys via SRY analysis. In 6 weeks, the kidneys of non-treated animals decreased in size, and the kidneys of the animals treated with adipose tissue-derived stem cells remained at normal size and display less deposition of type 1 collagen and FSP-1. The renal protection observed in animals treated with adipose tissue-derived stem cells was followed by a drop in serum levels of TNF-α, KC, RANTES, and IL-1a. Treatment with adipose tissue-derived stem cells after 6 weeks, when the animals already displayed established fibrosis, demonstrated an improvement in functional parameters and less fibrosis analyzed by Picrosirius stain, as well as a reduction of the expression of type 1 collagen and vimentin mRNA. Treatment with adipose tissue-derived stem cells may deter the progression of renal fibrosis by modulation of the early inflammatory response, likely via reduction of the epithelial

  3. Osteoinductive effects of glyceollins on adult mesenchymal stromal/stem cells from adipose tissue and bone marrow.

    PubMed

    Bateman, Marjorie E; Strong, Amy L; Hunter, Ryan S; Bratton, Melyssa R; Komati, Rajesh; Sridhar, Jayalakshmi; Riley, Kevin E; Wang, Guangdi; Hayes, Daniel J; Boue, Stephen M; Burow, Matthew E; Bunnell, Bruce A

    2017-04-15

    While current therapies for osteoporosis focus on reducing bone resorption, the development of therapies to regenerate bone may also be beneficial. Promising anabolic therapy candidates include phytoestrogens, such as daidzein, which effectively induce osteogenesis of adipose-derived stromal cells (ASCs) and bone marrow stromal cells (BMSCs). To investigate the effects of glyceollins, structural derivatives of daidzein, on osteogenesis of ASCs and BMSCs. Herein, the osteoinductive effects of glyceollin I and glyceollin II were assessed and compared to estradiol in ASCs and BMSCs. The mechanism by which glyceollin II induces osteogenesis was further examined. The ability of glyceollins to promote osteogenesis of ASCs and BMSCs was evaluated in adherent and scaffold cultures. Relative deposition of calcium was analyzed using Alizarin Red staining, Bichinchoninic acid Protein Assay, and Alamar Blue Assay. To further explore the mechanism by which glyceollin II exerts its osteoinductive effects, docking studies of glyceollin II, RNA isolation, cDNA synthesis, and quantitative RT-PCR (qPCR) were performed. In adherent cultures, ASCs and BMSCs treated with estradiol, glyceollin I, or glyceollin II demonstrated increased calcium deposition relative to vehicle-treated cells. During evaluation on PLGA scaffolds seeded with ASCs and BMSCs, glyceollin II was the most efficacious in inducing ASC and BMSC osteogenesis compared to estradiol and glyceollin I. Dose-response analysis in ASCs and BMSCs revealed that glyceollin II has the highest potency at 10nM in adherent cultures and 1µM in tissue scaffold cultures. At all doses, osteoinductive effects were attenuated by fulvestrant, suggesting that glyceollin II acts at least in part through estrogen receptor-mediated pathways to induce osteogenesis. Analysis of gene expression demonstrated that, similar to estradiol, glyceollin II induces upregulation of genes involved in osteogenic differentiation. The ability of glyceollin II

  4. Adipocytes Impair Efficacy of Antiretroviral Therapy

    PubMed Central

    Couturier, Jacob; Winchester, Lee C.; Suliburk, James W.; Wilkerson, Gregory K.; Podany, Anthony T.; Agarwal, Neeti; Chua, Corrine Ying Xuan; Nehete, Pramod N.; Nehete, Bharti P.; Grattoni, Alessandro; Sastry, K. Jagannadha; Fletcher, Courtney V.; Lake, Jordan E.; Balasubramanyan, Ashok; Lewis, Dorothy E.

    2018-01-01

    Adequate distribution of antiretroviral drugs to infected cells in HIV patients is critical for viral suppression. In humans and primates, HIV- and SIV-infected CD4 T cells in adipose tissues have recently been identified as reservoirs for infectious virus. To better characterize adipose tissue as a pharmacological sanctuary for HIV-infected cells, in vitro experiments were conducted to assess antiretroviral drug efficacy in the presence of adipocytes, and drug penetration in adipose tissue cells (stromal-vascular-fraction cells and mature adipocytes) was examined in treated humans and monkeys. Co-culture experiments between HIV-1-infected CD4 T cells and primary human adipocytes showed that adipocytes consistently reduced the antiviral efficacy of the nucleotide reverse transcriptase inhibitor tenofovir and its prodrug forms tenofovir disoproxil fumarate (TDF) and tenofovir alafenamide (TAF). In HIV-infected persons, LC-MS/MS analysis of intracellular lysates derived from adipose tissue stromal-vascular-fraction cells or mature adipocytes suggested that integrase inhibitors penetrate adipose tissue, whereas penetration of nucleoside/nucleotide reverse transcriptase inhibitors such as TDF, emtricitabine, abacavir, and lamivudine is restricted. The limited distribution and functions of key antiretroviral drugs within fat depots may contribute to viral persistence in adipose tissue. PMID:29630975

  5. Impaired autophagy activity is linked to elevated ER-stress and inflammation in aging adipose tissue

    PubMed Central

    Ghosh, Amiya Kumar; Mau, Theresa; O'Brien, Martin; Garg, Sanjay; Yung, Raymond

    2016-01-01

    Adipose tissue dysfunction in aging is associated with inflammation, metabolic syndrome and other diseases. We propose that impaired protein homeostasis due to compromised lysosomal degradation (micro-autophagy) might promote aberrant ER stress response and inflammation in aging adipose tissue. Using C57BL/6 mouse model, we demonstrate that adipose tissue-derived stromal vascular fraction (SVF) cells from old (18-20 months) mice have reduced expression of autophagy markers as compared to the younger (4-6 months) cohort. Elevated expressions of ER-stress marker CHOP and autophagy substrate SQSTM1/p62 are observed in old SVFs compared to young, when treated with either vehicle or with thapsigargin (Tg), an ER stress inducer. Treatment with bafilomycin A1 (Baf), a vacuolar-type H (+)-ATPase, or Tg elevated expressions of CHOP, and SQSTM1/p62 and LC-3-II, in 3T3-L1-preadipocytes. We also demonstrate impaired autophagy activity in old SVFs by analyzing increased accumulation of autophagy substrates LC3-II and p62. Compromised autophagy activity in old SVFs is correlated with enhanced release of pro-inflammatory cytokines IL-6 and MCP-1. Finally, SVFs from calorie restricted old mice (CR-O) have shown enhanced autophagy activity compared to ad libitum fed old mice (AL-O). Our results support the notion that diminished autophagy activity with aging contributes to increased adipose tissue ER stress and inflammation. PMID:27777379

  6. Bisphenol A enhances adipogenic differentiation of human adipose stromal/stem cells

    PubMed Central

    Ohlstein, Jason F; Strong, Amy L; McLachlan, John A; Gimble, Jeffrey M; Burow, Matthew E; Bunnell, Bruce A

    2016-01-01

    Exposure of humans to the endocrine disrupter bisphenol A (BPA) has been associated with increased weight and obesity. However, the mechanism(s) by which BPA increases adipose tissue in humans remains to be determined. The goal of this study was to determine the effects of BPA on adipogenesis of cultured human adipose stromal/stem cells (ASCs), precursors to mature adipocytes. ASCs from three donors were cultured for either 14 or 21 days in adipogenic differentiation media containing increasing concentrations of BPA (100 pM–10 μM). The extent of adipogenic differentiation in the ASCs was assessed by staining with Oil Red O to visualize adipogenic differentiation and then quantified by extraction and optical density measurement of the retained dye. BPA significantly enhanced adipogenesis at a concentration of 1 μM after 21 days of culture. Additionally, we found that BPA increased transcription of the estrogen receptor (ER (ESR1)) and that treatment with the ER antagonist ICI 182 780, blocked the effects of BPA, indicating that BPA may act via an ER-mediated pathway. The results of molecular analyses indicated that the expression of the adipogenesis-associated genes dual leucine zipper-bearing kinase (DLK (MAP3K12)), IGF1, CCAAT/enhancer-binding protein alpha (C/EBPα (CEBPA)), peroxisome proliferator-activated receptor gamma (PPARγ (PPARG)), and lipoprotein lipase (LPL) was temporally accelerated and increased by BPA. In summary, these results indicate that BPA significantly enhances adipogenesis in ASCs through an ER-mediated pathway at physiologically relevant concentrations. PMID:25143472

  7. The gene expression profile of non-cultured, highly purified human adipose tissue pericytes: Transcriptomic evidence that pericytes are stem cells in human adipose tissue

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Silva Meirelles, Lindolfo da, E-mail: lindolfomeirelles@gmail.com; Laboratory for Stem Cells and Tissue Engineering, PPGBioSaúde, Lutheran University of Brazil, Av. Farroupilha 8001, 92425-900 Canoas, RS; Deus Wagatsuma, Virgínia Mara de

    Pericytes (PCs) are a subset of perivascular cells that can give rise to mesenchymal stromal cells (MSCs) when culture-expanded, and are postulated to give rise to MSC-like cells during tissue repair in vivo. PCs have been suggested to behave as stem cells (SCs) in situ in animal models, although evidence for this role in humans is lacking. Here, we analyzed the transcriptomes of highly purified, non-cultured adipose tissue (AT)-derived PCs (ATPCs) to detect gene expression changes that occur as they acquire MSC characteristics in vitro, and evaluated the hypothesis that human ATPCs exhibit a gene expression profile compatible with anmore » AT SC phenotype. The results showed ATPCs are non-proliferative and express genes characteristic not only of PCs, but also of AT stem/progenitor cells. Additional analyses defined a gene expression signature for ATPCs, and revealed putative novel ATPC markers. Almost all AT stem/progenitor cell genes differentially expressed by ATPCs were not expressed by ATMSCs or culture-expanded ATPCs. Genes expressed by ATMSCs but not by ATPCs were also identified. These findings strengthen the hypothesis that PCs are SCs in vascularized tissues, highlight gene expression changes they undergo as they assume an MSC phenotype, and provide new insights into PC biology. - Highlights: • Non-cultured adipose tissue-derived human pericytes (ncATPCs) exhibit a distinctive gene expression signature. • ncATPCs express key adipose tissue stem cell genes previously described in vivo in mice. • ncATPCs express message for anti-proliferative and antiangiogenic molecules. • Most ncATPC-specific transcripts are absent in culture-expanded pericytes or ATMSCs • Gene expression changes ncATPCs undergo as they acquire a cultured ATMSC phenotype are pointed out.« less

  8. Comparative analysis of the immunomodulatory capacities of human bone marrow- and adipose tissue-derived mesenchymal stromal cells from the same donor.

    PubMed

    Valencia, Jaris; Blanco, Belén; Yáñez, Rosa; Vázquez, Miriam; Herrero Sánchez, Carmen; Fernández-García, María; Rodríguez Serrano, Concepción; Pescador, David; Blanco, Juan F; Hernando-Rodríguez, Miriam; Sánchez-Guijo, Fermín; Lamana, María Luisa; Segovia, José Carlos; Vicente, Ángeles; Del Cañizo, Consuelo; Zapata, Agustín G

    2016-10-01

    The immunomodulatory properties of mesenchymal stromal cells (MSCs), together with their tissue regenerative potential, make them interesting candidates for clinical application. In the current study, we analyzed the in vitro immunomodulatory effects of MSCs derived from bone marrow (BM-MSCs) and from adipose tissue (AT-MSCs) obtained from the same donor on both innate and acquired immunity cells. BM-MSCs and AT-MSCs were expanded to fourth or fifth passage and co-cultured with T cells, monocytes or natural killer (NK) cells isolated from human peripheral blood and stimulated in vitro. The possible differing impact of MSCs obtained from distinct sources on phenotype, cell proliferation and differentiation, cytokine production and function of these immune cells was comparatively analyzed. BM-MSCs and AT-MSCs induced a similar decrease in NK-cell proliferation, cytokine secretion and expression of both activating receptors and cytotoxic molecules. However, only BM-MSCs significantly reduced NK-cell cytotoxic activity, although both MSC populations showed the same susceptibility to NK-cell-mediated lysis. AT-MSCs were more potent in inhibiting dendritic-cell (DC) differentiation than BM-MSC, but both MSC populations similarly reduced the ability of DCs to induce CD4(+) T-cell proliferation and cytokine production. BM-MSCs and AT-MSCs induced a similar decrease in T-cell proliferation and production of inflammatory cytokines after activation. AT-MSCs and BM-MSCs from the same donor had similar immunomodulatory capacity on both innate and acquired immunity cells. Thus, other variables, such as accessibility of samples or the frequency of MSCs in the tissue should be considered to select the source of MSC for cell therapy. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  9. Adipose Tissue-Derived Mesenchymal Stromal Cells Protect Mice Infected with Trypanosoma cruzi from Cardiac Damage through Modulation of Anti-parasite Immunity

    PubMed Central

    Mesquita, Fernanda C. P.; Brasil, Guilherme V.; Rocha, Nazareth N.; Takiya, Christina M.; Lima, Ana Paula C. A.; Campos de Carvalho, Antonio C.; Goldenberg, Regina S.; Carvalho, Adriana B.

    2015-01-01

    Background Chagas disease, caused by the protozoan Trypanosoma cruzi (T.cruzi), is a complex disease endemic in Central and South America. It has been gathering interest due to increases in non-vectorial forms of transmission, especially in developed countries. The objective of this work was to investigate if adipose tissue-derived mesenchymal stromal cells (ASC) can alter the course of the disease and attenuate pathology in a mouse model of chagasic cardiomyopathy. Methodology/Principal Findings ASC were injected intraperitoneally at 3 days post-infection (dpi). Tracking by bioluminescence showed that cells remained in the abdominal cavity for up to 9 days after injection and most of them migrated to the abdominal or subcutaneous fat, an early parasite reservoir. ASC injection resulted in a significant reduction in blood parasitemia, which was followed by a decrease in cardiac tissue inflammation, parasitism and fibrosis at 30 dpi. At the same time point, analyses of cytokine release in cells isolated from the heart and exposed to T. cruzi antigens indicated an anti-inflammatory response in ASC-treated animals. In parallel, splenocytes exposed to the same antigens produced a pro-inflammatory response, which is important for the control of parasite replication, in placebo and ASC-treated groups. However, splenocytes from the ASC group released higher levels of IL-10. At 60 dpi, magnetic resonance imaging revealed that right ventricular (RV) dilation was prevented in ASC-treated mice. Conclusions/Significance In conclusion, the injection of ASC early after T. cruzi infection prevents RV remodeling through the modulation of immune responses. Lymphoid organ response to the parasite promoted the control of parasite burden, while the heart, a target organ of Chagas disease, was protected from damage due to an improved control of inflammation in ASC-treated mice. PMID:26248209

  10. Adipose Tissue-Derived Mesenchymal Stromal Cells Protect Mice Infected with Trypanosoma cruzi from Cardiac Damage through Modulation of Anti-parasite Immunity.

    PubMed

    Mello, Debora B; Ramos, Isalira P; Mesquita, Fernanda C P; Brasil, Guilherme V; Rocha, Nazareth N; Takiya, Christina M; Lima, Ana Paula C A; Campos de Carvalho, Antonio C; Goldenberg, Regina S; Carvalho, Adriana B

    2015-01-01

    Chagas disease, caused by the protozoan Trypanosoma cruzi (T. cruzi), is a complex disease endemic in Central and South America. It has been gathering interest due to increases in non-vectorial forms of transmission, especially in developed countries. The objective of this work was to investigate if adipose tissue-derived mesenchymal stromal cells (ASC) can alter the course of the disease and attenuate pathology in a mouse model of chagasic cardiomyopathy. ASC were injected intraperitoneally at 3 days post-infection (dpi). Tracking by bioluminescence showed that cells remained in the abdominal cavity for up to 9 days after injection and most of them migrated to the abdominal or subcutaneous fat, an early parasite reservoir. ASC injection resulted in a significant reduction in blood parasitemia, which was followed by a decrease in cardiac tissue inflammation, parasitism and fibrosis at 30 dpi. At the same time point, analyses of cytokine release in cells isolated from the heart and exposed to T. cruzi antigens indicated an anti-inflammatory response in ASC-treated animals. In parallel, splenocytes exposed to the same antigens produced a pro-inflammatory response, which is important for the control of parasite replication, in placebo and ASC-treated groups. However, splenocytes from the ASC group released higher levels of IL-10. At 60 dpi, magnetic resonance imaging revealed that right ventricular (RV) dilation was prevented in ASC-treated mice. In conclusion, the injection of ASC early after T. cruzi infection prevents RV remodeling through the modulation of immune responses. Lymphoid organ response to the parasite promoted the control of parasite burden, while the heart, a target organ of Chagas disease, was protected from damage due to an improved control of inflammation in ASC-treated mice.

  11. Construction of a Corneal Stromal Equivalent with SMILE-Derived Lenticules and Fibrin Glue.

    PubMed

    Yin, Houfa; Qiu, Peijin; Wu, Fang; Zhang, Wei; Teng, Wenqi; Qin, Zhenwei; Li, Chao; Zhou, Jiaojie; Fang, Zhi; Tang, Qiaomei; Fu, Qiuli; Ma, Jian; Yang, Yabo

    2016-09-21

    The scarcity of corneal tissue to treat deep corneal defects and corneal perforations remains a challenge. Currently, small incision lenticule extraction (SMILE)-derived lenticules appear to be a promising alternative for the treatment of these conditions. However, the thickness and toughness of a single piece of lenticule are limited. To overcome these limitations, we constructed a corneal stromal equivalent with SMILE-derived lenticules and fibrin glue. In vitro cell culture revealed that the corneal stromal equivalent could provide a suitable scaffold for the survival and proliferation of corneal epithelial cells, which formed a continuous pluristratified epithelium with the expression of characteristic markers. Finally, anterior lamellar keratoplasty in rabbits demonstrated that the corneal stromal equivalent with decellularized lenticules and fibrin glue could repair the anterior region of the stroma, leading to re-epithelialization and recovery of both transparency and ultrastructural organization. Corneal neovascularization, graft degradation, and corneal rejection were not observed within 3 months. Taken together, the corneal stromal equivalent with SMILE-derived lenticules and fibrin glue appears to be a safe and effective alternative for the repair of damage to the anterior cornea, which may provide new avenues in the treatment of deep corneal defects or corneal perforations.

  12. Neural Stem Cells Derived Directly from Adipose Tissue.

    PubMed

    Petersen, Eric D; Zenchak, Jessica R; Lossia, Olivia V; Hochgeschwender, Ute

    2018-05-01

    Neural stem cells (NSCs) are characterized as self-renewing cell populations with the ability to differentiate into the multiple tissue types of the central nervous system. These cells can differentiate into mature neurons, astrocytes, and oligodendrocytes. This category of stem cells has been shown to be a promisingly effective treatment for neurodegenerative diseases and neuronal injury. Most treatment studies with NSCs in animal models use embryonic brain-derived NSCs. This approach presents both ethical and feasibility issues for translation to human patients. Adult tissue is a more practical source of stem cells for transplantation therapies in humans. Some adult tissues such as adipose tissue and bone marrow contain a wide variety of stem cell populations, some of which have been shown to be similar to embryonic stem cells, possessing many pluripotent properties. Of these stem cell populations, some are able to respond to neuronal growth factors and can be expanded in vitro, forming neurospheres analogous to cells harvested from embryonic brain tissue. In this study, we describe a method for the collection and culture of cells from adipose tissue that directly, without going through intermediates such as mesenchymal stem cells, results in a population of NSCs that are able to be expanded in vitro and be differentiated into functional neuronal cells. These adipose-derived NSCs display a similar phenotype to those directly derived from embryonic brain. When differentiated into neurons, cells derived from adipose tissue have spontaneous spiking activity with network characteristics similar to that of neuronal cultures.

  13. 45S5-Bioglass®-Based 3D-Scaffolds Seeded with Human Adipose Tissue-Derived Stem Cells Induce In Vivo Vascularization in the CAM Angiogenesis Assay

    PubMed Central

    Handel, Marina; Hammer, Timo R.; Nooeaid, Patcharakamon; Boccaccini, Aldo R.

    2013-01-01

    Poor vascularization is the key limitation for long-term acceptance of large three-dimensional (3D) tissue engineering constructs in regenerative medicine. 45S5 Bioglass® was investigated given its potential for applications in bone engineering. Since native Bioglass® shows insufficient angiogenic properties, we used a collagen coating, to seed human adipose tissue-derived stem cells (hASC) confluently onto 3D 45S5 Bioglass®-based scaffolds. To investigate vascularization by semiquantitative analyses, these biofunctionalized scaffolds were then subjected to in vitro human umbilical vein endothelial cells formation assays, and were also investigated in the chorioallantoic membrane (CAM) angiogenesis model, an in vivo angiogenesis assay, which uses the CAM of the hen's egg. In their native, nonbiofunctionalized state, neither Bioglass®-based nor biologically inert fibrous polypropylene control scaffolds showed angiogenic properties. However, significant vascularization was induced by hASC-seeded scaffolds (Bioglass® and polypropylene) in the CAM angiogenesis assay. Biofunctionalized scaffolds also showed enhanced tube lengths, compared to unmodified scaffolds or constructs seeded with fibroblasts. In case of biologically inert hernia meshes, the quantification of vascular endothelial growth factor secretion as the key angiogenic stimulus strongly correlated to the tube lengths and vessel numbers in all models. This correlation proved the CAM angiogenesis assay to be a suitable semiquantitative tool to characterize angiogenic effects of larger 3D implants. In addition, our results suggest that combinations of suitable scaffold materials, such as 45S5 Bioglass®, with hASC could be a promising approach for future tissue engineering applications. PMID:23837884

  14. The effect of diabetes on the wound healing potential of adipose-tissue derived stem cells.

    PubMed

    Kim, Sue Min; Kim, Yun Ho; Jun, Young Joon; Yoo, Gyeol; Rhie, Jong Won

    2016-03-01

    To investigate whether diabetes mellitus affects the wound-healing-promoting potential of adipose tissue-derived stem cells, we designed a wound-healing model using diabetic mice. We compared the degree of wound healing between wounds treated with normal adipose tissue-derived stem cells and wounds treated with diabetic adipose tissue-derived stem cells. We evaluated the wound-healing rate, the epithelial tongue distance, the area of granulation tissue, the number of capillary and the number of Ki-67-stained cells. The wound-healing rate was significantly higher in the normal adipose tissue-derived stem cells group than in the diabetic adipose tissue-derived stem cells group; it was also significantly higher in the normal adipose tissue-derived stem cells group than in the control group. Although the diabetic adipose tissue-derived stem cells group showed a better wound-healing rate than the control group, the difference was not statistically significant. Similar trends were observed for the other parameters examined: re-epithelisation and keratinocyte proliferation; granulation tissue formation; and dermal regeneration. However, with regard to the number of capillary, diabetic adipose tissue-derived stem cells retained their ability to promote neovasculisation and angiogenesis. These results reflect the general impairment of the therapeutic potential of diabetic adipose tissue-derived stem cells in vivo. © 2016 Medicalhelplines.com Inc and John Wiley & Sons Ltd.

  15. Adipose-Derived Cell Construct Stabilizes Heart Function and Increases Microvascular Perfusion in an Established Infarct

    PubMed Central

    Nguyen, Quang T.; Touroo, Jeremy S.; Aird, Allison L.; Chang, Raymond C.; Ng, Chin K.; Hoying, James B.; Williams, Stuart K.

    2013-01-01

    We have previously shown that myocardial infarction (MI) immediately treated with an epicardial construct containing stromal vascular fraction (SVF) from adipose tissue preserved microvascular function and left ventricle contractile mechanisms. In order to evaluate a more clinically relevant condition, we investigated the cardiac recovery potential of an SVF construct implanted onto an established infarct. SVF cells were isolated from rat adipose tissue, plated on Vicryl, and cultured for 14 days. Fischer-344 rats were separated into MI groups: (a) 6-week MI (MI), (b) 6-week MI treated with an SVF construct at 2 weeks (MI SVF), (c) 6-week MI with Vicryl construct at 2 weeks (MI Vicryl), and (d) MI 2wk (time point of intervention). Emax, an indicator of systolic performance and contractile function, was lower in the MI and MI Vicryl versus MI SVF. Positron emission tomography imaging (18F-fluorodeoxyglucose) revealed a decreased percentage of relative infarct volume in the MI SVF versus MI and MI Vicryl. Total vessel count and percentage of perfusion assessed via immunohistochemistry were both increased in the infarct region of MI SVF versus MI and MI Vicryl. Overall cardiac function, percentage of relative infarct, and percentage of perfusion were similar between MI SVF and MI 2wk; however, total vessel count increased after SVF treatment. These data suggest that SVF treatment of an established infarct stabilizes the heart at the time point of intervention by preventing a worsening of cardiac performance and infarcted volume, and is associated with increased microvessel perfusion in the area of established infarct. PMID:24106337

  16. [Role of chronic inflammation in adipose tissue in the pathophysiology of obesity].

    PubMed

    Suganami, Takayoshi; Ogawa, Yoshihiro

    2013-02-01

    Obesity may be viewed as a chronic low-grade inflammatory disease as well as a metabolic disease. Evidence has accumulated suggesting that chronic inflammation in adipose tissue leads to dramatic changes in number and cell type of stromal cells during the course of obesity, which is referred to as"adipose tissue remodeling". Among stromal cells, macrophages in obese adipose tissue are considered to be crucial for adipose tissue inflammation, which results in dysregulated adipocytokine production and ectopic fat accumulation. Understanding the molecular mechanism underlying adipose tissue inflammation would contribute to the identification of novel therapeutic strategies to prevent or treat obesity-induced metabolic derangements.

  17. Adipose tissue-derived stem cells inhibit neointimal formation in a paracrine fashion in rat femoral artery.

    PubMed

    Takahashi, Masao; Suzuki, Etsu; Oba, Shigeyoshi; Nishimatsu, Hiroaki; Kimura, Kenjiro; Nagano, Tetsuo; Nagai, Ryozo; Hirata, Yasunobu

    2010-02-01

    Subcutaneous adipose tissue contains a lot of stem cells [adipose-derived stem cells (ASCs)] that can differentiate into a variety of cell lineages. In this study, we isolated ASCs from Wistar rats and examined whether ASCs would efficiently differentiate into vascular endothelial cells (ECs) in vitro. We also administered ASCs in a wire injury model of rat femoral artery and examined their effects. ASCs expressed CD29 and CD90, but not CD34, suggesting that ASCs resemble bone marrow-derived mesenchymal stem cells. When induced to differentiate into ECs with endothelial growth medium (EGM), ASCs expressed Flt-1, but not Flk-1 or mature EC markers such as CD31 and vascular endothelial cadherin. ASCs produced angiopoietin-1 when they were cultured in EGM. ASCs stimulated the migration of EC, as assessed by chemotaxis assay. When ASCs that were cultured in EGM were injected in the femoral artery, the ASCs potently and significantly inhibited neointimal formation without being integrated in the endothelial layer. EGM-treated ASCs significantly suppressed neointimal formation even when they were administered from the adventitial side. ASC administration significantly promoted endothelial repair. These results suggested that although ASCs appear to have little capacity to differentiate into mature ECs, ASCs have the potential to secrete paracrine factors that stimulate endothelial repair. Our results also suggested that ASCs inhibited neointimal formation via their paracrine effect of stimulation of EC migration in situ rather than the direct integration into the endothelial layer.

  18. Polymeric film of 6-arm-poly(ethylene glycol) amine graphene oxide with poly (ε-caprolactone): Adherence and growth of adipose derived mesenchymal stromal cells culture on rat bladder

    NASA Astrophysics Data System (ADS)

    Durán, Marcela; Durán, Nelson; Luzo, Angela C. M.; Duarte, Adriana S. S.; Volpe, Bruno B.; Ceragioli, Helder J.; Andrade, Patricia F.; De Souza, Joel G.; Fávaro, Wagner J.

    2017-06-01

    Nanotechnology has been more present in different fields related to health. The need to find a durable material, of easy use, and which does not interfere significantly in the growth and differentiation of stem cells for the construction of a scaffold for use in urologic surgery, with the purpose of reducing infections, regeneration times and even graft rejection during reconstitution in patients with urethral stricture was conducted a broad survey of information about this and came to the consensus of this project: using graphene oxide, a widely studied nanomaterials which has been presenting numerous beneficial results when in contact with the adipose-derived stem cells. Advanced techniques for the growth, differentiation and proliferation of adipose-derived stem cells were used, as well as the characterization of graphene oxide sheets. For this study, it was prepared the graphene oxide/6 ARM-Poly (ethylene glycol) amine films with poly (ε-caprolactone). The graphene suspension in organic solvent was prepared by using an ultrasonicator bath and subsequently, the film was formed by solvent evaporation. Total characterization of graphene oxide/6 ARM-PEG-amine/ poly (ε-caprolactone) film was carried out. It was tested growth and adhesion of adipose-derived stem cells on the film, as well as, were verified the histopathological effects of this scaffold when implanted in the urinary bladder to repair the lesion. Our results demonstrated that this scaffold with adipose-derived stem cells enhanced the repair in rat urinary bladder defect model, resulting in a regular bladder. Improved organized muscle bundles and urothelial layer were observed in animals treated with this scaffold with adipose-derived stem cells compared with those treated only suture thread or scaffold. Thus, our biomaterial could be suitable for tissue engineered urinary tract reconstruction.

  19. Wound-healing potential of human umbilical cord blood-derived mesenchymal stromal cells in vitro--a pilot study.

    PubMed

    You, Hi-Jin; Namgoong, Sik; Han, Seung-Kyu; Jeong, Seong-Ho; Dhong, Eun-Sang; Kim, Woo-Kyung

    2015-11-01

    Our previous studies demonstrated that human bone marrow-derived mesenchymal stromal cells have great potential for wound healing. However, it is difficult to clinically utilize cultured stem cells. Recently, human umbilical cord blood-derived mesenchymal stromal cells (hUCB-MSCs) have been commercialized for cartilage repair as a first cell therapy product that uses allogeneic stem cells. Should hUCB-MSCs have a superior effect on wound healing as compared with fibroblasts, which are the main cell source in current cell therapy products for wound healing, they may possibly replace fibroblasts. The purpose of this in vitro study was to compare the wound-healing activity of hUCB-MSCs with that of fibroblasts. This study was particularly designed to compare the effect of hUCB-MSCs on diabetic wound healing with those of allogeneic and autologous fibroblasts. Healthy (n = 5) and diabetic (n = 5) fibroblasts were used as the representatives of allogeneic and autologous fibroblasts for diabetic patients in the control group. Human UCB-MSCs (n = 5) were used in the experimental group. Cell proliferation, collagen synthesis and growth factor (basic fibroblast growth factor, vascular endothelial growth factor and transforming growth factor-β) production were compared among the three cell groups. Human UCB-MSCs produced significantly higher amounts of vascular endothelial growth factor and basic fibroblast growth factor when compared with both fibroblast groups. Human UCB-MSCs were superior to diabetic fibroblasts but not to healthy fibroblasts in collagen synthesis. There were no significant differences in cell proliferation and transforming growth factor-β production. Human UCB-MSCs may have greater capacity for diabetic wound healing than allogeneic or autologous fibroblasts, especially in angiogenesis. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  20. Can Human Embryonic Stem Cell-Derived Stromal Cells Serve a Starting Material for Myoblasts?

    PubMed Central

    Ando, Yu; Saito, Marie; Machida, Masakazu; Yoshida-Noro, Chikako; Akutsu, Hidenori; Takahashi, Masataka

    2017-01-01

    A large number of myocytes are necessary to treat intractable muscular disorders such as Duchenne muscular dystrophy with cell-based therapies. However, starting materials for cellular therapy products such as myoblasts, marrow stromal cells, menstrual blood-derived cells, and placenta-derived cells have a limited lifespan and cease to proliferate in vitro. From the viewpoints of manufacturing and quality control, cells with a long lifespan are more suitable as a starting material. In this study, we generated stromal cells for future myoblast therapy from a working cell bank of human embryonic stem cells (ESCs). The ESC-derived CD105+ cells with extensive in vitro proliferation capability exhibited myogenesis and genetic stability in vitro. These results imply that ESC-derived CD105+ cells are another cell source for myoblasts in cell-based therapy for patients with genetic muscular disorders. Since ESCs are immortal, mesenchymal stromal cells generated from ESCs can be manufactured at a large scale in one lot for pharmaceutical purposes. PMID:28706537

  1. Construction of a Corneal Stromal Equivalent with SMILE-Derived Lenticules and Fibrin Glue

    PubMed Central

    Yin, Houfa; Qiu, Peijin; Wu, Fang; Zhang, Wei; Teng, Wenqi; Qin, Zhenwei; Li, Chao; Zhou, Jiaojie; Fang, Zhi; Tang, Qiaomei; Fu, Qiuli; Ma, Jian; Yang, Yabo

    2016-01-01

    The scarcity of corneal tissue to treat deep corneal defects and corneal perforations remains a challenge. Currently, small incision lenticule extraction (SMILE)-derived lenticules appear to be a promising alternative for the treatment of these conditions. However, the thickness and toughness of a single piece of lenticule are limited. To overcome these limitations, we constructed a corneal stromal equivalent with SMILE-derived lenticules and fibrin glue. In vitro cell culture revealed that the corneal stromal equivalent could provide a suitable scaffold for the survival and proliferation of corneal epithelial cells, which formed a continuous pluristratified epithelium with the expression of characteristic markers. Finally, anterior lamellar keratoplasty in rabbits demonstrated that the corneal stromal equivalent with decellularized lenticules and fibrin glue could repair the anterior region of the stroma, leading to re-epithelialization and recovery of both transparency and ultrastructural organization. Corneal neovascularization, graft degradation, and corneal rejection were not observed within 3 months. Taken together, the corneal stromal equivalent with SMILE-derived lenticules and fibrin glue appears to be a safe and effective alternative for the repair of damage to the anterior cornea, which may provide new avenues in the treatment of deep corneal defects or corneal perforations. PMID:27651001

  2. Pulmonary Retention of Adipose Stromal Cells Following Intravenous Delivery Is Markedly Altered in the Presence of ARDS.

    PubMed

    Lu, Hongyan; Cook, Todd; Poirier, Christophe; Merfeld-Clauss, Stephanie; Petrache, Irina; March, Keith L; Bogatcheva, Natalia V

    2016-01-01

    Transplantation of mesenchymal stromal cells (MSCs) has been shown to effectively prevent lung injury in several preclinical models of acute respiratory distress syndrome (ARDS). Since MSC therapy is tested in clinical trials for ARDS, there is an increased need to define the dynamics of cell trafficking and organ-specific accumulation. We examined how the presence of ARDS changes retention and organ-specific distribution of intravenously delivered MSCs isolated from subcutaneous adipose tissue [adipose-derived stem cells (ADSCs)]. This type of cell therapy was previously shown to ameliorate ARDS pathology. ARDS was triggered by lipopolysaccharide (LPS) aspiration, 4 h after which 300,000 murine CRE + ADSCs were delivered intravenously. The distribution of ADSCs in the lungs and other organs was assessed by real-time polymerase chain reaction (PCR) of genomic DNA. As anticipated, the majority of delivered ADSCs accumulated in the lungs of both control and LPS-challenged mice, with minor amounts distributed to the liver, kidney, spleen, heart, and brain. Interestingly, within 2 h following ADSC administration, LPS-challenged lungs retained significantly lower levels of ADSCs compared to control lungs (∼7% vs. 15% of the original dose, respectively), whereas the liver, kidney, spleen, and brain of ARDS-affected animals retained significantly higher numbers of ADSCs compared to control animals. In contrast, 48 h later, only LPS-challenged lungs continued to retain ADSCs (∼3% of the original dose), whereas the lungs of control animals and nonpulmonary organs in either control or ARDS mice had no detectable levels of ADSCs. Our data suggest that the pulmonary microenvironment during ARDS may lessen the pulmonary capillary occlusion by MSCs immediately following cell delivery while facilitating pulmonary retention of the cells.

  3. Oxaloacetate and adipose stromal cells-conditional medium synergistically protected potassium/serum deprivation-induced neuronal apoptosis.

    PubMed

    Liu, Qingpeng; Zhao, Gang; Zhou, Changwei; Farlow, Martin R; Du, Yansheng; Xu, Guangxu; Gu, Huiying

    2017-01-01

    Adipose stromal cells conditioned media (ASC-CM) protect neurons in a variety of neuronal death models including potassium/serum deprivation-induced neuronal apoptosis. In this study, we found that ASC-CM contained glutamate oxaloacetate transaminase and its substrate, oxaloacetate (OAA) directly protected cerebellar granule neurons (CGN) from apoptosis induced by serum and potassium deprivation. Additionally, OAA inhibited serum and potassium deprivation-induced caspase 3 activation. ASC-CM and OAA in combination had a synergistic neuroprotective effect. Clearly, different from ASC-CM-induced neuroprotection, OAA-induced neuroprotection was Akt- independent but JNK-dependent. These data establish a mechanistic basis supporting that the application of ASC-CM for neuroprotective treatments could be significantly enhanced by addition of OAA. Copyright © 2016 Elsevier Inc. All rights reserved.

  4. Transcriptome-derived stromal and immune scores infer clinical outcomes of patients with cancer.

    PubMed

    Liu, Wei; Ye, Hua; Liu, Ying-Fu; Xu, Chao-Qun; Zhong, Yue-Xian; Tian, Tian; Ma, Shi-Wei; Tao, Huan; Li, Ling; Xue, Li-Chun; He, Hua-Qin

    2018-04-01

    The stromal and immune cells that form the tumor microenvironment serve a key role in the aggressiveness of tumors. Current tumor-centric interpretations of cancer transcriptome data ignore the roles of stromal and immune cells. The aim of the present study was to investigate the clinical utility of stromal and immune cells in tissue-based transcriptome data. The 'Estimation of STromal and Immune cells in MAlignant Tumor tissues using Expression data' (ESTIMATE) algorithm was used to probe diverse cancer datasets and the fraction of stromal and immune cells in tumor tissues was scored. The association between the ESTIMATE scores and patient survival data was asessed; it was indicated that the two scores have implications for patient survival, metastasis and recurrence. Analysis of a colorectal cancer progression dataset revealed that decreased levels immune cells could serve an important role in cancer progression. The results of the present study indicated that trasncriptome-derived stromal and immune scores may be a useful indicator of cancer prognosis.

  5. Effect of gender and adiposity on in vivo vascular function in young African Americans.

    PubMed

    Dass, Namrata; Kilakkathi, Sindhu; Obi, Brittaney; Moosreiner, Andrea; Krishnaswami, Shanthi; Widlansky, Michael E; Kidambi, Srividya

    2017-05-01

    The relationship between obesity and high blood pressure is not as strong among African Americans (AA) as compared to Caucasians. We designed the current study to determine the effect of adiposity on vascular endothelial function (a harbinger of hypertension) among young healthy AA without additional cardiovascular disease risk factors. A total of 108 AA subjects (46 women) between the ages of 18 and 45 years were recruited. All the subjects were normotensive, nonsmokers, and normoglycemic. Anthropometric and cardiovascular disease risk factor measurements (lipid, insulin resistance, and inflammatory markers) were obtained. Vascular endothelial function was measured by brachial artery flow-mediated dilation (FMD). Adiposity distribution was measured by using magnetic resonance imaging scan. There were no gender differences in age and levels of blood pressure, lipids, insulin resistance, and inflammatory markers. Women had higher total body fat percentage and higher peripheral adiposity compared to men. We observed that total and central adiposity did not correlate significantly with brachial artery FMD in women (r = -0.12 and r = 0.23, respectively; P = NS). However, in men, waist circumference was positively associated with FMD (r = 0.3, P ≤ .05). Hyperemic flow was negatively correlated significantly with total and central adiposity in men (r = -0.34 and r = -0.48, respectively; P < .05), but not in women (r = -0.26 and r = 0.03, respectively; P = NS). Our study suggests that increased adiposity may pose greater risk to AA men compared to AA women by adversely affecting resistance vessel function (as measured by hyperemic flow). Larger studies are necessary to validate these findings. Copyright © 2017 American Society of Hypertension. Published by Elsevier Inc. All rights reserved.

  6. The influence of thiazolidinediones on adipogenesis in vitro and in vivo: potential modifiers of intramuscular adipose tissue deposition in meat animals.

    PubMed

    Hausman, G J; Poulos, S P; Pringle, T D; Azain, M J

    2008-04-01

    Thiazolidinediones (TZD) are insulin sensitizing agents currently used for the treatment of type 2 diabetes and are widely used as adipogenic agents because they are ligands of peroxisome proliferator-activated receptor gamma (PPARgamma), a key adipogenic transcription factor. In vivo and in vitro studies of TZD as potential modifiers of intramuscular or marbling adipogenesis are reviewed. Thiazolidinedione-induced adipogenesis has been reported in numerous cell culture systems, including rodent, human, bovine, and porcine adipose tissue stromal-vascular (S-V) cell cultures. Studies of porcine S-V cell cultures derived from semitendinosus muscle show that TZD can potentially modify intramuscular or marbling adipogenesis. Preadipocyte recruitment was TZD-dependent in muscle S-V cultures but TZD-independent in adipose S-V cultures. There appear to be differences between adipocytes in muscle and subcutaneous adipose tissue, reminiscent of differences observed in adipocytes from different adipose tissue depots. Troglitazone, a TZD, induces marbling adipogenesis without inhibiting myogenesis when cells are grown on laminin precoated culture dishes. Additionally, troglitazone treatment does not increase lipid content in porcine adipose tissue or muscle S-V cell cultures. Thiazolidinedione treatment increases lipid content of muscle in rodents and humans; however, rosiglitazone treatment for 49 d in pigs did not influence muscle lipid content and meat quality, but several significant changes in muscle fatty acid composition were observed. Although timing of treatment with TZD needs to be optimized, evidence suggests these compounds may enhance marbling deposition in swine.

  7. In vitro mesenchymal trilineage differentiation and extracellular matrix production by adipose and bone marrow derived adult equine multipotent stromal cells on a collagen scaffold.

    PubMed

    Xie, Lin; Zhang, Nan; Marsano, Anna; Vunjak-Novakovic, Gordana; Zhang, Yanru; Lopez, Mandi J

    2013-12-01

    Directed differentiation of adult multipotent stromal cells (MSC) is critical for effective treatment strategies. This study was designed to evaluate the capability of equine MSC from bone marrow (BMSC) and adipose tissue (ASC) on a type I collagen (COLI) scaffold to undergo chondrogenic, osteogenic and adipogenic differentiation and form extracellular matrix (ECM) in vitro. Following determination of surface antigen expression, MSC were loaded into scaffolds in a perfusion bioreactor and loading efficiency was quantified. Cell-scaffold constructs were assessed after loading and 7, 14 and 21 days of culture in stromal or induction medium. Cell number was determined with DNA content, cell viability and spatial uniformity with confocal laser microscopy and cell phenotype and matrix production with light and scanning electron microscopy and mRNA levels. The MSC were positive for CD29 (>90 %), CD44 (>99 %), and CD105 (>60 %). Loading efficiencies were >70 %. The ASC and BMSC cell numbers on scaffolds were affected by culture in induction medium differently. Viable cells remained uniformly distributed in scaffolds for up to 21 days and could be directed to differentiate or to maintain an MSC phenotype. Micro- and ultrastructure showed lineage-specific cell and ECM changes. Lineage-specific mRNA levels differed between ASC and BMSC with induction and changed with time. Based on these results, equine ASC and BMSC differentiate into chondrogenic, osteogenic and adipogenic lineages and form ECM similarly on COLI scaffolds. The collected data supports the potential for equine MSC-COLI constructs to support diverse equine tissue formation for controlled biological studies.

  8. Evaluation of 2 Purification Methods for Isolation of Human Adipose-Derived Stem Cells Based on Red Blood Cell Lysis With Ammonium Chloride and Hypotonic Sodium Chloride Solution.

    PubMed

    Li, Sheng-Hong; Liao, Xuan; Zhou, Tian-En; Xiao, Li-Ling; Chen, Yuan-Wen; Wu, Fan; Wang, Jing-Ru; Cheng, Biao; Song, Jian-Xing; Liu, Hong-Wei

    2017-01-01

    The present study was conducted to compare 2 purification methods for isolation of human adipose-derived stromal vascular fraction or stem cells (ADSCs) based on red blood cell (RBC) lysis with 155 mM ammonium chloride (NH4Cl) and hypotonic sodium chloride (NaCl) solution, and try to develop a safe, convenient, and cost-effective purification method for clinical applications. Adipose-derived stem cells and RBC were harvested from the fatty and fluid portions of liposuction aspirates, respectively. The suitable concentration of hypotonic NaCl solution on RBC lysis for purification of ADSCs was developed by RBC osmotic fragility test and flow cytometry analysis. The effects of 155 mM NH4Cl or 0.3% NaCl solution on ADSCs proliferation and RBC lysis efficiency were examined by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide assay and lysis efficiency test, respectively. In addition, the adipogenic and osteogenic capabilities, phenotype and genetic stability of ADSCs were evaluated by oil red staining, alkaline phosphatase activity measurement, flow cytometry, and karyotype analysis, respectively. Sodium chloride solution in 0.3% concentration effectively removed RBCs and did not influence the survival of ADSCs in the 10-minute incubation time. The lysis efficiency did not differ significantly between 0.3% NaCl and 155 mM NH4Cl. Moreover, the adipogenic and osteogenic capabilities, surface marker expression and karyotype of the ADSCs were not affected by lysis solutions or by lysis per se. However, the proliferation capacity in the 0.3% NaCl group was superior to that in 155 mM NH4Cl group. Our data suggest that 0.3% NaCl solution is useful for isolating ADSCs from liposuction aspirate for clinical applications with safety, convenience, and cost-effect.

  9. Adipocytes impair efficacy of antiretroviral therapy.

    PubMed

    Couturier, Jacob; Winchester, Lee C; Suliburk, James W; Wilkerson, Gregory K; Podany, Anthony T; Agarwal, Neeti; Xuan Chua, Corrine Ying; Nehete, Pramod N; Nehete, Bharti P; Grattoni, Alessandro; Sastry, K Jagannadha; Fletcher, Courtney V; Lake, Jordan E; Balasubramanyam, Ashok; Lewis, Dorothy E

    2018-06-01

    Adequate distribution of antiretroviral drugs to infected cells in HIV patients is critical for viral suppression. In humans and primates, HIV- and SIV-infected CD4 T cells in adipose tissues have recently been identified as reservoirs for infectious virus. To better characterize adipose tissue as a pharmacological sanctuary for HIV-infected cells, in vitro experiments were conducted to assess antiretroviral drug efficacy in the presence of adipocytes, and drug penetration in adipose tissue cells (stromal-vascular-fraction cells and mature adipocytes) was examined in treated humans and monkeys. Co-culture experiments between HIV-1-infected CD4 T cells and primary human adipocytes showed that adipocytes consistently reduced the antiviral efficacy of the nucleotide reverse transcriptase inhibitor tenofovir and its prodrug forms tenofovir disoproxil fumarate (TDF) and tenofovir alafenamide (TAF). In HIV-infected persons, LC-MS/MS analysis of intracellular lysates derived from adipose tissue stromal-vascular-fraction cells or mature adipocytes suggested that integrase inhibitors penetrate adipose tissue, whereas penetration of nucleoside/nucleotide reverse transcriptase inhibitors such as TDF, emtricitabine, abacavir, and lamivudine is restricted. The limited distribution and functions of key antiretroviral drugs within fat depots may contribute to viral persistence in adipose tissue. Copyright © 2018 Elsevier B.V. All rights reserved.

  10. Validation of osteogenic properties of Cytochalasin D by high-resolution RNA-sequencing in mesenchymal stem cells derived from bone marrow and adipose tissues.

    PubMed

    Samsonraj, Rebekah; Paradise, Christopher R; Dudakovic, Amel; Sen, Buer; Nair, Asha A; Dietz, Allan B; Deyle, David R; Cool, Simon M; Rubin, Janet; van Wijnen, Andre

    2018-06-08

    Differentiation of mesenchymal stromal/stem cells (MSCs) involves a series of molecular signals and gene transcription events required for attaining cell lineage commitment. Modulation of the actin cytoskeleton using cytochalasin D (CytoD) drives osteogenesis at early time points in bone marrow-derived MSCs, and also initiates a robust osteogenic differentiation program in adipose-derived MSCs. To understand the molecular basis for these pronounced effects on osteogenic differentiation, we investigated global changes in gene expression in CytoD-treated murine and human MSCs by high-resolution RNA-sequencing (RNA-seq) analysis. A three-way bioinformatic comparison between human adipose-derived, human bone marrow-derived and mouse bone marrow-derived MSCs revealed significant upregulation of genes linked to extracellular matrix organization, cell adhesion and bone metabolism. As anticipated, the activation of these differentiation related genes is accompanied by a downregulation of nuclear and cell cycle-related genes presumably reflecting cytostatic effects of CytoD. We also identified eight novel CytoD activated genes - VGLL4, ARHGAP24, KLHL24, RCBTB2, BDH2, SCARF2, ACAD10, HEPH - which are commonly upregulated across the two species and tissue sources of our MSC samples. We selected the Hippo-pathway related VGLL4 gene, which encodes the transcriptional co-factor Vestigial-like 4, for further study because this pathway is linked to osteogenesis. VGLL4 siRNA depletion reduces mineralization of adipose-derived MSCs during CytoD-induced osteogenic differentiation. Together, our RNA-seq analyses suggest that while the stimulatory effects of CytoD on osteogenesis are pleiotropic and depend on the biological state of the cell type, a small group of genes including VGLL4 may contribute to MSC commitment towards the bone lineage.

  11. Substance P enhances proliferation and paracrine potential of adipose-derived stem cells in vitro

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kim, Suna; Piao, Jiyuan; Son, Youngsook

    Stem cells have tremendous promise to treat intractable diseases. Notably, adipose-derived stem cells (ADSCs) are actively being investigated because of ease of sampling and high repopulation capacity in vitro. ADSCs can exert a therapeutic effect through differentiation and paracrine potential, and these actions have been proven in many diseases, including cutaneous and inflammatory diseases. Transplantation of ADSCs necessitates therapeutic quantities and thus, long term ex vivo culture of ADSCs. However, this procedure can impair the activity of ADSCs and provoke cellular senescence, leading to low efficacy in vivo. Accordingly, strategies to restore cellular activity and inhibit senescence of stem cells during ex vivo culturemore » are needed for stem cell-based therapies. This study evaluated a potential supplementary role of Substance P (SP) in ADSC ex vivo culture. After confirming that the ADSC cell cycle was damaged by passage 6 (p6), ADSCs at p6 were cultured with SP, and their proliferation rates, cumulative cell numbers, cytokine profiles, and impact on T/endothelial cells were assessed. Long-term culture weakened proliferation ability and secretion of the cytokines, transforming growth factor-beta 1 (TGF-beta1), vascular endothelial growth factor (VEGF), and stromal cell derived factor-1 alpha (SDF-1alpha) in ADSCs. However, SP treatment reduced the population doubling time (PDT), enabling gain of a sufficient number of ADSCs at early passages. In addition, SP restored cytokine secretion, enhancing the ADSC-mediated paracrine effect on T cell and human umbilical vein endothelial cells (HUVECs). Taken together, these results suggest that SP can retain the therapeutic effect of ADSCs by elevating their proliferative and paracrine potential in ex vivo culture. - Highlights: • Long-term culture of ADSCs leads to cell senescence. • Paracrine potential of ADSC decreases as passage number increases. • SP enhances the weakened proliferation

  12. Leptin produced by obese adipose stromal/stem cells enhances proliferation and metastasis of estrogen receptor positive breast cancers.

    PubMed

    Strong, Amy L; Ohlstein, Jason F; Biagas, Brandi A; Rhodes, Lyndsay V; Pei, Dorothy T; Tucker, H Alan; Llamas, Claire; Bowles, Annie C; Dutreil, Maria F; Zhang, Shijia; Gimble, Jeffrey M; Burow, Matthew E; Bunnell, Bruce A

    2015-08-19

    The steady increase in the incidence of obesity among adults has been paralleled with higher levels of obesity-associated breast cancer. While recent studies have suggested that adipose stromal/stem cells (ASCs) isolated from obese women enhance tumorigenicity, the mechanism(s) by which this occurs remains undefined. Evidence suggests that increased adiposity results in increased leptin secretion from adipose tissue, which has been shown to increased cancer cell proliferation. Previously, our group demonstrated that ASCs isolated from obese women (obASCs) also express higher levels of leptin relative to ASCs isolated from lean women (lnASCs) and that this obASC-derived leptin may account for enhanced breast cancer cell growth. The current study investigates the impact of inhibiting leptin expression in lnASCs and obASCs on breast cancer cell (BCC) growth and progression. Estrogen receptor positive (ER+) BCCs were co-cultured with leptin shRNA lnASCs or leptin shRNA obASCs and changes in the proliferation, migration, invasion, and gene expression of BCCs were investigated. To assess the direct impact of leptin inhibition in obASCs on BCC proliferation, MCF7 cells were injected alone or mixed with control shRNA obASCs or leptin shRNA obASCs into SCID/beige mice. ER+ BCCs were responsive to obASCs during direct co-culture, whereas lnASCs were unable to increase ER(+) BCC growth. shRNA silencing of leptin in obASCs negated the enhanced proliferative effects of obASC on BCCs following direct co-culture. BCCs co-cultured with obASCs demonstrated enhanced expression of epithelial-to-mesenchymal transition (EMT) and metastasis genes (SERPINE1, MMP-2, and IL-6), while BCCs co-cultured with leptin shRNA obASCs did not display similar levels of gene induction. Knockdown of leptin significantly reduced tumor volume and decreased the number of metastatic lesions to the lung and liver. These results correlated with reduced expression of both SERPINE1 and MMP-2 in tumors formed

  13. Analysis of type II diabetes mellitus adipose-derived stem cells for tissue engineering applications

    PubMed Central

    Minteer, Danielle Marie; Young, Matthew T; Lin, Yen-Chih; Over, Patrick J; Rubin, J Peter; Gerlach, Jorg C

    2015-01-01

    To address the functionality of diabetic adipose-derived stem cells in tissue engineering applications, adipose-derived stem cells isolated from patients with and without type II diabetes mellitus were cultured in bioreactor culture systems. The adipose-derived stem cells were differentiated into adipocytes and maintained as functional adipocytes. The bioreactor system utilizes a hollow fiber–based technology for three-dimensional perfusion of tissues in vitro, creating a model in which long-term culture of adipocytes is feasible, and providing a potential tool useful for drug discovery. Daily metabolic activity of the adipose-derived stem cells was analyzed within the medium recirculating throughout the bioreactor system. At experiment termination, tissues were extracted from bioreactors for immunohistological analyses in addition to gene and protein expression. Type II diabetic adipose-derived stem cells did not exhibit significantly different glucose consumption compared to adipose-derived stem cells from patients without type II diabetes (p > 0.05, N = 3). Expression of mature adipocyte genes was not significantly different between diabetic/non-diabetic groups (p > 0.05, N = 3). Protein expression of adipose tissue grown within all bioreactors was verified by Western blotting.The results from this small-scale study reveal adipose-derived stem cells from patients with type II diabetes when removed from diabetic environments behave metabolically similar to the same cells of non-diabetic patients when cultured in a three-dimensional perfusion bioreactor, suggesting that glucose transport across the adipocyte cell membrane, the hindrance of which being characteristic of type II diabetes, is dependent on environment. The presented observation describes a tissue-engineered tool for long-term cell culture and, following future adjustments to the culture environment and increased sample sizes, potentially for anti-diabetic drug testing. PMID:26090087

  14. Human mesenchymal stromal cell-derived extracellular vesicles attenuate aortic aneurysm formation and macrophage activation via microRNA-147.

    PubMed

    Spinosa, Michael; Lu, Guanyi; Su, Gang; Bontha, Sai Vineela; Gehrau, Ricardo; Salmon, Morgan D; Smith, Joseph R; Weiss, Mark L; Mas, Valeria R; Upchurch, Gilbert R; Sharma, Ashish K

    2018-05-29

    The formation of an abdominal aortic aneurysm (AAA) is characterized by inflammation, macrophage infiltration, and vascular remodeling. In this study, we tested the hypothesis that mesenchymal stromal cell (MSC)-derived extracellular vesicles (EVs) immunomodulate aortic inflammation, to mitigate AAA formation via modulation of microRNA-147. An elastase-treatment model of AAA was used in male C57BL/6 wild-type (WT) mice. Administration of EVs in elastase-treated WT mice caused a significant attenuation of aortic diameter and mitigated proinflammatory cytokines, inflammatory cell infiltration, an increase in smooth muscle cell α-actin expression, and a decrease in elastic fiber disruption, compared with untreated mice. A 10-fold up-regulation of microRNA (miR)-147, a key mediator of macrophage inflammatory responses, was observed in murine aortic tissue in elastase-treated mice compared with controls on d 14. EVs derived from MSCs transfected with miR-147 mimic, but not with miR-147 inhibitor, attenuated aortic diameter, inflammation, and leukocyte infiltration in elastase-treated mice. In vitro studies of human aortic tissue explants and murine-derived CD11b + macrophages induced proinflammatory cytokines after elastase treatment, and the expression was attenuated by cocultures with EVs transfected with miR-147 mimic, but not with miR-147 inhibitor. Thus, our findings define a critical role of MSC-derived EVs in attenuation of aortic inflammation and macrophage activation via miR-147 during AAA formation.-Spinosa, M., Lu, G., Su, G., Bontha, S. V., Gehrau, R., Salmon, M. D., Smith, J. R., Weiss, M. L., Mas, V. R., Upchurch, G. R., Sharma, A. K. Human mesenchymal stromal cell-derived extracellular vesicles attenuate aortic aneurysm formation and macrophage activation via microRNA-147.

  15. Male Bovine GH Transgenic Mice Have Decreased Adiposity With an Adipose Depot-Specific Increase in Immune Cell Populations

    PubMed Central

    Benencia, Fabian; Harshman, Stephanie; Duran-Ortiz, Silvana; Lubbers, Ellen R.; List, Edward O.; Householder, Lara; Al-Naeeli, Mawadda; Liang, Xiaoyu; Welch, Lonnie; Kopchick, John J.

    2015-01-01

    White adipose tissue (WAT) is composed of mature adipocytes and a stromal vascular fraction (SVF), which contains a variety of cells, including immune cells that vary among the different WAT depots. Growth hormone (GH) impacts immune function and adiposity in an adipose depot-specific manner. However, its effects on WAT immune cell populations remain unstudied. Bovine GH transgenic (bGH) mice are commonly used to study the in vivo effects of GH. These giant mice have an excess of GH action, impaired glucose metabolism, decreased adiposity, increased lean mass, and a shortened lifespan. Therefore, the purpose of this study was to characterize the WAT depot-specific differences in immune cell populations in the presence of excess GH in vivo. Three WAT depots were assessed: inguinal (sc), epididymal (EPI), and mesenteric (MES). Subcutaneous and MES bGH WAT depots showed a significantly higher number of total SVF cells, yet only MES bGH WAT had higher leukocyte counts compared with control samples. By means of flow cytometry analysis of the SVF, we detected greater macrophage and regulatory T-cell infiltration in sc and MES bGH WAT depots compared with controls. However, no differences were observed in the EPI WAT depot. RNA-sequencing confirmed significant alterations in pathways related to T-cell infiltration and activation in the sc depot with fewer significant changes in the EPI bGH WAT depot. These findings collectively point to a previously unrecognized role for GH in influencing the distribution of WAT immune cell populations in a depot-specific manner. PMID:25521584

  16. Male bovine GH transgenic mice have decreased adiposity with an adipose depot-specific increase in immune cell populations.

    PubMed

    Benencia, Fabian; Harshman, Stephanie; Duran-Ortiz, Silvana; Lubbers, Ellen R; List, Edward O; Householder, Lara; Al-Naeeli, Mawadda; Liang, Xiaoyu; Welch, Lonnie; Kopchick, John J; Berryman, Darlene E

    2015-05-01

    White adipose tissue (WAT) is composed of mature adipocytes and a stromal vascular fraction (SVF), which contains a variety of cells, including immune cells that vary among the different WAT depots. Growth hormone (GH) impacts immune function and adiposity in an adipose depot-specific manner. However, its effects on WAT immune cell populations remain unstudied. Bovine GH transgenic (bGH) mice are commonly used to study the in vivo effects of GH. These giant mice have an excess of GH action, impaired glucose metabolism, decreased adiposity, increased lean mass, and a shortened lifespan. Therefore, the purpose of this study was to characterize the WAT depot-specific differences in immune cell populations in the presence of excess GH in vivo. Three WAT depots were assessed: inguinal (sc), epididymal (EPI), and mesenteric (MES). Subcutaneous and MES bGH WAT depots showed a significantly higher number of total SVF cells, yet only MES bGH WAT had higher leukocyte counts compared with control samples. By means of flow cytometry analysis of the SVF, we detected greater macrophage and regulatory T-cell infiltration in sc and MES bGH WAT depots compared with controls. However, no differences were observed in the EPI WAT depot. RNA-sequencing confirmed significant alterations in pathways related to T-cell infiltration and activation in the sc depot with fewer significant changes in the EPI bGH WAT depot. These findings collectively point to a previously unrecognized role for GH in influencing the distribution of WAT immune cell populations in a depot-specific manner.

  17. Weight-dependent changes of immune system in adipose tissue: Importance of leptin

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Caspar-Bauguil, S.; Groupe de Recherche et d'Etude en Nutrition; Cousin, B.

    2006-07-15

    Ancestral lymphoid cells reside in adipose tissues, and their numbers are highly altered in obesity. Leptin, production of which is correlated to fat mass, is strongly involved in the relationships between adipose tissues and immune system. We investigated in epididymal (EPI) and inguinal (ING) fat pads to determine whether 1) lymphocyte phenotypes were correlated to the tissue weight and 2) leptin was involved in such relationships. Immunohistological analyses revealed a tight relationship between the T and NK lymphocytes of the stromal vascular fraction and adipocytes. We identified a significant negative and positive correlation between EPI weight and the percentage ofmore » NK and total T cells respectively by cytofluorometric analyses. The NK and ancestral {gamma}{delta} T cell contents were directly dependent of leptin since they increased significantly in high-fat (HF) diet mice but not in leptin-deficient (ob/ob) mice as compared to control. By contrast, the {alpha}{beta} T cell content seemed independent of leptin because their percentages increased significantly with the EPI weight whatever the type of mice (control, HF, ob/ob). The present study suggests that adipose tissues present, according to their localization, different immunological mechanisms that might be involved in the regulation of adipose cells functions and proliferations.« less

  18. Aging exacerbates obesity-induced oxidative stress and inflammation in perivascular adipose tissue in mice: a paracrine mechanism contributing to vascular redox dysregulation and inflammation.

    PubMed

    Bailey-Downs, Lora C; Tucsek, Zsuzsanna; Toth, Peter; Sosnowska, Danuta; Gautam, Tripti; Sonntag, William E; Csiszar, Anna; Ungvari, Zoltan

    2013-07-01

    Obesity in the elderly individuals is increasing at alarming rates and there is evidence suggesting that elderly individuals are more vulnerable to the deleterious cardiovascular effects of obesity than younger individuals. However, the specific mechanisms through which aging and obesity interact to promote the development of cardiovascular disease remain unclear. The present study was designed to test the hypothesis that aging exacerbates obesity-induced inflammation in perivascular adipose tissue, which contributes to increased vascular oxidative stress and inflammation in a paracrine manner. To test this hypothesis, we assessed changes in the secretome, reactive oxygen species production, and macrophage infiltration in periaortic adipose tissue of young (7 month old) and aged (24 month old) high-fat diet-fed obese C57BL/6 mice. High-fat diet-induced vascular reactive oxygen species generation significantly increased in aged mice, which was associated with exacerbation of endothelial dysfunction and vascular inflammation. In young animals, high-fat diet-induced obesity promoted oxidative stress in the perivascular adipose tissue, which was associated with a marked proinflammatory shift in the profile of secreted cytokines and chemokines. Aging exacerbated obesity-induced oxidative stress and inflammation and significantly increased macrophage infiltration in periaortic adipose tissue. Using cultured arteries isolated from young control mice, we found that inflammatory factors secreted from the perivascular fat tissue of obese aged mice promote significant prooxidative and proinflammatory phenotypic alterations in the vascular wall, mimicking the aging phenotype. Overall, our findings support an important role for localized perivascular adipose tissue inflammation in exacerbation of vascular oxidative stress and inflammation in aging, an effect that likely enhances the risk for development of cardiovascular diseases from obesity in the elderly individuals.

  19. Aging Exacerbates Obesity-Induced Oxidative Stress and Inflammation in Perivascular Adipose Tissue in Mice: A Paracrine Mechanism Contributing to Vascular Redox Dysregulation and Inflammation

    PubMed Central

    Bailey-Downs, Lora C.; Tucsek, Zsuzsanna; Toth, Peter

    2013-01-01

    Obesity in the elderly individuals is increasing at alarming rates and there is evidence suggesting that elderly individuals are more vulnerable to the deleterious cardiovascular effects of obesity than younger individuals. However, the specific mechanisms through which aging and obesity interact to promote the development of cardiovascular disease remain unclear. The present study was designed to test the hypothesis that aging exacerbates obesity-induced inflammation in perivascular adipose tissue, which contributes to increased vascular oxidative stress and inflammation in a paracrine manner. To test this hypothesis, we assessed changes in the secretome, reactive oxygen species production, and macrophage infiltration in periaortic adipose tissue of young (7 month old) and aged (24 month old) high-fat diet–fed obese C57BL/6 mice. High-fat diet–induced vascular reactive oxygen species generation significantly increased in aged mice, which was associated with exacerbation of endothelial dysfunction and vascular inflammation. In young animals, high-fat diet–induced obesity promoted oxidative stress in the perivascular adipose tissue, which was associated with a marked proinflammatory shift in the profile of secreted cytokines and chemokines. Aging exacerbated obesity-induced oxidative stress and inflammation and significantly increased macrophage infiltration in periaortic adipose tissue. Using cultured arteries isolated from young control mice, we found that inflammatory factors secreted from the perivascular fat tissue of obese aged mice promote significant prooxidative and proinflammatory phenotypic alterations in the vascular wall, mimicking the aging phenotype. Overall, our findings support an important role for localized perivascular adipose tissue inflammation in exacerbation of vascular oxidative stress and inflammation in aging, an effect that likely enhances the risk for development of cardiovascular diseases from obesity in the elderly individuals

  20. Myostatin Attenuation In Vivo Reduces Adiposity, but Activates Adipogenesis.

    PubMed

    Li, Naisi; Yang, Qiyuan; Walker, Ryan G; Thompson, Thomas B; Du, Min; Rodgers, Buel D

    2016-01-01

    A potentially novel approach for treating obesity includes attenuating myostatin as this increases muscle mass and decreases fat mass. Notwithstanding, conflicting studies report that myostatin stimulates or inhibits adipogenesis and it is unknown whether reduced adiposity with myostatin attenuation results from changes in fat deposition or adipogenesis. We therefore quantified changes in the stem, transit amplifying and progenitor cell pool in white adipose tissue (WAT) and brown adipose tissue (BAT) using label-retaining wild-type and mstn(-/-) (Jekyll) mice. Muscle mass was larger in Jekyll mice, WAT and BAT mass was smaller and label induction was equal in all tissues from both wild-type and Jekyll mice. The number of label-retaining cells, however, dissipated quicker in WAT and BAT of Jekyll mice and was only 25% and 17%, respectively, of wild-type cell counts 1 month after induction. Adipose cell density was significantly higher in Jekyll mice and increased over time concomitant with label-retaining cell disappearance, which is consistent with enhanced expansion and differentiation of the stem, transit amplifying and progenitor pool. Stromal vascular cells from Jekyll WAT and BAT differentiated into mature adipocytes at a faster rate than wild-type cells and although Jekyll WAT cells also proliferated quicker in vitro, those from BAT did not. Differentiation marker expression in vitro, however, suggests that mstn(-/-) BAT preadipocytes are far more sensitive to the suppressive effects of myostatin. These results suggest that myostatin attenuation stimulates adipogenesis in vivo and that the reduced adiposity in mstn(-/-) animals results from nutrient partitioning away from fat and in support of muscle.

  1. Adipose-derived Mesenchymal Stem Cells and Their Reparative Potential in Ischemic Heart Disease.

    PubMed

    Badimon, Lina; Oñate, Blanca; Vilahur, Gemma

    2015-07-01

    Adipose tissue has long been considered an energy storage and endocrine organ; however, in recent decades, this tissue has also been considered an abundant source of mesenchymal cells. Adipose-derived stem cells are easily obtained, show a strong capacity for ex vivo expansion and differentiation to other cell types, release a large variety of angiogenic factors, and have immunomodulatory properties. Thus, adipose tissue is currently the focus of considerable interest in the field of regenerative medicine. In the context of coronary heart disease, numerous experimental studies have supported the safety and efficacy of adipose-derived stem cells in the setting of myocardial infarction. These results have encouraged the clinical use of these stem cells, possibly prematurely. Indeed, the presence of cardiovascular risk factors, such as hypertension, coronary disease, diabetes mellitus, and obesity, alter and reduce the functionality of adipose-derived stem cells, putting in doubt the efficacy of their autologous implantation. In the present article, white adipose tissue is described, the stem cells found in this tissue are characterized, and the use of these cells is discussed according to the preclinical and clinical trials performed so far. Copyright © 2015 Sociedad Española de Cardiología. Published by Elsevier España, S.L.U. All rights reserved.

  2. Partial regeneration of uterine horns in rats through adipose-derived stem cell sheets.

    PubMed

    Sun, Huijun; Lu, Jie; Li, Bo; Chen, Shuqiang; Xiao, Xifeng; Wang, Jun; Wang, Jingjing; Wang, Xiaohong

    2018-06-20

    Severe uterine damage and infection lead to intrauterine adhesions, which result in hypomenorrhea, amenorrhea and infertility. Cell sheet engineering has shown great promise in clinical applications. Adipose-derived stem cells (ADSCs) are emerging as an alternative source of stem cells for cell-based therapies. In the present study, we investigated the feasibility of applying ADSCs as seed cells to form scaffold-free cell sheet. Data showed that ADSC sheets expressed higher levels of FGF, Col I, TGFβ and VEGF than ADSCs in suspension, while increased expression of this gene set was associated with stemness, including Nanog, Oct4 and Sox2. We then investigated the therapeutic effects of 3D ADSCs sheet on regeneration in a rat model. We found that ADSCs were mainly detected in the basal layer of the regenerating endometrium in the cell sheet group at 21 days after transplantation. Additionally, some ADSCs differentiated into stromal-like cells. Moreover, ADSC sheets transplanted into partially excised uteri promoted regeneration of the endometrium cells, muscle cells and stimulated angiogenesis, and also resulted in better pregnancy outcomes. Therefore, ADSC sheet therapy shows considerable promise as a new treatment for severe uterine damage.

  3. Human Adipose-Derived Stem Cells Labeled with Plasmonic Gold Nanostars for Cellular Tracking and Photothermal Cancer Cell Ablation.

    PubMed

    Shammas, Ronnie L; Fales, Andrew M; Crawford, Bridget M; Wisdom, Amy J; Devi, Gayathri R; Brown, David A; Vo-Dinh, Tuan; Hollenbeck, Scott T

    2017-04-01

    Gold nanostars are unique nanoplatforms that can be imaged in real time and transform light energy into heat to ablate cells. Adipose-derived stem cells migrate toward tumor niches in response to chemokines. The ability of adipose-derived stem cells to migrate and integrate into tumors makes them ideal vehicles for the targeted delivery of cancer nanotherapeutics. To test the labeling efficiency of gold nanostars, undifferentiated adipose-derived stem cells were incubated with gold nanostars and a commercially available nanoparticle (Qtracker), then imaged using two-photon photoluminescence microscopy. The effects of gold nanostars on cell phenotype, proliferation, and viability were assessed with flow cytometry, 3-(4,5-dimethylthiazolyl-2)-2,5-diphenyltetrazolium bromide metabolic assay, and trypan blue, respectively. Trilineage differentiation of gold nanostar-labeled adipose-derived stem cells was induced with the appropriate media. Photothermolysis was performed on adipose-derived stem cells cultured alone or in co-culture with SKBR3 cancer cells. Efficient uptake of gold nanostars occurred in adipose-derived stem cells, with persistence of the luminescent signal over 4 days. Labeling efficiency and signal quality were greater than with Qtracker. Gold nanostars did not affect cell phenotype, viability, or proliferation, and exhibited stronger luminescence than Qtracker throughout differentiation. Zones of complete ablation surrounding the gold nanostar-labeled adipose-derived stem cells were observed following photothermolysis in both monoculture and co-culture models. Gold nanostars effectively label adipose-derived stem cells without altering cell phenotype. Once labeled, photoactivation of gold nanostar-labeled adipose-derived stem cells ablates neighboring cancer cells, demonstrating the potential of adipose-derived stem cells as a vehicle for the delivery of site-specific cancer therapy.

  4. Decidualized Human Endometrial Stromal Cells Mediate Hemostasis, Angiogenesis, and Abnormal Uterine Bleeding

    PubMed Central

    Lockwood, Charles J.; Krikun, Graciela; Hickey, Martha; Huang, S. Joseph; Schatz, Frederick

    2011-01-01

    Factor VII binds trans-membrane tissue factor to initiate hemostasis by forming thrombin. Tissue factor expression is enhanced in decidualized human endometrial stromal cells during the luteal phase. Long-term progestin only contraceptives elicit: 1) abnormal uterine bleeding from fragile vessels at focal bleeding sites, 2) paradoxically high tissue factor expression at bleeding sites; 3) reduced endometrial blood flow promoting local hypoxia and enhancing reactive oxygen species levels; and 4) aberrant angiogenesis reflecting increased stromal cell-expressed vascular endothelial growth factor, decreased Angiopoietin-1 and increased endothelial cell-expressed Angiopoietin-2. Aberrantly high local vascular permeability enhances circulating factor VII to decidualized stromal cell-expressed tissue factor to generate excess thrombin. Hypoxia-thrombin interactions augment expression of vascular endothelial growth factor and interleukin-8 by stromal cells. Thrombin, vascular endothelial growth factor and interlerukin-8 synergis-tically augment angiogenesis in a milieu of reactive oxygen species-induced endothelial cell activation. The resulting enhanced vessel fragility promotes abnormal uterine bleeding. PMID:19208784

  5. Patterns of adiposity, vascular phenotypes and cognitive function in the 1946 British Birth Cohort.

    PubMed

    Masi, Stefano; Georgiopoulos, Georgios; Khan, Tauseef; Johnson, William; Wong, Andrew; Charakida, Marietta; Whincup, Peter; Hughes, Alun D; Richards, Marcus; Hardy, Rebecca; Deanfield, John

    2018-05-28

    The relationship between long-term exposure to whole body or central obesity and cognitive function, as well as its potential determinants, remain controversial. In this study, we assessed (1) the potential impact of 30 years exposure to different patterns of whole body and central adiposity on cognitive function at 60-64 years, (2) whether trajectories of central adiposity can provide additional information on later cognitive function compared to trajectories of whole body adiposity, and (3) the influence of vascular phenotypes on these associations. The study included 1249 participants from the prospective cohort MRC National Survey of Health and Development. Body mass index (BMI), waist circumference (WC), and vascular (carotid intima-media thickness, carotid-femoral pulse wave velocity) and cognitive function (memory, processing speed, reaction time) data, at 60-64 years, were used to assess the associations between different patterns of adult WC or BMI (from 36 years of age) and late midlife cognitive performance, as well as the proportion of this association explained by cardiovascular phenotypes. Longer exposure to elevated WC was related to lower memory performance (p < 0.001 for both) and longer choice reaction time (p = 0.003). A faster gain of WC between 36 and 43 years of age was associated with the largest change in reaction time and memory test (P < 0.05 for all). Similar associations were observed when patterns of WC were substituted with patterns of BMI, but when WC and BMI were included in the same model, only patterns of WC remained significantly associated with cognitive function. Participants who dropped one BMI category and maintained a lower BMI had similar memory performance to those of normal weight during the whole follow-up. Conversely, those who dropped and subsequently regained one BMI category had a memory function similar to those with 30 years exposure to elevated BMI. Adjustment for vascular phenotypes, levels of

  6. Ultrasound -Assisted Gene Transfer to Adipose Tissue-Derived Stem/Progenitor Cells (ASCs)

    NASA Astrophysics Data System (ADS)

    Miyamoto, Yoshitaka; Ueno, Hitomi; Hokari, Rei; Yuan, Wenji; Kuno, Shuichi; Kakimoto, Takashi; Enosawa, Shin; Negishi, Yoichi; Yoshinaka, Kiyoshi; Matsumoto, Yoichiro; Chiba, Toshio; Hayashi, Shuji

    2011-09-01

    In recent years, multilineage adipose tissue-derived stem cells (ASCs) have become increasingly attractive as a promising source for cell transplantation and regenerative medicine. Particular interest has been expressed in the potential to make tissue stem cells, such as ASCs and marrow stromal cells (MSCs), differentiate by gene transfection. Gene transfection using highly efficient viral vectors such as adeno- and sendai viruses have been developed for this purpose. Sonoporation, or ultrasound (US)-assisted gene transfer, is an alternative gene manipulation technique which employs the creation of a jet stream by ultrasonic microbubble cavitation. Sonoporation using non-viral vectors is expected to be a much safer, although less efficient, tool for prospective clinical gene therapy. In this report, we assessed the efficacy of the sonoporation technique for gene transfer to ASCs. We isolated and cultured adipocyets from mouse adipose tissue. ASCs that have the potential to differentiate with transformation into adipocytes or osteoblasts were obtained. Using the US-assisted system, plasmid DNA containing beta-galactosidase (beta-Gal) and green fluorescent protein (GFP) genes were transferred to the ASCs. For this purpose, a Sonopore 4000 (NEPAGENE Co.) and a Sonazoid (Daiichi Sankyo Co.) instrument were used in combination. ASCs were subjected to US (3.1 MHz, 50% duty cycle, burst rate 2.0 Hz, intensity 1.2 W/cm2, exposure time 30 sec). We observed that the gene was more efficiently transferred with increased concentrations of plasmid DNA (5-150 μg/mL). However, further optimization of the US parameters is required, as the gene transfer efficiency was still relatively low. In conclusion, we herein demonstrate that a gene can be transferred to ASCs using our US-assisted system. In regenerative medicine, this system might resolve the current issues surrounding the use of viral vectors for gene transfer.

  7. Decrease of Perivascular Adipose Tissue Browning Is Associated With Vascular Dysfunction in Spontaneous Hypertensive Rats During Aging.

    PubMed

    Kong, Ling-Ran; Zhou, Yan-Ping; Chen, Dong-Rui; Ruan, Cheng-Chao; Gao, Ping-Jin

    2018-01-01

    Functional perivascular adipose tissue (PVAT) is necessary to maintain vascular physiology through both mechanical support and endocrine or paracrine ways. PVAT shows a brown adipose tissue (BAT)-like feature and the browning level of PVAT is dependent on the anatomic location and species. However, it is not clear whether PVAT browning is involved in the vascular tone regulation in spontaneously hypertensive rats (SHRs). In the present study, we aimed to illustrate the effect of aging on PVAT browning and subsequent vasomotor reaction in SHRs. Herein we utilized histological staining and western blot to detect the characteristics of thoracic PVAT (tPVAT) in 8-week-old and 16-week-old SHR and Wistar-Kyoto (WKY) rats. We also detected vascular reactivity analysis to determine the effect of tPVAT on vasomotor reaction during aging. The results showed that tPVAT had a similar phenotype to BAT, including smaller adipocyte size and positive uncoupling protein-1 (UCP1) staining. Interestingly, the tPVAT of 8-week-old SHR showed increased BAT phenotypic marker expression compared to WKY, whereas the browning level of tPVAT had a more dramatic decrease from 8 to 16 weeks of age in SHR than age-matched WKY rats. The vasodilation effect of tPVAT on aortas had no significant difference in 8-week-old WKY and SHR, whereas this effect is obviously decreased in 16-week-old SHR compared to WKY. In contrast, tPVAT showed a similar vasoconstriction effect in 8- or 16-week-old WKY and SHR rats. Moreover, we identified an important vasodilator adenosine, which regulates adipocyte browning and may be a potential PVAT-derived relaxing factor. Adenosine is dramatically decreased from 8 to 16 weeks of age in the tPVAT of SHR. In summary, aging is associated with a decrease of tPVAT browning and adenosine production in SHR rats. These may result in attenuated vasodilation effect of the tPVAT in SHR during aging.

  8. Exendin-4 enhances the migration of adipose-derived stem cells to neonatal rat ventricular cardiomyocyte-derived conditioned medium via the phosphoinositide 3-kinase/Akt-stromal cell-derived factor-1α/CXC chemokine receptor 4 pathway

    PubMed Central

    ZHOU, HAO; YANG, JUNJIE; XIN, TING; ZHANG, TAO; HU, SHUNYIN; ZHOU, SHANSHAN; CHEN, GUANGHUI; CHEN, YUNDAI

    2015-01-01

    Adipose-derived stem cells (ADSCs) are considered a suitable source of cells for the repair of tissue following acute myocardial infarction (AMI); however, the transplantation efficiency of ADSCs remains low. Therefore, identification of an efficient method to enhance the migration of engrafted cells to the target site is required. The present study used exendin-4 (Ex-4), a glucagon-like peptide-1 receptor agonist, to optimize the migratory capacity of ADSCs. The aim was to determine the effect and mechanisms of Ex-4 on the migration of ADSCs to neonatal rat ventricular cardiomyocyte-derived conditioned medium (NRVC-CM). The ADSCs and cardiomyocytes were cultured in vitro. Following incubation of the ADSCs with Ex-4, cell proliferation was measured using an MTT assay and the expression levels of CXC chemokine receptor 4 (CXCR4) were investigated by reverse transctiption quantitative polymerase chain reaction (RT-qPCR), western blot analysis and flow cytometry. In addition, the expression levels of stromal cell-derived factor-1α (SDF-1α) were evaluated in the NRVC-CM treated with Ex-4 by ELISA, RT-qPCR and western blot analysis. The migration of the ADSCs to the NRVC-CM was examined using a Transwell assay. Changes in the protein expression levels of phosphorylated (p−)Akt were examined in the two types of cell by western blot analysis. The results suggested that Ex-4 promoted the proliferation and expression of CXCR4 in the ADSCs, increased the secretion of SDF-1α in the cardiomyocytes and increased the expression levels of p-Akt in both cells. However, the alterations to the SDF-1α/C XC R4 cascade in the cells were abrogated following pretreatment with LY-294002, a phosphoinositide 3-kinase(PI3K) inhibitor. Furthermore, a Transwell migration assay revealed marked translocation of the ADSCs through the membranes, towards the NRVC-CM, following treatment with Ex-4. However, these effects were reduced significantly by pretreatment of the cells with the SDF-1

  9. Hydrogel derived from decellularized porcine adipose tissue as a promising biomaterial for soft tissue augmentation.

    PubMed

    Tan, Qiu-Wen; Zhang, Yi; Luo, Jing-Cong; Zhang, Di; Xiong, Bin-Jun; Yang, Ji-Qiao; Xie, Hui-Qi; Lv, Qing

    2017-06-01

    Decellularized extracellular matrix (ECM) scaffolds from human adipose tissue, characterized by impressive adipogenic induction ability, are promising for soft tissue augmentation. However, scaffolds from autologous human adipose tissue are limited by the availability of tissue resources and the time necessary for scaffold fabrication. The objective of the current study was to investigate the adipogenic properties of hydrogels of decellularized porcine adipose tissue (HDPA). HDPA induced the adipogenic differentiation of human adipose-derived stem cells (ADSCs) in vitro, with significantly increased expression of adipogenic genes. Subcutaneous injection of HDPA in immunocompetent mice induced host-derived adipogenesis without cell seeding, and adipogenesis was significantly enhanced with ADSCs seeding. The newly formed adipocytes were frequently located on the basal side in the non-seeding group, but this trend was not observed in the ADSCs seeding group. Our results indicated that, similar to human adipose tissue, the ECM scaffold derived from porcine adipose tissue could provide an adipogenic microenvironment for adipose tissue regeneration and is a promising biomaterial for soft tissue augmentation. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1756-1764, 2017. © 2017 Wiley Periodicals, Inc.

  10. Canine Platelet Lysate Is Inferior to Fetal Bovine Serum for the Isolation and Propagation of Canine Adipose Tissue- and Bone Marrow-Derived Mesenchymal Stromal Cells

    PubMed Central

    Russell, Keith A.; Gibson, Thomas W. G.; Chong, Andrew; Co, Carmon; Koch, Thomas G.

    2015-01-01

    Background Mesenchymal stromal cells (MSC) are increasingly investigated for their clinical utility in dogs. Fetal bovine serum (FBS) is a common culture supplement used for canine MSC expansion. However, FBS content is variable, its clinical use carries risk of an immune response, and its cost is increasing due to global demand. Platelet lysate (PL) has proven to be a suitable alternative to FBS for expansion of human MSC. Hypothesis and Objectives We hypothesized that canine adipose tissue (AT) and bone marrow (BM) MSC could be isolated and expanded equally in PL and FBS at conventionally-used concentrations with differentiation of these MSC unaffected by choice of supplement. Our objectives were to evaluate the use of canine PL in comparison with FBS at four stages: 1) isolation, 2) proliferation, 3) spontaneous differentiation, and 4) directed differentiation. Results 1) Medium with 10% PL was unable to isolate MSC. 2) MSC, initially isolated in FBS-supplemented media, followed a dose-dependent response with no significant difference between PL and FBS cultures at up to 20% (AT) or 30% (BM) enrichment. Beyond these respective peaks, proliferation fell in PL cultures only, while a continued dose-dependent proliferation response was noted in FBS cultures. 3) Further investigation indicated PL expansion culture was inducing spontaneous adipogenesis in concentrations as low as 10% and as early as 4 days in culture. 4) MSC isolated in FBS, but expanded in either FBS or PL, maintained ability to undergo directed adipogenesis and osteogenesis, but not chondrogenesis. Conclusions/Significance Canine PL did not support establishment of MSC colonies from AT and BM, nor expansion of MSC, which appear to undergo spontaneous adipogenesis in response to PL exposure. In vivo studies are warranted to determine if concurrent use of MSC with any platelet-derived products such as platelet-rich plasma are associated with synergistic, neutral or antagonistic effects. PMID:26353112

  11. Pigment epithelium derived factor inhibits the growth of human endometrial implants in nude mice and of ovarian endometriotic stromal cells in vitro.

    PubMed

    Sun, Yanmei; Che, Xuan; Zhu, Libo; Zhao, Mengdan; Fu, Guofang; Huang, Xiufeng; Xu, Hong; Hu, Fuqiang; Zhang, Xinmei

    2012-01-01

    Angiogenesis is a prerequisite for the formation and development of endometriosis. Pigment epithelium derived factor (PEDF) is a natural inhibitor of angiogenesis. We previously demonstrated a reduction of PEDF in the peritoneal fluid, serum and endometriotic lesions from women with endometriosis compared with women without endometriosis. Here, we aim to investigate the inhibitory effect of PEDF on human endometriotic cells in vivo and in vitro. We found that PEDF markedly inhibited the growth of human endometrial implants in nude mice and of ovarian endometriotic stromal cells in vitro by up-regulating PEDF expression and down-regulating vascular endothelial growth factor (VEGF) expression. Moreover, apoptotic index was significantly increased in endometriotic lesions in vivo and endometriotic stromal cells in vitro when treated with PEDF. In mice treated with PEDF, decreased microvessel density labeled by Von Willebrand factor but not by α-Smooth Muscle Actin was observed in endometriotic lesions. And it showed no increase in PEDF expression of the ovary and uterus tissues. These findings suggest that PEDF gene therapy may be a new treatment for endometriosis.

  12. Pigment Epithelium Derived Factor Inhibits the Growth of Human Endometrial Implants in Nude Mice and of Ovarian Endometriotic Stromal Cells In Vitro

    PubMed Central

    Sun, Yanmei; Che, Xuan; Zhu, Libo; Zhao, Mengdan; Fu, Guofang; Huang, Xiufeng; Xu, Hong; Hu, Fuqiang; Zhang, Xinmei

    2012-01-01

    Angiogenesis is a prerequisite for the formation and development of endometriosis. Pigment epithelium derived factor (PEDF) is a natural inhibitor of angiogenesis. We previously demonstrated a reduction of PEDF in the peritoneal fluid, serum and endometriotic lesions from women with endometriosis compared with women without endometriosis. Here, we aim to investigate the inhibitory effect of PEDF on human endometriotic cells in vivo and in vitro. We found that PEDF markedly inhibited the growth of human endometrial implants in nude mice and of ovarian endometriotic stromal cells in vitro by up-regulating PEDF expression and down-regulating vascular endothelial growth factor (VEGF) expression. Moreover, apoptotic index was significantly increased in endometriotic lesions in vivo and endometriotic stromal cells in vitro when treated with PEDF. In mice treated with PEDF, decreased microvessel density labeled by Von Willebrand factor but not by α-Smooth Muscle Actin was observed in endometriotic lesions. And it showed no increase in PEDF expression of the ovary and uterus tissues. These findings suggest that PEDF gene therapy may be a new treatment for endometriosis. PMID:23028859

  13. Beneficial effects of calcitriol on hypertension, glucose intolerance, impairment of endothelium-dependent vascular relaxation, and visceral adiposity in fructose-fed hypertensive rats.

    PubMed

    Chou, Chu-Lin; Pang, Cheng-Yoong; Lee, Tony J F; Fang, Te-Chao

    2015-01-01

    Besides regulating calcium homeostasis, the effects of vitamin D on vascular tone and metabolic disturbances remain scarce in the literature despite an increase intake with high-fructose corn syrup worldwide. We investigated the effects of calcitriol, an active form of vitamin D, on vascular relaxation, glucose tolerance, and visceral fat pads in fructose-fed rats. Male Wistar-Kyoto rats were divided into 4 groups (n = 6 per group). Group Con: standard chow diet for 8 weeks; Group Fru: high-fructose diet (60% fructose) for 8 weeks; Group Fru-HVD: high-fructose diet as Group Fru, high-dose calcitriol treatment (20 ng / 100 g body weight per day) 4 weeks after the beginning of fructose feeding; and Group Fru-LVD: high-fructose diet as Group Fru, low-dose calcitriol treatment (10 ng / 100 g body weight per day) 4 weeks after the beginning of fructose feeding. Systolic blood pressure was measured twice a week by the tail-cuff method. Blood was examined for serum ionized calcium, phosphate, creatinine, glucose, triglycerides, and total cholesterol. Intra-peritoneal glucose intolerance test, aortic vascular reactivity, the weight of visceral fat pads, adipose size, and adipose angiotensin II levels were analyzed at the end of the study. The results showed that the fructose-fed rats significantly developed hypertension, impaired glucose tolerance, heavier weight and larger adipose size of visceral fat pads, and raised adipose angiotensin II expressions compared with the control rats. High- and low-dose calcitriol reduced modestly systolic blood pressure, increased endothelium-dependent aortic relaxation, ameliorated glucose intolerance, reduced the weight and adipose size of visceral fat pads, and lowered adipose angiotensin II expressions in the fructose-fed rats. However, high-dose calcitriol treatment mildly increased serum ionized calcium levels (1.44 ± 0.05 mmol/L). These results suggest a protective role of calcitriol treatment on endothelial function, glucose

  14. Myostatin Attenuation In Vivo Reduces Adiposity, but Activates Adipogenesis

    PubMed Central

    Li, Naisi; Yang, Qiyuan; Walker, Ryan G.; Thompson, Thomas B.; Du, Min

    2016-01-01

    A potentially novel approach for treating obesity includes attenuating myostatin as this increases muscle mass and decreases fat mass. Notwithstanding, conflicting studies report that myostatin stimulates or inhibits adipogenesis and it is unknown whether reduced adiposity with myostatin attenuation results from changes in fat deposition or adipogenesis. We therefore quantified changes in the stem, transit amplifying and progenitor cell pool in white adipose tissue (WAT) and brown adipose tissue (BAT) using label-retaining wild-type and mstn−/− (Jekyll) mice. Muscle mass was larger in Jekyll mice, WAT and BAT mass was smaller and label induction was equal in all tissues from both wild-type and Jekyll mice. The number of label-retaining cells, however, dissipated quicker in WAT and BAT of Jekyll mice and was only 25% and 17%, respectively, of wild-type cell counts 1 month after induction. Adipose cell density was significantly higher in Jekyll mice and increased over time concomitant with label-retaining cell disappearance, which is consistent with enhanced expansion and differentiation of the stem, transit amplifying and progenitor pool. Stromal vascular cells from Jekyll WAT and BAT differentiated into mature adipocytes at a faster rate than wild-type cells and although Jekyll WAT cells also proliferated quicker in vitro, those from BAT did not. Differentiation marker expression in vitro, however, suggests that mstn−/− BAT preadipocytes are far more sensitive to the suppressive effects of myostatin. These results suggest that myostatin attenuation stimulates adipogenesis in vivo and that the reduced adiposity in mstn−/− animals results from nutrient partitioning away from fat and in support of muscle. PMID:26580671

  15. Role of innate immune receptors TLR2 and TLR4 as mediators of the inflammatory reaction in human visceral adipose tissue.

    PubMed

    Fusaru, Ana Marina; Stănciulescu, Camelia Elena; Surlin, V; Taisescu, C; Bold, Adriana; Pop, O T; Baniţă, Ileana Monica; Crăiţoiu, Stefania; Pisoschi, Cătălina Gabriela

    2012-01-01

    White adipose tissue from different locations is characterized by significant differences in the structure of adipocyte "secretoma". Fat accumulation in the central-visceral depots is usually associated with a chronic inflammatory state, which is complicated by the metabolic syndrome. Recently, the adipose tissue was emerged to have an essential role in the innate immunity, adipocytes being considered effector cells due to the presence of the Toll-like receptors (TLRs). In this study, we compared the expression of TNF-α, TLR2 and TLR4 in peripheral-subcutaneous and central-peritoneal adipose depots in three different conditions - lean, obese and obese diabetic - using immunohistochemistry. Our results suggest a correlation between the incidence of the stromal vascular cells and adipocytes TNF-α and TLR4 in the visceral depots in strong correlation with adipose tissue expansion. TLR2 positive cells were seen in the peripheral depots from all groups without any association with fat accumulation. These results focus on the existence of a new pathogenic pathway, the activation of TLR4, for the involvement of visceral adipose tissue in the activation and maintenance of the inflammatory cascade in obesity.

  16. SDF-1 improves wound healing ability of glucocorticoid-treated adipose tissue-derived mesenchymal stem cells.

    PubMed

    Kato, Toshiki; Khanh, Vuong Cat; Sato, Kazutoshi; Takeuchi, Kosuke; Carolina, Erica; Yamashita, Toshiharu; Sugaya, Hisashi; Yoshioka, Tomokazu; Mishima, Hajime; Ohneda, Osamu

    2017-11-18

    Glucocorticoids cause the delayed wound healing by suppressing inflammation that is required for wound healing process. Adipose tissue-derived mesenchymal stem cells (AT-MSCs) play an important role for wound healing by their cytokine productions including stromal derived factor 1 (SDF-1). However, it has not been clear how glucocorticoids affect the wound healing ability of AT-MSCs. In this study, we found that glucocorticoid downregulated SDF-1 expression in AT-MSCs. In addition, glucocorticoid-treated AT-MSCs induced less migration of inflammatory cells and impaired wound healing capacity compared with glucocorticoid-untreated AT-MSCs. Of note, prostaglandin E2 (PGE2) synthesis-related gene expression was downregulated by glucocorticoid and PGE2 treatment rescued not only SDF-1 expression in the presence of glucocorticoid but also their wound healing capacity in vivo. Furthermore, we found SDF-1-overexpressed AT-MSCs restored wound healing capacity even after treatment of glucocorticoid. Consistent with the results obtained from glucocorticoid-treated AT-MSCs, we found that AT-MSCs isolated from steroidal osteonecrosis donors (sAT-MSCs) who received chronic glucocorticoid therapy showed less SDF-1 expression and impaired wound healing capacity compared with traumatic osteonecrosis donor-derived AT-MSCs (nAT-MSCs). Moreover, the SDF-1 level was also reduced in plasma derived from steroidal osteonecrosis donors compared with traumatic osteonecrosis donors. These results provide the evidence that concomitant application of AT-MSCs with glucocorticoid shows impaired biological modulatory effects that induce impaired wound healing. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. Neuroprotective effects of intravitreally transplanted adipose tissue and bone marrow-derived mesenchymal stem cells in an experimental ocular hypertension model.

    PubMed

    Emre, Esra; Yüksel, Nurşen; Duruksu, Gökhan; Pirhan, Dilara; Subaşi, Cansu; Erman, Gülay; Karaöz, Erdal

    2015-05-01

    The purpose of this study was to investigate the neuroprotective effects of bone marrow bone marrow-derived and adipose tissue-derived mesenchymal stromal cells (MSCs) that were intravitreally transplanted in an experimental ocular hypertension (OHT) model. An OHT rat model was generated by means of intracameral injection of hyaluronic acid into the anterior chamber. MSCs labeled with green fluorescence protein were transplanted intravitreally 1 week after OHT induction. At the end of the second and fourth weeks, retinal ganglion cells were visualized with the use of a flat-mount retina method and were evaluated by means of immunofluorescence staining against green fluorescence protein, vimentin, CD105, and cytokines (interleukin [IL]-1Ra, prostaglandin E2 receptor, IL-6, transforming growth factor-β1, interferon-γ and tumor necrosis factor-α). The retinal ganglion cell numbers per area were significantly improved in stem cell-treated OHT groups compared with that in the non-treated OHT group (P < 0.05). The results of immunohistochemical analyses indicated that a limited number of stem cells had integrated into the ganglion cell layer and the inner nuclear layer. The number of cells expressing proinflammatory cytokines (interferon-γ and tumor necrosis factor-α) decreased in the MSC-transferred group compared with that in the OHT group after 4 weeks (P < 0.01). On the other hand, IL-1Ra and prostaglandin E2 receptor expressions were increased in the rat bone marrow-derived MSC group but were more significant in the rat adipose tissue-derived MSC group (P < 0.01). After intravitreal transplantation, MSCs showed a neuroprotective effect in the rat OHT model. Therefore, MSCs promise an alternative therapy approach for functional recovery in the treatment of glaucoma. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  18. White adipose tissue cells and the progression of cachexia: inflammatory pathways

    PubMed Central

    Neves, Rodrigo X.; Rosa‐Neto, José Cesar; Yamashita, Alex S.; Matos‐Neto, Emidio M.; Riccardi, Daniela M. R.; Lira, Fabio S.; Batista, Miguel L.

    2015-01-01

    Abstract Background Cachexia is a systemic syndrome leading to body wasting, systemic inflammation, and to metabolic chaos. It is a progressive condition, and little is known about its dynamics. Detection of the early signs of the disease may lead to the attenuation of the associated symptoms. The white adipose tissue is an organ with endocrine functions, capable of synthesising and secreting a plethora of proteins, including cytokines, chemokines, and adipokines. It is well established that different adipose tissue depots demonstrate heterogeneous responses to physiological and pathological stimuli. The present study aimed at providing insight into adipocyte involvement in inflammation along the progression of cachexia. Methods Eight‐weeks‐old male rats were subcutaneously inoculated with a Walker 256 carcinosarcoma cell suspension (2 × 107 cells in 1.0 mL; tumour‐bearing, T) or Phosphate‐buffered saline (control, C). The retroperitoneal, epididymal, and mesenteric adipose pads were excised on Days 0, 7, and 14 post‐tumour cell injection, and the adipocytes were isolated. Results Mesenteric and epididymal adipocytes showed up‐regulation of IL‐1β protein expression and activation of the inflammasome pathway, contributing for whole tissue inflammation. The stromal vascular fraction of the retroperitoneal adipose tissue, on the other hand, seems to be the major contributor for the inflammation in this specific pad. Conclusion Adipocytes seem to play a relevant role in the establishment of white adipose tissue inflammation, through the activation of the NF‐κB and inflammasome pathways. In epididymal adipocytes, induction of the inflammasome may be detected already on Day 7 post‐tumour cell inoculation. PMID:27493872

  19. Comparative study of adipose-derived stem cells and bone marrow-derived stem cells in similar microenvironmental conditions

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Guneta, Vipra; Tan, Nguan Soon; KK Research Centre, KK Women's and Children Hospital, 100 Bukit Timah Road, Singapore 229899

    Mesenchymal stem cells (MSCs), which were first isolated from the bone marrow, are now being extracted from various other tissues in the body, including the adipose tissue. The current study presents systematic evidence of how the adipose tissue-derived stem cells (ASCs) and bone marrow-derived mesenchymal stem cells (Bm-MSCs) behave when cultured in specific pro-adipogenic microenvironments. The cells were first characterized and identified as MSCs in terms of their morphology, phenotypic expression, self-renewal capabilities and multi-lineage potential. Subsequently, the proliferation and gene expression profiles of the cell populations cultured on two-dimensional (2D) adipose tissue extracellular matrix (ECM)-coated tissue culture plastic (TCP)more » and in three-dimensional (3D) AlgiMatrix® microenvironments were analyzed. Overall, it was found that adipogenesis was triggered in both cell populations due to the presence of adipose tissue ECM. However, in 3D microenvironments, ASCs and Bm-MSCs were predisposed to the adipogenic and osteogenic lineages respectively. Overall, findings from this study will contribute to ongoing efforts in adipose tissue engineering as well as provide new insights into the role of the ECM and cues provided by the immediate microenvironment for stem cell differentiation. - Highlights: • Native adipose tissue ECM coated on 2D TCP triggers adipogenesis in both ASCs and Bm-MSCs. • A 3D microenvironment with similar stiffness to adipose tissue induces adipogenic differentiation of ASCs. • ASCs cultured in 3D alginate scaffolds exhibit predisposition to adipogenesis. • Bm-MSCs cultured in 3D alginate scaffolds exhibit predisposition to osteogenesis. • The native microenvironment of the cells affects their differentiation behaviour in vitro.« less

  20. Impairment of adipose tissue in Prader-Willi syndrome rescued by growth hormone treatment.

    PubMed

    Cadoudal, T; Buléon, M; Sengenès, C; Diene, G; Desneulin, F; Molinas, C; Eddiry, S; Conte-Auriol, F; Daviaud, D; Martin, P G P; Bouloumié, A; Salles, J-P; Tauber, M; Valet, P

    2014-09-01

    Prader-Willi syndrome (PWS) results from abnormalities in the genomic imprinting process leading to hypothalamic dysfunction with an alteration of growth hormone (GH) secretion. PWS is associated with early morbid obesity and short stature which can be efficiently improved with GH treatment. Our aims were to highlight adipose tissue structural and functional impairments in children with PWS and to study the modifications of those parameters on GH treatment. Plasma samples and adipose tissue biopsies were obtained from 23 research centers in France coordinated by the reference center for PWS in Toulouse, France. Lean controls (n=33), non-syndromic obese (n=53), untreated (n=26) and GH-treated PWS (n=43) children were enrolled in the study. Adipose tissue biopsies were obtained during scheduled surgeries from 15 lean control, 7 untreated and 8 GH-treated PWS children. Children with PWS displayed higher insulin sensitivity as shown by reduced glycemia, insulinemia and HOMA-IR compared with non-syndromic obese children. In contrast, plasma inflammatory cytokines such as TNF-α, MCP-1 and IL-8 were increased in PWS. Analysis of biopsies compared with control children revealed decreased progenitor cell content in the stromal vascular fraction of adipose tissue and an impairment of lipolytic response to β-adrenergic agonist in PWS adipocytes. Interestingly, both of these alterations in PWS seem to be ameliorated on GH treatment. Herein, we report adipose tissue dysfunctions in children with PWS which may be partially restored by GH treatment.

  1. Effects of insulin, triiodothyronine and fat soluble vitamins on adipocyte differentiation and LPL gene expression in the stromal-vascular cells of red sea bream, Pagrus major.

    PubMed

    Oku, Hiromi; Tokuda, Masaharu; Okumura, Takuji; Umino, Tetsuya

    2006-07-01

    Various kinds of hormones including insulin, triiodothyronine (T(3)) and fat-soluble vitamins have been proposed as mediators of adipocyte differentiation in mammals. To investigate the factors which are responsible for fish adipocyte differentiation, we developed a serum-free culture system of stromal-vascular cells of red sea bream adipose tissue and examined the effects of bovine insulin, T(3), and fat-soluble vitamins (all-trans retinoic acid, retinyl acetate and 1,25-dihydroxyvitamin D(3)) on the differentiation-linked expression of the lipoprotein lipase (LPL) gene. As assessed by the increase in LPL gene expression after 3 day cultivation, like in mammalian adipocytes, insulin enhanced the adipocyte differentiation in a concentration-dependent manner. During 2 week cultivation, bovine insulin promoted lipid accumulation in differentiating adipocytes concentration-dependently until the terminal differentiation. These results indicate that the differentiation of fish adipocytes is inducible by insulin alone. T(3) alone had no effect but enhanced the differentiation-linked LPL gene expression in the presence of insulin. Fat-soluble vitamins, unlike in mammalian adipocytes, did not show any significant effects. The method developed in this study should be of interest for the characterization of factors involved in fish adipocyte differentiation.

  2. Des-acyl ghrelin inhibits the capacity of macrophages to stimulate the expression of aromatase in breast adipose stromal cells.

    PubMed

    Au, CheukMan C; Docanto, Maria M; Zahid, Heba; Raffaelli, Francesca-Maria; Ferrero, Richard L; Furness, John B; Brown, Kristy A

    2017-06-01

    Des-acyl ghrelin is the unacylated form of the well-characterized appetite-stimulating hormone ghrelin. It affects a number of physiological processes, including increasing adipose lipid accumulation and inhibiting adipose tissue inflammation. Breast adipose tissue inflammation in obesity is associated with an increase in the expression of the estrogen biosynthetic enzyme, aromatase, and is hypothesized to create a hormonal milieu conducive to tumor growth. We previously reported that des-acyl ghrelin inhibits the expression and activity of aromatase in isolated human adipose stromal cells (ASCs), the main site of aromatase expression in the adipose tissue. The current study aimed to examine the effect of des-acyl ghrelin on the capacity of mouse macrophages (RAW264.7 cells) and human adipose tissue macrophages (ATMs) to stimulate aromatase expression in primary human breast ASCs. RAW264.7 cells were treated with 0, 10 and 100pM des-acyl ghrelin following activation with phorbol 12-myristate 13-acetate, and cells and conditioned media were collected after 6 and 24h. The effect of des-acyl ghrelin on macrophage polarization was examined by assessing mRNA expression of pro-inflammatory M1-specific marker Cd11c and anti-inflammatory M2-specific marker Cd206, as well as expression of Tnf and Ptgs2, known mediators of the macrophage-dependent stimulation of aromatase. TNF protein in conditioned media was assessed by ELISA. The effect of RAW264.7 and ATM-conditioned media on aromatase expression in ASCs was assessed after 6h. Results demonstrate des-acyl ghrelin significantly increases the expression of Cd206 and suppresses the expression of Cd11c, Tnf and Ptgs2 in activated RAW264.7 cells. Treatment of RAW264.7 and ATMs with des-acyl ghrelin also significantly reduces the capacity of these cells to stimulate aromatase transcript expression in human breast ASCs. Overall, these findings suggest that in addition to direct effects on aromatase in ASCs, des-acyl ghrelin also

  3. The effect of adipose derived stromal cells on oxidative stress level, lung emphysema and white blood cells of guinea pigs model of chronic obstructive pulmonary disease

    PubMed Central

    2014-01-01

    Background Chronic obstructive pulmonary disease (COPD) is a worldwide epidemic disease and a major cause of death and disability. The present study aimed to elucidate pharmacological effects of adipose derived stromal cells (ASCs) on pathological and biochemical factors in a guinea pig model of COPD. Guinea pigs were randomized into 5 groups including: Control, COPD, COPD + intratracheal delivery of PBS as a vehicle (COPD-PBS), COPD + intratracheal delivery of ASCs (COPD-ITASC) and COPD + intravenous injection of ASCs (COPD-IVASC). COPD was induced by exposing animals to cigarette smoke for 3 months. Cell therapy was performed immediately after the end of animal exposure to cigarette smoke and 14 days after that, white blood cells, oxidative stress indices and pathological changes of the lung were measured. Results Compared with control group, emphysema was clearly observed in the COPD and COPD-PBS groups (p < 0.001). Lung histopathologic changes of COPD-ITASC and COPD-IVASC groups showed non-significant improvement compared to COPD-PBS group. The COPD-ITASC group showed a significant increase in total WBC compared to COPD-PBS group but there was not a significant increase in this regard in COPD-IVASC group. The differential WBC showed no significant change in number of different types of leukocytes. The serum level of malondialdehyde (MDA) significantly decreased but thiol groups of broncho-alveolar lavage fluid (BALF) increased in both cell treated groups (p < 0.05 for all cases). Weight of animals decreased during smoke exposure and improved after PBS or cell therapy. However, no significant change was observed between the groups receiving PBS and the ones receiving ASCs. Conclusion Cell therapy with ASCs can help in reducing oxidative damage during smoking which may collectively hold promise in attenuation of the severity of COPD although the lung structural changes couldn’t be ameliorated with these pharmacological therapeutic methods. PMID:24495506

  4. The effect of adipose derived stromal cells on oxidative stress level, lung emphysema and white blood cells of guinea pigs model of chronic obstructive pulmonary disease.

    PubMed

    Ghorbani, Ahmad; Feizpour, Azadeh; Hashemzahi, Milad; Gholami, Lila; Hosseini, Mahmoud; Soukhtanloo, Mohammad; Vafaee Bagheri, Farzaneh; Khodaei, Esmaeil; Mohammadian Roshan, Nema; Boskabady, Mohammad Hossein

    2014-02-04

    Chronic obstructive pulmonary disease (COPD) is a worldwide epidemic disease and a major cause of death and disability. The present study aimed to elucidate pharmacological effects of adipose derived stromal cells (ASCs) on pathological and biochemical factors in a guinea pig model of COPD. Guinea pigs were randomized into 5 groups including: Control, COPD, COPD + intratracheal delivery of PBS as a vehicle (COPD-PBS), COPD + intratracheal delivery of ASCs (COPD-ITASC) and COPD + intravenous injection of ASCs (COPD-IVASC). COPD was induced by exposing animals to cigarette smoke for 3 months. Cell therapy was performed immediately after the end of animal exposure to cigarette smoke and 14 days after that, white blood cells, oxidative stress indices and pathological changes of the lung were measured. Compared with control group, emphysema was clearly observed in the COPD and COPD-PBS groups (p < 0.001). Lung histopathologic changes of COPD-ITASC and COPD-IVASC groups showed non-significant improvement compared to COPD-PBS group. The COPD-ITASC group showed a significant increase in total WBC compared to COPD-PBS group but there was not a significant increase in this regard in COPD-IVASC group. The differential WBC showed no significant change in number of different types of leukocytes. The serum level of malondialdehyde (MDA) significantly decreased but thiol groups of broncho-alveolar lavage fluid (BALF) increased in both cell treated groups (p < 0.05 for all cases). Weight of animals decreased during smoke exposure and improved after PBS or cell therapy. However, no significant change was observed between the groups receiving PBS and the ones receiving ASCs. Cell therapy with ASCs can help in reducing oxidative damage during smoking which may collectively hold promise in attenuation of the severity of COPD although the lung structural changes couldn't be ameliorated with these pharmacological therapeutic methods.

  5. Long-term in-vivo tumorigenic assessment of human culture-expanded adipose stromal/stem cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    MacIsaac, Zoe Marie, E-mail: zmm4a@virgina.edu; Shang, Hulan, E-mail: shanghulan@gmail.com; Agrawal, Hitesh, E-mail: hiteshdos@hotmail.com

    2012-02-15

    After more than a decade of extensive experimentation, the promise of stem cells to revolutionize the field of medicine has negotiated their entry into clinical trial. Adipose tissue specifically holds potential as an attainable and abundant source of stem cells. Currently undergoing investigation are adipose stem cell (ASC) therapies for diabetes and critical limb ischemia, among others. In the enthusiastic pursuit of regenerative therapies, however, questions remain regarding ASC persistence and migration, and, importantly, their safety and potential for neoplasia. To date, assays of in vivo ASC activity have been limited by early end points. We hypothesized that with time,more » ASCs injected subcutaneously undergo removal by normal tissue turnover and homeostasis, and by the host's immune system. In this study, a high dose of culture expanded ASCs was formulated and implanted as multicellular aggregates into immunocompromised mice, which were maintained for over one year. Animals were monitored for toxicity, and surviving cells quantified at study endpoint. No difference in growth/weight or lifespan was found between cell-treated and vehicle treated animals, and no malignancies were detected in treated animals. Moreover, real-time PCR for a human specific sequence, ERV-3, detected no persistent ASCs. With the advent of clinical application, clarification of currently enigmatic stem cell properties has become imperative. Our study represents the longest duration determination of stem cell activity in vivo, and contributes strong evidence in support of the safety of adipose derived stem cell applications. -- Highlights: Black-Right-Pointing-Pointer Adipose stem cells promise novel clinical therapies. Black-Right-Pointing-Pointer Before clinical translation, safety profiles must be further elucidated. Black-Right-Pointing-Pointer Subcutaneously injected non-autologous adipose stem cells do not form tumors. Black-Right-Pointing-Pointer Subcutaneously injected non

  6. Creation and Transplantation of an Adipose-derived Stem Cell (ASC) Sheet in a Diabetic Wound-healing Model.

    PubMed

    Kato, Yuka; Iwata, Takanori; Washio, Kaoru; Yoshida, Toshiyuki; Kuroda, Hozue; Morikawa, Shunichi; Hamada, Mariko; Ikura, Kazuki; Kaibuchi, Nobuyuki; Yamato, Masayuki; Okano, Teruo; Uchigata, Yasuko

    2017-08-04

    Artificial skin has achieved considerable therapeutic results in clinical practice. However, artificial skin treatments for wounds in diabetic patients with impeded blood flow or with large wounds might be prolonged. Cell-based therapies have appeared as a new technique for the treatment of diabetic ulcers, and cell-sheet engineering has improved the efficacy of cell transplantation. A number of reports have suggested that adipose-derived stem cells (ASCs), a type of mesenchymal stromal cell (MSC), exhibit therapeutic potential due to their relative abundance in adipose tissue and their accessibility for collection when compared to MSCs from other tissues. Therefore, ASCs appear to be a good source of stem cells for therapeutic use. In this study, ASC sheets from the epididymal adipose fat of normal Lewis rats were successfully created using temperature-responsive culture dishes and normal culture medium containing ascorbic acid. The ASC sheets were transplanted into Zucker diabetic fatty (ZDF) rats, a rat model of type 2 diabetes and obesity, that exhibit diminished wound healing. A wound was created on the posterior cranial surface, ASC sheets were transplanted into the wound, and a bilayer artificial skin was used to cover the sheets. ZDF rats that received ASC sheets had better wound healing than ZDF rats without the transplantation of ASC sheets. This approach was limited because ASC sheets are sensitive to dry conditions, requiring the maintenance of a moist wound environment. Therefore, artificial skin was used to cover the ASC sheet to prevent drying. The allogenic transplantation of ASC sheets in combination with artificial skin might also be applicable to other intractable ulcers or burns, such as those observed with peripheral arterial disease and collagen disease, and might be administered to patients who are undernourished or are using steroids. Thus, this treatment might be the first step towards improving the therapeutic options for diabetic wound

  7. Expression of receptors for atrial natriuretic peptide on the murine bone marrow-derived stromal cells.

    PubMed

    Agui, T; Yamada, T; Legros, G; Nakajima, T; Clark, M; Peschel, C; Matsumoto, K

    1992-05-01

    Atrial natriuretic peptide (ANP) receptors were identified on both murine bone marrow-derived stromal cell lines A-3 and ALC and primary cultured cells using [125I]ANP binding assays and Northern blot analyses. The binding of [125I] ANP to the stromal cells was rapid, saturable, and of high affinity. The dissociation constants between ANP and its receptors on these cells showed no difference among cell types, while maximal binding capacity values were different among cell types. Competitive inhibition of [125I]ANP binding with C-atrial natriuretic factor, specific for ANP clearance receptor (ANPR-C), revealed that most of [125I]ANP-binding sites corresponded to ANPR-C. Northern blotting data corroborated that bone marrow-derived stromal cells expressed ANPR-C. However, in ALC cells, ANP biological receptors (either ANPR-A or ANPR-B), the mol wt of which is approximately 130K, were detected, and cGMP was accumulated after stimulation with ANP. On the other hand, in another stromal cell clone, A-3 cells, the expression of biological receptor was not detected in the affinity cross-linking and competitive inhibition experiments using [125I]ANP. However, A-3 cells accumulated cGMP by responding to ANPR-B-specific ligand, C-type natriuretic peptide. These results suggest that ALC cells equally express ANPR-A and ANPR-B, while A-3 cells express ANPR-B dominantly. Although the physiological roles of these receptors in the bone marrow is still not resolved, ANP is expected to play a role in the regulation of stromal cell functions in bone marrow.

  8. The osteogenic differentiation of SSEA-4 sub-population of human adipose derived stem cells using silicate nanoplatelets.

    PubMed

    Mihaila, Silvia M; Gaharwar, Akhilesh K; Reis, Rui L; Khademhosseini, Ali; Marques, Alexandra P; Gomes, Manuela E

    2014-11-01

    How to surpass in vitro stem cell differentiation, reducing cell manipulation, and lead the in situ regeneration process after transplantation, remains to be unraveled in bone tissue engineering (bTE). Recently, we showed that the combination of human bone marrow stromal cells with bioactive silicate nanoplatelets (sNPs) promotes the osteogenic differentiation without the use of standard osteogenic inductors. Even more, using SSEA-4(+) cell-subpopulations (SSEA-4(+)hASCs) residing within the adipose tissue, as a single-cellular source to obtain relevant cell types for bone regeneration, was also proposed. Herein, sNPs were used to promote the osteogenic differentiation of SSEA-4(+)hASCs. The interactions between SSEA-4(+)hASCs and sNPs, namely the internalization pathway and effect on cells osteogenic differentiation, were evaluated. SNPs below 100 μg/mL showed high cytocompatibility and fast internalization via clathrin-mediated pathway. SNPs triggered an overexpression of osteogenic-related markers (RUNX2, osteopontin, osteocalcin) accompanied by increased alkaline phosphatase activity and deposition of a predominantly collagen-type I matrix. Consequently, a robust matrix mineralization was achieved, covering >90% of the culturing surface area. Overall, we demonstrated the high osteogenic differentiation potential of SSEA-4(+)hASCs, further enhanced by the addition of sNPs in a dose dependent manner. This strategy endorses the combination of an adipose-derived cell-subpopulation with inorganic compounds to achieve bone matrix-analogs with clinical relevance. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. Arthroscopic Harvest of Adipose-Derived Mesenchymal Stem Cells From the Infrapatellar Fat Pad.

    PubMed

    Dragoo, Jason L; Chang, Wenteh

    2017-11-01

    The successful isolation of adipose-derived mesenchymal stem cells (ADSCs) from the arthroscopically harvested infrapatellar fat pad (IFP) would provide orthopaedic surgeons with an autologous solution for regenerative procedures. To demonstrate the quantity and viability of the mesenchymal stem cell population arthroscopically harvested from the IFP as well as the surrounding synovium. Descriptive laboratory study. The posterior border of the IFP, including the surrounding synovial tissue, was harvested arthroscopically from patients undergoing anterior cruciate ligament reconstruction. Tissue was then collected in an AquaVage adipose canister, followed by fat fractionization using syringe emulsification and concentration with an AdiPrep device. In the laboratory, the layers of tissue were separated and then digested with 0.3% type I collagenase. The pelleted stromal vascular fraction (SVF) cells were then immediately analyzed for viability, mesenchymal cell surface markers by fluorescence-activated cell sorting, and clonogenic capacity. After culture expansion, the metabolic activity of the ADSCs was assessed by an AlamarBlue assay, and the multilineage differentiation capability was tested. The transition of surface antigens from the SVF toward expanded ADSCs at passage 2 was further evaluated. SVF cells were successfully harvested with a mean yield of 4.86 ± 2.64 × 10 5 cells/g of tissue and a mean viability of 69.03% ± 10.75%, with ages ranging from 17 to 52 years (mean, 35.14 ± 13.70 years; n = 7). The cultured ADSCs composed a mean 5.85% ± 5.89% of SVF cells with a mean yield of 0.33 ± 0.42 × 10 5 cells/g of tissue. The nonhematopoietic cells (CD45 - ) displayed the following surface antigens as a percentage of the viable population: CD44 + (52.21% ± 4.50%), CD73 + CD90 + CD105 + (19.20% ± 17.04%), and CD44 + CD73 + CD90 + CD105 + (15.32% ± 15.23%). There was also a significant increase in the expression of ADSC markers CD73 (96.97% ± 1.72%; P

  10. Proteomic Profiling of Tissue-Engineered Blood Vessel Walls Constructed by Adipose-Derived Stem Cells

    PubMed Central

    Wang, Chen; Guo, Fangfang; Zhou, Heng; Zhang, Yun; Xiao, Zhigang

    2013-01-01

    Adipose-derived stem cells (ASCs) can differentiate into smooth muscle cells and have been engineered into elastic small diameter blood vessel walls in vitro. However, the mechanisms involved in the development of three-dimensional (3D) vascular tissue remain poorly understood. The present study analyzed protein expression profiles of engineered blood vessel walls constructed by human ASCs using methods of two-dimensional gel electrophoresis (2DE) and mass spectrometry (MS). These results were compared to normal arterial walls. A total of 1701±15 and 1265±26 protein spots from normal and engineered blood vessel wall extractions were detected by 2DE, respectively. A total of 20 spots with at least 2.0-fold changes in expression were identified, and 38 differently expressed proteins were identified by 2D electrophoresis and ion trap MS. These proteins were classified into seven functional categories: cellular organization, energy, signaling pathway, enzyme, anchored protein, cell apoptosis/defense, and others. These results demonstrated that 2DE, followed by ion trap MS, could be successfully utilized to characterize the proteome of vascular tissue, including tissue-engineered vessels. The method could also be employed to achieve a better understanding of differentiated smooth muscle protein expression in vitro. These results provide a basis for comparative studies of protein expression in vascular smooth muscles of different origin and could provide a better understanding of the mechanisms of action needed for constructing blood vessels that exhibit properties consistent with normal blood vessels. PMID:22963350

  11. Proteomic profiling of tissue-engineered blood vessel walls constructed by adipose-derived stem cells.

    PubMed

    Wang, Chen; Guo, Fangfang; Zhou, Heng; Zhang, Yun; Xiao, Zhigang; Cui, Lei

    2013-02-01

    Adipose-derived stem cells (ASCs) can differentiate into smooth muscle cells and have been engineered into elastic small diameter blood vessel walls in vitro. However, the mechanisms involved in the development of three-dimensional (3D) vascular tissue remain poorly understood. The present study analyzed protein expression profiles of engineered blood vessel walls constructed by human ASCs using methods of two-dimensional gel electrophoresis (2DE) and mass spectrometry (MS). These results were compared to normal arterial walls. A total of 1701±15 and 1265±26 protein spots from normal and engineered blood vessel wall extractions were detected by 2DE, respectively. A total of 20 spots with at least 2.0-fold changes in expression were identified, and 38 differently expressed proteins were identified by 2D electrophoresis and ion trap MS. These proteins were classified into seven functional categories: cellular organization, energy, signaling pathway, enzyme, anchored protein, cell apoptosis/defense, and others. These results demonstrated that 2DE, followed by ion trap MS, could be successfully utilized to characterize the proteome of vascular tissue, including tissue-engineered vessels. The method could also be employed to achieve a better understanding of differentiated smooth muscle protein expression in vitro. These results provide a basis for comparative studies of protein expression in vascular smooth muscles of different origin and could provide a better understanding of the mechanisms of action needed for constructing blood vessels that exhibit properties consistent with normal blood vessels.

  12. Adipose-derived stem cells for cartilage regeneration - moving towards clinical applicability

    PubMed Central

    2013-01-01

    Despite multiple methods of treatment and a wealth of research in the field of regenerative medicine focusing on cartilage defects, the management of cartilage injuries remains a challenge. A recent study by Van Pham and colleagues proposes a method for preconditioning autologous adipose-derived stem cells. Their study offers evidence about the increased proliferative and chondrogenetic capabilities of platelet-rich plasma-treated adipose-derived stem cells and the increased efficiency of these in treating articular cartilage defects in mice. Even though the method needs further elaboration and the composition of the repair tissue requires investigation, the results are promising for the design of clinically acceptable cell therapies aimed at cartilage regeneration. PMID:24079605

  13. Secretion of SDF-1α by bone marrow-derived stromal cells enhances skin wound healing of C57BL/6 mice exposed to ionizing radiation

    PubMed Central

    Landry, Yannick; Lê, Oanh; Mace, Kimberly A; Restivo, Terry E; Beauséjour, Christian M

    2010-01-01

    Abstract Patients treated for cancer therapy using ionizing radiation (IR) have delayed tissue repair and regeneration. The mechanisms mediating these defects remain largely unknown at present, thus limiting the development of therapeutic approaches. Using a wound healing model, we here investigate the mechanisms by which IR exposure limits skin regeneration. Our data show that induction of the stromal cell-derived growth factor 1α (SDF-1α) is severely impaired in the wounded skin of irradiated, compared to non-irradiated, mice. Hence, we evaluated the potential of bone marrow-derived multipotent stromal cells (MSCs), which secrete high levels of SDF-1α, to improve skin regeneration in irradiated mice. Injection of MSCs into the wound margin led to remarkable enhancement of skin healing in mice exposed to IR. Injection of irradiated MSCs into the wound periphery of non-irradiated mice delayed wound closure, also suggesting an important role for the stromal microenvironment in skin repair. The beneficial actions of MSCs were mainly paracrine, as the cells did not differentiate into keratinocytes. Specific knockdown of SDF-1α expression led to drastically reduced efficiency of MSCs in improving wound closure, indicating that SDF-1α secretion by MSCs is largely responsible for their beneficial action. We also found that one mechanism by which SDF-1α enhances wound closure likely involves increased skin vascularization. Our findings collectively indicate that SDF-1α is an important deregulated cytokine in irradiated wounded skin, and that the decline in tissue regeneration potential following IR can be reversed, given adequate microenvironmental support PMID:19725920

  14. Identification and validation of multiple cell surface markers of clinical-grade adipose-derived mesenchymal stromal cells as novel release criteria for good manufacturing practice-compliant production.

    PubMed

    Camilleri, Emily T; Gustafson, Michael P; Dudakovic, Amel; Riester, Scott M; Garces, Catalina Galeano; Paradise, Christopher R; Takai, Hideki; Karperien, Marcel; Cool, Simon; Sampen, Hee-Jeong Im; Larson, A Noelle; Qu, Wenchun; Smith, Jay; Dietz, Allan B; van Wijnen, Andre J

    2016-08-11

    Clinical translation of mesenchymal stromal cells (MSCs) necessitates basic characterization of the cell product since variability in biological source and processing of MSCs may impact therapeutic outcomes. Although expression of classical cell surface markers (e.g., CD90, CD73, CD105, and CD44) is used to define MSCs, identification of functionally relevant cell surface markers would provide more robust release criteria and options for quality control. In addition, cell surface expression may distinguish between MSCs from different sources, including bone marrow-derived MSCs and clinical-grade adipose-derived MSCs (AMSCs) grown in human platelet lysate (hPL). In this work we utilized quantitative PCR, flow cytometry, and RNA-sequencing to characterize AMSCs grown in hPL and validated non-classical markers in 15 clinical-grade donors. We characterized the surface marker transcriptome of AMSCs, validated the expression of classical markers, and identified nine non-classical markers (i.e., CD36, CD163, CD271, CD200, CD273, CD274, CD146, CD248, and CD140B) that may potentially discriminate AMSCs from other cell types. More importantly, these markers exhibit variability in cell surface expression among different cell isolates from a diverse cohort of donors, including freshly prepared, previously frozen, or proliferative state AMSCs and may be informative when manufacturing cells. Our study establishes that clinical-grade AMSCs expanded in hPL represent a homogeneous cell culture population according to classical markers,. Additionally, we validated new biomarkers for further AMSC characterization that may provide novel information guiding the development of new release criteria. Use of Autologous Bone Marrow Aspirate Concentrate in Painful Knee Osteoarthritis (BMAC): Clinicaltrials.gov NCT01931007 . Registered August 26, 2013. MSC for Occlusive Disease of the Kidney: Clinicaltrials.gov NCT01840540 . Registered April 23, 2013. Mesenchymal Stem Cell Therapy in Multiple

  15. Oxidative and inflammatory signals in obesity-associated vascular abnormalities.

    PubMed

    Reho, John J; Rahmouni, Kamal

    2017-07-15

    Obesity is associated with increased cardiovascular morbidity and mortality in part due to vascular abnormalities such as endothelial dysfunction and arterial stiffening. The hypertension and other health complications that arise from these vascular defects increase the risk of heart diseases and stroke. Prooxidant and proinflammatory signaling pathways as well as adipocyte-derived factors have emerged as critical mediators of obesity-associated vascular abnormalities. Designing treatments aimed specifically at improving the vascular dysfunction caused by obesity may provide an effective therapeutic approach to prevent the cardiovascular sequelae associated with excessive adiposity. In this review, we discuss the recent evidence supporting the role of oxidative stress and cytokines and inflammatory signals within the vasculature as well as the impact of the surrounding perivascular adipose tissue (PVAT) on the regulation of vascular function and arterial stiffening in obesity. In particular, we focus on the highly plastic nature of the vasculature in response to altered oxidant and inflammatory signaling and highlight how weight management can be an effective therapeutic approach to reduce the oxidative stress and inflammatory signaling and improve vascular function. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

  16. Tissue Engineering Strategies for Promoting Vascularized Bone Regeneration

    PubMed Central

    Almubarak, Sarah; Nethercott, Hubert; Freeberg, Marie; Beaudon, Caroline; Jha, Amit; Jackson, Wesley; Marcucio, Ralph; Miclau, Theodore; Healy, Kevin; Bahney, Chelsea

    2016-01-01

    This review focuses on current tissue engineering strategies for promoting vascularized bone regeneration. We review the role of angiogenic growth factors in promoting vascularized bone regeneration and discuss the different therapeutic strategies for controlled/sustained growth factor delivery. Next, we address the therapeutic uses of stem cells in vascularized bone regeneration. Specifically, this review addresses the concept of co-culture using osteogenic and vasculogenic stem cells, and how adipose derived stem cells compare to bone marrow derived mesenchymal stem cells in the promotion of angiogenesis. We conclude this review with a discussion of a novel approach to bone regeneration through a cartilage intermediate, and discuss why it has the potential to be more effective than traditional bone grafting methods. PMID:26608518

  17. One in vitro model for visceral adipose-derived fibroblasts in chronic inflammation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yue Guiping; Du Lirui; Xia Tao

    2005-08-05

    One pathogenesis of the obesity-associated complications is that consistent with increased body fat mass, the elevation of adipose tissue-derived cytokines inflicts a low-grade chronic inflammation, which ultimately leads to metabolic disorders. Adipocytes and macrophages in visceral adipose (VA) have been confirmed to contribute to the chronic inflammation; however, the role of the resident fibroblasts is still unknown. We established one VA fibroblast cell line, termed VAFC. Morphological analysis indicated that there were large numbers of pits at the cell plasma membrane. In vitro VAFC cells promoted bone marrow cells to differentiate into macrophages and protected them from apoptosis in themore » serum-free conditions. Additionally, they also interfered in lymphocytes proliferation. On the basis of these results, this cell line might be an in vitro model for understanding the role of adipose-derived fibroblasts in obesity-associated chronic inflammation.« less

  18. Breast Cancer and Estrogen Biosynthesis in Adipose Tissue

    DTIC Science & Technology

    1998-10-01

    transferred to a nitrocellulose mem - brane. The transferred proteins were subjected to a denaturation/rena- turation process and hybridized to the 32P...aromatase expression in adipose tissue has been recently observed to be regulated by mem - bers of the interleukin-6 (IL-6) cytokine family. Based on...shown in human adipose stromal cells that the stimulatory effects of serum on aromatase expression can be mimicked by mem - bers of the interleukin-6

  19. Cell Based Therapeutic Approach in Vascular Surgery: Application and Review

    PubMed Central

    Rocca, Aldo; Tafuri, Domenico; Paccone, Marianna; Giuliani, Antonio; Zamboli, Anna Ginevra Immacolata; Surfaro, Giuseppe; Paccone, Andrea; Compagna, Rita; Amato, Maurizo; Serra, Raffaele; Amato, Bruno

    2017-01-01

    Abstract Multipotent stem cells - such as mesenchymal stem/stromal cells and stem cells derived from different sources like vascular wall are intensely studied to try to rapidly translate their discovered features from bench to bedside. Vascular wall resident stem cells recruitment, differentiation, survival, proliferation, growth factor production, and signaling pathways transduced were analyzed. We studied biological properties of vascular resident stem cells and explored the relationship from several factors as Matrix Metalloproteinases (MMPs) and regulations of biological, translational and clinical features of these cells. In this review we described a translational and clinical approach to Adult Vascular Wall Resident Multipotent Vascular Stem Cells (VW-SCs) and reported their involvement in alternative clinical approach as cells based therapy in vascular disease like arterial aneurysms or peripheral arterial obstructive disease. PMID:29071303

  20. Chemically Defined and Xeno-Free Cryopreservation of Human Adipose-Derived Stem Cells

    PubMed Central

    López, Melany; Bollag, Roni J.; Yu, Jack C.; Isales, Carlos M.; Eroglu, Ali

    2016-01-01

    The stromal compartment of adipose tissue harbors multipotent cells known as adipose-derived stem cells (ASCs). These cells can differentiate into various lineages including osteogenic, chrondrogenic, adipogenic, and neurogenic; this cellular fraction may be easily obtained in large quantities through a clinically safe liposuction procedure. Therefore, ASCs offer exceptional opportunities for tissue engineering and regenerative medicine. However, current practices involving ASCs typically use fetal bovine serum (FBS)-based cryopreservation solutions that are associated with risks of immunological reactions and of transmitting infectious diseases and prions. To realize clinical applications of ASCs, serum- and xeno-free defined cryopreservation methods are needed. To this end, an animal product-free chemically defined cryopreservation medium was formulated by adding two antioxidants (reduced glutathione and ascorbic acid 2-phosphate), two polymers (PVA and ficoll), two permeating cryoprotectants (ethylene glycol and dimethylsulfoxide), a disaccharide (trehalose), and a calcium chelator (EGTA) to HEPES-buffered DMEM/F12. To limit the number of experimental groups, the concentration of trehalose, both polymers, and EGTA was fixed while the presence of the permeating CPAs and antioxidants was varied. ASCs suspended either in different versions of the defined medium or in the conventional undefined cryopreservation medium (10% dimethylsulfoxide+10% DMEM/F12+80% serum) were cooled to -70°C at 1°C/min before being plunged into liquid nitrogen. Samples were thawed either in air or in a water bath at 37°C. The presence of antioxidants along with 3.5% concentration of each penetrating cryoprotectant improved the freezing outcome to the level of the undefined cryopreservation medium, but the plating efficiency was still lower than that of unfrozen controls. Subsequently, increasing the concentration of both permeating cryoprotectants to 5% further improved the plating

  1. Human cord blood-derived platelet lysate enhances the therapeutic activity of adipose-derived mesenchymal stromal cells isolated from Crohn's disease patients in a mouse model of colitis.

    PubMed

    Forte, Dorian; Ciciarello, Marilena; Valerii, Maria Chiara; De Fazio, Luigia; Cavazza, Elena; Giordano, Rosaria; Parazzi, Valentina; Lazzari, Lorenza; Laureti, Silvio; Rizzello, Fernando; Cavo, Michele; Curti, Antonio; Lemoli, Roberto M; Spisni, Enzo; Catani, Lucia

    2015-09-09

    Due to their immunomodulatory properties, mesenchymal stromal cells (MSCs) have been used for auto-immune disease treatment. Crohn disease (CD) and ulcerative colitis are two major inflammatory bowel diseases (IBDs), resulting from pathological immune responses to environmental or microbial antigens. Preclinical and clinical studies have suggested that MSC-based cellular therapy hold promising potential for IBD treatment. However, open issues include the selection of the proper cell dose, the source and the optimal route of administration of MSCs for more effective results. Platelet lysate has gained clinical interest due to its efficacy in accelerating wound healing. Thus, we propose to combine the administration of MSCs with a human umbilical cord blood-derived platelet lysate (hCBPL) as a novel strategy to improve MSC-based therapy for IBD resolution. Colitis was induced in 8-week-old C57BL/6J mice by daily oral administration of dextran sulphate sodium (DSS) (1.5 % w/v in tap water) for 9 days. MSCs were isolated from adipose tissue of CD patients (adCD-MSCs), expanded in proliferation medium, resuspended in hCBPL or PBS and administrated via enema for three times (1 × 10(6) cells/mouse/time) every other day starting on day +7 from DSS induction. The colitis evolution was evaluated by daily monitoring of body weight, stool consistency and bleeding. Histopathological analysis was performed. Inflammatory cytokine plasma levels were determined. adCD-MSCs stained with lipophilic membrane dye Nile Red, were injected in DSS mice as described above. Colon section of mice sacrificed 24 hours after last cell administration, were analyzed by confocal microscopy. We found that adCD-MSCs could be easily isolated and expanded from CD patients. Upon injection, adCD-MSCs exerted a therapeutic effect on DSS-induced colitis. Moreover, hCBPL increased adCD-MSCs efficacy by significantly reducing colitis scores, extension of the colon inflamed area and plasma levels of

  2. Use of Adipose-Derived Mesenchymal Stem Cells in Keratoconjunctivitis Sicca in a Canine Model

    PubMed Central

    Villatoro, Antonio J.; Fernández, Viviana; Rico-Llanos, Gustavo A.; Becerra, José; Andrades, José A.

    2015-01-01

    Keratoconjunctivitis sicca (KCS) or dry eye disease (DED) is an immune-mediated multifactorial disease, with high level of prevalence in humans and dogs. Our aim in this study was to investigate the therapeutic effects of allogeneic adipose-derived mesenchymal stromal cells (Ad-MSCs) implanted around the lacrimal glands in 12 dogs (24 eyes) with KCS, which is refractory to current available treatments. Schirmer tear test (STT) and ocular surface integrity were assessed at 0 (before treatment), 3, 6, and 9 months after treatment. Average STT values and all clinical signs showed a statistically significant change (P < 0.001) during the follow-up with reduction in all ocular parameters scored: ocular discharge, conjunctival hyperaemia, and corneal changes, and there were no signs of regression or worsening. Implanted cells were well tolerated and were effective reducing clinical signs of KCS with a sustained effect during the study period. None of the animals showed systemic or local complications during the study. To our knowledge, this is the first time in literature that implantation of allogeneic Ad-MSCs around lacrimal glands has been found as an effective therapeutic alternative to treat dogs with KCS. These results could reinforce a good effective solution to be extrapolated to future studies in human. PMID:25802852

  3. Inflammation and Fibrosis in Perirenal Adipose Tissue of Patients With Aldosterone-Producing Adenoma.

    PubMed

    Wu, Chunyan; Zhang, Huijian; Zhang, Jiajun; Xie, Cuihua; Fan, Cunxia; Zhang, Hongbin; Wu, Peng; Wei, Qiang; Tan, Wanlong; Xu, Lingling; Wang, Ling; Xue, Yaoming; Guan, Meiping

    2018-01-01

    The prevalence of primary aldosteronism is much higher than previously thought. Recent studies have shown that primary aldosteronism is related to a higher risk of cardiovascular events. However, the underlying mechanism is not yet clear. Here we investigate the characteristics, including inflammation, fibrosis, and adipokine expression, of adipose tissues from different deposits in patients with aldosterone-producing adenoma (APA). Inflammation and fibrosis changes were evaluated in perirenal and subcutaneous adipose tissues obtained from patients with APA (n = 16), normotension (NT; n = 10), and essential hypertension (EH; n = 5) undergoing laparoscopic surgery. We also evaluated the effect of aldosterone in isolated human perirenal adipose tissue stromal vascular fraction (SVF) cells and investigated the effect of aldosterone in mouse 3T3-L1 and brown preadipocytes. Compared with the EH group, significantly higher levels of interleukin-6 (IL-6) and tumor necrosis factor-α messenger RNA (mRNA) and protein were observed in perirenal adipose tissue of patients with APA. Expression of genes related to fibrosis and adipogenesis in perirenal adipose tissue was notably higher in patients with APA than in patients with NT and EH. Aldosterone significantly induced IL-6 and fibrosis gene mRNA expression in differentiated SVF cells. Aldosterone treatment enhanced mRNA expression of genes associated with inflammation and fibrosis and stimulated differentiation of 3T3-L1 and brown preadipocytes. In conclusion, these data indicate that high aldosterone in patients with APA may induce perirenal adipose tissue dysfunction and lead to inflammation and fibrosis, which may be involved in the high risk of cardiovascular events observed in patients with primary aldosteronism. Copyright © 2018 Endocrine Society.

  4. Inhibition of M1 macrophage activation in adipose tissue by berberine improves insulin resistance.

    PubMed

    Ye, Lifang; Liang, Shu; Guo, Chao; Yu, Xizhong; Zhao, Juan; Zhang, Hao; Shang, Wenbin

    2016-12-01

    Insulin resistance is associated with a chronic inflammation in adipose tissue which is propagated by a phenotypic switch in adipose tissue macrophage (ATM) polarization. This study aimed to investigate whether berberine, the major alkaloid of rhizoma coptidis, can improve insulin resistance through inhibiting ATM activation and inflammatory response in adipose tissue. High-fat-diet induced obese mice were administered oral with berberine (50mg/kg/day) for 14days. ATMs were analysed using FACS and insulin resistance was evaluated. Expressions of pro-inflammatory cytokines and activation of inflammatory pathways were detected. The chemotaxis of macrophages was measured. Glucose consumption and insulin signalling of adipocytes were examined. Berberine significantly decreased F4/80 + /CD11c + /CD206 - cells in the stromal vascular fraction from adipose tissue and improved glucose tolerance in obsess mice. In addition, berberine reduced the elevated levels of serum TNF-α, IL-6 and MCP-1 and the expressions of TNF-α, IL-6 and MCP-1 and attenuated the phosphorylation of JNK and IKKβ and the expression of NF-κB p65 in the obese adipose tissue, Raw264.7 macrophages and 3T3-L1 adipocytes, respectively. The phosphorylation of IRS-1 (Ser307) was inhibited by berberine in adipose tissue and cultured adipocytes. The phosphorylation of AKT (Ser473) was increased in berberine-treated adipose tissue. Conditioned medium from adipocytes treated with berberine reduced the number of infiltrated macrophages. Berberine partly restored the impaired glucose consumption and the activation of IRS-1 (Ser307) in adipocytes induced by the activation of macrophages. Our findings imply that berberine improves insulin resistance by inhibiting M1 macrophage activation in adipose tissue. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Correlation of MRI-derived adipose tissue measurements and anthropometric markers with prevalent hypertension in the community.

    PubMed

    Lorbeer, Roberto; Rospleszcz, Susanne; Schlett, Christopher L; Heber, Sophia D; Machann, Jürgen; Thorand, Barbara; Meisinger, Christa; Heier, Margit; Peters, Annette; Bamberg, Fabian; Lieb, Wolfgang

    2018-07-01

    To compare the correlations of MRI-derived adipose tissue measurements and anthropometric markers, respectively, with prevalent hypertension in a community-based sample, free of clinical cardiovascular disease. MRI-derived adipose tissue measurements were obtained in 345 participants (143 women; age 39-73 years) of the KORA FF4 survey from Southern Germany using a 3-Tesla machine and included total adipose tissue (TAT), visceral adipose tissue (VAT), subcutaneous adipose tissue (SCAT), hepatic fat fraction (HFF), pancreatic fat fraction (PFF) as well as pericardial adipose tissue (PAT). In addition, the anthropometric markers body mass index, waist circumference, hip circumference, waist-hip ratio (WHR) and waist-height ratio (WHtR) as well as blood pressure measurements were obtained. The prevalence of hypertension was 33.6% (women: 28%, men: 38%). VAT and PAT had the highest area under the curve (AUC) values for identifying individuals with prevalent hypertension (AUC: 0.75; 0.73, respectively), whereas WHtR and waist circumference were best performing anthropometric markers (AUC: 0.72; 0.70, respectively). A 1SD increment of TAT was associated with the highest odd for hypertension in the age-adjusted and sex-adjusted model (OR = 2.20, 95% CI 1.67-2.91, P < 0.001) and in the fully adjusted model (OR = 1.97, 95% CI 1.45-2.66, P < 0.001). TAT was the only MRI-derived adipose tissue measurement that was associated with hypertension independently of the best performing anthropometric marker waist circumference in the fully adjusted model (OR = 1.93, 95% CI 1.00-3.72, P = 0.049). MRI-derived adipose tissue measurements perform similarly in identifying prevalent hypertension compared with anthropometric markers. Especially, TAT, VAT and PAT as well as WHtR and waist circumference were highly correlated with prevalent hypertension.

  6. Scaffold Composition Determines the Angiogenic Outcome of Cell-Based Vascular Endothelial Growth Factor Expression by Modulating Its Microenvironmental Distribution.

    PubMed

    Gaudiello, Emanuele; Melly, Ludovic; Cerino, Giulia; Boccardo, Stefano; Jalili-Firoozinezhad, Sasan; Xu, Lifen; Eckstein, Friedrich; Martin, Ivan; Kaufmann, Beat A; Banfi, Andrea; Marsano, Anna

    2017-12-01

    Delivery of genetically modified cells overexpressing Vascular Endothelial Growth Factor (VEGF) is a promising approach to induce therapeutic angiogenesis in ischemic tissues. The effect of the protein is strictly modulated by its interaction with the components of the extracellular matrix. Its therapeutic potential depends on a sustained but controlled release at the microenvironmental level in order to avoid the formation of abnormal blood vessels. In this study, it is hypothesized that the composition of the scaffold plays a key role in modulating the binding, hence the therapeutic effect, of the VEGF released by 3D-cell constructs. It is found that collagen sponges, which poorly bind VEGF, prevent the formation of localized hot spots of excessive concentration, therefore, precluding the development of aberrant angiogenesis despite uncontrolled expression by a genetically engineered population of adipose tissue-derived stromal cells. On the contrary, after seeding on VEGF-binding egg-white scaffolds, the same cell population caused aberrantly enlarged vascular structures after 14 d. Collagen-based engineered tissues also induced a safe and efficient angiogenesis in both the patch itself and the underlying myocardium in rat models. These findings open new perspectives on the control and the delivery of proangiogenic stimuli, and are fundamental for the vascularization of engineered tissues/organs. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  7. Noncultured Autologous Adipose-Derived Stem Cells Therapy for Chronic Radiation Injury

    PubMed Central

    Akita, Sadanori; Akino, Kozo; Hirano, Akiyoshi; Ohtsuru, Akira; Yamashita, Shunichi

    2010-01-01

    Increasing concern on chronic radiation injuries should be treated properly for life-saving improvement of wound management and quality of life. Recently, regenerative surgical modalities should be attempted with the use of noncultured autologous adipose-derived stem cells (ADSCs) with temporal artificial dermis impregnated and sprayed with local angiogenic factor such as basic fibroblast growth factor, and secondary reconstruction can be a candidate for demarcation and saving the donor morbidity. Autologous adipose-derived stem cells, together with angiogenic and mitogenic factor of basic fibroblast growth factor and an artificial dermis, were applied over the excised irradiated skin defect and tested for Patients who were uneventfully healed with minimal donor-site morbidity, which lasts more than 1.5 years. PMID:21151652

  8. Combined platelet and plasma derivatives enhance proliferation of stem/progenitor cells maintaining their differentiation potential.

    PubMed

    Muraglia, Anita; Todeschi, Maria Rosa; Papait, Andrea; Poggi, Alessandro; Spanò, Raffaele; Strada, Paolo; Cancedda, Ranieri; Mastrogiacomo, Maddalena

    2015-12-01

    Platelet derivatives have been proposed as alternatives to animal sera given that for cell therapy applications, the use of fetal bovine/calf serum (FBS/FCS) is subjected to severe limitations for safety and ethical concerns. We developed a cell culture medium additive obtained by the combination of two blood-derived standardized components. A platelet lysate (PL) and a platelet-poor plasma (PPP) were produced in a lyophilized form. Each component was characterized for its growth factor content (platelet-derived growth factor-BB/vascular endothelial growth factor). PL and PPP were used as single components or in combination in different ratio at cumulative 5% final concentration in the culture medium. The single components were less effective than the component combination. In primary cell cultures (bone marrow stromal cells, adipose derived adult stem cells, osteoblasts, chondrocytes, umbilical cord-derived mesenchymal stromal cells, lymphocytes), the PL/PPP supplement promoted an increased cell proliferation in respect to the standard FCS culture in a dose-dependent manner, maintaining the cell functionality, clonogenicity, phenotype and differentiative properties throughout the culture. At a different component ratio, the supplement was also used to support proliferation of a cell line (U-937). The PL/PPP supplement is an efficient cell culture medium additive that can replace FCS to promote cell proliferation. It can outdo FCS, especially when adopted in primary cultures from tissue biopsies. Moreover, the dual component nature of the supplement allows the researcher to determine the more appropriate ratio of the two components for the nutritional and functional requirements of the cell type of interest. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  9. Extra-pancreatic invasion induces lipolytic and fibrotic changes in the adipose microenvironment, with released fatty acids enhancing the invasiveness of pancreatic cancer cells

    PubMed Central

    Okumura, Takashi; Ohuchida, Kenoki; Sada, Masafumi; Abe, Toshiya; Endo, Sho; Koikawa, Kazuhiro; Iwamoto, Chika; Miura, Daisuke; Mizuuchi, Yusuke; Moriyama, Taiki; Nakata, Kohei; Miyasaka, Yoshihiro; Manabe, Tatsuya; Ohtsuka, Takao; Nagai, Eishi; Mizumoto, Kazuhiro; Oda, Yoshinao; Hashizume, Makoto; Nakamura, Masafumi

    2017-01-01

    Pancreatic cancer progression involves components of the tumor microenvironment, including stellate cells, immune cells, endothelial cells, and the extracellular matrix. Although peripancreatic fat is the main stromal component involved in extra-pancreatic invasion, its roles in local invasion and metastasis of pancreatic cancer remain unclear. This study investigated the role of adipose tissue in pancreatic cancer progression using genetically engineered mice (Pdx1-Cre; LSL-KrasG12D; Trp53R172H/+) and an in vitro model of organotypic fat invasion. Mice fed a high fat diet had significantly larger primary pancreatic tumors and a significantly higher rate of distant organ metastasis than mice fed a standard diet. In the organotypic fat invasion model, pancreatic cancer cell clusters were smaller and more elongated in shape and showed increased fibrosis. Adipose tissue-derived conditioned medium enhanced pancreatic cancer cell invasiveness and gemcitabine resistance, as well as inducing morphologic changes in cancer cells and increasing the numbers of lipid droplets in their cytoplasm. The concentrations of oleic, palmitoleic, and linoleic acids were higher in adipose tissue-derived conditioned medium than in normal medium, with these fatty acids significantly enhancing the migration of cancer cells. Mature adipocytes were smaller and the concentration of fatty acids in the medium higher when these cells were co-cultured with cancer cells. These findings indicate that lipolytic and fibrotic changes in peripancreatic adipose tissue enhance local invasiveness and metastasis via adipocyte-released fatty acids. Inhibition of fatty acid uptake by cancer cells may be a novel therapy targeting interactions between cancer and stromal cells. PMID:28407685

  10. The Cladophora glomerata Enriched by Biosorption Process in Cr(III) Improves Viability, and Reduces Oxidative Stress and Apoptosis in Equine Metabolic Syndrome Derived Adipose Mesenchymal Stromal Stem Cells (ASCs) and Their Extracellular Vesicles (MV's).

    PubMed

    Marycz, Krzysztof; Michalak, Izabela; Kocherova, Ievgeniia; Marędziak, Monika; Weiss, Christine

    2017-12-08

    This study investigated in vitro effects of freshwater alga Cladophora glomerata water extract enriched during a biosorption process in Cr(III) trivalent chromium and chromium picolinate on adipose-derived mesenchymal stromal stem cells (ASCs) and extracellular microvesicles (MVs) in equine metabolic syndrome-affected horses. Chemical characterisation of natural Cladophora glomerata was performed with special emphasis on: vitamin C, vitamin E, total phenols, fatty acids, free and protein-bound amino acids as well as measured Cr in algal biomass. To examine the influence of Cladophora glomerata water extracts, in vitro viability, oxidative stress factor accumulation, apoptosis, inflammatory response, biogenesis of mitochondria, autophagy in ASCs of EMS and secretory activity manifested by MV release were investigated. For this purpose, various methods of molecular biology and microscopic observations (i.e., immunofluorescence staining, SEM, TEM, FIB observations, mRNA and microRNA expression by RT-qPCR) were applied. The extract of Cladophora glomerata enriched with Cr(III) ions reduced apoptosis and inflammation in ASCs of EMS horses through improvement of mitochondrial dynamics, decreasing of PDK4 expression and reduction of endoplastic reticulum stress. Moreover, it was found, that Cladophora glomerata and Cr(III) induce antioxidative protection coming from enhanced SOD activity Therefore, Cladophora glomerata enriched with Cr(III) ions might become an interesting future therapeutic agent in the pharmacological treatment of EMS horses.

  11. Successful isolation of viable adipose-derived stem cells from human adipose tissue subject to long-term cryopreservation: positive implications for adult stem cell-based therapeutics in patients of advanced age.

    PubMed

    Devitt, Sean M; Carter, Cynthia M; Dierov, Raia; Weiss, Scott; Gersch, Robert P; Percec, Ivona

    2015-01-01

    We examined cell isolation, viability, and growth in adipose-derived stem cells harvested from whole adipose tissue subject to different cryopreservation lengths (2-1159 days) from patients of varying ages (26-62 years). Subcutaneous abdominal adipose tissue was excised during abdominoplasties and was cryopreserved. The viability and number of adipose-derived stem cells isolated were measured after initial isolation and after 9, 18, and 28 days of growth. Data were analyzed with respect to cryopreservation duration and patient age. Significantly more viable cells were initially isolated from tissue cryopreserved <1 year than from tissue cryopreserved >2 years, irrespective of patient age. However, this difference did not persist with continued growth and there were no significant differences in cell viability or growth at subsequent time points with respect to cryopreservation duration or patient age. Mesenchymal stem cell markers were maintained in all cohorts tested throughout the duration of the study. Consequently, longer cryopreservation negatively impacts initial live adipose-derived stem cell isolation; however, this effect is neutralized with continued cell growth. Patient age does not significantly impact stem cell isolation, viability, or growth. Cryopreservation of adipose tissue is an effective long-term banking method for isolation of adipose-derived stem cells in patients of varying ages.

  12. Human adipose-derived stem cell spheroid treated with photobiomodulation irradiation accelerates tissue regeneration in mouse model of skin flap ischemia.

    PubMed

    Park, In-Su; Chung, Phil-Sang; Ahn, Jin Chul; Leproux, Anais

    2017-11-01

    Skin flap grafting is a form of transplantation widely used in plastic surgery. However, ischemia/reperfusion injury is the main factor which reduces the survival rate of flaps following grafting. We investigated whether photobiomodulation (PBM) precondition prior to human adipose-derived stromal cell (hASC) spheroid (PBM-spheroid) transplantation improved skin tissue functional recovery by the stimulation of angiogenesis and tissue regeneration in skin flap of mice. The LED had an emission wavelength peaked at 660 ± 20 nm (6 J/cm 2 , 10 mW/cm 2 ). The expression of angiogenic growth factors in PBM-spheroid hASCs was much greater than that of not-PBM-treated spheroid or monolayer-cultured hASCs. From immunochemical staining analysis, the hASCs of PBM-spheroid were CD31 + , KDR + , and CD34 + , whereas monolayer-cultured hASCs were negative for these markers. To evaluate the therapeutic effect of hASC PBM-spheroid in vivo, PBS, monolayer-cultured hASCs, and not-PBM-spheroid were transplanted into a skin flap model. The animals were observed for 14 days. The PBM-spheroid hASCs transplanted into the skin flap ischemia differentiated into endothelial cells and remained differentiated. Transplantation of PBM-spheroid hASCs into the skin flap ischemia significantly elevated the density of vascular formations through angiogenic factors released by the skin flap ischemia and enhanced tissue regeneration at the lesion site. Consistent with these results, the transplantation of PBM-spheroid hASCs significantly improved functional recovery compared with PBS, monolayer-cultured hASCs, and not-PBM-spheroid treatment. These findings suggest that transplantation of PBM-spheroid hASCs may be an effective stem cell therapy for the treatment of skin flap ischemia.

  13. Excess Visceral Adipose Tissue Worsens the Vascular Endothelial Function in Patients with Type 2 Diabetes Mellitus.

    PubMed

    Kurozumi, Akira; Okada, Yosuke; Arao, Tadashi; Tanaka, Yoshiya

    Objective Visceral fat obesity and metabolic syndrome correlate with atherosclerosis in part due to insulin resistance and various other factors. The aim of this study was to determine the relationship between vascular endothelial dysfunction and excess visceral adipose tissue (VAT) in Japanese patients with type 2 diabetes mellitus (T2DM). Methods In 71 T2DM patients, the reactive hyperemia index (RHI) was measured using an Endo-PAT 2000, and VAT and subcutaneous adipose tissue (SAT) were measured via CT. We also measured various metabolic markers, including high-molecular-weight adiponectin (HMW-AN). Results VAT correlated negatively with the natural logarithm of RHI (L_RHI), the primary endpoint (p=0.042, r=-0.242). L_RHI did not correlate with SAT, VAT/SAT, abdominal circumference, homeostasis model assessment for insulin resistance, urinary C-peptide reactivity, HMW-AN, or alanine amino transferase, the secondary endpoints. A linear multivariate analysis via the forced entry method using age, sex, VAT, and smoking history as independent variables and L_RHI as the dependent variable revealed a lack of any determinants of L_RHI. Conclusion Excess VAT worsens the vascular endothelial function, represented by RHI which was analyzed using Endo-PAT, in Japanese patients with T2DM.

  14. The potential of chondrogenic pre-differentiation of adipose-derived mesenchymal stem cells for regeneration in harsh nucleus pulposus microenvironment.

    PubMed

    Wang, Jingkai; Tao, Yiqing; Zhou, Xiaopeng; Li, Hao; Liang, Chengzhen; Li, Fangcai; Chen, Qi-Xin

    2016-12-01

    Recent studies indicated that cell-based therapy could be a promising approach to treat intervertebral disc degeneration. Though the harsh microenvironment in disc is still challenging to implanted cells, it could be overcome by pre-conditioning graft cells before transplantation, suggested by previous literatures. Therefore, we designed this study to identify the potential effect of chondrogenic pre-differentiation on adipose-derived mesenchymal stem cells in intervertebral disc-like microenvironment, characterized by limited nutrition, acidic, and high osmosis in vitro. Adipose-derived mesenchymal stem cells of rat were divided into five groups, embedded in type II collagen scaffold, and cultured in chondrogenic differentiation medium for 0, 3, 7, 10, and 14 days. Then, the adipose-derived mesenchymal stem cells were implanted and cultured in intervertebral disc-like condition. The proliferation and differentiation of adipose-derived mesenchymal stem cells were evaluated by cell counting kit-8 test, real-time quantitative polymerase chain reaction, and Western blotting and immunofluorescence analysis. Analyzed by the first week in intervertebral disc-like condition, the results showed relatively greater proliferative capability and extracellular matrix synthesis ability of the adipose-derived mesenchymal stem cells pre-differentiated for 7 and 10 days than the control. We concluded that pre-differentiation of rat adipose-derived mesenchymal stem cells in chondrogenic culture medium for 7 to 10 days could promote the regeneration effect of adipose-derived mesenchymal stem cells in intervertebral disc-like condition, and the pre-differentiated cells could be a promising cell source for disc regeneration medicine.

  15. Mesenchymal Stromal Cell-Derived Extracellular Vesicles Protect the Fetal Brain After Hypoxia-Ischemia.

    PubMed

    Ophelders, Daan R M G; Wolfs, Tim G A M; Jellema, Reint K; Zwanenburg, Alex; Andriessen, Peter; Delhaas, Tammo; Ludwig, Anna-Kristin; Radtke, Stefan; Peters, Vera; Janssen, Leon; Giebel, Bernd; Kramer, Boris W

    2016-06-01

    Preterm neonates are susceptible to perinatal hypoxic-ischemic brain injury, for which no treatment is available. In a preclinical animal model of hypoxic-ischemic brain injury in ovine fetuses, we have demonstrated the neuroprotective potential of systemically administered mesenchymal stromal cells (MSCs). The mechanism of MSC treatment is unclear but suggested to be paracrine, through secretion of extracellular vesicles (EVs). Therefore, we investigated in this study the protective effects of mesenchymal stromal cell-derived extracellular vesicles (MSC-EVs) in a preclinical model of preterm hypoxic-ischemic brain injury. Ovine fetuses were subjected to global hypoxia-ischemia by transient umbilical cord occlusion, followed by in utero intravenous administration of MSC-EVs. The therapeutic effects of MSC-EV administration were assessed by analysis of electrophysiological parameters and histology of the brain. Systemic administration of MSC-EVs improved brain function by reducing the total number and duration of seizures, and by preserving baroreceptor reflex sensitivity. These functional protections were accompanied by a tendency to prevent hypomyelination. Cerebral inflammation remained unaffected by the MSC-EV treatment. Our data demonstrate that MSC-EV treatment might provide a novel strategy to reduce the neurological sequelae following hypoxic-ischemic injury of the preterm brain. Our study results suggest that a cell-free preparation comprising neuroprotective MSC-EVs could substitute MSCs in the treatment of preterm neonates with hypoxic-ischemic brain injury, thereby circumventing the potential risks of systemic administration of living cells. Bone marrow-derived mesenchymal stromal cells (MSCs) show promise in treating hypoxic-ischemic injury of the preterm brain. Study results suggest administration of extracellular vesicles, rather than intact MSCs, is sufficient to exert therapeutic effects and avoids potential concerns associated with administration

  16. Uninduced adipose-derived stem cells repair the defect of full-thickness hyaline cartilage.

    PubMed

    Zhang, Hai-Ning; Li, Lei; Leng, Ping; Wang, Ying-Zhen; Lv, Cheng-Yu

    2009-04-01

    To testify the effect of the stem cells derived from the widely distributed fat tissue on repairing full-thickness hyaline cartilage defects. Adipose-derived stem cells (ADSCs) were derived from adipose tissue and cultured in vitro. Twenty-seven New Zealand white rabbits were divided into three groups randomly. The cultured ADSCs mixed with calcium alginate gel were used to fill the full-thickness hyaline cartilage defects created at the patellafemoral joint, and the defects repaired with gel or without treatment served as control groups. After 4, 8 and 12 weeks, the reconstructed tissue was evaluated macroscopically and microscopically. Histological analysis and qualitative scoring were also performed to detect the outcome. Full thickness hyaline cartilage defects were repaired completely with ADSCs-derived tissue. The result was better in ADSCs group than the control ones. The microstructure of reconstructed tissue with ADSCs was similar to that of hyaline cartilage and contained more cells and regular matrix fibers, being better than other groups. Plenty of collagen fibers around cells could be seen under transmission electron microscopy. Statistical analysis revealed a significant difference in comparison with other groups at each time point (t equal to 4.360, P less than 0.01). These results indicate that stem cells derived from mature adipose without induction possess the ability to repair cartilage defects.

  17. [Isolation,culture and identification of adipose-derived stem cells from SD rat adipose tissues subjected to long-term cryopreservation].

    PubMed

    Liu, Qin; Wang, Liping; Chen, Fang; Zhang, Yi

    2017-02-01

    To study the feasibility of isolation and culture of adipose-derived stem cells( ADSCs) from SD rat adipose tissues subjected to long-term cryopreservation. We took inguinal fat pads from healthy SD rats. Adipose tissues were stored with 100 m L / L dimethyl sulfoxide( DMSO) combined with 900 m L / L fetal bovine serum( FBS) in liquid nitrogen. Three months later,the adipose tissues were resuscitated for the isolation and culture of ADSCs. The growth status and morphology were observed. The growth curve and cell surface markers CD29,CD45,CD90 of the 3rd passage cells were analyzed respectively by CCK-8 assay and immunocytochemistry. The 3rd passage cells were induced towards adipogenic lineages and osteogenic lineages by different inducers,and the resulting cells were examined separately by oil red O staining and alizarin red staining. The ADSCs obtained from SD rat adipose tissues subjected to long-term cryopreservation showed a spindle-shape appearance and had a good proliferation ability. The cell growth curve was typical "S " curve.Immunocytochemistry showed that the 3rd passage cells were positive for CD29 and CD90,while negative for CD45. The cells were positive for oil red O staining after adipogenic induction,and also positive for alizarin red staining after osteogenic induction. The ADSCs can be isolated from SD rat adipose tissues subjected to long-term cryopreservation.

  18. The Cladophora glomerata Enriched by Biosorption Process in Cr(III) Improves Viability, and Reduces Oxidative Stress and Apoptosis in Equine Metabolic Syndrome Derived Adipose Mesenchymal Stromal Stem Cells (ASCs) and Their Extracellular Vesicles (MV’s)

    PubMed Central

    Marycz, Krzysztof; Marędziak, Monika; Weiss, Christine

    2017-01-01

    This study investigated in vitro effects of freshwater alga Cladophora glomerata water extract enriched during a biosorption process in Cr(III) trivalent chromium and chromium picolinate on adipose-derived mesenchymal stromal stem cells (ASCs) and extracellular microvesicles (MVs) in equine metabolic syndrome-affected horses. Chemical characterisation of natural Cladophora glomerata was performed with special emphasis on: vitamin C, vitamin E, total phenols, fatty acids, free and protein-bound amino acids as well as measured Cr in algal biomass. To examine the influence of Cladophora glomerata water extracts, in vitro viability, oxidative stress factor accumulation, apoptosis, inflammatory response, biogenesis of mitochondria, autophagy in ASCs of EMS and secretory activity manifested by MV release were investigated. For this purpose, various methods of molecular biology and microscopic observations (i.e., immunofluorescence staining, SEM, TEM, FIB observations, mRNA and microRNA expression by RT-qPCR) were applied. The extract of Cladophora glomerata enriched with Cr(III) ions reduced apoptosis and inflammation in ASCs of EMS horses through improvement of mitochondrial dynamics, decreasing of PDK4 expression and reduction of endoplastic reticulum stress. Moreover, it was found, that Cladophora glomerata and Cr(III) induce antioxidative protection coming from enhanced SOD activity Therefore, Cladophora glomerata enriched with Cr(III) ions might become an interesting future therapeutic agent in the pharmacological treatment of EMS horses. PMID:29292726

  19. Inducing Heat Shock Proteins Enhances the Stemness of Frozen-Thawed Adipose Tissue-Derived Stem Cells.

    PubMed

    Shaik, Shahensha; Hayes, Daniel; Gimble, Jeffrey; Devireddy, Ram

    2017-04-15

    Extensive research has been performed to determine the effect of freezing protocol and cryopreservation agents on the viability of adipose tissue-derived stromal/stem cells (ASCs) as well as other cells. Unfortunately, the conclusion one may draw after decades of research utilizing fundamentally similar cryopreservation techniques is that a barrier exists, which precludes full recovery. We hypothesize that agents capable of inducing a subset of heat shock proteins (HSPs) and chaperones will reduce the intrinsic barriers to the post-thaw recovery of ASCs. ASCs were exposed to 43°C for 1 h to upregulate HSPs, and the temporal HSP expression profile postheat shock was determined by performing quantitative polymerase chain reaction (PCR) and western blotting assays. The expression levels of HSP70 and HSP32 were found to be maximum at 3 h after the heat shock, whereas HSP90 and HSP27 remain unchanged. The heat shocked ASCs cryopreserved during maximal HSPs expression exhibited increased post-thaw viability than the nonheat shocked samples. Histochemical staining and quantitative reverse transcription-PCR indicated that the ASC differentiation potential was retained. Thus, suggesting that the upregulation of HSPs before a freezing insult is beneficial to ASCs and a potential alternative to the use of harmful cryoprotective agents.

  20. Characterization of In Vitro Engineered Human Adipose Tissues: Relevant Adipokine Secretion and Impact of TNF-α

    PubMed Central

    Aubin, Kim; Safoine, Meryem; Proulx, Maryse; Audet-Casgrain, Marie-Alice; Côté, Jean-François; Têtu, Félix-André; Roy, Alphonse; Fradette, Julie

    2015-01-01

    Representative modelling of human adipose tissue functions is central to metabolic research. Tridimensional models able to recreate human adipogenesis in a physiological tissue-like context in vitro are still scarce. We describe the engineering of white adipose tissues reconstructed from their cultured adipose-derived stromal precursor cells. We hypothesize that these reconstructed tissues can recapitulate key functions of AT under basal and pro-inflammatory conditions. These tissues, featuring human adipocytes surrounded by stroma, were stable and metabolically active in long-term cultures (at least 11 weeks). Secretion of major adipokines and growth factors by the reconstructed tissues was determined and compared to media conditioned by human native fat explants. Interestingly, the secretory profiles of the reconstructed adipose tissues indicated an abundant production of leptin, PAI-1 and angiopoietin-1 proteins, while higher HGF levels were detected for the human fat explants. We next demonstrated the responsiveness of the tissues to the pro-inflammatory stimulus TNF-α, as reflected by modulation of MCP-1, NGF and HGF secretion, while VEGF and leptin protein expression did not vary. TNF-α exposure induced changes in gene expression for adipocyte metabolism-associated mRNAs such as SLC2A4, FASN and LIPE, as well as for genes implicated in NF-κB activation. Finally, this model was customized to feature adipocytes representative of progressive stages of differentiation, thereby allowing investigations using newly differentiated or more mature adipocytes. In conclusion, we produced tridimensional tissues engineered in vitro that are able to recapitulate key characteristics of subcutaneous white adipose tissue. These tissues are produced from human cells and their neo-synthesized matrix elements without exogenous or synthetic biomaterials. Therefore, they represent unique tools to investigate the effects of pharmacologically active products on human stromal cells

  1. Spontaneous hypertension occurs with adipose tissue dysfunction in perilipin-1 null mice.

    PubMed

    Zou, Liangqiang; Wang, Weiyi; Liu, Shangxin; Zhao, Xiaojing; Lyv, Ying; Du, Congkuo; Su, Xueying; Geng, Bin; Xu, Guoheng

    2016-02-01

    Perilipin-1 (Plin1) coats lipid droplets exclusively in adipocytes and regulates two principle functions of adipose tissue, triglyceride storage and hydrolysis, which are disrupted upon Plin1 deficiency. In the present study, we investigated the alterations in systemic metabolites and hormones, vascular function and adipose function in spontaneous hypertensive mice lacking perilipin-1 (Plin1-/-). Plin1-/- mice developed spontaneous hypertension without obvious alterations in systemic metabolites and hormones. Plin1 expressed only in adipose cells but not in vascular cells, so its ablation would have no direct effect in situ on blood vessels. Instead, Plin1-/- mice showed dysfunctions of perivascular adipose tissue (PVAT), a fat depot that anatomically surrounds systemic arteries and has an anticontractile effect. In Plin1-/- mice, aortic and mesenteric PVAT were reduced in mass and adipocyte derived relaxing factor secretion, but increased in basal lipolysis, angiotensin II secretion, macrophage infiltration and oxidative stress. Such multiple culprits impaired the anticontractile effect of PVAT to promote vasoconstriction of aortic and mesenteric arteries of Plin1-/- mice. Furthermore, arterial vessels of Plin1-/- mice showed increasing angiotensin II receptor type 1, monocyte chemotactic protein-1 and interlukin-6 expression, structural damage of endothelial and smooth muscle cells, along with impaired endothelium-dependent relaxation. Hypertension in Plin1-/- mice might occur as a deleterious consequence of PVAT dysfunction. This finding provides the direct evidence that links dysfunctional PVAT to vascular dysfunction and hypertension, particularly in pathophysiological states. This hypertensive mouse model might mimic and explain the hypertension occurring in patients with adipose tissue dysfunction, particularly with Plin1 mutations. Copyright © 2015 Elsevier B.V. All rights reserved.

  2. Human bone marrow harbors cells with neural crest-associated characteristics like human adipose and dermis tissues

    PubMed Central

    Coste, Cécile; Neirinckx, Virginie; Sharma, Anil; Agirman, Gulistan; Rogister, Bernard; Foguenne, Jacques; Lallemend, François

    2017-01-01

    Adult neural crest stem-derived cells (NCSC) are of extraordinary high plasticity and promising candidates for use in regenerative medicine. Several locations such as skin, adipose tissue, dental pulp or bone marrow have been described in rodent, as sources of NCSC. However, very little information is available concerning their correspondence in human tissues, and more precisely for human bone marrow. The main objective of this study was therefore to characterize NCSC from adult human bone marrow. In this purpose, we compared human bone marrow stromal cells to human adipose tissue and dermis, already described for containing NCSC. We performed comparative analyses in terms of gene and protein expression as well as functional characterizations. It appeared that human bone marrow, similarly to adipose tissue and dermis, contains NESTIN+ / SOX9+ / TWIST+ / SLUG+ / P75NTR+ / BRN3A+/ MSI1+/ SNAIL1+ cells and were able to differentiate into melanocytes, Schwann cells and neurons. Moreover, when injected into chicken embryos, all those cells were able to migrate and follow endogenous neural crest migration pathways. Altogether, the phenotypic characterization and migration abilities strongly suggest the presence of neural crest-derived cells in human adult bone marrow. PMID:28683107

  3. Novel clinical uses for cord blood derived mesenchymal stromal cells.

    PubMed

    Olson, Amanda L; McNiece, Ian K

    2015-06-01

    Regenerative medicine offers new hope for many debilitating diseases that result in damage to tissues and organs. The concept is straightforward with replacement of damaged cells with new functional cells. However, most tissues and organs are complex structures involving multiple cell types, supportive structures, a microenvironment producing cytokines and growth factors and a vascular system to supply oxygen and other nutrients. Therefore repair, particularly in the setting of ischemic damage, may require delivery of multiple cell types providing new vessel formation, a new microenvironment and functional cells. The field of stem cell biology has identified a number of stem cell sources including embryonic stem cells and adult stem cells that offer the potential to replace virtually all functional cells of the body. The focus of this article is a discussion of the potential of mesenchymal stromal cells (MSCs) from cord blood (CB) for regenerative medicine approaches. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  4. Comparative associations between anthropometric and bioelectric impedance analysis derived adiposity measures with blood pressure and hypertension in India: a cross-sectional analysis.

    PubMed

    Taing, Kevin Y; Farkouh, Michael E; Moineddin, Rahim; Tu, Jack V; Jha, Prabhat

    2017-01-01

    The utility of bioelectrical impedance analysis (BIA) derived adiposity measures as compared to anthropometric measures for the assessment of adiposity-related health risk is not clear. We aimed to clarify the relationships of BIA and anthropometric derived adipose measures with blood pressure and hypertension, and to compare the discriminative ability of the respective measures for hypertension. We used baseline data collected between 2015 and 2016 from the Indian Study on Health of Adults (ISHA), an ongoing population based cohort study in India ( N  = 5990; age 30-69 years). We examined and compared the associations and discriminative ability between anthropometric (body mass index, waist circumference, hip circumference, waist-hip ratio, waist-height ratio) and BIA (whole body and trunk fat percentage) derived adiposity measures with blood pressure components (systolic pressure, diastolic pressure, pulse pressure, mean arterial pressure, mid-blood pressure) and hypertension. Regardless of whether the adiposity measure was derived from BIA or anthropometry, all were strongly and positively associated with blood pressure and hypertension. For both men and women, the magnitude of association of BIA measures with blood pressure and hypertension were comparable to those of anthropometric measures. Further, the ability of BIA derived adiposity measures to distinguish between those with and without hypertension was similar to the discriminative ability of anthropometric measures. As compared to simple anthropometric measures, BIA derived estimates of adiposity provide no apparent advantage in the assessment of blood pressure and hypertension. The observed similarities between adiposity measures suggest that simple anthropometrics may be sufficient to assess adiposity and adiposity-related risks.

  5. Spontaneous In Vivo Chondrogenesis of Bone Marrow-Derived Mesenchymal Progenitor Cells by Blocking Vascular Endothelial Growth Factor Signaling.

    PubMed

    Marsano, Anna; Medeiros da Cunha, Carolina M; Ghanaati, Shahram; Gueven, Sinan; Centola, Matteo; Tsaryk, Roman; Barbeck, Mike; Stuedle, Chiara; Barbero, Andrea; Helmrich, Uta; Schaeren, Stefan; Kirkpatrick, James C; Banfi, Andrea; Martin, Ivan

    2016-12-01

    : Chondrogenic differentiation of bone marrow-derived mesenchymal stromal/stem cells (MSCs) can be induced by presenting morphogenetic factors or soluble signals but typically suffers from limited efficiency, reproducibility across primary batches, and maintenance of phenotypic stability. Considering the avascular and hypoxic milieu of articular cartilage, we hypothesized that sole inhibition of angiogenesis can provide physiological cues to direct in vivo differentiation of uncommitted MSCs to stable cartilage formation. Human MSCs were retrovirally transduced to express a decoy soluble vascular endothelial growth factor (VEGF) receptor-2 (sFlk1), which efficiently sequesters endogenous VEGF in vivo, seeded on collagen sponges and immediately implanted ectopically in nude mice. Although naïve cells formed vascularized fibrous tissue, sFlk1-MSCs abolished vascular ingrowth into engineered constructs, which efficiently and reproducibly developed into hyaline cartilage. The generated cartilage was phenotypically stable and showed no sign of hypertrophic evolution up to 12 weeks. In vitro analyses indicated that spontaneous chondrogenic differentiation by blockade of angiogenesis was related to the generation of a hypoxic environment, in turn activating the transforming growth factor-β pathway. These findings suggest that VEGF blockade is a robust strategy to enhance cartilage repair by endogenous or grafted mesenchymal progenitors. This article outlines the general paradigm of controlling the fate of implanted stem/progenitor cells by engineering their ability to establish specific microenvironmental conditions rather than directly providing individual morphogenic cues. Chondrogenic differentiation of mesenchymal stromal/stem cells (MSCs) is typically targeted by morphogen delivery, which is often associated with limited efficiency, stability, and robustness. This article proposes a strategy to engineer MSCs with the capacity to establish specific

  6. Direct transplantation of native pericytes from adipose tissue: A new perspective to stimulate healing in critical size bone defects.

    PubMed

    König, Matthias A; Canepa, Daisy D; Cadosch, Dieter; Casanova, Elisa; Heinzelmann, Michael; Rittirsch, Daniel; Plecko, Michael; Hemmi, Sonja; Simmen, Hans-Peter; Cinelli, Paolo; Wanner, Guido A

    2016-01-01

    Fractures with a critical size bone defect (e.g., open fracture with segmental bone loss) are associated with high rates of delayed union and non-union. The prevention and treatment of these complications remain a serious issue in trauma and orthopaedic surgery. Autologous cancellous bone grafting is a well-established and widely used technique. However, it has drawbacks related to availability, increased morbidity and insufficient efficacy. Mesenchymal stromal cells can potentially be used to improve fracture healing. In particular, human fat tissue has been identified as a good source of multilineage adipose-derived stem cells, which can be differentiated into osteoblasts. The main issue is that mesenchymal stromal cells are a heterogeneous population of progenitors and lineage-committed cells harboring a broad range of regenerative properties. This heterogeneity is also mirrored in the differentiation potential of these cells. In the present study, we sought to test the possibility to enrich defined subpopulations of stem/progenitor cells for direct therapeutic application without requiring an in vitro expansion. We enriched a CD146+NG2+CD45- population of pericytes from freshly isolated stromal vascular fraction from mouse fat tissue and tested their osteogenic differentiation capacity in vitro and in vivo in a mouse model for critical size bone injury. Our results confirm the ability of enriched CD146+NG2+CD45- cells to efficiently generate osteoblasts in vitro, to colonize cancellous bone scaffolds and to successfully contribute to regeneration of large bone defects in vivo. This study represents proof of principle for the direct use of enriched populations of cells with stem/progenitor identity for therapeutic applications. Copyright © 2015 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  7. Human Adipose-Derived Mesenchymal Stem Cells Cryopreservation and Thawing Decrease α4-Integrin Expression.

    PubMed

    Irioda, Ana Carolina; Cassilha, Rafael; Zocche, Larissa; Francisco, Julio Cesar; Cunha, Ricardo Correa; Ferreira, Priscila Elias; Guarita-Souza, Luiz Cesar; Ferreira, Reginaldo Justino; Mogharbel, Bassam Felipe; Garikipati, Venkata Naga Srikanth; Souza, Daiany; Beltrame, Mirian Perlingeiro; de Carvalho, Katherine Athayde Teixeira

    2016-01-01

    Aim. The effects of cryopreservation on adipose tissue-derived mesenchymal stem cells are not clearly documented, as there is a growing body of evidence about the importance of adipose-derived mesenchymal stem cells for regenerative therapies. The aim of this study was to analyze human adipose tissue-derived mesenchymal stem cells phenotypic expression (CD34, CD45, CD73, CD90, CD105, and CD49d), colony forming unit ability, viability, and differentiation potential before and after cryopreservation. Materials and Methods. 12 samples of the adipose tissue were collected from a healthy donor using the liposuction technique. The cell isolation was performed by enzymatic digestion and then the cells were cultured up to passage 2. Before and after cryopreservation the immunophenotype, cellular viability analysis by flow cytometer, colony forming units ability, differentiation potential into adipocytes and osteoblasts as demonstrated by Oil Red O and Alizarin Red staining, respectively. Results. The immunophenotypic markers expression was largely preserved, and their multipotency was maintained. However, after cryopreservation, the cells decreased α4-integrin expression (CD49d), cell viability, and number of colony forming units. Conclusions. These findings suggest that ADMSC transplanted after cryopreservation might compromise the retention of transplanted cells in the host tissue. Therefore, further studies are warranted to standardize protocols related to cryopreservation to attain full benefits of stem cell therapy.

  8. Mechanisms of vasculogenesis in 3D fibrin matrices mediated by the interaction of adipose-derived stem cells and endothelial cells.

    PubMed

    Rohringer, Sabrina; Hofbauer, Pablo; Schneider, Karl H; Husa, Anna-Maria; Feichtinger, Georg; Peterbauer-Scherb, Anja; Redl, Heinz; Holnthoner, Wolfgang

    2014-10-01

    Vascularization of tissue-engineered constructs is essential to provide sufficient nutrient supply and hemostasis after implantation into target sites. Co-cultures of adipose-derived stem cells (ASC) with outgrowth endothelial cells (OEC) in fibrin gels were shown to provide an effective possibility to induce vasculogenesis in vitro. However, the mechanisms of the interaction between these two cell types remain unclear so far. The aim of this study was to evaluate differences of direct and indirect stimulation of ASC-induced vasculogenesis, the influence of ASC on network stabilization and molecular mechanisms involved in vascular structure formation. Endothelial cells (EC) were embedded in fibrin gels either containing non-coated or ASC-coated microcarrier beads as well as ASC alone. Moreover, EC-seeded constructs incubated with ASC-conditioned medium were used in addition to constructs with ASC seeded on top. Vascular network formation was visualized by green fluorescent protein expressing cells or immunostaining for CD31 and quantified. RT-qPCR of cells derived from co-cultures in fibrin was performed to evaluate changes in the expression of EC marker genes during the first week of culture. Moreover, angiogenesis-related protein levels were measured by performing angiogenesis proteome profiler arrays. The results demonstrate that proximity of endothelial cells and ASC is required for network formation and ASC stabilize EC networks by developing pericyte characteristics. We further showed that ASC induce controlled vessel growth by secreting pro-angiogenic and regulatory proteins. This study reveals angiogenic protein profiles involved in EC/ASC interactions in fibrin matrices and confirms the usability of OEC/ASC co-cultures for autologous vascular tissue engineering.

  9. Three-dimensional spheroid cell culture of umbilical cord tissue-derived mesenchymal stromal cells leads to enhanced paracrine induction of wound healing.

    PubMed

    Santos, Jorge M; Camões, Sérgio P; Filipe, Elysse; Cipriano, Madalena; Barcia, Rita N; Filipe, Mariana; Teixeira, Mariana; Simões, Sandra; Gaspar, Manuela; Mosqueira, Diogo; Nascimento, Diana S; Pinto-do-Ó, Perpétua; Cruz, Pedro; Cruz, Helder; Castro, Matilde; Miranda, Joana P

    2015-05-09

    The secretion of trophic factors by mesenchymal stromal cells has gained increased interest given the benefits it may bring to the treatment of a variety of traumatic injuries such as skin wounds. Herein, we report on a three-dimensional culture-based method to improve the paracrine activity of a specific population of umbilical cord tissue-derived mesenchymal stromal cells (UCX®) towards the application of conditioned medium for the treatment of cutaneous wounds. A UCX® three-dimensional culture model was developed and characterized with respect to spheroid formation, cell phenotype and cell viability. The secretion by UCX® spheroids of extracellular matrix proteins and trophic factors involved in the wound-healing process was analysed. The skin regenerative potential of UCX® three-dimensional culture-derived conditioned medium (CM3D) was also assessed in vitro and in vivo against UCX® two-dimensional culture-derived conditioned medium (CM2D) using scratch and tubulogenesis assays and a rat wound splinting model, respectively. UCX® spheroids kept in our three-dimensional system remained viable and multipotent and secreted considerable amounts of vascular endothelial growth factor A, which was undetected in two-dimensional cultures, and higher amounts of matrix metalloproteinase-2, matrix metalloproteinase-9, hepatocyte growth factor, transforming growth factor β1, granulocyte-colony stimulating factor, fibroblast growth factor 2 and interleukin-6, when compared to CM2D. Furthermore, CM3D significantly enhanced elastin production and migration of keratinocytes and fibroblasts in vitro. In turn, tubulogenesis assays revealed increased capillary maturation in the presence of CM3D, as seen by a significant increase in capillary thickness and length when compared to CM2D, and increased branching points and capillary number when compared to basal medium. Finally, CM3D-treated wounds presented signs of faster and better resolution when compared to untreated and CM

  10. Sea cucumbers with an anti-inflammatory effect on endothelial cells and subcutaneous but not on epicardial adipose tissue.

    PubMed

    Mena-Bueno, Sara; Atanasova, Miroslava; Fernández-Trasancos, Ángel; Paradela-Dobarro, Beatriz; Bravo, Susana B; Álvarez, Ezequiel; Fernández, Ángel L; Carrera, Iván; González-Juanatey, José R; Eiras, Sonia

    2016-02-01

    epicardial adipose tissue (EAT) from patients with coronary artery disease (CAD) contains higher levels of inflammatory proteins and lower adiponectin levels than subcutaneous adipose tissue (SAT), enhancing the progression of atherosclerosis. Since products from sea cucumber have anti-inflammatory properties, we investigated its effect on EAT, SAT and endothelial cells. stromal cells or explants from EAT and SAT were obtained from patients with cardiovascular disease. Extracts were obtained after hydrolysis by food-grade enzymes at different times. Proteins were identified by LC-MALDI mass spectrometry. Adipogenesis and adiponectin induction were determined on stromal cells in the presence/absence of extracts. The bioavailability of the extracts was tested on a Caco-2 cell culture model in vitro. The bioavailable fraction was probed on endothelial cells and EAT or SAT explants. Vascular cell adhesion protein (VCAM-1), intercellular adhesion molecule (ICAM-1), IL-6 and adiponectin were determined by real time polymerase chain reaction (RT-PCR). our results showed that H. forskali and P. tremulus extracts contained compounds with anti-oxidant and anti-inflammatory properties. The bioavailable fraction of P. tremulus reduced VCAM-1 (p < 0.01) and IL-6 (p < 0.05) expression levels in endothelial cells while bioavailable compounds from H. forskali decreased ICAM-1 expression in SAT (p < 0.05). No effect was observed on EAT. these results suggest that sea cucumber extracts might be used for the prevention of endothelial cells and SAT inflammation.

  11. Tracking of adipose tissue-derived progenitor cells using two magnetic nanoparticle types

    NASA Astrophysics Data System (ADS)

    Kasten, Annika; Siegmund, Birte J.; Grüttner, Cordula; Kühn, Jens-Peter; Frerich, Bernhard

    2015-04-01

    Magnetic resonance imaging (MRI) is to be considered as an emerging detection technique for cell tracking experiments to evaluate the fate of transplanted progenitor cells and develop successful cell therapies for tissue engineering. Adipose tissue engineering using adipose tissue-derived progenitor cells has been advocated for the cure of soft tissue defects or for persistent soft tissue augmentation. Adipose tissue-derived progenitor cells were differentiated into the adipogenic lineage and labeled with two different types of magnetic iron oxide nanoparticles in varying concentrations which resulted in a concentration-dependent reduction of gene expression of adipogenic differentiation markers, adiponectin and fatty acid-binding protein 4 (FABP4), whereas the metabolic activity was not altered. As a result, only low nanoparticle concentrations for labeling were used for in vivo experiments. Cells were seeded onto collagen scaffolds and subcutaneously implanted into severe combined immunodeficient (SCID) mice. At 24 h as well as 28 days after implantation, MRI analyses were performed visualizing nanoparticle-labeled cells using T2-weighted sequences. The quantification of absolute volume of the scaffolds revealed a decrease of volume over time in all experimental groups. The distribution of nanoparticle-labeled cells within the scaffolds varied likewise over time.

  12. The development and endocrine functions of adipose tissue

    USDA-ARS?s Scientific Manuscript database

    White adipose tissue is a mesenchymal tissue that begins developing in the fetus. Classically known for storing the body’s fuel reserves, adipose tissue is now recognized as an endocrine organ. As such, the secretions from adipose tissue are known to affect several systems such as the vascular and...

  13. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chu, Yijing; Tang, Huijuan; Guo, Yan

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOCmore » cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs.« less

  14. Optimized adipose tissue engineering strategy based on a neo-mechanical processing method.

    PubMed

    He, Yunfan; Lin, Maohui; Wang, Xuecen; Guan, Jingyan; Dong, Ziqing; Feng, Lu; Xing, Malcolm; Feng, Chuanbo; Li, Xiaojian

    2018-05-26

    Decellularized adipose tissue (DAT) represents a promising scaffold for adipose tissue engineering. However, the unique and prolonged lipid removal process required for adipose tissue can damage extracellular matrix (ECM) constituents. Moreover, inadequate vascularization limits the recellularization of DAT in vivo. We proposed a neo-mechanical protocol for rapidly breaking adipocytes and removing lipid content from adipose tissue. The lipid-depleted adipose tissue was then subjected to a fast and mild decellularization to fabricate high-quality DAT (M-DAT). Adipose liquid extract (ALE) derived from this mechanical process was collected and incorporated into M-DAT to further optimize in vivo recellularization. Ordinary DAT was fabricated and served as a control. This developed strategy was evaluated based on decellularization efficiency, ECM quality, and recellularization efficiency. Angiogenic factor components and angiogenic potential of ALE were evaluated in vivo and in vitro. M-DAT achieved the same decellularization efficiency, but exhibited better retention of ECM components and recellularization, compared to those with ordinary DAT. Protein quantification revealed considerable levels of angiogenic factors (basic fibroblast growth factor, epidermal growth factor, transforming growth factor-β1, and vascular endothelial growth factor) in ALE. ALE promoted tube formation in vitro and induced intense angiogenesis in M-DAT in vivo; furthermore, higher expression of the adipogenic factor PPARγ and greater numbers of adipocytes were evident following ALE treatment, compared to those in the M-DAT group. Mechanical processing of adipose tissue led to the production of high-quality M-DAT and angiogenic factor-enriched ALE. The combination of ALE and M-DAT could be a promising strategy for engineered adipose tissue construction. This article is protected by copyright. All rights reserved. © 2018 by the Wound Healing Society.

  15. Adipose Tissue Is a Neglected Viral Reservoir and an Inflammatory Site during Chronic HIV and SIV Infection

    PubMed Central

    Damouche, Abderaouf; Huot, Nicolas; Dejucq-Rainsford, Nathalie; Satie, Anne-Pascale; Mélard, Adeline; David, Ludivine; Gommet, Céline; Ghosn, Jade; Noel, Nicolas; Pourcher, Guillaume; Martinez, Valérie; Benoist, Stéphane; Béréziat, Véronique; Cosma, Antonio; Favier, Benoit; Vaslin, Bruno; Rouzioux, Christine; Capeau, Jacqueline; Müller-Trutwin, Michaela; Dereuddre-Bosquet, Nathalie; Le Grand, Roger; Lambotte, Olivier; Bourgeois, Christine

    2015-01-01

    Two of the crucial aspects of human immunodeficiency virus (HIV) infection are (i) viral persistence in reservoirs (precluding viral eradication) and (ii) chronic inflammation (directly associated with all-cause morbidities in antiretroviral therapy (ART)-controlled HIV-infected patients). The objective of the present study was to assess the potential involvement of adipose tissue in these two aspects. Adipose tissue is composed of adipocytes and the stromal vascular fraction (SVF); the latter comprises immune cells such as CD4+ T cells and macrophages (both of which are important target cells for HIV). The inflammatory potential of adipose tissue has been extensively described in the context of obesity. During HIV infection, the inflammatory profile of adipose tissue has been revealed by the occurrence of lipodystrophies (primarily related to ART). Data on the impact of HIV on the SVF (especially in individuals not receiving ART) are scarce. We first analyzed the impact of simian immunodeficiency virus (SIV) infection on abdominal subcutaneous and visceral adipose tissues in SIVmac251 infected macaques and found that both adipocytes and adipose tissue immune cells were affected. The adipocyte density was elevated, and adipose tissue immune cells presented enhanced immune activation and/or inflammatory profiles. We detected cell-associated SIV DNA and RNA in the SVF and in sorted CD4+ T cells and macrophages from adipose tissue. We demonstrated that SVF cells (including CD4+ T cells) are infected in ART-controlled HIV-infected patients. Importantly, the production of HIV RNA was detected by in situ hybridization, and after the in vitro reactivation of sorted CD4+ T cells from adipose tissue. We thus identified adipose tissue as a crucial cofactor in both viral persistence and chronic immune activation/inflammation during HIV infection. These observations open up new therapeutic strategies for limiting the size of the viral reservoir and decreasing low-grade chronic

  16. Effect of Diabetes Mellitus on Adipocyte-Derived Stem Cells in Rat.

    PubMed

    Jumabay, Medet; Moon, Jeremiah H; Yeerna, Huwate; Boström, Kristina I

    2015-11-01

    Diabetes mellitus affects the adipose tissue and mesenchymal stem cells derived from the adipose stroma and other tissues. Previous reports suggest that bone morphogenetic protein 4 (BMP4) is involved in diabetic complications, at the same time playing an important role in the maintenance of stem cells. In this study, we used rats transgenic for human islet amyloid polypeptide (HIP rats), a model of type 2 diabetes, to study the effect of diabetes on adipocyte-derived stem cells, referred to as dedifferentiated fat (DFAT) cells. Our results show that BMP4 expression in inguinal adipose tissue is significantly increased in HIP rats compared to controls, whereas matrix Gla protein (MGP), an inhibitor of BMP4 is decreased as determined by quantitative PCR, and immunofluorescence. In addition, adipose vascularity and expression of multiple endothelial cell markers was increased in the diabetic tissue, visualized by immunofluorescence for endothelial markers. The endothelial markers co-localized with the enhanced BMP4 expression, suggesting that vascular cells play a role BMP4 induction. The DFAT cells are multipotent stem cells derived from white mature adipocytes that undergo endothelial and adipogenic differentiation. DFAT cells prepared from the inguinal adipose tissue in HIP rats exhibited enhanced proliferative capacity compared to wild type. In addition, their ability to undergo both endothelial cell and adipogenic lineage differentiation was enhanced, as well as their response to BMP4, as assessed by lineage marker expression. We conclude that the DFAT cells are affected by diabetic changes and may contribute to the adipose dysfunction in diabetes. © 2015 Wiley Periodicals, Inc.

  17. Direct comparison of progenitor cells derived from adipose, muscle, and bone marrow from wild-type or craniosynostotic rabbits

    PubMed Central

    GM, Cooper; EL, Lensie; JJ, Cray; MR, Bykowski; GE, DeCesare; MA, Smalley; MP, Mooney; PG, Campbell; JE, Losee

    2010-01-01

    Background Reports have identified cells capable of osteogenic differentiation in bone marrow, muscle, and adipose tissues, but there are few direct comparisons of these different cell-types. Also, few have investigated the potential connection between a tissue-specific pathology and cells derived from seemingly unrelated tissues. Here, we compare cells isolated from wild-type rabbits or rabbits with nonsyndromic craniosynostosis, defined as the premature fusion of one or more of the cranial sutures. Methods Cells were derived from bone marrow, adipose, and muscle of 10 day-old wild-type rabbits (WT; n=17) or from age-matched rabbits with familial nonsyndromic craniosynostosis (CS; n=18). Cells were stimulated with bone morphogenetic protein 4 (BMP4) and alkaline phosphatase expression and cell proliferation were assessed. Results In WT rabbits, cells derived from muscle had more alkaline phosphatase activity than cells derived from either adipose or bone marrow. The cells derived from CS rabbit bone marrow and muscle were significantly more osteogenic than WT. Adipose-derived cells demonstrated no significant differences. While muscle-derived cells were most osteogenic in WT rabbits, bone marrow-derived cells were most osteogenic in CS rabbits. Conclusions Results suggest that cells from different tissues have different potentials for differentiation. Furthermore, cells derived from rabbits with craniosynostosis were different from wild-type derived cells. Interestingly, cells derived from the craniosynostotic rabbits were not uniformly more responsive compared with wild-type cells, suggesting that specific tissue-derived cells may react differently in individuals with craniosynostosis. PMID:20871482

  18. Ischemic stroke and select adipose-derived and sex hormones: a review.

    PubMed

    Meadows, Kristy L

    2018-06-06

    Ischemic stroke is the fifth leading cause of death in the USA and is the leading cause of serious, long-term disability worldwide. The principle sex hormones (estrogen, progesterone, and testosterone), both endogenous and exogenous, have profound effects on various stroke outcomes and have become the focus of a number of studies evaluating risk factors and treatment options for ischemic stroke. In addition, the expression of other hormones that may influence stroke outcome, including select adipose-derived hormones (adiponectin, leptin, and ghrelin), can be regulated by sex hormones and are also the focus of several ischemic stroke studies. This review aims to summarize some of the preclinical and clinical studies investigating the principle sex hormones, as well as select adipose-derived hormones, as risk factors or potential treatments for ischemic stroke. In addition, the potential for relaxin, a lesser studied sex hormone, as a novel treatment option for ischemic stroke is explored.

  19. Comparison of three methods for the derivation of a biologic scaffold composed of adipose tissue extracellular matrix.

    PubMed

    Brown, Bryan N; Freund, John M; Han, Li; Rubin, J Peter; Reing, Janet E; Jeffries, Eric M; Wolf, Mathew T; Tottey, Stephen; Barnes, Christopher A; Ratner, Buddy D; Badylak, Stephen F

    2011-04-01

    Extracellular matrix (ECM)-based scaffold materials have been used successfully in both preclinical and clinical tissue engineering and regenerative medicine approaches to tissue reconstruction. Results of numerous studies have shown that ECM scaffolds are capable of supporting the growth and differentiation of multiple cell types in vitro and of acting as inductive templates for constructive tissue remodeling after implantation in vivo. Adipose tissue represents a potentially abundant source of ECM and may represent an ideal substrate for the growth and adipogenic differentiation of stem cells harvested from this tissue. Numerous studies have shown that the methods by which ECM scaffold materials are prepared have a dramatic effect upon both the biochemical and structural properties of the resultant ECM scaffold material as well as the ability of the material to support a positive tissue remodeling outcome after implantation. The objective of the present study was to characterize the adipose ECM material resulting from three methods of decellularization to determine the most effective method for the derivation of an adipose tissue ECM scaffold that was largely free of potentially immunogenic cellular content while retaining tissue-specific structural and functional components as well as the ability to support the growth and adipogenic differentiation of adipose-derived stem cells. The results show that each of the decellularization methods produced an adipose ECM scaffold that was distinct from both a structural and biochemical perspective, emphasizing the importance of the decellularization protocol used to produce adipose ECM scaffolds. Further, the results suggest that the adipose ECM scaffolds produced using the methods described herein are capable of supporting the maintenance and adipogenic differentiation of adipose-derived stem cells and may represent effective substrates for use in tissue engineering and regenerative medicine approaches to soft tissue

  20. Effects of nanoporous anodic titanium oxide on human adipose derived stem cells.

    PubMed

    Malec, Katarzyna; Góralska, Joanna; Hubalewska-Mazgaj, Magdalena; Głowacz, Paulina; Jarosz, Magdalena; Brzewski, Pawel; Sulka, Grzegorz D; Jaskuła, Marian; Wybrańska, Iwona

    The aim of current bone biomaterials research is to design implants that induce controlled, guided, successful, and rapid healing. Titanium implants are widely used in dental, orthopedic, and reconstructive surgery. A series of studies has indicated that cells can respond not only to the chemical properties of the biomaterial, but also, in particular, to the changes in surface topography. Nanoporous materials remain in focus of scientific queries due to their exclusive properties and broad applications. One such material is nanostructured titanium oxide with highly ordered, mutually perpendicular nanopores. Nanoporous anodic titanium dioxide (TiO 2 ) films were fabricated by a three-step anodization process in propan-1,2,3-triol-based electrolyte containing fluoride ions. Adipose-derived stem cells offer many interesting opportunities for regenerative medicine. The important goal of tissue engineering is to direct stem cell differentiation into a desired cell lineage. The influence of nanoporous TiO 2 with pore diameters of 80 and 108 nm on cell response, growth, viability, and ability to differentiate into osteoblastic lineage of human adipose-derived progenitors was explored. Cells were harvested from the subcutaneous abdominal fat tissue by a simple, minimally invasive, and inexpensive method. Our results indicate that anodic nanostructured TiO 2 is a safe and nontoxic biomaterial. In vitro studies demonstrated that the nanotopography induced and enhanced osteodifferentiation of human adipose-derived stem cells from the abdominal subcutaneous fat tissue.

  1. Construction of engineering adipose-like tissue in vivo utilizing human insulin gene-modified umbilical cord mesenchymal stromal cells with silk fibroin 3D scaffolds.

    PubMed

    Li, Shi-Long; Liu, Yi; Hui, Ling

    2015-12-01

    We evaluated the use of a combination of human insulin gene-modified umbilical cord mesenchymal stromal cells (hUMSCs) with silk fibroin 3D scaffolds for adipose tissue engineering. In this study hUMSCs were isolated and cultured. HUMSCs infected with Ade-insulin-EGFP were seeded in fibroin 3D scaffolds with uniform 50-60 µm pore size. Silk fibroin scaffolds with untransfected hUMSCs were used as control. They were cultured for 4 days in adipogenic medium and transplanted under the dorsal skins of female Wistar rats after the hUMSCs had been labelled with chloromethylbenzamido-1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (CM-Dil). Macroscopical impression, fluorescence observation, histology and SEM were used for assessment after transplantation at 8 and 12 weeks. Macroscopically, newly formed adipose tissue was observed in the experimental group and control group after 8 and 12 weeks. Fluorescence observation supported that the formed adipose tissue originated from seeded hUMSCs rather than from possible infiltrating perivascular tissue. Oil red O staining of newly formed tissue showed that there was substantially more tissue regeneration in the experimental group than in the control group. SEM showed that experimental group cells had more fat-like cells, whose volume was larger than that of the control group, and degradation of the silk fibroin scaffold was greater under SEM observation. This study provides significant evidence that hUMSCs transfected by adenovirus vector have good compatibility with silk fibroin scaffold, and adenoviral transfection of the human insulin gene can be used for the construction of tissue-engineered adipose. Copyright © 2013 John Wiley & Sons, Ltd.

  2. Platelets secrete stromal cell–derived factor 1α and recruit bone marrow–derived progenitor cells to arterial thrombi in vivo

    PubMed Central

    Massberg, Steffen; Konrad, Ildiko; Schürzinger, Katrin; Lorenz, Michael; Schneider, Simon; Zohlnhoefer, Dietlind; Hoppe, Katharina; Schiemann, Matthias; Kennerknecht, Elisabeth; Sauer, Susanne; Schulz, Christian; Kerstan, Sandra; Rudelius, Martina; Seidl, Stefan; Sorge, Falko; Langer, Harald; Peluso, Mario; Goyal, Pankaj; Vestweber, Dietmar; Emambokus, Nikla R.; Busch, Dirk H.; Frampton, Jon; Gawaz, Meinrad

    2006-01-01

    The accumulation of smooth muscle and endothelial cells is essential for remodeling and repair of injured blood vessel walls. Bone marrow–derived progenitor cells have been implicated in vascular repair and remodeling; however, the mechanisms underlying their recruitment to the site of injury remain elusive. Here, using real-time in vivo fluorescence microscopy, we show that platelets provide the critical signal that recruits CD34+ bone marrow cells and c-Kit+ Sca-1+ Lin− bone marrow–derived progenitor cells to sites of vascular injury. Correspondingly, specific inhibition of platelet adhesion virtually abrogated the accumulation of both CD34+ and c-Kit+ Sca-1+ Lin− bone marrow–derived progenitor cells at sites of endothelial disruption. Binding of bone marrow cells to platelets involves both P-selectin and GPIIb integrin on platelets. Unexpectedly, we found that activated platelets secrete the chemokine SDF-1α, thereby supporting further primary adhesion and migration of progenitor cells. These findings establish the platelet as a major player in the initiation of vascular remodeling, a process of fundamental importance for vascular repair and pathological remodeling after vascular injury. PMID:16618794

  3. Platelets secrete stromal cell-derived factor 1alpha and recruit bone marrow-derived progenitor cells to arterial thrombi in vivo.

    PubMed

    Massberg, Steffen; Konrad, Ildiko; Schürzinger, Katrin; Lorenz, Michael; Schneider, Simon; Zohlnhoefer, Dietlind; Hoppe, Katharina; Schiemann, Matthias; Kennerknecht, Elisabeth; Sauer, Susanne; Schulz, Christian; Kerstan, Sandra; Rudelius, Martina; Seidl, Stefan; Sorge, Falko; Langer, Harald; Peluso, Mario; Goyal, Pankaj; Vestweber, Dietmar; Emambokus, Nikla R; Busch, Dirk H; Frampton, Jon; Gawaz, Meinrad

    2006-05-15

    The accumulation of smooth muscle and endothelial cells is essential for remodeling and repair of injured blood vessel walls. Bone marrow-derived progenitor cells have been implicated in vascular repair and remodeling; however, the mechanisms underlying their recruitment to the site of injury remain elusive. Here, using real-time in vivo fluorescence microscopy, we show that platelets provide the critical signal that recruits CD34+ bone marrow cells and c-Kit+ Sca-1+ Lin- bone marrow-derived progenitor cells to sites of vascular injury. Correspondingly, specific inhibition of platelet adhesion virtually abrogated the accumulation of both CD34+ and c-Kit+ Sca-1+ Lin- bone marrow-derived progenitor cells at sites of endothelial disruption. Binding of bone marrow cells to platelets involves both P-selectin and GPIIb integrin on platelets. Unexpectedly, we found that activated platelets secrete the chemokine SDF-1alpha, thereby supporting further primary adhesion and migration of progenitor cells. These findings establish the platelet as a major player in the initiation of vascular remodeling, a process of fundamental importance for vascular repair and pathological remodeling after vascular injury.

  4. Human umbilical vein endothelial cells protect against hypoxic-ischemic damage in neonatal brain via stromal cell-derived factor 1/C-X-C chemokine receptor type 4.

    PubMed

    Wu, Chia-Ching; Chen, Yi-Chi; Chang, Ying-Chao; Wang, Lan-Wan; Lin, Yung-Chieh; Chiang, Yi-Lun; Ho, Chien-Jung; Huang, Chao-Ching

    2013-05-01

    Agents that protect against neurovascular damage provide a powerful neuroprotective strategy. Human umbilical vein endothelial cells (HUVECs) may be used to treat neonates with hypoxic-ischemia (HI) because of its autologous capability. We hypothesized that peripherally injected HUVECs entered the brain after HI, protected against neurovascular damage, and provided protection via stromal cell-derived factor 1/C-X-C chemokine receptor type 4 pathway in neonatal brain. Postpartum day 7 rat pups received intraperitoneal injections of low-passage HUVEC-P4, high-passage HUVEC-P8, or conditioned medium before and immediately after HI. HUVECs were transfected with adenovirus-green fluorescent protein for cell tracing. Oxygen-glucose deprivation was established by coculturing HUVEC-P4 with mouse neuroblastoma neuronal cells (Neuro-2a) and with mouse immortalized cerebral vascular endothelial cells (b.End3). HUVEC-P4-treated group had more blood levels of green fluorescent protein-positive cells than HUVEC-P8-treated group 3 hours postinjection. Intraperitoneally injected HUVEC-P4, but not HUVEC-P8, entered the cortex after HI and positioned closed to the neurons and microvessels. Compared with the condition medium-treated group, the HUVEC-P4-treated but not the HUVEC-P8-treated group showed significantly less neuronal apoptosis and blood-brain barrier damage and more preservation of microvessels in the cortex 24 hours after HI. On postpartum day 14, the HUVEC-P4-treated group showed significant neuroprotection compared with the condition medium-treated group. Stromal cell-derived factor 1 was upregulated in the ipsilateral cortex 3 hours after HI, and inhibiting the stromal cell-derived factor 1/C-X-C chemokine receptor type 4 reduced the protective effect of HUVEC-P4. In vitro transwell coculturing of HUVEC-P4 also significantly protected against oxygen-glucose deprivation cell death in neurons and endothelial cells. Cell therapy using HUVECs may provide a powerful

  5. Circadian changes in uterine artery and ovarian stromal blood flow after pituitary down-regulation.

    PubMed

    Chan, Carina C W; Ng, Ernest H Y; Tang, Oi-Shan; Ho, Pak-Chung

    2005-09-01

    To investigate changes in the uterine artery and ovarian stromal blood flow in relation to the time of the day after pituitary down-regulation during in vitro fertilization treatment. Thirteen women were recruited. The uterine artery blood flow was studied using pulsed color Doppler ultrasonography and the ovarian stromal blood flow was measured using three-dimensional power Doppler ultrasonography. Ultrasound scan examinations and blood pressure measurements were performed in the morning and evening. The diastolic and the mean arterial pressures were significantly higher in the evening. An increase in the uterine artery pulsatility index and resistance index in the evening was observed. The ovarian vascularization index, vascularization flow index, and right ovarian flow index were significantly lower in the evening. Despite the small sample size, we have demonstrated the presence of a diurnal change in uterine artery and ovarian stromal blood flow after pituitary down-regulation. Such changes may be related to the systemic change in the sympathetic system and hence vascular resistance. Future study regarding ovarian stromal blood flow should take into account the effect of the time of the day on the readings in order to avoid misleading interpretation of data.

  6. Isolation and characterization of canine perivascular stem/stromal cells for bone tissue engineering.

    PubMed

    James, Aaron W; Zhang, Xinli; Crisan, Mihaela; Hardy, Winters R; Liang, Pei; Meyers, Carolyn A; Lobo, Sonja; Lagishetty, Venu; Childers, Martin K; Asatrian, Greg; Ding, Catherine; Yen, Yu-Hsin; Zou, Erin; Ting, Kang; Peault, Bruno; Soo, Chia

    2017-01-01

    For over 15 years, human subcutaneous adipose tissue has been recognized as a rich source of tissue resident mesenchymal stem/stromal cells (MSC). The isolation of perivascular progenitor cells from human adipose tissue by a cell sorting strategy was first published in 2008. Since this time, the interest in using pericytes and related perivascular stem/stromal cell (PSC) populations for tissue engineering has significantly increased. Here, we describe a set of experiments identifying, isolating and characterizing PSC from canine tissue (N = 12 canine adipose tissue samples). Results showed that the same antibodies used for human PSC identification and isolation are cross-reactive with canine tissue (CD45, CD146, CD34). Like their human correlate, canine PSC demonstrate characteristics of MSC including cell surface marker expression, colony forming unit-fibroblast (CFU-F) inclusion, and osteogenic differentiation potential. As well, canine PSC respond to osteoinductive signals in a similar fashion as do human PSC, such as the secreted differentiation factor NEL-Like Molecule-1 (NELL-1). Nevertheless, important differences exist between human and canine PSC, including differences in baseline osteogenic potential. In summary, canine PSC represent a multipotent mesenchymogenic cell source for future translational efforts in tissue engineering.

  7. Invited review: Pre- and postnatal adipose tissue development in farm animals: from stem cells to adipocyte physiology.

    PubMed

    Louveau, I; Perruchot, M-H; Bonnet, M; Gondret, F

    2016-11-01

    Both white and brown adipose tissues are recognized to be differently involved in energy metabolism and are also able to secrete a variety of factors called adipokines that are involved in a wide range of physiological and metabolic functions. Brown adipose tissue is predominant around birth, except in pigs. Irrespective of species, white adipose tissue has a large capacity to expand postnatally and is able to adapt to a variety of factors. The aim of this review is to update the cellular and molecular mechanisms associated with pre- and postnatal adipose tissue development with a special focus on pigs and ruminants. In contrast to other tissues, the embryonic origin of adipose cells remains the subject of debate. Adipose cells arise from the recruitment of specific multipotent stem cells/progenitors named adipose tissue-derived stromal cells. Recent studies have highlighted the existence of a variety of those cells being able to differentiate into white, brown or brown-like/beige adipocytes. After commitment to the adipocyte lineage, progenitors undergo large changes in the expression of many genes involved in cell cycle arrest, lipid accumulation and secretory functions. Early nutrition can affect these processes during fetal and perinatal periods and can also influence or pre-determinate later growth of adipose tissue. How these changes may be related to adipose tissue functional maturity around birth and can influence newborn survival is discussed. Altogether, a better knowledge of fetal and postnatal adipose tissue development is important for various aspects of animal production, including neonatal survival, postnatal growth efficiency and health.

  8. Physiologically based microenvironment for in vitro neural differentiation of adipose-derived stem cells.

    PubMed

    Graziano, Adriana Carol Eleonora; Avola, Rosanna; Perciavalle, Vincenzo; Nicoletti, Ferdinando; Cicala, Gianluca; Coco, Marinella; Cardile, Venera

    2018-03-26

    The limited capacity of nervous system to promote a spontaneous regeneration and the high rate of neurodegenerative diseases appearance are keys factors that stimulate researches both for defining the molecular mechanisms of pathophysiology and for evaluating putative strategies to induce neural tissue regeneration. In this latter aspect, the application of stem cells seems to be a promising approach, even if the control of their differentiation and the maintaining of a safe state of proliferation should be troubled. Here, we focus on adipose tissue-derived stem cells and we seek out the recent advances on the promotion of their neural differentiation, performing a critical integration of the basic biology and physiology of adipose tissue-derived stem cells with the functional modifications that the biophysical, biomechanical and biochemical microenvironment induces to cell phenotype. The pre-clinical studies showed that the neural differentiation by cell stimulation with growth factors benefits from the integration with biomaterials and biophysical interaction like microgravity. All these elements have been reported as furnisher of microenvironments with desirable biological, physical and mechanical properties. A critical review of current knowledge is here proposed, underscoring that a real advance toward a stable, safe and controllable adipose stem cells clinical application will derive from a synergic multidisciplinary approach that involves material engineer, basic cell biology, cell and tissue physiology.

  9. Physiologically based microenvironment for in vitro neural differentiation of adipose-derived stem cells

    PubMed Central

    Graziano, Adriana Carol Eleonora; Avola, Rosanna; Perciavalle, Vincenzo; Nicoletti, Ferdinando; Cicala, Gianluca; Coco, Marinella; Cardile, Venera

    2018-01-01

    The limited capacity of nervous system to promote a spontaneous regeneration and the high rate of neurodegenerative diseases appearance are keys factors that stimulate researches both for defining the molecular mechanisms of pathophysiology and for evaluating putative strategies to induce neural tissue regeneration. In this latter aspect, the application of stem cells seems to be a promising approach, even if the control of their differentiation and the maintaining of a safe state of proliferation should be troubled. Here, we focus on adipose tissue-derived stem cells and we seek out the recent advances on the promotion of their neural differentiation, performing a critical integration of the basic biology and physiology of adipose tissue-derived stem cells with the functional modifications that the biophysical, biomechanical and biochemical microenvironment induces to cell phenotype. The pre-clinical studies showed that the neural differentiation by cell stimulation with growth factors benefits from the integration with biomaterials and biophysical interaction like microgravity. All these elements have been reported as furnisher of microenvironments with desirable biological, physical and mechanical properties. A critical review of current knowledge is here proposed, underscoring that a real advance toward a stable, safe and controllable adipose stem cells clinical application will derive from a synergic multidisciplinary approach that involves material engineer, basic cell biology, cell and tissue physiology. PMID:29588808

  10. Control of Human Endometrial Stromal Cell Motility by PDGF-BB, HB-EGF and Trophoblast-Secreted Factors

    PubMed Central

    Schwenke, Maren; Knöfler, Martin; Velicky, Philipp; Weimar, Charlotte H. E.; Kruse, Michelle; Samalecos, Annemarie; Wolf, Anja; Macklon, Nick S.; Bamberger, Ana-Maria; Gellersen, Birgit

    2013-01-01

    Human implantation involves extensive tissue remodeling at the fetal-maternal interface. It is becoming increasingly evident that not only trophoblast, but also decidualizing endometrial stromal cells are inherently motile and invasive, and likely contribute to the highly dynamic processes at the implantation site. The present study was undertaken to further characterize the mechanisms involved in the regulation of endometrial stromal cell motility and to identify trophoblast-derived factors that modulate migration. Among local growth factors known to be present at the time of implantation, heparin-binding epidermal growth factor-like growth factor (HB-EGF) triggered chemotaxis (directed locomotion), whereas platelet-derived growth factor (PDGF)-BB elicited both chemotaxis and chemokinesis (non-directed locomotion) of endometrial stromal cells. Supernatants of the trophoblast cell line AC-1M88 and of first trimester villous explant cultures stimulated chemotaxis but not chemokinesis. Proteome profiling for cytokines and angiogenesis factors revealed neither PDGF-BB nor HB-EGF in conditioned media from trophoblast cells or villous explants, while placental growth factor, vascular endothelial growth factor and PDGF-AA were identified as prominent secretory products. Among these, only PDGF-AA triggered endometrial stromal cell chemotaxis. Neutralization of PDGF-AA in trophoblast conditioned media, however, did not diminish chemoattractant activity, suggesting the presence of additional trophoblast-derived chemotactic factors. Pathway inhibitor studies revealed ERK1/2, PI3 kinase/Akt and p38 signaling as relevant for chemotactic motility, whereas chemokinesis depended primarily on PI3 kinase/Akt activation. Both chemotaxis and chemokinesis were stimulated upon inhibition of Rho-associated, coiled-coil containing protein kinase. The chemotactic response to trophoblast secretions was not blunted by inhibition of isolated signaling cascades, indicating activation of

  11. Human umbilical cord blood-derived mesenchymal stromal cells and small intestinal submucosa hydrogel composite promotes combined radiation-wound healing of mice.

    PubMed

    Lee, Changsun; Shim, Sehwan; Jang, Hyosun; Myung, Hyunwook; Lee, Janet; Bae, Chang-Hwan; Myung, Jae Kyung; Kim, Min-Jung; Lee, Seung Bum; Jang, Won-Suk; Lee, Sun-Joo; Kim, Hwi-Yool; Lee, Seung-Sook; Park, Sunhoo

    2017-09-01

    Mesenchymal stromal cells (MSCs) are a promising agent for treating impaired wound healing, and their therapeutic potential may be enhanced by employing extracellular matrix scaffolds as cell culture scaffolds or transplant cell carriers. Here, we evaluated the effect of human umbilical cord blood-derived (hUCB)-MSCs and a porcine small intestinal submucosa (SIS)-derived extracellular matrix scaffold in a combined radiation-wound mouse model of impaired wound healing. hUCB-MSCs and SIS hydrogel composite was applied to the excisional wound of whole-body irradiated mice. Assessment of wound closing and histological evaluation were performed in vivo. We also cultured hUCB-MSCs on SIS gel and examined the angiogenic effect of conditioned medium on irradiated human umbilical vein endothelial cells (HUVECs) in vitro. hUCB-MSCs and SIS hydrogel composite treatment enhanced wound healing and angiogenesis in the wound site of mice. Conditioned medium from hUCB-MSCs cultured on SIS hydrogel promoted the chemotaxis of irradiated HUVECs more than their proliferation. The secretion of angiogenic growth factors hepatocyte growth factor, vascular endothelial growth factor-A and angiopoietin-1 from hUCB-MSCs was significantly increased by SIS hydrogel, with HGF being the predominant angiogenic factor of irradiated HUVECs. Our results suggest that the wound healing effect of hUCB-MSCs is enhanced by SIS hydrogel via a paracrine factor-mediated recruitment of vascular endothelial cells in a combined radiation-wound mouse model. Copyright © 2017 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  12. Influence of smartphone Wi-Fi signals on adipose-derived stem cells.

    PubMed

    Lee, Sang-Soon; Kim, Hyung-Rok; Kim, Min-Sook; Park, Sanghoon; Yoon, Eul-Sik; Park, Seung-Ha; Kim, Deok-Woo

    2014-09-01

    The use of smartphones is expanding rapidly around the world, thus raising the concern of possible harmful effects of radiofrequency generated by smartphones. We hypothesized that Wi-Fi signals from smartphones may have harmful influence on adipose-derived stem cells (ASCs). An in vitro study was performed to assess the influence of Wi-Fi signals from smartphones. The ASCs were incubated under a smartphone connected to a Wi-Fi network, which was uploading files at a speed of 4.8 Mbps for 10 hours a day, for a total of 5 days. We constructed 2 kinds of control cells, one grown in 37°C and the other grown in 39°C. After 5 days of Wi-Fi exposure from the smartphone, the cells underwent cell proliferation assay, apoptosis assay, and flow cytometry analysis. Three growth factors, vascular endothelial growth factor, hepatocyte growth factor, and transforming growth factor-β, were measured from ASC-conditioned media. Cell proliferation rate was higher in Wi-Fi-exposed cells and 39°C control cells compared with 37°C control cells. Apoptosis assay, flow cytometry analysis, and growth factor concentrations showed no remarkable differences among the 3 groups. We could not find any harmful effects of Wi-Fi electromagnetic signals from smartphones. The increased proliferation of ASCs under the smartphone, however, might be attributable to the thermal effect.

  13. Cellular Therapy With Human Autologous Adipose-Derived Adult Stem Cells for Advanced Keratoconus.

    PubMed

    Alió Del Barrio, Jorge L; El Zarif, Mona; de Miguel, María P; Azaar, Albert; Makdissy, Norman; Harb, Walid; El Achkar, Ibrahim; Arnalich-Montiel, Francisco; Alió, Jorge L

    2017-08-01

    The aim of this phase 1 study was to preliminarily evaluate the safety and efficacy of autologous adipose-derived adult stem cell (ADASC) implantation within the corneal stroma of patients with advanced keratoconus. Five consecutive patients were selected. Autologous ADASCs were obtained by elective liposuction. ADASCs (3 × 10) contained in 1 mL saline were injected into the corneal stroma through a femtosecond-assisted 9.5-mm diameter lamellar pocket under topical anesthesia. Patients were reviewed at 1 day, 1 week, 1, 3, and 6 months postoperatively. Visual function, manifest refraction, slit-lamp biomicroscopy, intraocular pressure, endothelial cell density, corneal topography, corneal optical coherence tomography, and corneal confocal biomicroscopy were recorded. No intraoperative or postoperative complications were recorded, with full corneal transparency recovery within 24 hours. Four patients completed the full follow-up. All patients improved their visual function (mean: 1 line of unaided and spectacle-corrected distance vision and 2 lines of rigid contact lens distance vision). Manifest refraction and topographic keratometry remained stable. Corneal optical coherence tomography showed a mean improvement of 16.5 μm in the central corneal thickness, and new collagen production was observed as patchy hyperreflective areas at the level of the stromal pocket. Confocal biomicroscopy confirmed the survival of the implanted stem cells at the surgical plane. Intraocular pressure and endothelial cell density remained stable. Cellular therapy of the human corneal stroma in vivo with autologous ADASCs appears to be safe. Stem cells survive in vivo with intrastromal new collagen production. Future studies with larger samples are required to confirm these preliminary results.

  14. Adipose-derived stem cells retain their regenerative potential after methotrexate treatment

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Beane, Olivia S.; Fonseca, Vera C.; Darling, Eric M., E-mail: Eric_Darling@brown.edu

    In musculoskeletal tissues like bone, chemotherapy can impair progenitor cell differentiation and proliferation, resulting in decreased bone growth and mineralization throughout a patient's lifetime. In the current study, we investigated the effects of chemotherapeutics on adipose-derived stem cell (ASC) function to determine whether this cell source could be a candidate for repairing, or even preventing, chemotherapy-induced tissue damage. Dose-dependent proliferation rates of ASCs and normal human fibroblasts (NHFs) were quantified after treatment with cytarabine (CY), etoposide (ETO), methotrexate (MTX), and vincristine (VIN) using a fluorescence-based assay. The influence of MTX on the multipotency of ASCs and freshly isolated stromal vascularmore » fraction (SVF) cells was also evaluated using lineage-specific stains and spectrophotometry. ASC and NHF proliferation were equally inhibited by exposure to CY and ETO; however, when treated with MTX and VIN, ASCs exhibited greater resistance. This was especially apparent for MTX-treated samples, with ASC proliferation showing no inhibition for clinically relevant MTX doses ranging from 0.1 to 50 μM. Additional experiments revealed that the differentiation potential of ASCs was not affected by MTX treatment and that upregulation of dihydrofolate reductase possibly contributed to this response. Moreover, SVF cells, which include ASCs, exhibited similar resistance to MTX impairment, with respect to cellular proliferation, clonogenicity, and differentiation capability. Therefore, we have shown that the regenerative properties of ASCs resist the cytotoxicity of MTX, identifying these cells as a potential key for repairing musculoskeletal damage in patients undergoing chemotherapy. - Highlights: • Long-term effects of chemotherapeutics can include musculoskeletal dysfunction. • A screen of common drugs showed disparate effects on ASCs and fibroblasts. • One drug, methotrexate, did not impair ASC growth

  15. Comparison of Tumor- and Bone Marrow-Derived Mesenchymal Stromal/Stem Cells from Patients with High-Grade Osteosarcoma

    PubMed Central

    Le Nail, Louis-Romée; Brennan, Meadhbh; Rosset, Philippe; Piloquet, Philippe; Pichon, Olivier; Le Caignec, Cédric; Crenn, Vincent; Layrolle, Pierre; Hérault, Olivier; De Pinieux, Gonzague

    2018-01-01

    Osteosarcoma (OS) is suspected to originate from dysfunctional mesenchymal stromal/stem cells (MSC). We sought to identify OS-derived cells (OSDC) with potential cancer stem cell (CSC) properties by comparing OSDC to MSC derived from bone marrow of patients. This study included in vitro characterization with sphere forming assays, differentiation assays, cytogenetic analysis, and in vivo investigations of their tumorigenicity and tumor supportive capacities. Primary cell lines were isolated from nine high-grade OS samples. All primary cell lines demonstrated stromal cell characteristics. Compared to MSC, OSDC presented a higher ability to form sphere clones, indicating a potential CSC phenotype, and were more efficient at differentiation towards osteoblasts. None of the OSDC displayed the complex chromosome rearrangements typical of high grade OS and none of them induced tumors in immunodeficient mice. However, two OSDC demonstrated focused genomic abnormalities. Three out of seven, and six out of seven OSDC showed a supportive role on local tumor development, and on metastatic progression to the lungs, respectively, when co-injected with OS cells in nude mice. The observation of OS-associated stromal cells with rare genetic abnormalities and with the capacity to sustain tumor progression may have implications for future tumor treatments. PMID:29494553

  16. Two-step transplantation with adipose tissue-derived stem cells increases follicle survival by enhancing vascularization in xenografted frozen-thawed human ovarian tissue.

    PubMed

    Manavella, D D; Cacciottola, L; Pommé, S; Desmet, C M; Jordan, B F; Donnez, J; Amorim, C A; Dolmans, M M

    2018-06-01

    Do adipose tissue-derived stem cells (ASCs) enhance vascularization and follicle survival in xenografted ovarian tissue using a two-step transplantation approach? Higher rates of oxygenation and vascularization of ovarian tissue, as well as increased follicle survival rates, were detected in the early post-grafting period. ASCs have multilineage differentiation potential, proangiogenic properties and enhance vascularization in a peritoneal grafting site. Some studies suggest that using ASCs may improve ovarian tissue quality by enhancing graft angiogenesis. A total of 15 severe combined immunodeficient (SCID) mice were intraperitoneally grafted with frozen-thawed human ovarian tissue (OT) from five different patients. A peritoneal transplantation site had been previously prepared in a first step using either empty fibrin (Fi+OT group [n = 5]) or ASC-loaded fibrin (Fi/ASCs+OT group [n = 5]) for 14 days prior to grafting. Five mice underwent the standard one-step transplantation procedure and served as controls (OT group). Lithium phthalocyanine (LiPc) crystals were inserted into all grafted human ovarian tissue before transplantation. Levels of partial pressure of oxygen (pO2) in grafts were monitored in vivo by electron paramagnetic resonance (EPR) oximetry on Days 3 and 7. Samples for histology and immunohistochemistry (IHC) were collected after euthanizing the mice on Day 7 following EPR. One piece of ovarian tissue per patient was fixed for analysis to serve as non-grafted controls. Prospective experimental study conducted at the Gynecology Research Unit, Université Catholique de Louvain. All materials were used to perform pO2 measurements (EPR oximetry), histological (haematoxylin and eosin staining), immunohistochemistry (anti-mouse and human double CD34 and anti-human Ki-67) and TUNEL analyses. A significant increase in pO2 was observed in all groups between Days 3 and 7 (P < 0.001). A significantly higher pO2 level was observed in the Fi/ASCs+OT group

  17. Human Adipose Tissue Derived Extracellular Matrix and Methylcellulose Hydrogels Augments and Regenerates the Paralyzed Vocal Fold

    PubMed Central

    Kim, Eun Na; Sung, Myung Whun; Kwon, Tack-Kyun; Cho, Yong Woo; Kwon, Seong Keun

    2016-01-01

    Vocal fold paralysis results from various etiologies and can induce voice changes, swallowing complications, and issues with aspiration. Vocal fold paralysis is typically managed using injection laryngoplasty with fat or synthetic polymers. Injection with autologous fat has shown excellent biocompatibility. However, it has several disadvantages such as unpredictable resorption rate, morbidities associated with liposuction procedure which has to be done in operating room under general anesthesia. Human adipose-derived extracellular matrix (ECM) grafts have been reported to form new adipose tissue and have greater biostability than autologous fat graft. Here, we present an injectable hydrogel that is constructed from adipose tissue derived soluble extracellular matrix (sECM) and methylcellulose (MC) for use in vocal fold augmentation. Human sECM derived from adipose tissue was extracted using two major steps—ECM was isolated from human adipose tissue and was subsequently solubilized. Injectable sECM/MC hydrogels were prepared by blending of sECM and MC. Sustained vocal fold augmentation and symmetric vocal fold vibration were accomplished by the sECM/MC hydrogel in paralyzed vocal fold which were confirmed by laryngoscope, histology and a high-speed imaging system. There were increased number of collagen fibers and fatty granules at the injection site without significant inflammation or fibrosis. Overall, these results indicate that the sECM/MC hydrogel can enhance vocal function in paralyzed vocal folds without early resorption and has potential as a promising material for injection laryngoplasty for stable vocal fold augmentation which can overcome the shortcomings of autologous fat such as unpredictable duration and morbidity associated with the fat harvest. PMID:27768757

  18. Adipose tissue engineering: state of the art, recent advances and innovative approaches.

    PubMed

    Tanzi, Maria Cristina; Farè, Silvia

    2009-09-01

    Adipose tissue is a highly specialized connective tissue found either in white or brown forms, the white form being the most abundant in adult humans. Loss or damage of white adipose tissue due to aging or pathological conditions needs reconstructive approaches. To date, two main strategies are being investigated for generating functional adipose tissue: autologous tissue/cell transplantation and adipose tissue engineering. Free-fat transplantation rarely achieves sufficient tissue augmentation owing to delayed neovascularization, with subsequent cell necrosis and graft volume shrinkage. Tissue engineering approaches represent, instead, a more suitable alternative for adipose tissue regeneration; they can be performed either with in situ or de novo adipogenesis. In situ adipogenesis or transplantation of encapsulated cells can be useful in healing small-volume defects, whereas restoration of large defects, where vascularization and a rapid volumetric gain are strict requirements, needs de novo strategies with 3D scaffold/filling matrix combinations. For adipose tissue engineering, the use of adult mesenchymal stem cells (both adipose- and bone marrow-derived stem cells) or of preadipocytes is preferred to the use of mature adipocytes, which have low expandability and poor ability for volume retention. This review intends to assemble and describe recent work on this topic, critically presenting successes obtained and drawbacks faced to date.

  19. Adipose tissue macrophages in non-rodent mammals: a comparative study.

    PubMed

    Ampem, Grace; Azegrouz, Hind; Bacsadi, Árpád; Balogh, Lajos; Schmidt, Susanne; Thuróczy, Julianna; Röszer, Tamás

    2016-02-01

    The stromal vascular fraction (SVF) of adipose tissue in rodents and primates contains mesenchymal stem cells and immune cells. SVF cells have complex metabolic, immune and endocrine functions with biomedical impact. However, in other mammals, the amount of data on SVF stem cells is negligible and whether the SVF hosts immune cells is unknown. In this study, we show that the SVF is rich in immune cells, with a dominance of adipose tissue macrophages (ATMs) in cattle (Bos primigenius taurus), domestic goat (Capra aegagrus hircus), domestic sheep (Ovis aries), domestic cat (Felis catus) and domestic dog (Canis familiaris). ATMs of these species are granulated lysosome-rich cells with lamellipodial protrusions and express the lysosome markers acid phosphatase 5 (ACP-5) and Mac-3/Lamp-2. Using ACP-5 and Mac-3/Lamp-2 as markers, we additionally detected ATMs in other species, such as the domestic horse (Equus ferus caballus), wild boar (Sus scrofa) and red fox (Vulpes vulpes). Feline and canine ATMs also express the murine macrophage marker F4/80 antigen. In the lean condition, the alternative macrophage activation marker CD206 is expressed by feline and canine ATMs and arginase-1 by feline ATMs. Obesity is associated with interleukin-6 and interferon gamma expression and with overt tyrosine nitration in both feline and canine ATMs. This resembles the obesity-induced phenotype switch of murine and human ATMs. Thus, we show, for the first time, that the presence of ATMs is a general trait of mammals. The interaction between the adipose cells and SVF immune cells might be evolutionarily conserved among mammals.

  20. Elevated autophagy gene expression in adipose tissue of obese humans: A potential non-cell-cycle-dependent function of E2F1

    PubMed Central

    Haim, Yulia; Blüher, Matthias; Slutsky, Noa; Goldstein, Nir; Klöting, Nora; Harman-Boehm, Ilana; Kirshtein, Boris; Ginsberg, Doron; Gericke, Martin; Guiu Jurado, Esther; Kovsan, Julia; Tarnovscki, Tanya; Kachko, Leonid; Bashan, Nava; Gepner, Yiftach; Shai, Iris; Rudich, Assaf

    2015-01-01

    Autophagy genes' expression is upregulated in visceral fat in human obesity, associating with obesity-related cardio-metabolic risk. E2F1 (E2F transcription factor 1) was shown in cancer cells to transcriptionally regulate autophagy. We hypothesize that E2F1 regulates adipocyte autophagy in obesity, associating with endocrine/metabolic dysfunction, thereby, representing non-cell-cycle function of this transcription factor. E2F1 protein (N=69) and mRNA (N=437) were elevated in visceral fat of obese humans, correlating with increased expression of ATG5 (autophagy-related 5), MAP1LC3B/LC3B (microtubule-associated protein 1 light chain 3 β), but not with proliferation/cell-cycle markers. Elevated E2F1 mainly characterized the adipocyte fraction, whereas MKI67 (marker of proliferation Ki-67) was elevated in the stromal-vascular fraction of adipose tissue. In human visceral fat explants, chromatin-immunoprecipitation revealed body mass index (BMI)-correlated increase in E2F1 binding to the promoter of MAP1LC3B, but not to the classical cell cycle E2F1 target, CCND1 (cyclin D1). Clinically, omental fat E2F1 expression correlated with insulin resistance, circulating free-fatty-acids (FFA), and with decreased circulating ADIPOQ/adiponectin, associations attenuated by adjustment for autophagy genes. Overexpression of E2F1 in HEK293 cells enhanced promoter activity of several autophagy genes and autophagic flux, and sensitized to further activation of autophagy by TNF. Conversely, mouse embryonic fibroblast (MEF)-derived adipocytes from e2f1 knockout mice (e2f1−/−) exhibited lower autophagy gene expression and flux, were more insulin sensitive, and secreted more ADIPOQ. Furthermore, e2f1−/− MEF-derived adipocytes, and autophagy-deficient (by Atg7 siRNA) adipocytes were resistant to cytokines-induced decrease in ADIPOQ secretion. Jointly, upregulated E2F1 sensitizes adipose tissue autophagy to inflammatory stimuli, linking visceral obesity to adipose and systemic

  1. Toward angiogenesis of implanted bio-artificial liver using scaffolds with type I collagen and adipose tissue-derived stem cells

    PubMed Central

    Lee, Jae Geun; Bak, Seon Young; Nahm, Ji Hae; Lee, Sang Woo; Min, Seon Ok

    2015-01-01

    Backgrounds/Aims Stem cell therapies for liver disease are being studied by many researchers worldwide, but scientific evidence to demonstrate the endocrinologic effects of implanted cells is insufficient, and it is unknown whether implanted cells can function as liver cells. Achieving angiogenesis, arguably the most important characteristic of the liver, is known to be quite difficult, and no practical attempts have been made to achieve this outcome. We carried out this study to observe the possibility of angiogenesis of implanted bio-artificial liver using scaffolds. Methods This study used adipose tissue-derived stem cells that were collected from adult patients with liver diseases with conditions similar to the liver parenchyma. Specifically, microfilaments were used to create an artificial membrane and maintain the structure of an artificial organ. After scratching the stomach surface of severe combined immunocompromised (SCID) mice (n=4), artificial scaffolds with adipose tissue-derived stem cells and type I collagen were implanted. Expression levels of angiogenesis markers including vascular endothelial growth factor (VEGF), CD34, and CD105 were immunohistochemically assessed after 30 days. Results Grossly, the artificial scaffolds showed adhesion to the stomach and surrounding organs; however, there was no evidence of angiogenesis within the scaffolds; and VEGF, CD34, and CD105 expressions were not detected after 30 days. Conclusions Although implantation of cells into artificial scaffolds did not facilitate angiogenesis, the artificial scaffolds made with type I collagen helped maintain implanted cells, and surrounding tissue reactions were rare. Our findings indicate that type I collagen artificial scaffolds can be considered as a possible implantable biomaterial. PMID:26155277

  2. Toward angiogenesis of implanted bio-artificial liver using scaffolds with type I collagen and adipose tissue-derived stem cells.

    PubMed

    Lee, Jae Geun; Bak, Seon Young; Nahm, Ji Hae; Lee, Sang Woo; Min, Seon Ok; Kim, Kyung Sik

    2015-05-01

    Stem cell therapies for liver disease are being studied by many researchers worldwide, but scientific evidence to demonstrate the endocrinologic effects of implanted cells is insufficient, and it is unknown whether implanted cells can function as liver cells. Achieving angiogenesis, arguably the most important characteristic of the liver, is known to be quite difficult, and no practical attempts have been made to achieve this outcome. We carried out this study to observe the possibility of angiogenesis of implanted bio-artificial liver using scaffolds. This study used adipose tissue-derived stem cells that were collected from adult patients with liver diseases with conditions similar to the liver parenchyma. Specifically, microfilaments were used to create an artificial membrane and maintain the structure of an artificial organ. After scratching the stomach surface of severe combined immunocompromised (SCID) mice (n=4), artificial scaffolds with adipose tissue-derived stem cells and type I collagen were implanted. Expression levels of angiogenesis markers including vascular endothelial growth factor (VEGF), CD34, and CD105 were immunohistochemically assessed after 30 days. Grossly, the artificial scaffolds showed adhesion to the stomach and surrounding organs; however, there was no evidence of angiogenesis within the scaffolds; and VEGF, CD34, and CD105 expressions were not detected after 30 days. Although implantation of cells into artificial scaffolds did not facilitate angiogenesis, the artificial scaffolds made with type I collagen helped maintain implanted cells, and surrounding tissue reactions were rare. Our findings indicate that type I collagen artificial scaffolds can be considered as a possible implantable biomaterial.

  3. Cell-specific dysregulation of microRNA expression in obese white adipose tissue.

    PubMed

    Oger, Frédérik; Gheeraert, Celine; Mogilenko, Denis; Benomar, Yacir; Molendi-Coste, Olivier; Bouchaert, Emmanuel; Caron, Sandrine; Dombrowicz, David; Pattou, François; Duez, Hélène; Eeckhoute, Jérome; Staels, Bart; Lefebvre, Philippe

    2014-08-01

    Obesity is characterized by the excessive accumulation of dysfunctional white adipose tissue (WAT), leading to a strong perturbation of metabolic regulations. However, the molecular events underlying this process are not fully understood. MicroRNAs (miRNAs) are small noncoding RNAs acting as posttranscriptional regulators of gene expression in multiple tissues and organs. However, their expression and roles in WAT cell subtypes, which include not only adipocytes but also immune, endothelial, and mesenchymal stem cells as well as preadipocytes, have not been characterized. Design/Results: By applying differential miRNome analysis, we demonstrate that the expression of several miRNAs is dysregulated in epididymal WAT from ob/ob and high-fat diet-fed mice. Adipose tissue-specific down-regulation of miR-200a and miR-200b and the up-regulation of miR-342-3p, miR-335-5p, and miR-335-3p were observed. Importantly, a similarly altered expression of miR-200a and miR-200b was observed in obese diabetic patients. Furthermore, cell fractionation of mouse adipose tissue revealed that miRNAs are differentially expressed in adipocytes and in subpopulations from the stromal vascular fraction. Finally, integration of transcriptomic data showed that bioinformatically predicted miRNA target genes rarely showed anticorrelated expression with that of targeting miRNA, in contrast to experimentally validated target genes. Taken together, our data indicate that the dysregulated expression of miRNAs occurs in distinct cell types and is likely to affect cell-specific function(s) of obese WAT.

  4. Intracoronary Administration of Allogeneic Adipose Tissue-Derived Mesenchymal Stem Cells Improves Myocardial Perfusion But Not Left Ventricle Function, in a Translational Model of Acute Myocardial Infarction.

    PubMed

    Bobi, Joaquim; Solanes, Núria; Fernández-Jiménez, Rodrigo; Galán-Arriola, Carlos; Dantas, Ana Paula; Fernández-Friera, Leticia; Gálvez-Montón, Carolina; Rigol-Monzó, Elisabet; Agüero, Jaume; Ramírez, José; Roqué, Mercè; Bayés-Genís, Antoni; Sánchez-González, Javier; García-Álvarez, Ana; Sabaté, Manel; Roura, Santiago; Ibáñez, Borja; Rigol, Montserrat

    2017-05-03

    Autologous adipose tissue-derived mesenchymal stem cells (ATMSCs) therapy is a promising strategy to improve post-myocardial infarction outcomes. In a porcine model of acute myocardial infarction, we studied the long-term effects and the mechanisms involved in allogeneic ATMSCs administration on myocardial performance. Thirty-eight pigs underwent 50 minutes of coronary occlusion; the study was completed in 33 pigs. After reperfusion, allogeneic ATMSCs or culture medium (vehicle) were intracoronarily administered. Follow-ups were performed at short (2 days after acute myocardial infarction vehicle-treated, n=10; ATMSCs-treated, n=9) or long term (60 days after acute myocardial infarction vehicle-treated, n=7; ATMSCs-treated, n=7). At short term, infarcted myocardium analysis showed reduced apoptosis in the ATMSCs-treated animals (48.6±6% versus 55.9±5.7% in vehicle; P =0.017); enhancement of the reparative process with up-regulated vascular endothelial growth factor, granulocyte macrophage colony-stimulating factor, and stromal-derived factor-1α gene expression; and increased M2 macrophages (67.2±10% versus 54.7±10.2% in vehicle; P =0.016). In long-term groups, increase in myocardial perfusion at the anterior infarct border was observed both on day-7 and day-60 cardiac magnetic resonance studies in ATMSCs-treated animals, compared to vehicle (87.9±28.7 versus 57.4±17.7 mL/min per gram at 7 days; P =0.034 and 99±22.6 versus 43.3±14.7 22.6 mL/min per gram at 60 days; P =0.0001, respectively). At day 60, higher vascular density was detected at the border zone in the ATMSCs-treated animals (118±18 versus 92.4±24.3 vessels/mm 2 in vehicle; P =0.045). Cardiac magnetic resonance-measured left ventricular ejection fraction of left ventricular volumes was not different between groups at any time point. In this porcine acute myocardial infarction model, allogeneic ATMSCs-based therapy was associated with increased cardioprotective and reparative

  5. Mesenchymal Stem Cells: New Players in Retinopathy Therapy

    PubMed Central

    Rajashekhar, Gangaraju

    2014-01-01

    Retinopathies in human and animal models have shown to occur through loss of pericytes resulting in edema formation, excessive immature retinal angiogenesis, and neuronal apoptosis eventually leading to blindness. In recent years, the concept of regenerating terminally differentiated organs with a cell-based therapy has evolved. The cells used in these approaches are diverse and include tissue-specific endogenous stem cells, endothelial progenitor (EPC), embryonic stem cells, induced pluripotent stem cells (iPSC) and mesenchymal stem cells (MSC). Recently, MSC derived from the stromal fraction of adipose tissue have been shown to possess pluripotent differentiation potential in vitro. These adipose stromal cells (ASC) have been differentiated in a number of laboratories to osteogenic, myogenic, vascular, and adipocytic cell phenotypes. In vivo, ASC have been shown to have functional and phenotypic overlap with pericytes lining microvessels in adipose tissues. Furthermore, these cells either in paracrine mode or physical proximity with endothelial cells, promoted angiogenesis, improved ischemia–reperfusion, protected from myocardial infarction, and were neuroprotective. Owing to the easy isolation procedure and abundant supply, fat-derived ASC are a more preferred source of autologous mesenchymal cells compared to bone marrow MSC. In this review, we present evidence that these readily available ASC from minimally invasive liposuction will facilitate translation of ASC research into patients with retinal diseases in the near future. PMID:24795699

  6. Depressed levels of prostaglandin F2α in mice lacking Akr1b7 increase basal adiposity and predispose to diet-induced obesity.

    PubMed

    Volat, Fanny E; Pointud, Jean-Christophe; Pastel, Emilie; Morio, Béatrice; Sion, Benoit; Hamard, Ghislaine; Guichardant, Michel; Colas, Romain; Lefrançois-Martinez, Anne-Marie; Martinez, Antoine

    2012-11-01

    Negative regulators of white adipose tissue (WAT) expansion are poorly documented in vivo. Prostaglandin F(2α) (PGF(2α)) is a potent antiadipogenic factor in cultured preadipocytes, but evidence for its involvement in physiological context is lacking. We previously reported that Akr1b7, an aldo-keto reductase enriched in adipose stromal vascular fraction but absent from mature adipocytes, has antiadipogenic properties possibly supported by PGF(2α) synthase activity. To test whether lack of Akr1b7 could influence WAT homeostasis in vivo, we generated Akr1b7(-/-) mice in 129/Sv background. Akr1b7(-/-) mice displayed excessive basal adiposity resulting from adipocyte hyperplasia/hypertrophy and exhibited greater sensitivity to diet-induced obesity. Following adipose enlargement and irrespective of the diet, they developed liver steatosis and progressive insulin resistance. Akr1b7 loss was associated with decreased PGF(2α) WAT contents. Cloprostenol (PGF(2α) agonist) administration to Akr1b7(-/-) mice normalized WAT expansion by affecting both de novo adipocyte differentiation and size. Treatment of 3T3-L1 adipocytes and Akr1b7(-/-) mice with cloprostenol suggested that decreased adipocyte size resulted from inhibition of lipogenic gene expression. Hence, Akr1b7 is a major regulator of WAT development through at least two PGF(2α)-dependent mechanisms: inhibition of adipogenesis and lipogenesis. These findings provide molecular rationale to explore the status of aldo-keto reductases in dysregulations of adipose tissue homeostasis.

  7. Characterization of human adipose tissue-derived stem cells in vitro culture and in vivo differentiation in a temperature-sensitive chitosan/β- glycerophosphate/collagen hybrid hydrogel.

    PubMed

    Song, Kedong; Li, Liying; Yan, Xinyu; Zhang, Wen; Zhang, Yu; Wang, Yiwei; Liu, Tianqing

    2017-01-01

    In this study, the interaction of human adipose tissue-derived stem cells (ADSCs) with chitosan/β-glycerophosphate/collagen (C/GP/Co) hybrid hydrogel was test, followed by investigating the capability of engineered adipose tissue formation using this ADSCs seeded hydrogel. The ADSCs were harvested and mixed with a C/GP/Co hydrogel followed by a gelation at 37°C and an in vitro culture. The results showed that the ADSCs within C/GP/Co hydrogels achieved a 30% of expansion over 7days in culture medium and encapsulated cell in C/GP/Co hydrogel demonstrated a characteristic morphology with high viability over 5days. C/GP/Co hydrogel were subcutaneous injected into SD-rats to assess the biocompatibility. The induced ADSCs-C/GP/Co hydrogel and non-induced ADSCs-C/GP/Co hydrogel were subcutaneously injected into nude mice for detecting potential of adipogenic differentiation. It has shown that C/GP/Co hydrogel were well tolerated in SD rats where they had persisted over 4weeks post implantation. Histology analysis indicated that induced ADSCs-C/GP/Co hydrogel has a greater number of adipocytes and vascularized adipose tissues compared with non-induced ADSCs-C/GP/Co hydrogel. Crown Copyright © 2016. Published by Elsevier B.V. All rights reserved.

  8. Adipose-derived Stem Cells Stimulated with n-Butylidenephthalide Exhibit Therapeutic Effects in a Mouse Model of Parkinson's Disease.

    PubMed

    Chi, Kang; Fu, Ru-Huei; Huang, Yu-Chuen; Chen, Shih-Yin; Hsu, Ching-Ju; Lin, Shinn-Zong; Tu, Chi-Tang; Chang, Li-Hsun; Wu, Ping-An; Liu, Shih-Ping

    2018-03-01

    Parkinson's disease (PD) causes motor dysfunction and dopaminergic cell death. Drug treatments can effectively reduce symptoms but often cause unwanted side effects. Stem cell therapies using cell replacement or indirect beneficial secretomes have recently emerged as potential therapeutic strategies. Although various types of stem cells have been proposed as possible candidates, adipose-derived stem cells (ADSCs) are easily obtainable, more abundant, less ethically disputed, and able to differentiate into multiple cell lineages. However, treatment of PD using adult stem cells is known to be less efficacious than neuron or embryonic stem cell transplantation. Therefore, improved therapies are urgently needed. n-Butylidenephthalide (BP), which is extracted from Angelica sinensis, has been shown to have anti-inflammatory and neuroprotective effects. Indeed, we previously demonstrated that BP treatment of ADSCs enhances the expression of neurogenesis and homing factors such as nuclear receptor related 1 protein, stromal-derived factor 1, and brain-derived neurotrophic factor. In the present study, we examined the ability of BP-pretreated ADSC transplantation to improve PD motor symptoms and protect dopamine neurons in a mouse model of PD. We evaluated the results using neuronal behavior tests such as beam walking, rotarod, and locomotor activity tests. ADSCs with or without BP pretreatment were transplanted into the striatum. Our findings demonstrated that ADSC transplantation improved motor abilities with varied efficacies and that BP stimulation improved the therapeutic effects of transplantation. Dopaminergic cell numbers returned to normal in ADSC-transplanted mice after 22 d. In summary, stimulating ADSCs with BP improved PD recovery efficiency. Thus, our results provide important new strategies to improve stem cell therapies for neurodegenerative diseases in future studies.

  9. Tissue Source and Cell Expansion Condition Influence Phenotypic Changes of Adipose-Derived Stem Cells

    PubMed Central

    Mangum, Lauren H.; Stone, Randolph; Wrice, Nicole L.; Larson, David A.; Florell, Kyle F.; Christy, Barbara A.; Herzig, Maryanne C.; Cap, Andrew P.

    2017-01-01

    Stem cells derived from the subcutaneous adipose tissue of debrided burned skin represent an appealing source of adipose-derived stem cells (ASCs) for regenerative medicine. Traditional tissue culture uses fetal bovine serum (FBS), which complicates utilization of ASCs in human medicine. Human platelet lysate (hPL) is one potential xeno-free, alternative supplement for use in ASC culture. In this study, adipogenic and osteogenic differentiation in media supplemented with 10% FBS or 10% hPL was compared in human ASCs derived from abdominoplasty (HAP) or from adipose associated with debrided burned skin (BH). Most (95–99%) cells cultured in FBS were stained positive for CD73, CD90, CD105, and CD142. FBS supplementation was associated with increased triglyceride content and expression of adipogenic genes. Culture in hPL significantly decreased surface staining of CD105 by 31% and 48% and CD142 by 27% and 35% in HAP and BH, respectively (p < 0.05). Culture of BH-ASCs in hPL also increased expression of markers of osteogenesis and increased ALP activity. These data indicate that application of ASCs for wound healing may be influenced by ASC source as well as culture conditions used to expand them. As such, these factors must be taken into consideration before ASCs are used for regenerative purposes. PMID:29138638

  10. Kallikrein-related peptidase 4 induces cancer-associated fibroblast features in prostate-derived stromal cells.

    PubMed

    Kryza, Thomas; Silva, Lakmali M; Bock, Nathalie; Fuhrman-Luck, Ruth A; Stephens, Carson R; Gao, Jin; Samaratunga, Hema; Lawrence, Mitchell G; Hooper, John D; Dong, Ying; Risbridger, Gail P; Clements, Judith A

    2017-10-01

    The reciprocal communication between cancer cells and their microenvironment is critical in cancer progression. Although involvement of cancer-associated fibroblasts (CAF) in cancer progression is long established, the molecular mechanisms leading to differentiation of CAFs from normal fibroblasts are poorly understood. Here, we report that kallikrein-related peptidase-4 (KLK4) promotes CAF differentiation. KLK4 is highly expressed in prostate epithelial cells of premalignant (prostatic intraepithelial neoplasia) and malignant lesions compared to normal prostate epithelia, especially at the peristromal interface. KLK4 induced CAF-like features in the prostate-derived WPMY1 normal stromal cell line, including increased expression of alpha-smooth muscle actin, ESR1 and SFRP1. KLK4 activated protease-activated receptor-1 in WPMY1 cells increasing expression of several factors (FGF1, TAGLN, LOX, IL8, VEGFA) involved in prostate cancer progression. In addition, KLK4 induced WPMY1 cell proliferation and secretome changes, which in turn stimulated HUVEC cell proliferation that could be blocked by a VEGFA antibody. Importantly, the genes dysregulated by KLK4 treatment of WPMY1 cells were also differentially expressed between patient-derived CAFs compared to matched nonmalignant fibroblasts and were further increased by KLK4 treatment. Taken together, we propose that epithelial-derived KLK4 promotes tumour progression by actively promoting CAF differentiation in the prostate stromal microenvironment. © 2017 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.

  11. Inorganic Arsenic–Related Changes in the Stromal Tumor Microenvironment in a Prostate Cancer Cell–Conditioned Media Model

    PubMed Central

    Shearer, Joseph J.; Wold, Eric A.; Umbaugh, Charles S.; Lichti, Cheryl F.; Nilsson, Carol L.; Figueiredo, Marxa L.

    2015-01-01

    Background: The tumor microenvironment plays an important role in the progression of cancer by mediating stromal–epithelial paracrine signaling, which can aberrantly modulate cellular proliferation and tumorigenesis. Exposure to environmental toxicants, such as inorganic arsenic (iAs), has also been implicated in the progression of prostate cancer. Objective: The role of iAs exposure in stromal signaling in the tumor microenvironment has been largely unexplored. Our objective was to elucidate molecular mechanisms of iAs-induced changes to stromal signaling by an enriched prostate tumor microenvironment cell population, adipose-derived mesenchymal stem/stromal cells (ASCs). Results: ASC-conditioned media (CM) collected after 1 week of iAs exposure increased prostate cancer cell viability, whereas CM from ASCs that received no iAs exposure decreased cell viability. Cytokine array analysis suggested changes to cytokine signaling associated with iAs exposure. Subsequent proteomic analysis suggested a concentration-dependent alteration to the HMOX1/THBS1/TGFβ signaling pathway by iAs. These results were validated by quantitative reverse transcriptase–polymerase chain reaction (RT-PCR) and Western blotting, confirming a concentration-dependent increase in HMOX1 and a decrease in THBS1 expression in ASC following iAs exposure. Subsequently, we used a TGFβ pathway reporter construct to confirm a decrease in stromal TGFβ signaling in ASC following iAs exposure. Conclusions: Our results suggest a concentration-dependent alteration of stromal signaling: specifically, attenuation of stromal-mediated TGFβ signaling following exposure to iAs. Our results indicate iAs may enhance prostate cancer cell viability through a previously unreported stromal-based mechanism. These findings indicate that the stroma may mediate the effects of iAs in tumor progression, which may have future therapeutic implications. Citation: Shearer JJ, Wold EA, Umbaugh CS, Lichti CF, Nilsson CL

  12. Role of adipose-derived stem cells in wound healing.

    PubMed

    Hassan, Waqar Ul; Greiser, Udo; Wang, Wenxin

    2014-01-01

    Impaired wound healing remains a challenge to date and causes debilitating effects with tremendous suffering. Recent advances in tissue engineering approaches in the area of cell therapy have provided promising treatment options to meet the challenges of impaired skin wound healing such as diabetic foot ulcers. Over the last few years, stem cell therapy has emerged as a novel therapeutic approach for various diseases including wound repair and tissue regeneration. Several different types of stem cells have been studied in both preclinical and clinical settings such as bone marrow-derived stem cells, adipose-derived stem cells (ASCs), circulating angiogenic cells (e.g., endothelial progenitor cells), human dermal fibroblasts, and keratinocytes for wound healing. Adipose tissue is an abundant source of mesenchymal stem cells, which have shown an improved outcome in wound healing studies. ASCs are pluripotent stem cells with the ability to differentiate into different lineages and to secrete paracrine factors initiating tissue regeneration process. The abundant supply of fat tissue, ease of isolation, extensive proliferative capacities ex vivo, and their ability to secrete pro-angiogenic growth factors make them an ideal cell type to use in therapies for the treatment of nonhealing wounds. In this review, we look at the pathogenesis of chronic wounds, role of stem cells in wound healing, and more specifically look at the role of ASCs, their mechanism of action and their safety profile in wound repair and tissue regeneration. © 2014 by the Wound Healing Society.

  13. Adipose-derived adult stem cells: available technologies for potential clinical regenerative applications in dentistry.

    PubMed

    Catalano, Enrico; Cochis, Andrea; Varoni, Elena; Rimondini, Lia; Carrassi, Antonio; Azzimonti, Barbara

    2013-01-01

    Tissue homeostasis depends closely on the activity and welfare of adult stem cells. These cells represent a promising tool for biomedical research since they can aid in treatment and promote the regeneration of damaged organs in many human disorders. Adult stem cells indefinitely preserve their ability to self-renew and differentiate into various phenotypes; this capacity could be promoted in vitro by particular culture conditions (differentiation media) or spontaneously induced in vivo by exploiting the biochemical and mechanical properties of the tissue in which the stem cells are implanted. Among the different sources of adult stem cells, adipose tissue is an attractive possibility thanks to its ready availability and the standard extraction techniques at our disposal today. This review discusses the isolation, characterization, and differentiation of human adipose-derived adult stem cells, as well as regeneration strategies, therapeutic uses, and adverse effects of their delivery. In particular, since oral disorders (e.g., trauma, erosion, and chronic periodontitis) often cause the loss of dental tissue along with functional, phonetic, and aesthetic impairment, this review focuses on the application of human adipose-derived adult stem cells, alone or in combination with biomaterials, in treating oral diseases.

  14. Extracellular vesicles derived from mesenchymal stromal cells: a therapeutic option in respiratory diseases?

    PubMed

    Abreu, Soraia C; Weiss, Daniel J; Rocco, Patricia R M

    2016-04-14

    Extracellular vesicles (EVs) are plasma membrane-bound fragments released from several cell types, including mesenchymal stromal cells (MSCs), constitutively or under stimulation. EVs derived from MSCs and other cell types transfer molecules (such as DNA, proteins/peptides, mRNA, microRNA, and lipids) and/or organelles with reparative and anti-inflammatory properties to recipient cells. The paracrine anti-inflammatory effects promoted by MSC-derived EVs have attracted significant interest in the regenerative medicine field, including for potential use in lung injuries. In the present review, we describe the characteristics, biological activities, and mechanisms of action of MSC-derived EVs. We also review the therapeutic potential of EVs as reported in relevant preclinical models of acute and chronic respiratory diseases, such as pneumonia, acute respiratory distress syndrome, asthma, and pulmonary arterial hypertension. Finally, we discuss possible approaches for potentiating the therapeutic effects of MSC-derived EVs so as to enable use of this therapy in clinical practice.

  15. [Construction of injectable tissue engineered adipose tissue with fibrin glue scaffold and human adipose-derived stem cells transfected by lentivirus vector expressing hepatocyte growth factor].

    PubMed

    Zhu, Yuanzheng; Yi, Yangyan; Yang, Shuifa; Zhang, Jing; Wu, Shu; Wang, Zhaohui

    2017-09-01

    To discuss the possibility of constructing injectable tissue engineered adipose tissue, and to provide a new approach for repairing soft tissue defects. Human adipose-derived stem cells (hADSCs) were extracted from the lipid part of human liposuction aspirate by enzymatic digestion and identified by morphological observation, flow cytometry, and adipogenic induction. The hADSCs underwent transfection by lentivirus vector expressing hepatocyte growth factor and green fluorescent protein (HGF-GFP-LVs) of different multiplicity of infection (MOI, 10, 30, 50, and 100), the transfection efficiency was calculated to determine the optimum MOI. The hADSCs transfected by HGF-GFP-LVs of optimal MOI and being adipogenic inducted were combined with injectable fibrin glue scaffold, and were injected subcutaneously into the right side of the low back of 10 T-cell deficiency BALB/c female nude mice (transfected group); non-HGF-GFP-LVs transfected hADSCs (being adipogenic inducted) combined with injectable fibrin glue scaffold were injected subcutaneously into the left side of the low back (untransfected group); and injectable fibrin glue scaffold were injected subcutaneously into the middle part of the neck (blank control group); 0.4 mL at each point. Twelve weeks later the mice were killed and the implants were taken out. Gross observation, wet weight measurement, HE staining, GFP fluorescence labeling, and immunofluorescence staining were performed to assess the in vivo adipogenic ability of the seed cells and the neovascularization of the grafts. The cultured cells were identified as hADSCs. Poor transfection efficiency was observed in MOI of 10 and 30, the transfection efficiency of MOI of 50 and 100 was more than 80%, so the optimum MOI was 50. Adipose tissue-like new-born tissues were found in the injection sites of the transfected and untransfected groups after 12 weeks of injection, and no new-born tissues was found in the blank control group. The wet-weight of new

  16. Regenerative Repair of Damaged Meniscus with Autologous Adipose Tissue-Derived Stem Cells

    PubMed Central

    Pak, Jaewoo; Lee, Jung Hun; Lee, Sang Hee

    2014-01-01

    Mesenchymal stem cells (MSCs) are defined as pluripotent cells found in numerous human tissues, including bone marrow and adipose tissue. Such MSCs, isolated from bone marrow and adipose tissue, have been shown to differentiate into bone and cartilage, along with other types of tissues. Therefore, MSCs represent a promising new therapy in regenerative medicine. The initial treatment of meniscus tear of the knee is managed conservatively with nonsteroidal anti-inflammatory drugs and physical therapy. When such conservative treatment fails, an arthroscopic resection of the meniscus is necessary. However, the major drawback of the meniscectomy is an early onset of osteoarthritis. Therefore, an effective and noninvasive treatment for patients with continuous knee pain due to damaged meniscus has been sought. Here, we present a review, highlighting the possible regenerative mechanisms of damaged meniscus with MSCs (especially adipose tissue-derived stem cells (ASCs)), along with a case of successful repair of torn meniscus with significant reduction of knee pain by percutaneous injection of autologous ASCs into an adult human knee. PMID:24592390

  17. Parallels in Immunometabolic Adipose Tissue Dysfunction with Ageing and Obesity

    PubMed Central

    Trim, William; Turner, James E.; Thompson, Dylan

    2018-01-01

    Ageing, like obesity, is often associated with alterations in metabolic and inflammatory processes resulting in morbidity from diseases characterised by poor metabolic control, insulin insensitivity, and inflammation. Ageing populations also exhibit a decline in immune competence referred to as immunosenescence, which contributes to, or might be driven by chronic, low-grade inflammation termed “inflammageing”. In recent years, animal and human studies have started to uncover a role for immune cells within the stromal fraction of adipose tissue in driving the health complications that come with obesity, but relatively little work has been conducted in the context of immunometabolic adipose function in ageing. It is now clear that aberrant immune function within adipose tissue in obesity—including an accumulation of pro-inflammatory immune cell populations—plays a major role in the development of systemic chronic, low-grade inflammation, and limiting the function of adipocytes leading to an impaired fat handling capacity. As a consequence, these changes increase the chance of multiorgan dysfunction and disease onset. Considering the important role of the immune system in obesity-associated metabolic and inflammatory diseases, it is critically important to further understand the interplay between immunological processes and adipose tissue function, establishing whether this interaction contributes to age-associated immunometabolic dysfunction and inflammation. Therefore, the aim of this article is to summarise how the interaction between adipose tissue and the immune system changes with ageing, likely contributing to the age-associated increase in inflammatory activity and loss of metabolic control. To understand the potential mechanisms involved, parallels will be drawn to the current knowledge derived from investigations in obesity. We also highlight gaps in research and propose potential future directions based on the current evidence. PMID:29479350

  18. Mesenchymal Stromal Cells form Vascular Tubes when Placed in Fibrin Sealant and Accelerate Wound Healing in Vivo

    PubMed Central

    Mendez, Julio J.; Ghaedi, Mahboobe; Sivarapatna, Amogh; Dimitrievska, Sashka; Shao, Zhen; Osuji, Chinedum; Steinbacher, Derek M.; Leffell, David J.; Niklason, Laura E.

    2014-01-01

    Non-healing, chronic wounds are a growing public health problem and may stem from insufficient angiogenesis in affected sites. Here, we have developed a fibrin formulation that allows adipose-derived mesenchymal stromal cells (ADSCs) to form tubular structures in vitro. The tubular structures express markers of endothelium, including CD31 and VE-Cadherin, as well as the pericyte marker NG2. The ability for the MSCs to form tubular structures within the fibrin gels was directly dependent on the stoichiometric ratios of thrombin and fibrinogen and the resulting gel concentration, as well as on the presence of bFGF. Fibrin gel formulations that varied in stiffness were tested. ADSCs that are embedded in a stiff fibrin formulation express VE-cadherin and CD31 as shown by PCR, FACS and immunostaining. Confocal imaging analysis demonstrated that tubular structures formed, containing visible lumens, in the stiff fibrin gels in vitro. There was also a difference in the amounts of bFGF secreted by ADSCs grown in the stiffer gels as compared to softer gels. Additionally, hAT-MSCs gave rise to perfusable vessels that were VE-cadherin positive after subcutaneous injection into mice, whereas the softer fibrin formulation containing ADSCs did not. The application of ADSCs delivered in the stiff fibrin gels allowed for the wounds to heal more quickly, as assessed by wound size, amount of granulation tissue and collagen content. Interestingly, following 5 days of healing, the ADSCs remained within the fibrin gel and did not integrate into the granulation tissue of healing wounds in vivo. These data show that ADSCs are able to form tubular structures within fibrin gels, and may also contribute to faster wound healing, as compared with no treatment or to wounds treated with fibrin gels devoid of ADSCs. PMID:25433608

  19. Adipose Tissue Angiogenesis: Impact on Obesity and Type-2 Diabetes

    PubMed Central

    Corvera, Silvia; Gealekman, Olga

    2013-01-01

    The growth and function of tissues is critically dependent on their vascularization. Adipose tissue is capable of expanding many-fold during adulthood, therefore requiring the formation of new vasculature to supply growing and proliferating adipocytes. The expansion of the vasculature in adipose tissue occurs through angiogenesis, where new blood vessels develop from those pre-existing within the tissue. Inappropriate angiogenesis may underlie adipose tissue dysfunction in obesity, which in turn increases type-2 diabetes risk. In addition, genetic and developmental factors involved in vascular patterning may define the size and expandability of diverse adipose tissue depots, which are also associated with type-2 diabetes risk. Moreover, the adipose tissue vasculature appears to be the niche for pre-adipocyte precursors, and factors that affect angiogenesis may directly impact the generation of new adipocytes. Here we review recent advances on the basic mechanisms of angiogenesis, and on the role of angiogenesis in adipose tissue development and obesity. A substantial amount of data point to a deficit in adipose tissue angiogenesis as a contributing factor to insulin resistance and metabolic disease in obesity. These emerging findings support the concept of the adipose tissue vasculature as a source of new targets for metabolic disease therapies. PMID:23770388

  20. Anti-aging effect of adipose-derived stem cells in a mouse model of skin aging induced by D-galactose.

    PubMed

    Zhang, Shengchang; Dong, Ziqing; Peng, Zhangsong; Lu, Feng

    2014-01-01

    Glycation products accumulate during aging of slowly renewing tissue, including skin, and are suggested as an important mechanism underlying the skin aging process. Adipose-derived cells are widely used in the clinic to treat ischemic diseases and enhance wound healing. Interestingly, adipose-derived stem cells (ASCs) are also effective in anti-aging therapy, although the mechanism underlying their effects remains unknown. The purpose of the present study was to examine the anti-aging effect of ASCs in a D-galactose-induced aging animal model and to clarify the underlying mechanism. Six-week-old nude mice were subcutaneously injected with D-gal daily for 8 weeks. Two weeks after completion of treatment, mice were randomized to receive subcutaneous injections of 106 green fluorescent protein (GFP)-expressing ASCs, aminoguanidine (AG) or phosphate-buffered saline (PBS). Control mice received no treatment. We examined tissue histology and determined the activity of senescence-associated molecular markers such as superoxide dismutase (SOD) and malondialdehyde (MDA). Transplanted ASCs were detectable for 14 days and their GFP signal disappeared at day 28 after injection. ASCs inhibited advanced glycation end product (AGE) levels in our animal model as well as increased the SOD level and decreased the MDA level, all of which act to reverse the aging phenotype in a similar way to AG, an inhibitor of AGE formation. Furthermore, ASCs released angiogenic factors in vivo such as vascular endothelial growth factor, suggesting a skin trophic effect. These results demonstrate that ASCs may contribute to the regeneration of skin during aging. In addition, the data shows that ASCs provide a functional benefit by glycation suppression, antioxidation, and trophic effects in a mouse model of aging.

  1. Tumor necrosis a regulation of adipokine gene expression in neonatal adipose tissue

    USDA-ARS?s Scientific Manuscript database

    The neonatal period is also a time of significant stress and susceptibility to infection, conditions which favor the secretion of tumor necrosis a. The present study was designed to determine if TNFa can alter adipokine gene expression within the adipose tissue of neonatal swine. Primary stromal v...

  2. Cultured Human Adipose Tissue Pericytes and Mesenchymal Stromal Cells Display a Very Similar Gene Expression Profile

    PubMed Central

    Malta, Tathiane Maistro; de Deus Wagatsuma, Virgínia Mara; Palma, Patrícia Viana Bonini; Araújo, Amélia Goes; Ribeiro Malmegrim, Kelen Cristina; Morato de Oliveira, Fábio; Panepucci, Rodrigo Alexandre; Silva, Wilson Araújo; Kashima Haddad, Simone; Covas, Dimas Tadeu

    2015-01-01

    Mesenchymal stromal cells (MSCs) are cultured cells that can give rise to mature mesenchymal cells under appropriate conditions and secrete a number of biologically relevant molecules that may play an important role in regenerative medicine. Evidence indicates that pericytes (PCs) correspond to mesenchymal stem cells in vivo and can give rise to MSCs when cultured, but a comparison between the gene expression profiles of cultured PCs (cPCs) and MSCs is lacking. We have devised a novel methodology to isolate PCs from human adipose tissue and compared cPCs to MSCs obtained through traditional methods. Freshly isolated PCs expressed CD34, CD140b, and CD271 on their surface, but not CD146. Both MSCs and cPCs were able to differentiate along mesenchymal pathways in vitro, displayed an essentially identical surface immunophenotype, and exhibited the ability to suppress CD3+ lymphocyte proliferation in vitro. Microarray expression data of cPCs and MSCs formed a single cluster among other cell types. Further analyses showed that the gene expression profiles of cPCs and MSCs are extremely similar, although MSCs differentially expressed endothelial cell (EC)-specific transcripts. These results confirm, using the power of transcriptomic analysis, that PCs give rise to MSCs and suggest that low levels of ECs may persist in MSC cultures established using traditional protocols. PMID:26192741

  3. Adipose-Derived Mesenchymal Stem Cell Administration Does Not Improve Corneal Graft Survival Outcome

    PubMed Central

    Fuentes-Julián, Sherezade; Arnalich-Montiel, Francisco; Jaumandreu, Laia; Leal, Marina; Casado, Alfonso; García-Tuñon, Ignacio; Hernández-Jiménez, Enrique; López-Collazo, Eduardo; De Miguel, Maria P.

    2015-01-01

    The effect of local and systemic injections of mesenchymal stem cells derived from adipose tissue (AD-MSC) into rabbit models of corneal allograft rejection with either normal-risk or high-risk vascularized corneal beds was investigated. The models we present in this study are more similar to human corneal transplants than previously reported murine models. Our aim was to prevent transplant rejection and increase the length of graft survival. In the normal-risk transplant model, in contrast to our expectations, the injection of AD-MSC into the graft junction during surgery resulted in the induction of increased signs of inflammation such as corneal edema with increased thickness, and a higher level of infiltration of leukocytes. This process led to a lower survival of the graft compared with the sham-treated corneal transplants. In the high-risk transplant model, in which immune ocular privilege was undermined by the induction of neovascularization prior to graft surgery, we found the use of systemic rabbit AD-MSCs prior to surgery, during surgery, and at various time points after surgery resulted in a shorter survival of the graft compared with the non-treated corneal grafts. Based on our results, local or systemic treatment with AD-MSCs to prevent corneal rejection in rabbit corneal models at normal or high risk of rejection does not increase survival but rather can increase inflammation and neovascularization and break the innate ocular immune privilege. This result can be partially explained by the immunomarkers, lack of immunosuppressive ability and immunophenotypical secretion molecules characterization of AD-MSC used in this study. Parameters including the risk of rejection, the inflammatory/vascularization environment, the cell source, the time of injection, the immunosuppression, the number of cells, and the mode of delivery must be established before translating the possible benefits of the use of MSCs in corneal transplants to clinical practice. PMID

  4. RPL13A and EEF1A1 Are Suitable Reference Genes for qPCR during Adipocyte Differentiation of Vascular Stromal Cells from Patients with Different BMI and HOMA-IR.

    PubMed

    Gentile, Adriana-Mariel; Lhamyani, Said; Coín-Aragüez, Leticia; Oliva-Olivera, Wilfredo; Zayed, Hatem; Vega-Rioja, Antonio; Monteseirin, Javier; Romero-Zerbo, Silvana-Yanina; Tinahones, Francisco-José; Bermúdez-Silva, Francisco-Javier; El Bekay, Rajaa

    2016-01-01

    Real-time or quantitative PCR (qPCR) is a useful technique that requires reliable reference genes for data normalization in gene expression analysis. Adipogenesis is among the biological processes suitable for this technique. The selection of adequate reference genes is essential for qPCR gene expression analysis of human Vascular Stromal Cells (hVSCs) during their differentiation into adipocytes. To the best of our knowledge, there are no studies validating reference genes for the analyses of visceral and subcutaneous adipose tissue hVSCs from subjects with different Body Mass Index (BMI) and Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) index. The present study was undertaken to analyze this question. We first analyzed the stability of expression of five potential reference genes: CYC, GAPDH, RPL13A, EEF1A1, and 18S ribosomal RNA, during in vitro adipogenic differentiation, in samples from these types of patients. The expression of RPL13A and EEF1A1 was not affected by differentiation, thus being these genes the most stable candidates, while CYC, GAPDH, and 18S were not suitable for this sort of analysis. This work highlights that RPL13A and EEF1A1 are good candidates as reference genes for qPCR analysis of hVSCs differentiation into adipocytes from subjects with different BMI and HOMA-IR.

  5. RPL13A and EEF1A1 Are Suitable Reference Genes for qPCR during Adipocyte Differentiation of Vascular Stromal Cells from Patients with Different BMI and HOMA-IR

    PubMed Central

    Gentile, Adriana-Mariel; Lhamyani, Said; Coín-Aragüez, Leticia; Oliva-Olivera, Wilfredo; Zayed, Hatem; Vega-Rioja, Antonio; Monteseirin, Javier; Romero-Zerbo, Silvana-Yanina; Tinahones, Francisco-José; Bermúdez-Silva, Francisco-Javier; El Bekay, Rajaa

    2016-01-01

    Real-time or quantitative PCR (qPCR) is a useful technique that requires reliable reference genes for data normalization in gene expression analysis. Adipogenesis is among the biological processes suitable for this technique. The selection of adequate reference genes is essential for qPCR gene expression analysis of human Vascular Stromal Cells (hVSCs) during their differentiation into adipocytes. To the best of our knowledge, there are no studies validating reference genes for the analyses of visceral and subcutaneous adipose tissue hVSCs from subjects with different Body Mass Index (BMI) and Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) index. The present study was undertaken to analyze this question. We first analyzed the stability of expression of five potential reference genes: CYC, GAPDH, RPL13A, EEF1A1, and 18S ribosomal RNA, during in vitro adipogenic differentiation, in samples from these types of patients. The expression of RPL13A and EEF1A1 was not affected by differentiation, thus being these genes the most stable candidates, while CYC, GAPDH, and 18S were not suitable for this sort of analysis. This work highlights that RPL13A and EEF1A1 are good candidates as reference genes for qPCR analysis of hVSCs differentiation into adipocytes from subjects with different BMI and HOMA-IR. PMID:27304673

  6. Endogenous Stem Cells Were Recruited by Defocused Low-Energy Shock Wave in Treating Diabetic Bladder Dysfunction.

    PubMed

    Jin, Yang; Xu, Lina; Zhao, Yong; Wang, Muwen; Jin, Xunbo; Zhang, Haiyang

    2017-04-01

    Defocused low-energy shock wave (DLSW) has been shown effects on activating mesenchymal stromal cells (MSCs) in vitro. In this study, recruitment of endogenous stem cells was firstly examined as an important pathway during the healing process of diabetic bladder dysfunction (DBD) treated by DLSW in vivo. Neonatal rats received intraperitoneal injection of 5-ethynyl-2-deoxyuridine (EdU) and then DBD rat model was created by injecting streptozotocin. Four weeks later, DLSW treatment was performed. Afterward, their tissues were examined by histology. Meanwhile, adipose tissue-derived stem cells (ADSCs) were treated by DLSW in vitro. Results showed DLSW ameliorated voiding function of diabetic rats by recruiting EdU + Stro-1 + CD34 - endogenous stem cells to release abundant nerve growth factor (NGF) and vascular endothelial growth factor (VEGF). Some EdU + cells overlapped with staining of smooth muscle actin. After DLSW treatment, ADSCs showed higher migration ability, higher expression level of stromal cell-derived factor-1 and secreted more NGF and VEGF. In conclusion, DLSW could ameliorate DBD by recruiting endogenous stem cells. Beneficial effects were mediated by secreting NGF and VEGF, resulting into improved innervation and vascularization in bladder.

  7. Tracking of autologous adipose tissue-derived mesenchymal stromal cells with in vivo magnetic resonance imaging and histology after intralesional treatment of artificial equine tendon lesions--a pilot study.

    PubMed

    Geburek, Florian; Mundle, Kathrin; Conrad, Sabine; Hellige, Maren; Walliser, Ulrich; van Schie, Hans T M; van Weeren, René; Skutella, Thomas; Stadler, Peter M

    2016-02-01

    Adipose tissue-derived mesenchymal stromal cells (AT-MSCs) are frequently used to treat equine tendinopathies. Up to now, knowledge about the fate of autologous AT-MSCs after intralesional injection into equine superficial digital flexor tendons (SDFTs) is very limited. The purpose of this study was to monitor the presence of intralesionally injected autologous AT-MSCs labelled with superparamagnetic iron oxide (SPIO) nanoparticles and green fluorescent protein (GFP) over a staggered period of 3 to 9 weeks with standing magnetic resonance imaging (MRI) and histology. Four adult warmblood horses received a unilateral injection of 10 × 10(6) autologous AT-MSCs into surgically created front-limb SDFT lesions. Administered AT-MSCs expressed lentivirally transduced reporter genes for GFP and were co-labelled with SPIO particles in three horses. The presence of AT-MSCs in SDFTs was evaluated by repeated examinations with standing low-field MRI in two horses and post-mortem in all horses with Prussian blue staining, fluorescence microscopy and with immunofluorescence and immunohistochemistry using anti-GFP antibodies at 3, 5, 7 and 9 weeks after treatment. AT-MSCs labelled with SPIO particles were detectable in treated SDFTs during each MRI in T2*- and T1-weighted sequences until the end of the observation period. Post-mortem examinations revealed that all treated tendons contained high numbers of SPIO- and GFP-labelled cells. Standing low-field MRI has the potential to track SPIO-labelled AT-MSCs successfully. Histology, fluorescence microscopy, immunofluorescence and immunohistochemistry are efficient tools to detect labelled AT-MSCs after intralesional injection into surgically created equine SDFT lesions. Intralesional injection of 10 × 10(6) AT-MSCs leads to the presence of high numbers of AT-MSCs in and around surgically created tendon lesions for up to 9 weeks. Integration of injected AT-MSCs into healing tendon tissue is an essential pathway after intralesional

  8. Autologous fat grafting: use of closed syringe microcannula system for enhanced autologous structural grafting

    PubMed Central

    Alexander, Robert W; Harrell, David B

    2013-01-01

    Objectives Provide background for use of acquiring autologous adipose tissue as a tissue graft and source of adult progenitor cells for use in cosmetic plastic surgery. Discuss the background and mechanisms of action of closed syringe vacuum lipoaspiration, with emphasis on accessing adipose-derived mesenchymal/stromal cells and the stromal vascular fraction (SVF) for use in aesthetic, structural reconstruction and regenerative applications. Explain a proven protocol for acquiring high-quality autologous fat grafts (AFG) with use of disposable, microcannula systems. Design Explain the components and advantage of use of the patented super luer-lock and microcannulas system for use with the closed-syringe system. A sequential explanation of equipment selection for minimally traumatic lipoaspiration in small volumes is presented, including use of blunt injection cannulas to reduce risk of embolism. Results Thousands of AFG have proven safe and efficacious for lipoaspiration techniques for large and small structural fat grafting procedures. The importance and advantages of gentle harvesting of the adipose tissue complex has become very clear in the past 5 years. The closed-syringe system offers a minimally invasive, gentle system with which to mobilize subdermal fat tissues in a suspension form. Resulting total nuclear counting of undifferentiated cells of the adipose-derived -SVF suggests that the yield achieved is better than use of always-on, constant mechanical pump applied vacuum systems. Conclusion Use of a closed-syringe lipoaspiration system featuring disposable microcannulas offers a safe and effective means of harvesting small volumes of nonmanipulated adipose tissues and its accompanying progenitor cells within the SVF. Closed syringes and microcannulas are available as safe, sterile, disposable, compact systems for acquiring high-quality AFG. Presented is a detailed, step-by-step, proven protocol for performing quality autologous structural adipose

  9. Intrinsic Deregulation of Vascular Smooth Muscle and Myofibroblast Differentiation in Mesenchymal Stromal Cells from Patients with Systemic Sclerosis.

    PubMed

    Hegner, Björn; Schaub, Theres; Catar, Rusan; Kusch, Angelika; Wagner, Philine; Essin, Kirill; Lange, Claudia; Riemekasten, Gabriela; Dragun, Duska

    2016-01-01

    Obliterative vasculopathy and fibrosis are hallmarks of systemic sclerosis (SSc), a severe systemic autoimmune disease. Bone marrow-derived mesenchymal stromal cells (MSCs) from SSc patients may harbor disease-specific abnormalities. We hypothesized disturbed vascular smooth muscle cell (VSMC) differentiation with increased propensity towards myofibroblast differentiation in response to SSc-microenvironment defining growth factors and determined responsible mechanisms. We studied responses of multipotent MSCs from SSc-patients (SSc-MSCs) and healthy controls (H-MSCs) to long-term exposure to CTGF, b-FGF, PDGF-BB or TGF-β1. Differentiation towards VSMC and myofibroblast lineages was analyzed on phenotypic, biochemical, and functional levels. Intracellular signaling studies included analysis of TGF-β receptor regulation, SMAD, AKT, ERK1/2 and autocrine loops. VSMC differentiation towards both, contractile and synthetic VSMC phenotypes in response to CTGF and b-FGF was disturbed in SSc-MSCs. H-MSCs and SSc-MSCs responded equally to PDGF-BB with prototypic fibroblastic differentiation. TGF-β1 initiated myofibroblast differentiation in both cell types, yet with striking phenotypic and functional differences: In relation to H-MSC-derived myofibroblasts induced by TGF-β1, those obtained from SSc-MSCs expressed more contractile proteins, migrated towards TGF-β1, had low proliferative capacity, and secreted higher amounts of collagen paralleled by reduced MMP expression. Higher levels of TGF-β receptor 1 and enhanced canonical and noncanonical TGF-β signaling in SSc-MSCs accompanied aberrant differentiation response of SSc-MSCs in comparison to H-MSCs. Deregulated VSMC differentiation with a shift towards myofibroblast differentiation expands the concept of disturbed endogenous regenerative capacity of MSCs from SSc patients. Disease related intrinsic hyperresponsiveness to TGF-β1 with increased collagen production may represent one responsible mechanism. Better

  10. Amount of stroma is associated with mammographic density and stromal expression of oestrogen receptor in normal breast tissues.

    PubMed

    Gabrielson, Marike; Chiesa, Flaminia; Paulsson, Janna; Strell, Carina; Behmer, Catharina; Rönnow, Katarina; Czene, Kamila; Östman, Arne; Hall, Per

    2016-07-01

    Following female sex and age, mammographic density is considered one of the strongest risk factors for breast cancer. Despite the association between mammographic density and breast cancer risk, little is known about the underlying histology and biological basis of breast density. To better understand the mechanisms behind mammographic density we assessed morphology, proliferation and hormone receptor status in relation to mammographic density in breast tissues from healthy women. Tissues were obtained from 2012-2013 by ultrasound-guided core needle biopsy from 160 women as part of the Karma (Karolinska mammography project for risk prediction for breast cancer) project. Mammograms were collected through routine mammography screening and mammographic density was calculated using STRATUS. The histological composition, epithelial and stromal proliferation status and hormone receptor status were assessed through immunohistochemical staining. Higher mammographic density was significantly associated with a greater proportion of stromal and epithelial tissue and a lower proportion of adipose tissue. Epithelial expression levels of Ki-67, oestrogen receptor (ER) and progesterone receptor (PR) were not associated with mammographic density. Epithelial Ki-67 was associated with a greater proportion of epithelial tissue, and epithelial PR was associated with a greater proportion of stromal and a lower proportion of adipose tissue. Epithelial ER was not associated with any tissues. In contrast, expression of ER in the stroma was significantly associated with a greater proportion of stroma, and negatively associated with the amount of adipose tissue. High mammographic density is associated with higher amount of stroma and epithelium and less amount of fat, but is not associated with a change in epithelial proliferation or receptor status. Increased expressions of both epithelial PR and stromal ER are associated with a greater proportion of stroma, suggesting hormonal involvement

  11. Depressed Levels of Prostaglandin F2α in Mice Lacking Akr1b7 Increase Basal Adiposity and Predispose to Diet-Induced Obesity

    PubMed Central

    Volat, Fanny E.; Pointud, Jean-Christophe; Pastel, Emilie; Morio, Béatrice; Sion, Benoit; Hamard, Ghislaine; Guichardant, Michel; Colas, Romain; Lefrançois-Martinez, Anne-Marie; Martinez, Antoine

    2012-01-01

    Negative regulators of white adipose tissue (WAT) expansion are poorly documented in vivo. Prostaglandin F2α (PGF2α) is a potent antiadipogenic factor in cultured preadipocytes, but evidence for its involvement in physiological context is lacking. We previously reported that Akr1b7, an aldo-keto reductase enriched in adipose stromal vascular fraction but absent from mature adipocytes, has antiadipogenic properties possibly supported by PGF2α synthase activity. To test whether lack of Akr1b7 could influence WAT homeostasis in vivo, we generated Akr1b7−/− mice in 129/Sv background. Akr1b7−/− mice displayed excessive basal adiposity resulting from adipocyte hyperplasia/hypertrophy and exhibited greater sensitivity to diet-induced obesity. Following adipose enlargement and irrespective of the diet, they developed liver steatosis and progressive insulin resistance. Akr1b7 loss was associated with decreased PGF2α WAT contents. Cloprostenol (PGF2α agonist) administration to Akr1b7−/− mice normalized WAT expansion by affecting both de novo adipocyte differentiation and size. Treatment of 3T3-L1 adipocytes and Akr1b7−/− mice with cloprostenol suggested that decreased adipocyte size resulted from inhibition of lipogenic gene expression. Hence, Akr1b7 is a major regulator of WAT development through at least two PGF2α-dependent mechanisms: inhibition of adipogenesis and lipogenesis. These findings provide molecular rationale to explore the status of aldo-keto reductases in dysregulations of adipose tissue homeostasis. PMID:22851578

  12. Estrogen receptor 1 (ESR1) regulates VEGFA in adipose tissue.

    PubMed

    Fatima, L A; Campello, R S; Santos, R de Souza; Freitas, H S; Frank, A P; Machado, U F; Clegg, D J

    2017-12-01

    Vascular endothelial growth factor A (VEGFA) is a key factor in the regulation of angiogenesis in adipose tissue. Poor vascularization during adipose tissue proliferation causes fibrosis and local inflammation, and is associated with insulin resistance. It is known that 17-beta estradiol (E2) regulates adipose tissue function and VEGFA expression in other tissues; however, the ability of E2 to regulate VEGFA in adipose tissue is currently unknown. In this study, we showed that, in 3T3-L1 cells, E2 and the estrogen receptor 1 (ESR1) agonist PPT induced VEGFA expression, while ESR1 antagonist (MPP), and selective knockdown of ESR1 using siRNA decreased VEGFA and prevented the ability of E2 to modulate its expression. Additionally, we found that E2 and PPT induced the binding of hypoxia inducible factor 1 alpha subunit (HIF1A) in the VEGFA gene promoter. We further found that VEGFA expression was lower in inguinal and gonadal white adipose tissues of ESR1 total body knockout female mice compared to wild type mice. In conclusion, our data provide evidence of an important role for E2/ESR1 in modulating adipose tissue VEGFA, which is potentially important to enhance angiogenesis, reduce inflammation and improve adipose tissue function.

  13. Adipose Derived Stem Cells for Corneal Wound Healing after Laser Induced Corneal Lesions in Mice.

    PubMed

    Zeppieri, Marco; Salvetat, Maria Letizia; Beltrami, Antonio; Cesselli, Daniela; Russo, Rossella; Alcalde, Ignacio; Merayo-Lloves, Jesús; Brusini, Paolo; Parodi, Pier Camillo

    2017-12-05

    The aim of our study was to assess the clinical effectiveness of topical adipose derived stem cell (ADSC) treatment in laser induced corneal wounds in mice by comparing epithelial repair, inflammation, and histological analysis between treatment arms. Corneal lesions were performed on both eyes of 40 mice by laser induced photorefractive keratectomy. All eyes were treated with topical azythromycin bid for three days. Mice were divided in three treatment groups ( n = 20), which included: control, stem cells and basic serum; which received topical treatment three times daily for five consecutive days. Biomicroscope assessments and digital imaging were performed by two masked graders at 30, 54, 78, 100, and 172 h to analyze extent of fluorescein positive epithelial defect, corneal inflammation, etc. Immunohistochemical techniques were used in fixed eyes to assess corneal repair markers Ki67, α Smooth Muscle Actin (α-SMA) and E-Cadherin. The fluorescein positive corneal lesion areas were significantly smaller in the stem cells group on days 1 ( p < 0.05), 2 ( p < 0.02) and 3. The stem cell treated group had slightly better and faster re-epithelization than the serum treated group in the initial phases. Comparative histological data showed signs of earlier and better corneal repair in epithelium and stromal layers in stem cell treated eyes, which showed more epithelial layers and enhanced wound healing performance of Ki67, E-Cadherin, and α-SMA. Our study shows the potential clinical and histological advantages in the topical ADSC treatment for corneal lesions in mice.

  14. Adipose Derived Stem Cells for Corneal Wound Healing after Laser Induced Corneal Lesions in Mice

    PubMed Central

    Salvetat, Maria Letizia; Beltrami, Antonio; Cesselli, Daniela; Russo, Rossella; Merayo-Lloves, Jesús; Brusini, Paolo; Parodi, Pier Camillo

    2017-01-01

    The aim of our study was to assess the clinical effectiveness of topical adipose derived stem cell (ADSC) treatment in laser induced corneal wounds in mice by comparing epithelial repair, inflammation, and histological analysis between treatment arms. Corneal lesions were performed on both eyes of 40 mice by laser induced photorefractive keratectomy. All eyes were treated with topical azythromycin bid for three days. Mice were divided in three treatment groups (n = 20), which included: control, stem cells and basic serum; which received topical treatment three times daily for five consecutive days. Biomicroscope assessments and digital imaging were performed by two masked graders at 30, 54, 78, 100, and 172 h to analyze extent of fluorescein positive epithelial defect, corneal inflammation, etc. Immunohistochemical techniques were used in fixed eyes to assess corneal repair markers Ki67, α Smooth Muscle Actin (α-SMA) and E-Cadherin. The fluorescein positive corneal lesion areas were significantly smaller in the stem cells group on days 1 (p < 0.05), 2 (p < 0.02) and 3. The stem cell treated group had slightly better and faster re-epithelization than the serum treated group in the initial phases. Comparative histological data showed signs of earlier and better corneal repair in epithelium and stromal layers in stem cell treated eyes, which showed more epithelial layers and enhanced wound healing performance of Ki67, E-Cadherin, and α-SMA. Our study shows the potential clinical and histological advantages in the topical ADSC treatment for corneal lesions in mice. PMID:29206194

  15. Investigating the mincing method for isolation of adipose-derived stem cells from pregnant women fat.

    PubMed

    Li, Yuan-Sheng; Chen, Pao-Jen; Wu, Li-Wei; Chou, Pei-Wen; Sun, Li-Yi; Chiou, Tzyy-Wen

    2018-02-01

    The success of stem cell application in regenerative medicine, usually require a stable source of stem or progenitor cells. Fat tissue represents a good source of stem cells because it is rich in stem cells and there are fewer ethical issues related to the use of such stem cells, unlike embryonic stem cells. Therefore, there has been increased interest in adipose-derived stem cells (ADSCs) for tissue engineering applications. Here, we aim to provide an easy processing method for isolating adult stem cells from human adipose tissue harvested from the subcutaneous fat of the abdominal wall during gynecologic surgery. We used a homogenizer to mince fat and compared the results with those obtained from the traditional cut method involving a sterile scalpel and forceps. Our results showed that our method provides another stable and quality source of stem cells that could be used in cases with a large quantity of fat. Furthermore, we found that pregnancy adipose-derived stem cells (P-ADSCs) could be maintained in vitro for extended periods with a stable population doubling and low senescence levels. P-ADSCs could also differentiate in vitro into adipogenic, osteogenic, chondrogenic, and insulin-producing cells in the presence of lineage-specific induction factors. In conclusion, like human lipoaspirates, adipose tissues obtained from pregnant women contain multipotent cells with better proliferation and showed great promise for use in both stem cell banking studies as well as in stem cell therapy.

  16. Stromal cell-derived factor 2 is critical for Hsp90-dependent eNOS activation.

    PubMed

    Siragusa, Mauro; Fröhlich, Florian; Park, Eon Joo; Schleicher, Michael; Walther, Tobias C; Sessa, William C

    2015-08-18

    Endothelial nitric oxide synthase (eNOS) catalyzes the conversion of l-arginine and molecular oxygen into l-citrulline and nitric oxide (NO), a gaseous second messenger that influences cardiovascular physiology and disease. Several mechanisms regulate eNOS activity and function, including phosphorylation at Ser and Thr residues and protein-protein interactions. Combining a tandem affinity purification approach and mass spectrometry, we identified stromal cell-derived factor 2 (SDF2) as a component of the eNOS macromolecular complex in endothelial cells. SDF2 knockdown impaired agonist-stimulated NO synthesis and decreased the phosphorylation of eNOS at Ser(1177), a key event required for maximal activation of eNOS. Conversely, SDF2 overexpression dose-dependently increased NO synthesis through a mechanism involving Akt and calcium (induced with ionomycin), which increased the phosphorylation of Ser(1177) in eNOS. NO synthesis by iNOS (inducible NOS) and nNOS (neuronal NOS) was also enhanced upon SDF2 overexpression. We found that SDF2 was a client protein of the chaperone protein Hsp90, interacting preferentially with the M domain of Hsp90, which is the same domain that binds to eNOS. In endothelial cells exposed to vascular endothelial growth factor (VEGF), SDF2 was required for the binding of Hsp90 and calmodulin to eNOS, resulting in eNOS phosphorylation and activation. Thus, our data describe a function for SDF2 as a component of the Hsp90-eNOS complex that is critical for signal transduction in endothelial cells. Copyright © 2015, American Association for the Advancement of Science.

  17. Stromal cell–derived factor 2 is critical for Hsp90-dependent eNOS activation

    PubMed Central

    Siragusa, Mauro; Fröhlich, Florian; Park, Eon Joo; Schleicher, Michael; Walther, Tobias C.; Sessa, William C.

    2016-01-01

    Endothelial nitric oxide synthase (eNOS) catalyzes the conversion of l-arginine and molecular oxygen into l-citrulline and nitric oxide (NO), a gaseous second messenger that influences cardiovascular physiology and disease. Several mechanisms regulate eNOS activity and function, including phosphorylation at Ser and Thr residues and protein-protein interactions. Combining a tandem affinity purification approach and mass spectrometry, we identified stromal cell–derived factor 2 (SDF2) as a component of the eNOS macromolecular complex in endothelial cells. SDF2 knockdown impaired agonist-stimulated NO synthesis and decreased the phosphorylation of eNOS at Ser1177, a key event required for maximal activation of eNOS. Conversely, SDF2 overexpression dose-dependently increased NO synthesis through a mechanism involving Akt and calcium (induced with ionomycin), which increased the phosphorylation of Ser1177 in eNOS. NO synthesis by iNOS (inducible NOS) and nNOS (neuronal NOS) was also enhanced upon SDF2 overexpression. We found that SDF2 was a client protein of the chaperone protein Hsp90, interacting preferentially with the M domain of Hsp90, which is the same domain that binds to eNOS. In endothelial cells exposed to vascular endothelial growth factor (VEGF), SDF2 was required for the binding of Hsp90 and calmodulin to eNOS, resulting in eNOS phosphorylation and activation. Thus, our data describe a function for SDF2 as a component of the Hsp90-eNOS complex that is critical for signal transduction in endothelial cells. PMID:26286023

  18. VASCULAR RESPONSE TO TRAFFIC-DERIVED INHALATION IN HUMANS

    EPA Science Inventory

    By coordinating closely with Center Projects 1-3, we will determine whether specific aspects of traffic-derived exposure (primary vs. secondary organics, particulate vs. gases, spark-ignition vs. diesel engine vs. a mixture) enhance the human vascular response to pollutants. W...

  19. C1q/TNF-related protein 6 (CTRP6) links obesity to adipose tissue inflammation and insulin resistance.

    PubMed

    Lei, Xia; Seldin, Marcus M; Little, Hannah C; Choy, Nicholas; Klonisch, Thomas; Wong, G William

    2017-09-08

    Obesity is associated with chronic low-grade inflammation, and metabolic regulators linking obesity to inflammation have therefore received much attention. Secreted C1q/TNF-related proteins (CTRPs) are one such group of regulators that regulate glucose and fat metabolism in peripheral tissues and modulate inflammation in adipose tissue. We have previously shown that expression of CTRP6 is up-regulated in leptin-deficient mice and, conversely, down-regulated by the anti-diabetic drug rosiglitazone. Here, we provide evidence for a novel role of CTRP6 in modulating both inflammation and insulin sensitivity. We found that in obese and diabetic humans and mouse models, CTRP6 expression was markedly up-regulated in adipose tissue and that stromal vascular cells, such as macrophages, are a major CTRP6 source. Overexpressing mouse or human CTRP6 impaired glucose disposal in peripheral tissues in response to glucose and insulin challenge in wild-type mice. Conversely, Ctrp6 gene deletion improved insulin action and increased metabolic rate and energy expenditure in diet-induced obese mice. Mechanistically, CTRP6 regulates local inflammation and glucose metabolism by targeting macrophages and adipocytes, respectively. In cultured macrophages, recombinant CTRP6 dose-dependently up-regulated the expression and production of TNF-α. Conversely, CTRP6 deficiency reduced circulating inflammatory cytokines and pro-inflammatory macrophages in adipose tissue. CTRP6-overexpressing mice or CTRP6-treated adipocytes had reduced insulin-stimulated Akt phosphorylation and glucose uptake. In contrast, loss of CTRP6 enhanced insulin-stimulated Akt activation in adipose tissue. Together, these results establish CTRP6 as a novel metabolic/immune regulator linking obesity to adipose tissue inflammation and insulin resistance. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  20. Mammary Adipose Tissue-derived Lysophospholipids Promote Estrogen Receptor-negative Mammary Epithelial Cell Proliferation

    PubMed Central

    Volden, Paul A.; Skor, Maxwell N.; Johnson, Marianna B.; Singh, Puneet; Patel, Feenalie N.; McClintock, Martha K.; Brady, Matthew J.; Conzen, Suzanne D.

    2016-01-01

    Lysophosphatidic acid (LPA), acting in an autocrine or paracrine fashion through G protein-coupled receptors, has been implicated in many physiological and pathological processes including cancer. LPA is converted to lysophosphatidylcholine (LPC) by the secreted phospholipase, autotaxin (ATX). Although various cell types can produce ATX, adipocyte-derived ATX is believed to be the major source of circulating ATX and also to be the major regulator of plasma LPA. In addition to ATX, adipocytes secrete numerous other factors (adipokines); although several adipokines have been implicated in breast cancer biology, the contribution of mammary adipose tissue-derived LPC/ATX/LPA (LPA-axis) signaling to breast cancer is poorly understood. Using mammary fat-conditioned medium, we investigated the contribution of LPA signaling to mammary epithelial cancer cell biology and identified LPA signaling as a significant contributor to the oncogenic effects of the mammary adipose tissue secretome. To interrogate the role of mammary fat in the LPA-axis during breast cancer progression, we exposed mammary adipose tissue to secreted factors from estrogen receptor-negative mammary epithelial cell lines and monitored changes in the mammary fat pad LPA-axis. Our data indicate that bidirectional interactions between mammary cancer cells and mammary adipocytes alter the local LPA-axis and increase ATX expression in the mammary fat pad during breast cancer progression. Thus, the LPC/ATX/LPA axis may be a useful target for prevention in patients at risk of ER-negative breast cancer. PMID:26862086

  1. Fibronectin Deposition Participates in Extracellular Matrix Assembly and Vascular Morphogenesis

    PubMed Central

    Hielscher, Abigail; Ellis, Kim; Qiu, Connie; Porterfield, Josh; Gerecht, Sharon

    2016-01-01

    The extracellular matrix (ECM) has been demonstrated to facilitate angiogenesis. In particular, fibronectin has been documented to activate endothelial cells, resulting in their transition from a quiescent state to an active state in which the cells exhibit enhanced migration and proliferation. The goal of this study is to examine the role of polymerized fibronectin during vascular tubulogenesis using a 3 dimensional (3D) cell-derived de-cellularized matrix. A fibronectin-rich 3D de-cellularized ECM was used as a scaffold to study vascular morphogenesis of endothelial cells (ECs). Confocal analyses of several matrix proteins reveal high intra- and extra-cellular deposition of fibronectin in formed vascular structures. Using a small peptide inhibitor of fibronectin polymerization, we demonstrate that inhibition of fibronectin fibrillogenesis in ECs cultured atop de-cellularized ECM resulted in decreased vascular morphogenesis. Further, immunofluorescence and ultrastructural analyses reveal decreased expression of stromal matrix proteins in the absence of polymerized fibronectin with high co-localization of matrix proteins found in association with polymerized fibronectin. Evaluating vascular kinetics, live cell imaging showed that migration, migration velocity, and mean square displacement, are disrupted in structures grown in the absence of polymerized fibronectin. Additionally, vascular organization failed to occur in the absence of a polymerized fibronectin matrix. Consistent with these observations, we tested vascular morphogenesis following the disruption of EC adhesion to polymerized fibronectin, demonstrating that block of integrins α5β1 and αvβ3, abrogated vascular morphogenesis. Overall, fibronectin deposition in a 3D cell-derived de-cellularized ECM appears to be imperative for matrix assembly and vascular morphogenesis. PMID:26811931

  2. Adipose-Derived Stem Cells for Tissue Engineering and Regenerative Medicine Applications

    PubMed Central

    Dai, Ru; Wang, Zongjie; Samanipour, Roya; Koo, Kyo-in; Kim, Keekyoung

    2016-01-01

    Adipose-derived stem cells (ASCs) are a mesenchymal stem cell source with properties of self-renewal and multipotential differentiation. Compared to bone marrow-derived stem cells (BMSCs), ASCs can be derived from more sources and are harvested more easily. Three-dimensional (3D) tissue engineering scaffolds are better able to mimic the in vivo cellular microenvironment, which benefits the localization, attachment, proliferation, and differentiation of ASCs. Therefore, tissue-engineered ASCs are recognized as an attractive substitute for tissue and organ transplantation. In this paper, we review the characteristics of ASCs, as well as the biomaterials and tissue engineering methods used to proliferate and differentiate ASCs in a 3D environment. Clinical applications of tissue-engineered ASCs are also discussed to reveal the potential and feasibility of using tissue-engineered ASCs in regenerative medicine. PMID:27057174

  3. Adipogenesis of human adipose-derived stem cells within three-dimensional hollow fiber-based bioreactors.

    PubMed

    Gerlach, Jörg C; Lin, Yen-Chih; Brayfield, Candace A; Minteer, Danielle M; Li, Han; Rubin, J Peter; Marra, Kacey G

    2012-01-01

    To further differentiate adipose-derived stem cells (ASCs) into mature adipocytes and create three-dimensional (3D) adipose tissue in vitro, we applied multicompartment hollow fiber-based bioreactor technology with decentral mass exchange for more physiological substrate gradients and integral oxygenation. We hypothesize that a dynamic 3D perfusion in such a bioreactor will result in longer-term culture of human adipocytes in vitro, thus providing metabolically active tissue serving as a diagnostic model for screening drugs to treat diabetes. ASCs were isolated from discarded human abdominal subcutaneous adipose tissue and then inoculated into dynamic 3D culture bioreactors to undergo adipogenic differentiation. Insulin-stimulated glucose uptake from the medium was assessed with and without TNF-alpha. 3D adipose tissue was generated in the 3D-bioreactors. Immunohistochemical staining indicated that 3D-bioreactor culture displayed multiple mature adipocyte markers with more unilocular morphologies as compared with two-dimensional (2D) cultures. Results of real-time polymerase chain reaction showed 3D-bioreactor treatment had more efficient differentiation in fatty acid-binding protein 4 expression. Repeated insulin stimulation resulted in increased glucose uptake, with a return to baseline between testing. Importantly, TNF-alpha inhibited glucose uptake, an indication of the metabolic activity of the tissue. 3D bioreactors allow more mature adipocyte differentiation of ASCs compared with traditional 2D culture and generate adipose tissue in vitro for up to 2 months. Reproducible metabolic activity of the adipose tissue in the bioreactor was demonstrated, which is potentially useful for drug discovery. We present here, to the best of our knowledge for the first time, the development of a coherent 3D high density fat-like tissue consisting of unilocular structure from primary adipose stem cells in vitro.

  4. Adipogenesis of Human Adipose-Derived Stem Cells Within Three-Dimensional Hollow Fiber-Based Bioreactors

    PubMed Central

    Gerlach, Jörg C.; Lin, Yen-Chih; Brayfield, Candace A.; Minteer, Danielle M.; Li, Han; Rubin, J. Peter

    2012-01-01

    To further differentiate adipose-derived stem cells (ASCs) into mature adipocytes and create three-dimensional (3D) adipose tissue in vitro, we applied multicompartment hollow fiber-based bioreactor technology with decentral mass exchange for more physiological substrate gradients and integral oxygenation. We hypothesize that a dynamic 3D perfusion in such a bioreactor will result in longer-term culture of human adipocytes in vitro, thus providing metabolically active tissue serving as a diagnostic model for screening drugs to treat diabetes. ASCs were isolated from discarded human abdominal subcutaneous adipose tissue and then inoculated into dynamic 3D culture bioreactors to undergo adipogenic differentiation. Insulin-stimulated glucose uptake from the medium was assessed with and without TNF-alpha. 3D adipose tissue was generated in the 3D-bioreactors. Immunohistochemical staining indicated that 3D-bioreactor culture displayed multiple mature adipocyte markers with more unilocular morphologies as compared with two-dimensional (2D) cultures. Results of real-time polymerase chain reaction showed 3D-bioreactor treatment had more efficient differentiation in fatty acid-binding protein 4 expression. Repeated insulin stimulation resulted in increased glucose uptake, with a return to baseline between testing. Importantly, TNF-alpha inhibited glucose uptake, an indication of the metabolic activity of the tissue. 3D bioreactors allow more mature adipocyte differentiation of ASCs compared with traditional 2D culture and generate adipose tissue in vitro for up to 2 months. Reproducible metabolic activity of the adipose tissue in the bioreactor was demonstrated, which is potentially useful for drug discovery. We present here, to the best of our knowledge for the first time, the development of a coherent 3D high density fat-like tissue consisting of unilocular structure from primary adipose stem cells in vitro. PMID:21902468

  5. Regeneration of the oesophageal muscle layer from oesophagus acellular matrix scaffold using adipose-derived stem cells.

    PubMed

    Wang, Fang; Maeda, Yasuko; Zachar, Vladimir; Ansari, Tahera; Emmersen, Jeppe

    2018-06-14

    This study explored the feasibility of constructing a tissue engineered muscle layer in the oesophagus using oesophageal acellular matrix (OAM) scaffolds and human aortic smooth muscle cells (hASMCs) or human adipose-derived stem cells (hASCs). The second objective was to investigate the effect of hypoxic preconditioning of seeding cells on cell viability and migration depth. Our results demonstrated that hASMCs and hASCs could attach and adhere to the decellularized OAM scaffold and survive and proliferate for at least 7 days depending on the growth conditions. This indicates adipose-derived stem cells (ASCs) have the potential to substitute for smooth muscle cells (SMCs) in the construction of tissue engineered oesophageal muscle layers. Copyright © 2018 Elsevier Inc. All rights reserved.

  6. Rapid Selection of Mesenchymal Stem and Progenitor Cells in Primary Prostate Stromal Cultures

    PubMed Central

    Brennen, W. Nathaniel; Kisteman, L. Nelleke; Isaacs, John T.

    2016-01-01

    BACKGROUND Carcinoma-associated fibroblasts (CAFs) are a dominant component of the tumor microenvironment with pro-tumorigenic properties. Despite this knowledge, their physiologic origins remain poorly understood. Mesenchymal stem cells (MSCs) can be recruited from the bone marrow to areas of tissue damage and inflammation, including prostate cancer. MSCs can generate and have many overlapping properties with CAFs in preclinical models. METHODS Multiparameter flow cytometry and multipotent differentiation assays used to define MSCs in primary prostate stromal cultures derived from young (>25 yrs) organ donors and prostate cancer patients compared with bone marrow-derived stromal cultures. Population doubling times, population doublings, cell size, and differentiation potential determined under multiple culture conditions, including normoxia, hypoxia, and a variety of media. TGF-β measured by ELISA. RESULTS MSCs and stromal progenitors are not only present in normal and malignant prostate tissue, but are quickly selected for in primary stromal cultures derived from these tissues; becoming the dominant population within just a few passages. Growth potential inversely associated with TGF-β concentrations. All conditions generated populations with an average cell diameter >15 μm. All cultures tested had the ability to undergo osteogenic and chondrogenic differentiation, but unlike bone marrow-derived MSCs, primary stromal cultures derived from normal prostate tissue lack adipogenic differentiation potential. In contrast, a subset of stromal cultures derived from prostate cancer patients retain the ability to differentiate into adipocytes; a property that is significantly suppressed under hypoxic conditions in both bone marrow- and prostate-derived MSCs. CONCLUSIONS Primary prostate stromal cultures are highly enriched in cells with an MSC or stromal progenitor phenotype. The use of primary cultures such as these to study CAFs raises interesting implications when

  7. Rapid selection of mesenchymal stem and progenitor cells in primary prostate stromal cultures.

    PubMed

    Brennen, W Nathaniel; Kisteman, L Nelleke; Isaacs, John T

    2016-05-01

    Carcinoma-associated fibroblasts (CAFs) are a dominant component of the tumor microenvironment with pro-tumorigenic properties. Despite this knowledge, their physiologic origins remain poorly understood. Mesenchymal stem cells (MSCs) can be recruited from the bone marrow to areas of tissue damage and inflammation, including prostate cancer. MSCs can generate and have many overlapping properties with CAFs in preclinical models. Multiparameter flow cytometry and multipotent differentiation assays used to define MSCs in primary prostate stromal cultures derived from young (<25 yrs) organ donors and prostate cancer patients compared with bone marrow-derived stromal cultures. Population doubling times, population doublings, cell size, and differentiation potential determined under multiple culture conditions, including normoxia, hypoxia, and a variety of media. TGF-β measured by ELISA. MSCs and stromal progenitors are not only present in normal and malignant prostate tissue, but are quickly selected for in primary stromal cultures derived from these tissues; becoming the dominant population within just a few passages. Growth potential inversely associated with TGF-β concentrations. All conditions generated populations with an average cell diameter >15 µm. All cultures tested had the ability to undergo osteogenic and chondrogenic differentiation, but unlike bone marrow-derived MSCs, primary stromal cultures derived from normal prostate tissue lack adipogenic differentiation potential. In contrast, a subset of stromal cultures derived from prostate cancer patients retain the ability to differentiate into adipocytes; a property that is significantly suppressed under hypoxic conditions in both bone marrow- and prostate-derived MSCs. Primary prostate stromal cultures are highly enriched in cells with an MSC or stromal progenitor phenotype. The use of primary cultures such as these to study CAFs raises interesting implications when considering their overlapping

  8. Recloned dogs derived from adipose stem cells of a transgenic cloned beagle.

    PubMed

    Oh, Hyun Ju; Park, Jung Eun; Kim, Min Jung; Hong, So Gun; Ra, Jeong Chan; Jo, Jung Youn; Kang, Sung Keun; Jang, Goo; Lee, Byeong Chun

    2011-04-15

    A number of studies have postulated that efficiency in mammalian cloning is inversely correlated with donor cell differentiation status and may be increased by using undifferentiated cells as nuclear donors. Here, we attempted the recloning of dogs by nuclear transfer of canine adipose tissue-derived mesenchymal stem cells (cAd-MSCs) from a transgenic cloned beagle to determine if cAd-MSCs can be a suitable donor cell type. In order to isolate cAd-MSCs, adipose tissues were collected from a transgenic cloned beagle produced by somatic cell nuclear transfer (SCNT) of canine fetal fibroblasts modified genetically with a red fluorescent protein (RFP) gene. The cAd-MSCs expressed the RFP gene and cell-surface marker characteristics of MSCs including CD29, CD44 and thy1.1. Furthermore, cAd-MSCs underwent osteogenic, adipogenic, myogenic, neurogenic and chondrogenic differentiation when exposed to specific differentiation-inducing conditions. In order to investigate the developmental potential of cAd-MSCs, we carried out SCNT. Fused-couplets (82/109, 75.2%) were chemically activated and transferred into the uterine tube of five naturally estrus-synchronized surrogates. One of them (20%) maintained pregnancy and subsequently gave birth to two healthy cloned pups. The present study demonstrated for the first time the successful production of cloned beagles by nuclear transfer of cAd-MSCs. Another important outcome of the present study is the successful recloning of RFP-expressing transgenic cloned beagle pups by nuclear transfer of cells derived from a transgenic cloned beagle. In conclusion, the present study demonstrates that adipose stem cells can be a good nuclear donor source for dog cloning. Copyright © 2011 Elsevier Inc. All rights reserved.

  9. Isolation, identification and multipotential differentiation of mouse adipose tissue-derived stem cells.

    PubMed

    Taha, Masoumeh Fakhr; Hedayati, Vahideh

    2010-08-01

    Bone marrow and adipose tissue have provided two suitable sources of mesenchymal stem cells. Although previous studies have confirmed close similarities between bone marrow-derived stem cells (BM-MSCs) and adipose tissue-derived stem cells (ADSCs), the molecular phenotype of ADSCs is still poorly identified. In the present study, mouse ADSCs were isolated from the inguinal fat pad of 12-14 weeks old mice. Freshly isolated and three passaged ADSCs were analyzed for the expression of OCT4, Sca-1, c-kit and CD34 by RT-PCR. Three passaged ADSCs were analyzed by flow cytometry for the presence of CD11b, CD45, CD31, CD29 and CD44. Moreover, cardiogenic, adipogenic and neurogenic differentiation of ADSCs were induced in vitro. Freshly isolated ADSCs showed the expression of OCT4, Sca-1, c-kit and CD34, and two days cultured ADSCs were positively immunostained with anti-OCT4 monoclonal antibody. After three passages, the expression of OCT4, c-kit and CD34 eliminated, while the expression of Sca-1 showed a striking enhancement. These cells were identified positive for CD29 and CD44 markers, and they showed the lack of CD45 and CD31 expression. Three passaged ADSCs were differentiated to adipocyte-, cardiomyocyte- and neuron-like cells that were identified based on the positive staining with Sudan black, anti-cardiac troponin I antibody and anti-map-2 antibody, respectively. In conclusion, adipose tissue contains a stem cell population that seems to be a good multipotential cell candidate for the future cell replacement therapy. Copyright 2010 Elsevier Ltd. All rights reserved.

  10. Placenta-derived mesenchymal stromal cells and their exosomes exert therapeutic effects in Duchenne muscular dystrophy.

    PubMed

    Bier, Ariel; Berenstein, Peter; Kronfeld, Noam; Morgoulis, Daria; Ziv-Av, Amotz; Goldstein, Hodaya; Kazimirsky, Gila; Cazacu, Simona; Meir, Rinat; Popovtzer, Rachela; Dori, Amir; Brodie, Chaya

    2018-05-03

    Duchenne muscular dystrophy (DMD) is a degenerative lethal, X-linked disease of skeletal and cardiac muscles caused by mutations in the dystrophin gene. Cell therapy using different cell types, including mesenchymal stromal cells (MSCs), has been considered as a potential approach for the treatment of DMD. MSCs can be obtained from autologous sources such as bone marrow and adipose tissues or from allogeneic placenta and umbilical cord. The safety and therapeutic impact of these cells has been demonstrated in pre-clinical and clinical studies and their functions are attributed to paracrine effects that are mediated by secreted cytokines and extracellular vesicles. Here, we studied the therapeutic effects of placenta-derived MSCs (PL-MSCs) and their secreted exosomes using mouse and human myoblasts from healthy controls, Duchenne patients and mdx mice. Treatment of myoblasts with conditioned medium or exosomes secreted by PL-MSCs increased the differentiation of these cells and decreased the expression of fibrogenic genes in DMD patient myoblasts. In addition, these treatments also increased the expression of utrophin in these cells. Using a quantitative miR-29c reporter, we demonstrated that the PL-MSC effects were partly mediated by the transfer of exosomal miR-29c. Intramuscular transplantation of PL-MSCs in mdx mice resulted in decreased creatine kinase levels. PL-MSCs significantly decreased the expression of TGF-β and the level of fibrosis in the diaphragm and cardiac muscles, inhibited inflammation and increased utrophin expression. In vivo imaging analyses using MSCs labeled with gold nanoparticles or fluorescent dyes demonstrated localization of the cells in the muscle tissues up to 3 weeks post treatment. Altogether, these results demonstrate that PL-MSCs and their secreted exosomes have important clinical applications in cell therapy of DMD partly via the targeted delivery of exosomal miR-29c. Copyright © 2018. Published by Elsevier Ltd.

  11. Adipose tissue derived mesenchymal stem cells for musculoskeletal repair in veterinary medicine

    PubMed Central

    Arnhold, Stefan; Wenisch, Sabine

    2015-01-01

    Adipose tissue derived stem cells (ASCs) are mesenchymal stem cells which can be obtained from different adipose tissue sources within the body. It is an abundant cell pool, which is easy accessible and the cells can be obtained in large numbers, cultivated and expanded in vitro and prepared for tissue engineering approaches, especially for skeletal tissue repair. In the recent years this cell population has attracted a great amount of attention among researchers in human as well as in veterinary medicine. In the meantime ASCs have been well characterized and their use in regenerative medicine is very well established. This review focuses on the characterization of ASCs for their use for tissue engineering approaches especially in veterinary medicine and also highlights a selection of clinical trials on the basis of ASCs as the relevant cell source. PMID:25973326

  12. Adipose tissue derived mesenchymal stem cells for musculoskeletal repair in veterinary medicine.

    PubMed

    Arnhold, Stefan; Wenisch, Sabine

    2015-01-01

    Adipose tissue derived stem cells (ASCs) are mesenchymal stem cells which can be obtained from different adipose tissue sources within the body. It is an abundant cell pool, which is easy accessible and the cells can be obtained in large numbers, cultivated and expanded in vitro and prepared for tissue engineering approaches, especially for skeletal tissue repair. In the recent years this cell population has attracted a great amount of attention among researchers in human as well as in veterinary medicine. In the meantime ASCs have been well characterized and their use in regenerative medicine is very well established. This review focuses on the characterization of ASCs for their use for tissue engineering approaches especially in veterinary medicine and also highlights a selection of clinical trials on the basis of ASCs as the relevant cell source.

  13. Clinical-Grade Manufacturing of Therapeutic Human Mesenchymal Stem/Stromal Cells in Microcarrier-Based Culture Systems.

    PubMed

    Fernandes-Platzgummer, Ana; Carmelo, Joana G; da Silva, Cláudia Lobato; Cabral, Joaquim M S

    2016-01-01

    The therapeutic potential of mesenchymal stem/stromal cells (MSC) has triggered the need for high cell doses in a vast number of clinical applications. This demand requires the development of good manufacturing practices (GMP)-compliant ex vivo expansion protocols that should be effective to deliver a robust and reproducible supply of clinical-grade cells in a safe and cost-effective manner. Controlled stirred-tank bioreactor systems under xenogeneic (xeno)-free culture conditions offer ideal settings to develop and optimize cell manufacturing to meet the standards and needs of human MSC for cellular therapies. Herein we describe two microcarrier-based stirred culture systems using spinner flasks and controlled stirred-tank bioreactors under xeno-free conditions for the efficient ex vivo expansion of human bone marrow and adipose tissue-derived MSC.

  14. Chronic hyperinsulinemia promotes meta-inflammation and extracellular matrix deposition in adipose tissue: Implications of nitric oxide.

    PubMed

    Kumar, Durgesh; Shankar, Kripa; Patel, Saraswati; Gupta, Abhishek; Varshney, Salil; Gupta, Sanchita; Rajan, Sujith; Srivastava, Ankita; Vishwakarma, Achchhe Lal; Gaikwad, Anil N

    2018-05-10

    Various imperative studies support the notion that hyperinsulinemia (HI) itself serves as the common link between adipose tissue inflammation (ATI) and metabolic syndrome. However, the contribution of HI mediated ATI and its metabolic consequences are yet to be explored. We induced chronic HI per se in mice by administration of exogenous insulin for 8 weeks through mini-osmotic pumps. For the reduction of circulating insulin in response to excess calorie intake, we have partially ablated β-cells by using streptozotocin (STZ) in the diet-induced obesity (DIO) and genetic mice models (db/db). Flow cytometry analysis was performed for the quantification of immune cells in stromal vascular fraction (SVF) isolated from epididymal white adipose tissue (eWAT). Our studies demonstrated that chronic HI augmented ATI in terms of elevated pro-inflammatory cells (M1 macrophages and NK-cells) and suppressed anti-inflammatory cells (M2 macrophages, eosinophils and regulatory T-cells). These results were correlated with altered obesity-associated metabolic phenotype. Partial reduction of circulating insulin level attenuated excess calorie-induced ATI and improved insulin sensitivity. Mechanistically, an imbalance in M1 and M2 macrophage proportions in eWAT promoted iNOS (inducible nitric oxide synthase): arginase-1 imbalance that resulted into extracellular matrix (ECM) deposition and insulin resistance (IR) development. However, iNOS -/- mice were protected from HI-induced M1:M2 macrophage imbalance, ECM deposition and IR in adipose tissue. Overall, we conclude that chronic HI per se contributed in ATI and iNOS corroborated ECM deposition. Copyright © 2018. Published by Elsevier B.V.

  15. Isolation and characterization of porcine adipose tissue-derived adult stem cells.

    PubMed

    Williams, Kellie J; Picou, Alicia A; Kish, Sharon L; Giraldo, Angelica M; Godke, Robert A; Bondioli, Kenneth R

    2008-01-01

    Stem cell characteristics such as self-renewal, differentiation and expression of CD34 and CD44 stem cell markers have not been identified in porcine adipose tissue-derived adult stem (ADAS) cells. The objective of this study was to develop a protocol for the isolation and culture of porcine adipose tissue-derived cells and to determine stem cell-like characteristics. Primary cultures were established and cell cultures were maintained. Cloning capacity was determined using a ring cloning procedure. Primary cultures and clones were differentiated and stained for multiple differentiated phenotypes. CD34 and CD44 messenger ribonucleic acid (mRNA) was isolated and reverse transcriptase polymerase chain reaction was used to compare expression profiles. An average of 2,700,000 nucleated cells/ml was isolated; 26% were adherent, and cells completed a cell cycle approximately every 3.3 days. Ring cloning identified 19 colonies. Primary cultures and clones were determined to differentiate along osteogenic, adipogenic and chondrogenic tissue lineages. The mRNA expression profiles showed CD34 expression was higher for undifferentiated ADAS cells versus differentiated cell types and the CD34 expression level was lower than that of CD44 among differentiated cells. Improved culture conditions and defined cellular characteristics of these porcine ADAS cells have been identified. Porcine ADAS can self-renew, can differentiate into multiple tissue lineages and they express CD34. Copyright 2008 S. Karger AG, Basel.

  16. A Novel Biological Approach to Treat Chondromalacia Patellae

    PubMed Central

    Lee, Sang Hee

    2013-01-01

    Mesenchymal stem cells from several sources (bone marrow, synovial tissue, cord blood, and adipose tissue) can differentiate into variable parts (bones, cartilage, muscle, and adipose tissue), representing a promising new therapy in regenerative medicine. In animal models, mesenchymal stem cells have been used successfully to regenerate cartilage and bones. However, there have been no follow-up studies on humans treated with adipose-tissue-derived stem cells (ADSCs) for the chondromalacia patellae. To obtain ADSCs, lipoaspirates were obtained from lower abdominal subcutaneous adipose tissue. The stromal vascular fraction was separated from the lipoaspirates by centrifugation after treatment with collagenase. The stem-cell-containing stromal vascular fraction was mixed with calcium chloride-activated platelet rich plasma and hyaluronic acid, and this ADSCs mixture was then injected under ultrasonic guidance into the retro-patellar joints of all three patients. Patients were subjected to pre- and post-treatment magnetic resonance imaging (MRI) scans. Pre- and post-treatment subjective pain scores and physical therapy assessments measured clinical changes. One month after the injection of autologous ADSCs, each patient's pain improved 50–70%. Three months after the treatment, the patients' pain improved 80–90%. The pain improvement persisted over 1 year, confirmed by telephone follow ups. Also, all three patients did not report any serious side effects. The repeated magnetic resonance imaging scans at three months showed improvement of the damaged tissues (softened cartilages) on the patellae-femoral joints. In patients with chondromalacia patellae who have continuous anterior knee pain, percutaneous injection of autologous ADSCs may play an important role in the restoration of the damaged tissues (softened cartilages). Thus, ADSCs treatment presents a glimpse of a new promising, effective, safe, and non-surgical method of treatment for chondromalacia patellae

  17. A novel biological approach to treat chondromalacia patellae.

    PubMed

    Pak, Jaewoo; Lee, Jung Hun; Lee, Sang Hee

    2013-01-01

    Mesenchymal stem cells from several sources (bone marrow, synovial tissue, cord blood, and adipose tissue) can differentiate into variable parts (bones, cartilage, muscle, and adipose tissue), representing a promising new therapy in regenerative medicine. In animal models, mesenchymal stem cells have been used successfully to regenerate cartilage and bones. However, there have been no follow-up studies on humans treated with adipose-tissue-derived stem cells (ADSCs) for the chondromalacia patellae. To obtain ADSCs, lipoaspirates were obtained from lower abdominal subcutaneous adipose tissue. The stromal vascular fraction was separated from the lipoaspirates by centrifugation after treatment with collagenase. The stem-cell-containing stromal vascular fraction was mixed with calcium chloride-activated platelet rich plasma and hyaluronic acid, and this ADSCs mixture was then injected under ultrasonic guidance into the retro-patellar joints of all three patients. Patients were subjected to pre- and post-treatment magnetic resonance imaging (MRI) scans. Pre- and post-treatment subjective pain scores and physical therapy assessments measured clinical changes. One month after the injection of autologous ADSCs, each patient's pain improved 50-70%. Three months after the treatment, the patients' pain improved 80-90%. The pain improvement persisted over 1 year, confirmed by telephone follow ups. Also, all three patients did not report any serious side effects. The repeated magnetic resonance imaging scans at three months showed improvement of the damaged tissues (softened cartilages) on the patellae-femoral joints. In patients with chondromalacia patellae who have continuous anterior knee pain, percutaneous injection of autologous ADSCs may play an important role in the restoration of the damaged tissues (softened cartilages). Thus, ADSCs treatment presents a glimpse of a new promising, effective, safe, and non-surgical method of treatment for chondromalacia patellae.

  18. FNDC5 attenuates adipose tissue inflammation and insulin resistance via AMPK-mediated macrophage polarization in obesity.

    PubMed

    Xiong, Xiao-Qing; Geng, Zhi; Zhou, Bing; Zhang, Feng; Han, Ying; Zhou, Ye-Bo; Wang, Jue-Jin; Gao, Xing-Ya; Chen, Qi; Li, Yue-Hua; Kang, Yu-Ming; Zhu, Guo-Qing

    2018-06-01

    Obesity-induced chronic inflammation is critical in the pathogenesis of insulin resistance, and the recruitment and proinflammatory activation of adipose tissue macrophages (ATMs) is important for the development of this process. Here, we examined the effects of fibronectin type III domain-containing 5 (FNDC5) on inflammation and insulin resistance in high-fat diet-induced obese mice. Male wild-type (WT) and FNDC5 -/- mice were fed with standard chow (Ctrl) or high fat diet (HFD) for 20 weeks to induce obesity and insulin resistance. Firstly, effects of FNDC5 gene deletion on obesity, insulin resistance, macrophage accumulation and polarization and adipose tissue inflammation were determined in mice. Secondly, the macrophage polarity shift was further examined with flow cytometry in isolated stromal vascular fraction (SVF). Thirdly, the effects of exogenous FNDC5 on lipopolysaccharide (LPS)-induced macrophage polarization, inflammation and the underlying signaling mechanism were investigated in RAW264.7 macrophages and primary mouse peritoneal cavity macrophages (PMs). Finally, the therapeutic effects of FNDC5 overexpression were examined in HFD-induced obese WT and FNDC5 -/- mice. FNDC5 gene deletion aggravated obesity, insulin resistance, fat accumulation and inflammation accompanied with enhanced AMPK inhibition, macrophages recruitment and M1 polarization in mice fed with HFD. Exogenous FNDC5 inhibited LPS-induced M1 macrophage polarization and inflammatory cytokine production via AMPK phosphorylation in both RAW264.7 macrophages and PMs. FNDC5 overexpression attenuated insulin resistance, AMPK inhibition, M1 macrophage polarization and inflammatory cytokine production in adipose tissue of obese WT and FNDC5 -/- mice. FNDC5 attenuates adipose tissue inflammation and insulin resistance via AMPK-mediated macrophage polarization in HFD-induced obesity. FNDC5 plays several beneficial roles in obesity and may be used as a therapeutic regimen for preventing

  19. Three-Dimensional Vascular Network Assembly From Diabetic Patient-Derived Induced Pluripotent Stem Cells.

    PubMed

    Chan, Xin Yi; Black, Rebecca; Dickerman, Kayla; Federico, Joseph; Lévesque, Mathieu; Mumm, Jeff; Gerecht, Sharon

    2015-12-01

    In diabetics, hyperglycemia results in deficient endothelial progenitors and cells, leading to cardiovascular complications. We aim to engineer 3-dimensional (3D) vascular networks in synthetic hydrogels from type 1 diabetes mellitus (T1D) patient-derived human-induced pluripotent stem cells (hiPSCs), to serve as a transformative autologous vascular therapy for diabetic patients. We validated and optimized an adherent, feeder-free differentiation procedure to derive early vascular cells (EVCs) with high portions of vascular endothelial cadherin-positive cells from hiPSCs. We demonstrate similar differentiation efficiency from hiPSCs derived from healthy donor and patients with T1D. T1D-hiPSC-derived vascular endothelial cadherin-positive cells can mature to functional endothelial cells-expressing mature markers: von Willebrand factor and endothelial nitric oxide synthase are capable of lectin binding and acetylated low-density lipoprotein uptake, form cords in Matrigel and respond to tumor necrosis factor-α. When embedded in engineered hyaluronic acid hydrogels, T1D-EVCs undergo morphogenesis and assemble into 3D networks. When encapsulated in a novel hypoxia-inducible hydrogel, T1D-EVCs respond to low oxygen and form 3D networks. As xenografts, T1D-EVCs incorporate into developing zebrafish vasculature. Using our robust protocol, we can direct efficient differentiation of T1D-hiPSC to EVCs. Early endothelial cells derived from T1D-hiPSC are functional when mature. T1D-EVCs self-assembled into 3D networks when embedded in hyaluronic acid and hypoxia-inducible hydrogels. The capability of T1D-EVCs to assemble into 3D networks in engineered matrices and to respond to a hypoxic microenvironment is a significant advancement for autologous vascular therapy in diabetic patients and has broad importance for tissue engineering. © 2015 American Heart Association, Inc.

  20. Cyclosporine-assisted adipose-derived mesenchymal stem cell therapy to mitigate acute kidney ischemia–reperfusion injury

    PubMed Central

    2013-01-01

    Introduction This study tested the hypothesis that cyclosporine (CsA)-supported syngeneic adipose-derived mesenchymal stem cell (ADMSC) therapy offered superior attenuation of acute ischemia–reperfusion (IR) kidney injury to either therapy alone. Methods Adult Sprague–Dawley rats (n = 40) were equally divided into group 1 (sham controls), group 2 (IR injury), group 3 (IR + CsA (20 mg/kg at 1 and 24 hours after procedure)), group 4 (syngeneic ADMSC (1.2×106) at 1, 6 and 24 hours after procedure), and group 5 (IR + CsA-ADMSC). Results By 72 hours after the IR procedure, the creatinine level and the ratio of urine protein to creatinine were highest in group 2 and lowest in group 1, and significantly higher in groups 3 and 4 than in group 5 (all P <0.05 for inter-group comparisons), but showed no differences between groups 3 and 4 (P >0.05). The inflammatory biomarkers at mRNA (matrix metalloproteinase-9, RANTES, TNF-α), protein (TNF-α, NF-κB, intercellular adhesion molecule-1, platelet-derived growth factor), and cellular (CD68+) levels of IR kidney showed a similar pattern compared with that of creatinine in all groups (all P <0.05 for inter-group comparisons). The protein expressions of oxidative stress (oxidized protein), reactive oxygen species (NADPH oxidases NOX-1, NOX-2), apoptosis (Bcl-2–associated X protein, caspase-3 and poly(ADP-ribose) polymerase) and DNA damage (phosphorylated H2A histone family member X-positive, proliferating cell nuclear antigen-positive cells) markers exhibited a pattern similar to that of inflammatory mediators amongst all groups (all P <0.05 for inter-group comparisons). Expressions of antioxidant biomarkers at cellular (glutathione peroxidase, glutathione reductase, heme oxygenase-1 (HO-1)) and protein (NADPH dehydrogenase (quinone)-1, HO-1, endothelial nitric oxide synthase) levels, and endothelial progenitor cell markers (C-X-C chemokine receptor type 4-positive, stromal cell-derived factor-1α-positive) were lowest in

  1. Cyclosporine-assisted adipose-derived mesenchymal stem cell therapy to mitigate acute kidney ischemia-reperfusion injury.

    PubMed

    Chen, Yen-Ta; Yang, Chih-Chau; Zhen, Yen-Yi; Wallace, Christopher Glenn; Yang, Jenq-Lin; Sun, Cheuk-Kwan; Tsai, Tzu-Hsien; Sheu, Jiunn-Jye; Chua, Sarah; Chang, Chia-Lo; Cho, Chung-Lung; Leu, Steve; Yip, Hon-Kan

    2013-05-31

    This study tested the hypothesis that cyclosporine (CsA)-supported syngeneic adipose-derived mesenchymal stem cell (ADMSC) therapy offered superior attenuation of acute ischemia-reperfusion (IR) kidney injury to either therapy alone. Adult Sprague-Dawley rats (n = 40) were equally divided into group 1 (sham controls), group 2 (IR injury), group 3 (IR + CsA (20 mg/kg at 1 and 24 hours after procedure)), group 4 (syngeneic ADMSC (1.2×106) at 1, 6 and 24 hours after procedure), and group 5 (IR + CsA-ADMSC). By 72 hours after the IR procedure, the creatinine level and the ratio of urine protein to creatinine were highest in group 2 and lowest in group 1, and significantly higher in groups 3 and 4 than in group 5 (all P <0.05 for inter-group comparisons), but showed no differences between groups 3 and 4 (P >0.05). The inflammatory biomarkers at mRNA (matrix metalloproteinase-9, RANTES, TNF-α), protein (TNF-α, NF-κB, intercellular adhesion molecule-1, platelet-derived growth factor), and cellular (CD68+) levels of IR kidney showed a similar pattern compared with that of creatinine in all groups (all P <0.05 for inter-group comparisons). The protein expressions of oxidative stress (oxidized protein), reactive oxygen species (NADPH oxidases NOX-1, NOX-2), apoptosis (Bcl-2-associated X protein, caspase-3 and poly(ADP-ribose) polymerase) and DNA damage (phosphorylated H2A histone family member X-positive, proliferating cell nuclear antigen-positive cells) markers exhibited a pattern similar to that of inflammatory mediators amongst all groups (all P <0.05 for inter-group comparisons). Expressions of antioxidant biomarkers at cellular (glutathione peroxidase, glutathione reductase, heme oxygenase-1 (HO-1)) and protein (NADPH dehydrogenase (quinone)-1, HO-1, endothelial nitric oxide synthase) levels, and endothelial progenitor cell markers (C-X-C chemokine receptor type 4-positive, stromal cell-derived factor-1α-positive) were lowest in groups 1 and 2, higher in groups 3

  2. Roles of Perivascular Adipose Tissue in the Pathogenesis of Atherosclerosis

    PubMed Central

    Tanaka, Kimie; Sata, Masataka

    2018-01-01

    Traditionally, it is believed that white adipose tissues serve as energy storage, heat insulation, and mechanical cushion, whereas non-shivering thermogenesis occurs in brown adipose tissue. Recent evidence revealed that adipose tissue secretes many types of cytokines, called as adipocytokines, which modulate glucose metabolism, lipid profile, appetite, fibrinolysis, blood pressure, and inflammation. Most of the arteries are surrounded by perivascular adipose tissue (PVAT). PVAT has been thought to be simply a structurally supportive tissue for vasculature. However, recent studies showed that PVAT influences vasodilation and vasocontraction, suggesting that PVAT regulates vascular tone and diameter. Adipocytokines secreted from PVAT appear to have direct access to the adjacent arterial wall by diffusion or via vasa vasorum. In fact, PVAT around atherosclerotic lesions and mechanically-injured arteries displayed inflammatory cytokine profiles, suggesting that PVAT functions to promote vascular lesion formation. Many clinical studies revealed that increased accumulation of epicardial adipose tissue (EAT), which surrounds coronary arteries, is associated with coronary artery disease. In this review article, we will summarize recent findings about potential roles of PVAT in the pathogenesis of atherosclerosis, particularly focusing on a series of basic and clinical studies from our laboratory. PMID:29487532

  3. Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sawada, Keigo; Takedachi, Masahide, E-mail: takedati@dent.osaka-u.ac.jp; Yamamoto, Satomi

    Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response.more » ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. - Highlights: • ADMPC-derived humoral factors stimulate cytodifferentiation of HPDLs. • ADMPCs secret growth factors including IGFBP6, VEGF and HGF. • IGFBP6 is involved in the promotion effect of ADMPC-CM on HPDL cytodifferentiation.« less

  4. Chemotaxis of primitive hematopoietic cells in response to stromal cell–derived factor-1

    PubMed Central

    Jo, Deog-Yeon; Rafii, Shahin; Hamada, Tsuneyoshi; Moore, Malcolm A.S.

    2000-01-01

    Stromal cell–derived factor-1 (SDF-1) provides a potent chemotactic stimulus for CD34+ hematopoietic cells. We cultured mobilized peripheral blood (PB) and umbilical cord blood (CB) for up to 5 weeks and examined the migratory activity of cobblestone area–forming cells (CAFCs) and long-term culture–initiating cells (LTC-ICs) in a transwell assay. In this system, SDF-1 or MS-5 marrow stromal cells placed in the lower chamber induced transmembrane and transendothelial migration by 2- and 5-week-old CAFCs and LTC-ICs in 3 hours. Transmigration was blocked by preincubation of input CD34+ cells with antibody to CXCR4. Transendothelial migration of CB CAFCs and LTC-ICs was higher than that of PB. We expanded CD34+ cells from CB in serum-free medium with thrombopoietin, flk-2 ligand, and c-kit ligand, with or without IL-3 and found that CAFCs cultured in the absence of IL-3 had a chemotactic response equivalent to noncultured cells, even after 5 weeks. However, addition of IL-3 to the culture reduced this response by 20–50%. These data indicate that SDF-1 induces chemotaxis of primitive hematopoietic cells signaling through CXCR4 and that the chemoattraction could be downmodulated by culture ex vivo. PMID:10619866

  5. Nitric oxide regulates tumor cell cross-talk with stromal cells in the tumor microenvironment of the liver.

    PubMed

    Decker, Ningling Kang; Abdelmoneim, Soha S; Yaqoob, Usman; Hendrickson, Helen; Hormes, Joe; Bentley, Mike; Pitot, Henry; Urrutia, Raul; Gores, Greg J; Shah, Vijay H

    2008-10-01

    Tumor progression is regulated through paracrine interactions between tumor cells and stromal cells in the microenvironment, including endothelial cells and myofibroblasts. Nitric oxide (NO) is a key molecule in the regulation of tumor-microenvironment interactions, although its precise role is incompletely defined. By using complementary in vitro and in vivo approaches, we studied the effect of endothelial NO synthase (eNOS)-derived NO on liver tumor growth and metastasis in relation to adjacent stromal myofibroblasts and matrix because liver tumors maintain a rich, vascular stromal network enriched with phenotypically heterogeneous myofibroblasts. Mice with an eNOS deficiency developed liver tumors more frequently in response to carcinogens compared with control animals. In a surgical model of pancreatic cancer liver metastasis, eNOS overexpression in the tumor microenvironment attenuated both the number and size of tumor implants. NO promoted anoikis of tumor cells in vitro and limited their invasive capacity. Because tumor cell anoikis and invasion are both regulated by myofibroblast-derived matrix, we explored the effect of NO on tumor cell protease expression. Both microarray and Western blot analysis revealed eNOS-dependent down-regulation of the matrix protease cathepsin B within tumor cells, and silencing of cathepsin B attenuated tumor cell invasive capacity in a similar manner to that observed with eNOS overexpression. Thus, a NO gradient within the tumor microenvironment influences tumor progression through orchestrated molecular interactions between tumor cells and stroma.

  6. Directing bone marrow-derived stromal cell function with mechanics.

    PubMed

    Potier, E; Noailly, J; Ito, K

    2010-03-22

    Because bone marrow-derived stromal cells (BMSCs) are able to generate many cell types, they are envisioned as source of regenerative cells to repair numerous tissues, including bone, cartilage, and ligaments. Success of BMSC-based therapies, however, relies on a number of methodological improvements, among which better understanding and control of the BMSC differentiation pathways. Since many years, the biochemical environment is known to govern BMSC differentiation, but more recent evidences show that the biomechanical environment is also directing cell functions. Using in vitro systems that aim to reproduce selected components of the in vivo mechanical environment, it was demonstrated that mechanical loadings can affect BMSC proliferation and improve the osteogenic, chondrogenic, or myogenic phenotype of BMSCs. These effects, however, seem to be modulated by parameters other than mechanics, such as substrate nature or soluble biochemical environment. This paper reviews and discusses recent experimental data showing that despite some knowledge limitation, mechanical stimulation already constitutes an additional and efficient tool to drive BMSC differentiation. Copyright (c) 2009 Elsevier Ltd. All rights reserved.

  7. Human umbilical cord mesenchymal stromal cells suppress MHC class II expression on rat vascular endothelium and prolong survival time of cardiac allograft

    PubMed Central

    Qiu, Ying; Yun, Mark M; Han, Xia; Zhao, Ruidong; Zhou, Erxia; Yun, Sheng

    2014-01-01

    Background: Human umbilical cord mesenchymal stromal cells (UC-MSCs) have low immunogenicity and immune regulation. To investigate immunomodulatory effects of human UC-MSCs on MHC class II expression and allograft, we transplanted heart of transgenic rats with MHC class II expression on vascular endothelium. Methods: UC-MSCs were obtained from human umbilical cords and confirmed with flow cytometry analysis. Transgenic rat line was established using the construct of human MHC class II transactivator gene (CIITA) under mouse ICAM-2 promoter control. The induced MHC class II expression on transgenic rat vascular endothelial cells (VECs) was assessed with immunohistological staining. And the survival time of cardiac allograft was compared between the recipients with and without UC-MSC transfusion. Results: Flow cytometry confirmed that the human UC-MSCs were positive for CD29, CD44, CD73, CD90, CD105, CD271, and negative for CD34 and HLA-DR. Repeated infusion of human UC-MSCs reduced MHC class II expression on vascular endothelia of transplanted hearts, and increased survival time of allograft. The UC-MSCs increased regulatory cytokines IL10, transforming growth factor (TGF)-β1 and suppressed proinflammatory cytokines IL2 and IFN-γ in vivo. The UC-MSC culture supernatant had similar effects on cytokine expression, and decreased lymphocyte proliferation in vitro. Conclusions: Repeated transfusion of the human UC-MSCs reduced MHC class II expression on vascular endothelia and prolonged the survival time of rat cardiac allograft. PMID:25126177

  8. Stromal-Based Signatures for the Classification of Gastric Cancer.

    PubMed

    Uhlik, Mark T; Liu, Jiangang; Falcon, Beverly L; Iyer, Seema; Stewart, Julie; Celikkaya, Hilal; O'Mahony, Marguerita; Sevinsky, Christopher; Lowes, Christina; Douglass, Larry; Jeffries, Cynthia; Bodenmiller, Diane; Chintharlapalli, Sudhakar; Fischl, Anthony; Gerald, Damien; Xue, Qi; Lee, Jee-Yun; Santamaria-Pang, Alberto; Al-Kofahi, Yousef; Sui, Yunxia; Desai, Keyur; Doman, Thompson; Aggarwal, Amit; Carter, Julia H; Pytowski, Bronislaw; Jaminet, Shou-Ching; Ginty, Fiona; Nasir, Aejaz; Nagy, Janice A; Dvorak, Harold F; Benjamin, Laura E

    2016-05-01

    Treatment of metastatic gastric cancer typically involves chemotherapy and monoclonal antibodies targeting HER2 (ERBB2) and VEGFR2 (KDR). However, reliable methods to identify patients who would benefit most from a combination of treatment modalities targeting the tumor stroma, including new immunotherapy approaches, are still lacking. Therefore, we integrated a mouse model of stromal activation and gastric cancer genomic information to identify gene expression signatures that may inform treatment strategies. We generated a mouse model in which VEGF-A is expressed via adenovirus, enabling a stromal response marked by immune infiltration and angiogenesis at the injection site, and identified distinct stromal gene expression signatures. With these data, we designed multiplexed IHC assays that were applied to human primary gastric tumors and classified each tumor to a dominant stromal phenotype representative of the vascular and immune diversity found in gastric cancer. We also refined the stromal gene signatures and explored their relation to the dominant patient phenotypes identified by recent large-scale studies of gastric cancer genomics (The Cancer Genome Atlas and Asian Cancer Research Group), revealing four distinct stromal phenotypes. Collectively, these findings suggest that a genomics-based systems approach focused on the tumor stroma can be used to discover putative predictive biomarkers of treatment response, especially to antiangiogenesis agents and immunotherapy, thus offering an opportunity to improve patient stratification. Cancer Res; 76(9); 2573-86. ©2016 AACR. ©2016 American Association for Cancer Research.

  9. Three-dimensional micro computed tomography analysis of the lung vasculature and differential adipose proteomics in the Sugen/hypoxia rat model of pulmonary arterial hypertension.

    PubMed

    Shields, Kelly J; Verdelis, Kostas; Passineau, Michael J; Faight, Erin M; Zourelias, Lee; Wu, Changgong; Chong, Rong; Benza, Raymond L

    2016-12-01

    Pulmonary arterial hypertension (PAH) is a rare disease characterized by significant vascular remodeling. The obesity epidemic has produced great interest in the relationship between small visceral adipose tissue depots producing localized inflammatory conditions, which may link metabolism, innate immunity, and vascular remodeling. This study used novel micro computed tomography (microCT) three-dimensional modeling to investigate the degree of remodeling of the lung vasculature and differential proteomics to determine small visceral adipose dysfunction in rats with severe PAH. Sprague-Dawley rats were subjected to a subcutaneous injection of vascular endothelial growth factor receptor blocker (Sugen 5416) with subsequent hypoxia exposure for 3 weeks (SU/hyp). At 12 weeks after hypoxia, microCT analysis showed a decrease in the ratio of vascular to total tissue volume within the SU/hyp group (mean ± standard deviation: 0.27 ± 0.066; P = 0.02) with increased vascular separation (0.37 ± 0.062 mm; P = 0.02) when compared with the control (0.34 ± 0.084 and 0.30 ± 0.072 mm). Differential proteomics detected an up-regulation of complement protein 3 (C3; SU/hyp∶control ratio = 2.86) and the adipose tissue-specific fatty acid binding protein-4 (FABP4, 2.66) in the heart adipose of the SU/hyp. Significant remodeling of the lung vasculature validates the efficacy of the SU/hyp rat for modeling human PAH. The upregulation of C3 and FABP4 within the heart adipose implicates small visceral adipose dysfunction. C3 has been associated with vascular stiffness, and FABP4 suppresses peroxisome proliferator-activated receptor, which is a major regulator of adipose function and known to be downregulated in PAH. These findings reveal that small visceral adipose tissue within the SU/hyp model provides mechanistic links for vascular remodeling and adipose dysfunction in the pathophysiology of PAH.

  10. Ginsenoside Rg1 and platelet-rich fibrin enhance human breast adipose-derived stem cells function for soft tissue regeneration

    PubMed Central

    Li, Hong-Mian; Peng, Qi-Liu; Huang, Min-Hong; Li, De-Quan; Liang, Yi-Dan; Chi, Gang-Yi; Li, De-Hui; Yu, Bing-Chao; Huang, Ji-Rong

    2016-01-01

    Adipose-derived stem cells (ASCs) can be used to repair soft tissue defects, wounds, burns, and scars and to regenerate various damaged tissues. The cell differentiation capacity of ASCs is crucial for engineered adipose tissue regeneration in reconstructive and plastic surgery. We previously reported that ginsenoside Rg1 (G-Rg1 or Rg1) promotes proliferation and differentiation of ASCs in vitro and in vivio. Here we show that both G-Rg1 and platelet-rich fibrin (PRF) improve the proliferation, differentiation, and soft tissue regeneration capacity of human breast adipose-derived stem cells (HBASCs) on collagen type I sponge scaffolds in vitro and in vivo. Three months after transplantation, tissue wet weight, adipocyte number, intracellular lipid, microvessel density, and gene and protein expression of VEGF, HIF-1α, and PPARγ were higher in both G-Rg1- and PRF-treated HBASCs than in control grafts. More extensive new adipose tissue formation was evident after treatment with G-Rg1 or PRF. In summary, G-Rg1 and/or PRF co-administration improves the function of HBASCs for soft tissue regeneration engineering. PMID:27191987

  11. Human adipose tissue-derived mesenchymal stem cells differentiate into insulin, somatostatin, and glucagon expressing cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Timper, Katharina; Seboek, Dalma; Eberhardt, Michael

    2006-03-24

    Mesenchymal stem cells (MSC) from mouse bone marrow were shown to adopt a pancreatic endocrine phenotype in vitro and to reverse diabetes in an animal model. MSC from human bone marrow and adipose tissue represent very similar cell populations with comparable phenotypes. Adipose tissue is abundant and easily accessible and could thus also harbor cells with the potential to differentiate in insulin producing cells. We isolated human adipose tissue-derived MSC from four healthy donors. During the proliferation period, the cells expressed the stem cell markers nestin, ABCG2, SCF, Thy-1 as well as the pancreatic endocrine transcription factor Isl-1. The cellsmore » were induced to differentiate into a pancreatic endocrine phenotype by defined culture conditions within 3 days. Using quantitative PCR a down-regulation of ABCG2 and up-regulation of pancreatic developmental transcription factors Isl-1, Ipf-1, and Ngn3 were observed together with induction of the islet hormones insulin, glucagon, and somatostatin.« less

  12. Alginate-Encapsulation for the Improved Hypothermic Preservation of Human Adipose-Derived Stem Cells

    PubMed Central

    Swioklo, Stephen; Constantinescu, Andrei

    2016-01-01

    Despite considerable progress within the cell therapy industry, unmet bioprocessing and logistical challenges associated with the storage and distribution of cells between sites of manufacture and the clinic exist. We examined whether hypothermic (4°C–23°C) preservation of human adipose-derived stem cells could be improved through their encapsulation in 1.2% calcium alginate. Alginate encapsulation improved the recovery of viable cells after 72 hours of storage. Viable cell recovery was highly temperature-dependent, with an optimum temperature of 15°C. At this temperature, alginate encapsulation preserved the ability for recovered cells to attach to tissue culture plastic on rewarming, further increasing its effect on total cell recovery. On attachment, the cells were phenotypically normal, displayed normal growth kinetics, and maintained their capacity for trilineage differentiation. The number of cells encapsulated (up to 2 × 106 cells per milliliter) did not affect viable cell recovery nor did storage of encapsulated cells in a xeno-free, serum-free,current Good Manufacturing Practice-grade medium. We present a simple, low-cost system capable of enhancing the preservation of human adipose-derived stem cells stored at hypothermic temperatures, while maintaining their normal function. The storage of cells in this manner has great potential for extending the time windows for quality assurance and efficacy testing, distribution between the sites of manufacture and the clinic, and reducing the wastage associated with the limited shelf life of cells stored in their liquid state. Significance Despite considerable advancement in the clinical application of cell-based therapies, major logistical challenges exist throughout the cell therapy supply chain associated with the storage and distribution of cells between the sites of manufacture and the clinic. A simple, low-cost system capable of preserving the viability and functionality of human adipose-derived stem

  13. Myeloma-derived macrophage inhibitory factor regulates bone marrow stromal cell-derived IL-6 via c-MYC.

    PubMed

    Piddock, Rachel E; Marlein, Christopher R; Abdul-Aziz, Amina; Shafat, Manar S; Auger, Martin J; Bowles, Kristian M; Rushworth, Stuart A

    2018-05-16

    Multiple myeloma (MM) remains an incurable malignancy despite the recent advancements in its treatment. The protective effects of the niche in which it develops has been well documented; however, little has been done to investigate the MM cell's ability to 're-program' cells within its environment to benefit disease progression. Here, we show that MM-derived macrophage migratory inhibitory factor (MIF) stimulates bone marrow stromal cells to produce the disease critical cytokines IL-6 and IL-8, prior to any cell-cell contact. Furthermore, we provide evidence that this IL-6/8 production is mediated by the transcription factor cMYC. Pharmacological inhibition of cMYC in vivo using JQ1 led to significantly decreased levels of serum IL-6-a highly positive prognostic marker in MM patients. Our presented findings show that MM-derived MIF causes BMSC secretion of IL-6 and IL-8 via BMSC cMYC. Furthermore, we show that the cMYC inhibitor JQ1 can reduce BMSC secreted IL-6 in vivo, irrespective of tumor burden. These data provide evidence for the clinical evaluation of both MIF and cMYC inhibitors in the treatment of MM.

  14. Activin B regulates adipose-derived mesenchymal stem cells to promote skin wound healing via activation of the MAPK signaling pathway.

    PubMed

    Zhang, Lei; Xu, Pengcheng; Wang, Xueer; Zhang, Min; Yan, Yuan; Chen, Yinghua; Zhang, Lu; Zhang, Lin

    2017-06-01

    Adipose-derived stem cells (ADSCs) are multipotent stromal cells that can differentiate into a variety of cell types, including skin cells, and they can provide an abundant source of cells for skin tissue engineering and skin wound healing. The purpose of this study is to explore the therapeutic effects of activin B in combination with ADSCs and the possible signaling mechanism. In this study, we found that activin B was able to promote ADSC migration by inducing actin stress fiber formation in vitro. In vivo, activin B in combination with ADSCs was capable of enhancing α-SMA expression and wound closure. This combined treatment also promoted fibroblast and keratinocyte proliferation and accelerated re-epithelialization and collagen deposition. Moreover, activin B in combination with ADSCs boosted angiogenesis in the wound area. Further study of the mechanism revealed that activation of JNK and ERK signaling, but not p38 signaling, were required for activin B-induced ADSC actin stress fiber formation and cell migration. These results showed that activin B was able to activate JNK and ERK signaling pathways to induce actin stress fiber formation and ADSC migration to promote wound healing. These results suggest that combined treatment with activin B and ADSCs is a promising therapeutic strategy for the management of serious skin wounds. Copyright © 2017. Published by Elsevier Ltd.

  15. Human adipose-derived stem cells ameliorate repetitive behavior, social deficit and anxiety in a VPA-induced autism mouse model.

    PubMed

    Ha, Sungji; Park, Hyunjun; Mahmood, Usman; Ra, Jeong Chan; Suh, Yoo-Hun; Chang, Keun-A

    2017-01-15

    Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder characterized by impairments in social interaction and communication, and patients often display co-occurring repetitive behaviors. Although the global prevalence of ASD has increased over time, the etiology and treatments for ASD are poorly understood. Recently, some researchers have suggested that stem cells have therapeutic potential for ASD. Thus, in the present study, we investigated the therapeutic effects of human adipose-derived stem cells (hASCs), a kind of autologous mesenchymal stem cells (MSCs) isolated from adipose tissue, on valproic acid (VPA)-induced autism model mice. Human ASCs were injected into the neonatal pups (P2 or P3) intraventricularly and then we evaluated major behavior symptoms of ASD. VPA-treated mice showed increased repetitive behaviors, decreased social interactions and increased anxiety but these autistic behaviors were ameliorated through transplantation of hASCs. In addition, hASCs transplantation restored the alteration of phosphatase and tensin homolog (PTEN) expression and p-AKT/AKT ratio in the brains of VPA-induced ASD model mice. The decreased level of vascular endothelial growth factor (VEGF) and interleukin 10 (IL-10) by VPA were rescued in the brains of the hASC-injected VPA mice. With these results, we experimentally found hASCs' therapeutic effects on autistic phenotypes in a ASD model mice for the first time. This animal model system can be used to elucidate further mechanisms of therapeutic effects of hASCs in ASD. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Carotenoids in Adipose Tissue Biology and Obesity.

    PubMed

    Bonet, M Luisa; Canas, Jose A; Ribot, Joan; Palou, Andreu

    2016-01-01

    Cell, animal and human studies dealing with carotenoids and carotenoid derivatives as nutritional regulators of adipose tissue biology with implications for the etiology and management of obesity and obesity-related metabolic diseases are reviewed. Most studied carotenoids in this context are β-carotene, cryptoxanthin, astaxanthin and fucoxanthin, together with β-carotene-derived retinoids and some other apocarotenoids. Studies indicate an impact of these compounds on essential aspects of adipose tissue biology including the control of adipocyte differentiation (adipogenesis), adipocyte metabolism, oxidative stress and the production of adipose tissue-derived regulatory signals and inflammatory mediators. Specific carotenoids and carotenoid derivatives restrain adipogenesis and adipocyte hypertrophy while enhancing fat oxidation and energy dissipation in brown and white adipocytes, and counteract obesity in animal models. Intake, blood levels and adipocyte content of carotenoids are reduced in human obesity. Specifically designed human intervention studies in the field, though still sparse, indicate a beneficial effect of carotenoid supplementation in the accrual of abdominal adiposity. In summary, studies support a role of specific carotenoids and carotenoid derivatives in the prevention of excess adiposity, and suggest that carotenoid requirements may be dependent on body composition.

  17. Peripheral Motor and Sensory Nerve Conduction following Transplantation of Undifferentiated Autologous Adipose Tissue-Derived Stem Cells in a Biodegradable U.S. Food and Drug Administration-Approved Nerve Conduit.

    PubMed

    Klein, Silvan M; Vykoukal, Jody; Li, De-Pei; Pan, Hui-Lin; Zeitler, Katharina; Alt, Eckhard; Geis, Sebastian; Felthaus, Oliver; Prantl, Lukas

    2016-07-01

    Conduits preseeded with either Schwann cells or stem cells differentiated into Schwann cells demonstrated promising results for the outcome of nerve regeneration in nerve defects. The concept of this trial combines nerve repair by means of a commercially available nerve guidance conduit and preseeding with autologous, undifferentiated, adipose tissue-derived stem cells. Adipose tissue-derived stem cells were harvested from rats and subsequently seeded onto a U.S. Food and Drug Administration-approved type I collagen conduit. Sciatic nerve gaps 10 mm in length were created, and nerve repair was performed by the transplantation of either conduits preseeded with autologous adipose tissue-derived stem cells or acellular (control group) conduits. After 6 months, the motor and sensory nerve conduction velocity were assessed. Nerves were removed and examined by hematoxylin and eosin, van Gieson, and immunohistochemistry (S100 protein) staining for the quality of axonal regeneration. Nerve gaps treated with adipose tissue-derived stem cells showed superior nerve regeneration, reflected by higher motor and sensory nerve conduction velocity values. The motor and sensory nerve conduction velocity were significantly greater in nerves treated with conduits preseeded with adipose tissue-derived stem cells than in nerves treated with conduits alone (p < 0.05). Increased S100 immunoreactivity was detected for the adipose tissue-derived stem cell group. In this group, axon arrangement inside the conduits was more organized. Transplantation of adipose tissue-derived stem cells significantly improves motor and sensory nerve conduction velocity in peripheral nerve gaps. Preseeded conduits showed a more organized axon arrangement inside the conduit in comparison with nerve conduits alone. The approach used here could readily be translated into a clinical therapy. Therapeutic, V.

  18. Angiogenic Synergistic Effect of Adipose-Derived Stromal Cell Spheroids with Low-Level Light Therapy in a Model of Acute Skin Flap Ischemia.

    PubMed

    Park, In-Su; Chung, Phil-Sang; Ahn, Jin Chul

    2016-01-01

    Human adipose-derived mesenchymal stem cells (hASCs) are an attractive cell source for tissue engineering. However, one obstacle to this approach is that the transplanted hASC population can decline rapidly in the recipient tissue. The aim of this study was to investigate the effects of low-level light therapy (LLLT) on transplanted spheroid hASCs in skin flaps of mice. hASCs were cultured in monolayers or spheroids. LLLT, hASCs, spheroids and spheroids transplanted with LLLT were applied to the skin flaps. Healing of the skin flaps was assessed by gross evaluation and by hematoxylin and eosin staining and elastin van Gieson staining. Compared with the spheroid group, skin flap healing was enhanced in the spheroid + LLLT group, including the neovascularization and regeneration of skin appendages. The survival of hASCs was enhanced by decreased apoptosis of hASCs in the skin flaps of the spheroid + LLLT group. The secretion of growth factors was stimulated in the spheroid + LLLT group compared with the ASC and spheroid groups. These data suggest that LLLT was an effective biostimulator of spheroid hASCs in the skin flaps, enhancing the survival of hASCs and stimulating the secretion of growth factors. © 2016 S. Karger AG, Basel.

  19. Stromal laminin chain distribution in normal, hyperplastic and malignant oral mucosa: relation to myofibroblast occurrence and vessel formation.

    PubMed

    Franz, Marcus; Wolheim, Anke; Richter, Petra; Umbreit, Claudia; Dahse, Regine; Driemel, Oliver; Hyckel, Peter; Virtanen, Ismo; Kosmehl, Hartwig; Berndt, Alexander

    2010-04-01

    The contribution of stromal laminin chain expression to malignant potential, tumour stroma reorganization and vessel formation in oral squamous cell carcinoma (OSCC) is not fully understood. Therefore, the expression of the laminin chains alpha2, alpha3, alpha4, alpha5 and gamma2 in the stromal compartment/vascular structures in OSCC was analysed. Frozen tissue of OSCC (9x G1, 24x G2, 8x G3) and normal (2x)/hyperplastic (11x) oral mucosa was subjected to laminin chain and alpha-smooth muscle actin (ASMA) immunohistochemistry. Results were correlated to tumour grade. The relation of laminin chain positive vessels to total vessel number was assessed by immunofluorescence double labelling with CD31. Stromal laminin alpha2 chain significantly decreases and alpha3, alpha4, alpha5 and gamma2 chains and also ASMA significantly increase with rising grade. The amount of stromal alpha3, alpha4 and gamma2 chains significantly increased with rising ASMA positivity. There is a significant decrease in alpha3 chain positive vessels with neoplastic transformation. Mediated by myofibroblasts, OSCC development is associated with a stromal up-regulation of laminin isoforms possibly contributing to a migration promoting microenvironment. A vascular basement membrane reorganization concerning alpha3 and gamma2 chain laminins during tumour angioneogenesis is suggested.

  20. Adipose-derived mesenchymal stem cells accelerate nerve regeneration and functional recovery in a rat model of recurrent laryngeal nerve injury.

    PubMed

    Li, Yun; Xu, Wen; Cheng, Li-Yu

    2017-09-01

    Medialization thyroplasty or injection laryngoplasty for unilateral vocal fold paralysis cannot restore mobility of the vocal fold. Recent studies have shown that transplantation of mesenchymal stem cells is effective in the repair of nerve injuries. This study investigated whether adipose-derived stem cell transplantation could repair recurrent laryngeal nerve injury. Rat models of recurrent laryngeal nerve injury were established by crushing with micro forceps. Adipose-derived mesenchymal stem cells (ADSCs; 8 × 10 5 ) or differentiated Schwann-like adipose-derived mesenchymal stem cells (dADSCs; 8 × 10 5 ) or extracellular matrix were injected at the site of injury. At 2, 4 and 6 weeks post-surgery, a higher density of myelinated nerve fiber, thicker myelin sheath, improved vocal fold movement, better recovery of nerve conduction capacity and reduced thyroarytenoid muscle atrophy were found in ADSCs and dADSCs groups compared with the extracellular matrix group. The effects were more pronounced in the ADSCs group than in the dADSCs group. These experimental results indicated that ADSCs transplantation could be an early interventional strategy to promote regeneration after recurrent laryngeal nerve injury.

  1. 3D printed scaffolds of calcium silicate-doped β-TCP synergize with co-cultured endothelial and stromal cells to promote vascularization and bone formation.

    PubMed

    Deng, Yuan; Jiang, Chuan; Li, Cuidi; Li, Tao; Peng, Mingzheng; Wang, Jinwu; Dai, Kerong

    2017-07-17

    Synthetic bone scaffolds have potential application in repairing large bone defects, however, inefficient vascularization after implantation remains the major issue of graft failure. Herein, porous β-tricalcium phosphate (β-TCP) scaffolds with calcium silicate (CS) were 3D printed, and pre-seeded with co-cultured human umbilical cord vein endothelial cells (HUVECs) and human bone marrow stromal cells (hBMSCs) to construct tissue engineering scaffolds with accelerated vascularization and better bone formation. Results showed that in vitro β-TCP scaffolds doped with 5% CS (5%CS/β-TCP) were biocompatible, and stimulated angiogenesis and osteogenesis. The results also showed that 5%CS/β-TCP scaffolds not only stimulated co-cultured cells angiogenesis on Matrigel, but also stimulated co-cultured cells to form microcapillary-like structures on scaffolds, and promoted migration of BMSCs by stimulating co-cultured cells to secrete PDGF-BB and CXCL12 into the surrounding environment. Moreover, 5%CS/β-TCP scaffolds enhanced vascularization and osteoinduction in comparison with β-TCP, and synergized with co-cultured cells to further increase early vessel formation, which was accompanied by earlier and better ectopic bone formation when implanted subcutaneously in nude mice. Thus, our findings suggest that porous 5%CS/β-TCP scaffolds seeded with co-cultured cells provide new strategy for accelerating tissue engineering scaffolds vascularization and osteogenesis, and show potential as treatment for large bone defects.

  2. Different Procoagulant Activity of Therapeutic Mesenchymal Stromal Cells Derived from Bone Marrow and Placental Decidua.

    PubMed

    Moll, Guido; Ignatowicz, Lech; Catar, Rusan; Luecht, Christian; Sadeghi, Behnam; Hamad, Osama; Jungebluth, Philipp; Dragun, Duska; Schmidtchen, Artur; Ringdén, Olle

    2015-10-01

    While therapeutic mesenchymal stromal/stem cells (MSCs) have usually been obtained from bone marrow, perinatal tissues have emerged as promising new sources of cells for stromal cell therapy. In this study, we present a first safety follow-up on our clinical experience with placenta-derived decidual stromal cells (DSCs), used as supportive immunomodulatory and regenerative therapy for patients with severe complications after allogeneic hematopoietic stem cell transplantation (HSCT). We found that DSCs are smaller, almost half the volume of MSCs, which may favor microvascular passage. DSCs also show different hemocompatibility, with increased triggering of the clotting cascade after exposure to human blood and plasma in vitro. After infusion of DSCs in HSCT patients, we observed a weak activation of the fibrinolytic system, but the other blood activation markers remained stable, excluding major adverse events. Expression profiling identified differential levels of key factors implicated in regulation of hemostasis, such as a lack of prostacyclin synthase and increased tissue factor expression in DSCs, suggesting that these cells have intrinsic blood-activating properties. The stronger triggering of the clotting cascade by DSCs could be antagonized by optimizing the cell graft reconstitution before infusion, for example, by use of low-dose heparin anticoagulant in the cell infusion buffer. We conclude that DSCs are smaller and have stronger hemostatic properties than MSCs, thus triggering stronger activation of the clotting system, which can be antagonized by optimizing the cell graft preparation before infusion. Our results highlight the importance of hemocompatibility safety testing for every novel cell therapy product before clinical use, when applied using systemic delivery.

  3. Tendon tissue engineering: Adipose 1 derived stem cell and GDF-5 mediated regeneration using electrospun matrix systems

    PubMed Central

    James, R; Kumbar, S G; Laurencin, C T; Balian, G; Chhabra, A B

    2011-01-01

    Tendon tissue engineering with a biomaterial scaffold that mimics the tendon extracellular matrix (ECM) and is biomechanically suitable when combined with readily available autologous cells may provide successful regeneration of defects in tendon. Current repair strategies using suitable autografts and freeze-dried allografts lead to a slow repair process that is sub-optimal and fails to restore function, particularly in difficult clinical situations such as zone II flexor tendon injuries of the hand. We have investigated the effect of GDF-5 on cell proliferation and gene expression by primary rat adipose-derived stromal cells (ADSCs) that were cultured on poly(DL-lactide-co-glycolide) PLAGA fiber scaffold and compared to PLAGA 2D film scaffold. The electrospun scaffold mimics the collagen fiber bundles present in native tendon tissue, and supports the adhesion and proliferation of multipotent ADSCs. Gene expression of scleraxis, the neotendon marker was upregulated 7 – 8 fold at 1 week with GDF-5 treatment when cultured on 3D electrospun scaffold, and was significantly higher at 2 weeks compared to 2D films with or without GDF-5 treatment. Expression of the genes that encode the major tendon ECM protein, collagen type I, was increased by 4 fold starting at 1 week on treatment with 100ng/mL GDF-5, and at all time points the expression was significantly higher compared to 2D films irrespective of GDF-5 treatment. Thus stimulation with GDF-5 can modulate primary ADSCs on PLAGA fiber scaffold to produce a soft, collagenous musculoskeletal tissue that fulfills the need for tendon regeneration. PMID:21436509

  4. Decellularized silk fibroin scaffold primed with adipose mesenchymal stromal cells improves wound healing in diabetic mice.

    PubMed

    Navone, Stefania Elena; Pascucci, Luisa; Dossena, Marta; Ferri, Anna; Invernici, Gloria; Acerbi, Francesco; Cristini, Silvia; Bedini, Gloria; Tosetti, Valentina; Ceserani, Valentina; Bonomi, Arianna; Pessina, Augusto; Freddi, Giuliano; Alessandrino, Antonio; Ceccarelli, Piero; Campanella, Rolando; Marfia, Giovanni; Alessandri, Giulio; Parati, Eugenio Agostino

    2014-01-14

    Silk fibroin (SF) scaffolds have been shown to be a suitable substrate for tissue engineering and to improve tissue regeneration when cellularized with mesenchymal stromal cells (MSCs). We here demonstrate, for the first time, that electrospun nanofibrous SF patches cellularized with human adipose-derived MSCs (Ad-MSCs-SF), or decellularized (D-Ad-MSCs-SF), are effective in the treatment of skin wounds, improving skin regeneration in db/db diabetic mice. The conformational and structural analyses of SF and D-Ad-MSCs-SF patches were performed by scanning electron microscopy, confocal microscopy, Fourier transform infrared spectroscopy and differential scanning calorimetry. Wounds were performed by a 5 mm punch biopsy tool on the mouse's back. Ad-MSCs-SF and D-Ad-MSCs-SF patches were transplanted and the efficacy of treatments was assessed by measuring the wound closure area, by histological examination and by gene expression profile. We further investigated the in vitro angiogenic properties of Ad-MSCs-SF and D-Ad-MSCs-SF patches by affecting migration of human umbilical vein endothelial cells (HUVECs), keratinocytes (KCs) and dermal fibroblasts (DFs), through the aortic ring assay and, finally, by evaluating the release of angiogenic factors. We found that Ad-MSCs adhere and grow on SF, maintaining their phenotypic mesenchymal profile and differentiation capacity. Conformational and structural analyses on SF and D-Ad-MSCs-SF samples, showed that sterilization, decellularization, freezing and storing did not affect the SF structure. When grafted in wounds of diabetic mice, both Ad-MSCs-SF and D-Ad-MSCs-SF significantly improved tissue regeneration, reducing the wound area respectively by 40% and 35%, within three days, completing the process in around 10 days compared to 15-17 days of controls. RT2 gene profile analysis of the wounds treated with Ad-MSCs-SF and D-Ad-MSCs-SF showed an increment of genes involved in angiogenesis and matrix remodeling. Finally, Ad

  5. Gap junction communication between uterine stromal cells plays a critical role in pregnancy-associated neovascularization and embryo survival.

    PubMed

    Laws, Mary J; Taylor, Robert N; Sidell, Neil; DeMayo, Francesco J; Lydon, John P; Gutstein, David E; Bagchi, Milan K; Bagchi, Indrani C

    2008-08-01

    In the uterus, the formation of new maternal blood vessels in the stromal compartment at the time of embryonic implantation is critical for the establishment and maintenance of pregnancy. Although uterine angiogenesis is known to be influenced by the steroid hormones estrogen (E) and progesterone (P), the underlying molecular pathways remain poorly understood. Here, we report that the expression of connexin 43 (Cx43), a major gap junction protein, is markedly enhanced in response to E in uterine stromal cells surrounding the implanted embryo during the early phases of pregnancy. Conditional deletion of the Cx43 gene in these stromal cells and the consequent disruption of their gap junctions led to a striking impairment in the development of new blood vessels within the stromal compartment, resulting in the arrest of embryo growth and early pregnancy loss. Further analysis of this phenotypical defect revealed that loss of Cx43 expression resulted in aberrant differentiation of uterine stromal cells and impaired production of several key angiogenic factors, including the vascular endothelial growth factor (Vegf). Ablation of CX43 expression in human endometrial stromal cells in vitro led to similar findings. Collectively, these results uncovered a unique link between steroid hormone-regulated cell-cell communication within the pregnant uterus and the development of an elaborate vascular network that supports embryonic growth. Our study presents the first evidence that Cx43-type gap junctions play a critical and conserved role in modulating stromal differentiation, and regulate the consequent production of crucial paracrine signals that control uterine neovascularization during implantation.

  6. Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells.

    PubMed

    Sawada, Keigo; Takedachi, Masahide; Yamamoto, Satomi; Morimoto, Chiaki; Ozasa, Masao; Iwayama, Tomoaki; Lee, Chun Man; Okura, Hanayuki; Matsuyama, Akifumi; Kitamura, Masahiro; Murakami, Shinya

    2015-08-14

    Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response. ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. Copyright © 2015 Elsevier Inc. All rights reserved.

  7. Omega-3-derived mediators counteract obesity-induced adipose tissue inflammation.

    PubMed

    Titos, Esther; Clària, Joan

    2013-12-01

    Chronic low-grade inflammation in adipose tissue has been recognized as a key step in the development of obesity-associated complications. In obesity, the accumulation of infiltrating macrophages in adipose tissue and their phenotypic switch to M1-type dysregulate inflammatory adipokine production leading to obesity-linked insulin resistance. Resolvins are potent anti-inflammatory and pro-resolving mediators endogenously generated from omega-3 fatty acids that act as "stop-signals" of the inflammatory response promoting the resolution of inflammation. Recently, a deficit in the production of these endogenous anti-inflammatory signals has been demonstrated in obese adipose tissue. The restoration of their levels by either exogenous administration of these mediators or feeding omega-3-enriched diets, improves the inflammatory status of adipose tissue and ameliorates metabolic dysfunction. Here, we review the current knowledge on the role of these endogenous autacoids in the resolution of adipose tissue inflammation with special emphasis on their functional actions on macrophages. Copyright © 2013 Elsevier Inc. All rights reserved.

  8. AMP-Activated Protein Kinase (AMPK) Regulates Energy Metabolism through Modulating Thermogenesis in Adipose Tissue

    PubMed Central

    Wu, Lingyan; Zhang, Lina; Li, Bohan; Jiang, Haowen; Duan, Yanan; Xie, Zhifu; Shuai, Lin; Li, Jia; Li, Jingya

    2018-01-01

    Obesity occurs when excess energy accumulates in white adipose tissue (WAT), whereas brown adipose tissue (BAT), which is specialized in dissipating energy through thermogenesis, potently counteracts obesity. White adipocytes can be converted to thermogenic “brown-like” cells (beige cells; WAT browning) under various stimuli, such as cold exposure. AMP-activated protein kinase (AMPK) is a crucial energy sensor that regulates energy metabolism in multiple tissues. However, the role of AMPK in adipose tissue function, especially in the WAT browning process, is not fully understood. To illuminate the effect of adipocyte AMPK on energy metabolism, we generated Adiponectin-Cre-driven adipose tissue-specific AMPK α1/α2 KO mice (AKO). These AKO mice were cold intolerant and their inguinal WAT displayed impaired mitochondrial integrity and biogenesis, and reduced expression of thermogenic markers upon cold exposure. High-fat-diet (HFD)-fed AKO mice exhibited increased adiposity and exacerbated hepatic steatosis and fibrosis and impaired glucose tolerance and insulin sensitivity. Meanwhile, energy expenditure and oxygen consumption were markedly decreased in the AKO mice both in basal conditions and after stimulation with a β3-adrenergic receptor agonist, CL 316,243. In contrast, we found that in HFD-fed obese mouse model, chronic AMPK activation by A-769662 protected against obesity and related metabolic dysfunction. A-769662 alleviated HFD-induced glucose intolerance and reduced body weight gain and WAT expansion. Notably, A-769662 increased energy expenditure and cold tolerance in HFD-fed mice. A-769662 treatment also induced the browning process in the inguinal fat depot of HFD-fed mice. Likewise, A-769662 enhanced thermogenesis in differentiated inguinal stromal vascular fraction (SVF) cells via AMPK signaling pathway. In summary, a lack of adipocyte AMPKα induced thermogenic impairment and obesity in response to cold and nutrient-overload, respectively

  9. Sustained release of adipose-derived stem cells by thermosensitive chitosan/gelatin hydrogel for therapeutic angiogenesis.

    PubMed

    Cheng, Nai-Chen; Lin, Wei-Jhih; Ling, Thai-Yen; Young, Tai-Horng

    2017-03-15

    Adipose-derived stem cells (ASCs) secrete several angiogenic growth factors and can be applied to treat ischemic tissue. However, transplantation of dissociated ASCs has frequently resulted in rapid cell death. Therefore, we aimed to develop a thermosensitive chitosan/gelatin hydrogel that is capable of ASC sustained release for therapeutic angiogenesis. By blending gelatin in the chitosan thermosensitive hydrogel, we significantly enhanced the viability of the encapsulated ASCs. During in vitro culturing, the gradual degradation of gelatin led to sustained release of ASCs from the chitosan/gelatin hydrogel. In vitro wound healing assays revealed significantly faster cell migration by co-culturing fibroblasts with ASCs encapsulated in chitosan/gelatin hydrogel compared to pure chitosan hydrogels. Additionally, significantly higher concentrations of vascular endothelial growth factor were found in the supernatant of ASC-encapsulated chitosan/gelatin hydrogels. Co-culturing SVEC4-10 endothelial cells with ASC-encapsulated chitosan/gelatin hydrogels resulted in significantly more tube-like structures, indicating the hydrogel's potential in promoting angiogenesis. Chick embryo chorioallantoic membrane assay and mice wound healing model showed significantly higher capillary density after applying ASC-encapsulated chitosan/gelatin hydrogel. Relative to ASC alone or ASC-encapsulated chitosan hydrogel, more ASCs were also found in the wound tissue on post-wounding day 5 after applying ASC-encapsulated chitosan/gelatin hydrogel. Therefore, chitosan/gelatin thermosensitive hydrogels not only maintain ASC survival, they also enable sustained release of ASCs for therapeutic angiogenesis applications, thereby exhibiting great clinical potential in treating ischemic diseases. Adipose-derived stem cells (ASCs) exhibit great potential to treat ischemic diseases. However, poor delivery methods lead to low cellular survival or dispersal of cells from target sites. In this study, we

  10. Cinnamon extract regulates plasma levels of adipose-derived factors and expression of multiple genes related to carbohydrate metabolism and lipogenesis in adipose tissue of fructose-fed rats

    USDA-ARS?s Scientific Manuscript database

    We reported previously that a dietary cinnamon extract (CE) improves systemic insulin sensitivity and dyslipidemia by enhancing insulin signaling. In the present study, we examined the effects of CE on several biomarkers including plasma levels of adipose-derived adipokines, and the potential molec...

  11. Stem cell-derived vascular endothelial cells and their potential application in regenerative medicine

    USDA-ARS?s Scientific Manuscript database

    Although a 'vascular stem cell' population has not been identified or generated, vascular endothelial and mural cells (smooth muscle cells and pericytes) can be derived from currently known pluripotent stem cell sources, including human embryonic stem cells and induced pluripotent stem cells. We rev...

  12. Hemangioblastomas: histogenesis of the stromal cell studied by immunocytochemistry.

    PubMed

    Jurco, S; Nadji, M; Harvey, D G; Parker, J C; Font, R L; Morales, A R

    1982-01-01

    Twenty-one cases of hemangioblastoma from the cerebellum, spinal cord and retina were studied using the unlabeled antibody peroxidase-antiperoxidase technique with antibodies directed against glial fibrillary acidic protein (GFAP) and factor VIII related antigen (VIIIR:Ag). In 19 of 21 cases studied with anti-GFAP, astrocytes were identified peripherally, and in 13 cases they were found centrally within the tumor. In no instance did stromal cells react positively for GFAP. Sixteen cases with anti-VIIIR:Ag antibody were examined, and in all cases many stromal cells showed positive staining. It is concluded that the stromal cells were of endothelial origin. The occasional stromal cells that other investigators have identified as reacting positively for GFAP may represent stromal cells capable of ingesting extracellular GFAP derived from reactive astrocytes within the tumor, or they may be lipidized astrocytes.

  13. Effectiveness of autologous serum as an alternative to fetal bovine serum in adipose-derived stem cell engineering.

    PubMed

    Choi, Jaehoon; Chung, Jee-Hyeok; Kwon, Geun-Yong; Kim, Ki-Wan; Kim, Sukwha; Chang, Hak

    2013-09-01

    In cell culture, medium supplemented with fetal bovine serum is commonly used, and it is widely known that fetal bovine serum supplies an adequate environment for culture and differentiation of stem cells. Nevertheless, the use of xenogeneic serum can cause several problems. We compared the effects of four different concentrations of autologous serum (1, 2, 5, and 10%) on expansion and adipogenic differentiation of adipose-derived stem cells using 10% fetal bovine serum as a control. The stem cells were grafted on nude mice and the in vivo differentiation capacity was evaluated. The isolation of adipose-derived stem cells was successful irrespective of the culture medium. The proliferation potential was statistically significant at passage 2, as follows: 10% autologous serum > 10% fetal bovine serum = 5% autologous serum > 2% autologous serum = 1% autologous serum. The differentiation capacity appeared statistically significant at passage 4, as follows: 10% fetal bovine serum > 10% autologous serum = 5% autologous serum > 2% autologous serum = 1% autologous serum. Ten percent autologous serum and 10% fetal bovine serum had greater differentiation capacity than 1 and 2% autologous serum in vivo, and no significant difference was observed between the groups at ≥ 5% concentration at 14 weeks. In conclusion, 10% autologous serum was at least as effective as 10% fetal bovine serum with respect to the number of adipose-derived stem cells at the end of both isolation and expansion, whereas 1 and 2% autologous serum was inferior.

  14. The Stromal Vascular Fraction From Fat Tissue in the Treatment of Osteochondral Knee Defect: Case Report

    PubMed Central

    Salikhov, Ramil Z.; Masgutov, Ruslan F.; Chekunov, Mikhail A.; Tazetdinova, Leysan G.; Masgutova, Galina; Teplov, Oleg V.; Galimov, Damir; Plakseichuk, Yuri; Yagudin, Ramil; Pankov, Igor O.; Rizvanov, Albert

    2018-01-01

    In this study we applied autologous fat tissue stromal vascular fraction (SVF) cells in combination with microfracturing technique in a 36-year-old man with an osteochondral lesion of the medial femoral condyle 8 months after the injury. Cell material was generated by fat tissue liposuction from the anterior abdominal wall with subsequent extraction of the SVF and injected through a mini-arthrotomy portal with subsequent fibrin sealant fixation. The follow-up period was 2 years. Clinical score improved from 23 to 96 according to IKDC and from 10 to 90 according to EQ-VAS at 24 months follow-up. Magnetic resonance imaging (MRI) before the surgery revealed an osteochondral lesion with development of significant trabecular edema that remained unchanged for 6 months despite conservative treatment. MRI 1 and 2 years after the surgery showed the recovery of the damaged cartilage thickness with somewhat uneven structure and a decrease in the trabecular edema of the femoral condyle. The use of SVF cells with fibrin sealant fixation might be a promising approach in the treatment of osteochondral joint lesions. Further studies are required. PMID:29900170

  15. Human adipose-derived stem cells: definition, isolation, tissue-engineering applications.

    PubMed

    Nae, S; Bordeianu, I; Stăncioiu, A T; Antohi, N

    2013-01-01

    Recent researches have demonstrated that the most effective repair system of the body is represented by stem cells - unspecialized cells, capable of self-renewal through successive mitoses, which have also the ability to transform into different cell types through differentiation. The discovery of adult stem cells represented an important step in regenerative medicine because they no longer raises ethical or legal issues and are more accessible. Only in 2002, stem cells isolated from adipose tissue were described as multipotent stem cells. Adipose tissue stem cells benefits in tissue engineering and regenerative medicine are numerous. Development of adipose tissue engineering techniques offers a great potential in surpassing the existing limits faced by the classical approaches used in plastic and reconstructive surgery. Adipose tissue engineering clinical applications are wide and varied, including reconstructive, corrective and cosmetic procedures. Nowadays, adipose tissue engineering is a fast developing field, both in terms of fundamental researches and medical applications, addressing issues related to current clinical pathology or trauma management of soft tissue injuries in different body locations.

  16. Kojyl cinnamate ester derivatives promote adiponectin production during adipogenesis in human adipose tissue-derived mesenchymal stem cells.

    PubMed

    Rho, Ho Sik; Hong, Soo Hyun; Park, Jongho; Jung, Hyo-Il; Park, Young-Ho; Lee, John Hwan; Shin, Song Seok; Noh, Minsoo

    2014-05-01

    The subcutaneous fat tissue mass gradually decreases with age, and its regulation is a strategy to develop anti-aging compounds to ameliorate the photo-aging of human skin. The adipogenesis of human adipose tissue-mesenchymal stem cells (hAT-MSCs) can be used as a model to discover novel anti-aging compounds. Cinnamomum cassia methanol extracts were identified as adipogenesis-promoting agents by natural product library screening. Cinnamates, the major chemical components of Cinnamomum cassia extracts, promoted adipogenesis in hAT-MSCs. We synthesized kojyl cinnamate ester derivatives to improve the pharmacological activity of cinnamates. Structure-activity studies of kojyl cinnamate derivatives showed that both the α,β-unsaturated carbonyl ester group and the kojic acid moiety play core roles in promoting adiponectin production during adipogenesis in hAT-MSCs. We conclude that kojyl cinnamate ester derivatives provide novel pharmacophores that can regulate adipogenesis in hAT-MSCs. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. Generation of an iPS cell line from bone marrow derived mesenchymal stromal cells from an elderly patient.

    PubMed

    Megges, Matthias; Geissler, Sven; Duda, Georg N; Adjaye, James

    2015-11-01

    An induced pluripotent stem cell line was generated from primary human bone marrow derived mesenchymal stromal cells of a 74 year old donor using retroviruses harboring OCT4, SOX2, KLF4 and c-MYC in combination with the following inhibitors TGFβ receptor-SB 431542, MEK-PD325901, and p53-Pifithrin α. Pluripotency was confirmed both in vitro and in vivo. Copyright © 2015 Elsevier B.V. All rights reserved.

  18. Adipose-derived stem cells seeded in Pluronic F-127 hydrogel promotes diabetic wound healing.

    PubMed

    Kaisang, Lin; Siyu, Wang; Lijun, Fan; Daoyan, Pan; Xian, Cory J; Jie, Shen

    2017-09-01

    Chronic nonhealing wound is a multifactorial complication of diabetes that results specifically as a consequence of impaired angiogenesis and currently lacks in effective treatments. Although a stem cell-based therapy may provide a novel treatment to augment diabetic wound healing, inferior cell survival at the diabetic skin wound is one of the key causes that are responsible for the low efficacy of the stem cell therapy. In this work, we used an injectable, biocompatible, and thermosensitive hydrogel Pluronic F-127 to encapsulate allogeneic nondiabetic adipose-derived stem cells (ADSCs) and topically applied the cells to a full-thickness cutaneous wound in the streptozotocin-induced diabetic model in rats. The cells seeded in the hydrogel enhanced angiogenesis (CD31 marker) and promoted the cell proliferation (Ki67 marker) at the wound site and significantly accelerated wound closure, which was accompanied by facilitated regeneration of granulation tissue. Consistently, levels of the messenger RNA expression of key angiogenesis growth factor, vascular endothelial growth factor, and key wound healing growth factor, transforming growth factor beta 1, were also upregulated in the cell-treated wounds when compared with untreated wounds. The results indicated that the transplantation of allogeneic ADSCs via the hydrogel improves the efficiency of cell delivery and optimizes the performance of ADSCs for augmenting diabetic wound healing. In conclusion, this ADSC-based therapy may provide a novel therapeutic strategy for the treatment of nonhealing diabetic foot ulcers. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Effective Treatment of Traumatic Brain Injury in Rowett Nude Rats with Stromal Vascular Fraction Transplantation.

    PubMed

    Berman, Sean; Uhlendorf, Toni L; Berman, Mark; Lander, Elliot B

    2018-06-18

    Traumatic brain injury (TBI) affects 1.9 million Americans, including blast TBI that is the signature injury of the Iraq and Afghanistan wars. Our project investigated whether stromal vascular fraction (SVF) can assist in post-TBI recovery. We utilized strong acoustic waves (5.0 bar) to induce TBI in the cortex of adult Rowett Nude (RNU) rats. One hour post-TBI, harvested human SVF (500,000 cells suspended in 0.5 mL lactated Ringers) was incubated with Q-Tracker cell label and administered into tail veins of RNU rats. For comparison, we utilized rats that received SVF 72 h post-TBI, and a control group that received lactated Ringers solution. Rotarod and water maze assays were used to monitor motor coordination and spatial memories. Rats treated immediately after TBI showed no signs of motor skills and memory regression. SVF treatment 72 h post-TBI enabled the rats maintain their motor skills, while controls treated with lactated Ringers were 25% worse statistically in both assays. Histological analysis showed the presence of Q-dot labeled human cells near the infarct in both SVF treatment groups; however, labeled cells were twice as numerous in the one hour group. Our study suggests that immediate treatment with SVF would serve as potential therapeutic agents in TBI.

  20. Associations between accelerometer-derived physical activity and regional adiposity in young men and women.

    PubMed

    Smith, H A; Storti, K L; Arena, V C; Kriska, A M; Gabriel, K K Pettee; Sutton-Tyrrell, K; Hames, K C; Conroy, M B

    2013-06-01

    Empirical evidence supports an inverse relationship between physical activity (PA) and adiposity, but studies using detailed measures of both are scarce. The relationship between regional adiposity and accelerometer-derived PA in men and women are described. Cross-sectional analysis included 253 participants from a weight loss study limited to ages 20-45 years and BMI 25-39.9 kg m(-2) . PA data were collected with accelerometers and expressed as total accelerometer counts and average amount of time per day accumulated in different intensity levels [sedentary, light-, and moderate-to-vigorous intensity PA (MVPA)]. Accumulation of time spent above 100 counts was expressed as total active time. Computed tomography (CT) was used to measure abdominal and adipose tissue (AT). Multivariate linear regression analyses were used to assess the relationship between regional adiposity (dependent variable) and the various PA levels (independent variable), and were executed separately for men and women, adjusting for wear time, age, race, education, and BMI. Among males, light activity was inversely associated with total AT (β = -0.19; P = 0.02) as well as visceral AT (VAT) (β = -0.30; P = 0.03). Among females sedentary time was positively associated with VAT (β = 0.11; P = 0.04) and total active time was inversely associated with VAT (β = -0.12; P = 0.04). Findings from this study suggest that PA intensity level may influence regional adiposity differently in men and women. Additional research is needed in larger samples to clarify the difference in these associations by sex, create recommendations for the frequency, duration and intensity of PA needed to target fat deposits, and determine if these recommendations should differ by sex. Copyright © 2013 The Obesity Society.

  1. The corneal fibrosis response to epithelial-stromal injury

    PubMed Central

    Torricelli, Andre A. M.; Santhanam, Abirami; Wu, Jiahui; Singh, Vivek; Wilson, Steven E.

    2014-01-01

    The corneal wound healing response, including the development of stromal opacity in some eyes, is a process that often leads to scarring that occurs after injury, surgery or infection to the cornea. Immediately after epithelial and stromal injury, a complex sequence of processes contributes to wound repair and regeneration of normal corneal structure and function. In some corneas, however, often depending on the type and extent of injury, the response may also lead to the development of mature vimentin+ α-smooth muscle actin+ desmin+ myofibroblasts. Myofibroblasts are specialized fibroblastic cells generated in the cornea from keratocyte-derived or bone marrow-derived precursor cells. The disorganized extracellular matrix components secreted by myofibroblasts, in addition to decreased expression of corneal crystallins in these cells, are central biological processes that result in corneal stromal fibrosis associated with opacity or “haze”. Several factors are associated with myofibroblast generation and haze development after PRK surgery in rabbits, a reproducible model of scarring, including the amount of tissue ablated, which may relate to the extent of keratocyte apoptosis in the early response to injury, irregularity of stromal surface after surgery, and changes on corneal stromal proteoglycans, but normal regeneration of the epithelial basement membrane (EBM) appears to be a critical factor determining whether a cornea heals with relative transparency or vision-limiting stromal opacity. Structural and functional abnormalities of the regenerated EBM facilitate prolonged entry of epithelium-derived growth factors such as transforming growth factor β (TGF-β) and platelet-derived growth factor (PDGF) into the stroma that both drive development of mature myofibroblasts from precursor cells and lead to persistence of the cells in the anterior stroma. A major discovery that has contributed to our understanding of haze development is that keratocytes and

  2. Stromal signatures in endometrioid endometrial carcinomas.

    PubMed

    Espinosa, Iñigo; Catasus, Lluis; D' Angelo, Emanuela; Mozos, Ana; Pedrola, Nuria; Bértolo, Cristina; Ferrer, Irene; Zannoni, Gian Franco; West, Robert B; van de Rijn, Matt; Matias-Guiu, Xavier; Prat, Jaime

    2014-04-01

    The pattern of myometrial invasion in endometrioid endometrial carcinomas varies considerably; ie, from widely scattered glands and cell nests, often associated with a fibromyxoid stromal reaction (desmoplasia) and/or a lymphocytic infiltrate, to invasive glands with little or no stromal response. Recently, two distinct stromal signatures derived from a macrophage response (colony-stimulating factor 1, CSF1) and a fibroblastic response (desmoid-type fibromatosis, DTF) were identified in breast carcinomas and correlated with clinicopathologic features including outcome. In this study, we explored whether these stromal signatures also apply to endometrioid carcinomas and how their expression patterns correlated with morphologic changes. We studied the stromal signatures both by immunohistochemistry and in situ hybridization in 98 primary endometrioid carcinomas with (87 cases) and without (11 cases) myometrial invasion as well as in the corresponding regional lymph nodes metatases of 9 myoinvasive tumors. Desmoplasia correlated positively with the DTF expression signature. Likewise, mononuclear infiltrates were found in the stroma of tumors expressing CSF1. Twenty-four out of eighty-seven (27%) myoinvasive endometrioid carcinomas were positive for the macrophage signature and thirteen out of eighty-seven (15%) expressed the fibroblast signature. Eleven additional cases were positive for both DTF and CSF1 signatures (11/87; 13%). However, over half of the cases (39/87; 45%) and the majority of the non-myoinvasive tumors (8/11; 73%) failed to express any of the two stromal signatures. The macrophage response (CSF1) was associated with higher tumor grade, lymphovascular invasion, and PIK3CA mutations (P<0.05). There was a concordance in the expression of the CSF1 signature in the primary tumors and their corresponding lymph node metastases. This study is the first characterization of stromal signatures in endometrioid carcinomas. Our findings shed new light on the

  3. Does an injection of a stromal vascular fraction containing adipose-derived mesenchymal stem cells influence the outcomes of marrow stimulation in osteochondral lesions of the talus? A clinical and magnetic resonance imaging study.

    PubMed

    Kim, Yong Sang; Lee, Ho Jin; Choi, Yun Jin; Kim, Yong Il; Koh, Yong Gon

    2014-10-01

    Marrow stimulation for the treatment of osteochondral lesions of the talus (OLTs) is controversial in patients with poor prognostic factors of OLTs. Currently, mesenchymal stem cells (MSCs) are expected to biologically augment the treatment of OLTs. To compare the clinical and magnetic resonance imaging (MRI) outcomes between an injection of MSCs with marrow stimulation and marrow stimulation alone in patients with OLTs. Cohort study; Level of evidence, 3. A total of 49 patients (50 ankles) with OLTs underwent follow-up MRI after arthroscopic treatment. Among these 50 ankles, 26 underwent marrow stimulation alone (conventional group), and 24 underwent marrow stimulation with an injection of a stromal vascular fraction (SVF) containing MSCs (MSC group). Clinical outcomes were evaluated according to the visual analog scale (VAS) for pain, American Orthopaedic Foot and Ankle Society (AOFAS) Ankle-Hindfoot Scale, and Tegner activity scale. The magnetic resonance observation of cartilage repair tissue (MOCART) score was used for the MRI evaluation of repaired lesions. The mean VAS score, AOFAS score, and Tegner score improved from 7.1 ± 1.2, 68.5 ± 5.6, and 3.4 ± 0.6 to 3.9 ± 0.8, 78.3 ± 4.9, and 3.5 ± 0.8, respectively, in the conventional group and from 7.1 ± 0.8, 67.7 ± 4.7, and 3.4 ± 0.5 to 3.2 ± 0.8, 83.3 ± 7.0, and 3.9 ± 0.7, respectively, in the MSC group. All clinical outcomes, including the VAS, AOFAS, and Tegner scores, improved significantly in the MSC group compared with the conventional group (P = .003, .009, and .041, respectively). There was a significant difference (P = .037) in the mean MOCART score between the conventional and MSC groups (49.4 ± 16.6 vs 62.1 ± 21.8, respectively), and significant correlations of the MOCART score with clinical outcomes were found in both groups (P < .05). Patient age (≥46.1 years), large lesion size (≥151.2 mm(2)), and the presence of subchondral cysts were associated with a worse MOCART score in the

  4. The role of SDF-1-CXCR4/CXCR7 axis in biological behaviors of adipose tissue-derived mesenchymal stem cells in vitro

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Li, Qiang; Zhang, Aijun; Tao, Changbo

    2013-11-22

    Highlights: •SDF-1 pretreating increased the levels of CXCR4, CXCR7 in ADSCs. •SDF-1 improved cells paracrine migration and proliferation abilities. •CXCR4 and CXCR7 could function in ADSCs paracrine, migration and proliferation. -- Abstract: Numerous studies have reported that CXCR4 and CXCR7 play an essential, but differential role in stromal cell-derived factor-1 (SDF-1)-inducing cell chemotaxis, viability and paracrine actions of BMSCs. Adipose tissue-derived mesenchymal stem cells (ADSCs) have been suggested to be potential seed cells for clinical application instead of bone marrow derived stroma cell (BMSCs). However, the function of SDF-1/CXCR4 and SDF-1/CXCR7 in ADSCs is not well understood. This study wasmore » designed to analyze the effect of SDF-1/CXCR4 and SDF-1/CXCR7 axis on ADSCs biological behaviors in vitro. Using Flow cytometry and Western blot methods, we found for the first time that CXCR4/CXCR7 expression was increased after treatment with SDF-1 in ADSCs. SDF-1 promoted ADSCs paracrine, proliferation and migration abilities. CXCR4 or CXCR7 antibody suppressed ADSCs paracrine action induced by SDF-1. The migration of ADSCs can be abolished by CXCR4 antibody, while the proliferation of ADSCs was only downregulated by CXCR7 antibody. Our study indicated that the angiogenesis of ADSCs is, at least partly, mediated by SDF-1/CXCR4 and SDF-1/CXCR7 axis. However, only binding of SDF-1/CXCR7 was required for proliferation of ADSCs, and CXCR7 was required for migration of ADSCs induced by SDF-1. Our studies provide evidence that the activation of either axis may be helpful to improve the effectiveness of ADSCs-based stem cell therapy.« less

  5. Fibroblast growth factor-2 stimulates adipogenic differentiation of human adipose-derived stem cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Kakudo, Natsuko; Shimotsuma, Ayuko; Kusumoto, Kenji

    2007-07-27

    Adipose-derived stem cells (ASCs) have demonstrated a capacity for differentiating into a variety of lineages, including bone, cartilage, or fat, depending on the inducing stimuli and specific growth and factors. It is acknowledged that fibroblast growth factor-2 (FGF-2) promotes chondrogenic and inhibits osteogenic differentiation of ASCs, but thorough investigations of its effects on adipogenic differentiation are lacking. In this study, we demonstrate at the cellular and molecular levels the effect of FGF-2 on adipogenic differentiation of ASCs, as induced by an adipogenic hormonal cocktail consisting of 3-isobutyl-1-methylxanthine (IBMX), dexamethasone, insulin, and indomethacin. FGF-2 significantly enhances the adipogenic differentiation of humanmore » ASCs. Furthermore, in cultures receiving FGF-2 before adipogenic induction, mRNA expression of peroxisome proliferator-activated receptor {gamma}2 (PPAR{gamma}2), a key transcription factor in adipogenesis, was upregulated. The results of FGF-2 supplementation suggest the potential applications of FGF-2 and ASCs in adipose tissue regeneration.« less

  6. Effects of in vitro low oxygen tension preconditioning of adipose stromal cells on their in vivo chondrogenic potential: application in cartilage tissue repair.

    PubMed

    Portron, Sophie; Merceron, Christophe; Gauthier, Olivier; Lesoeur, Julie; Sourice, Sophie; Masson, Martial; Fellah, Borhane Hakim; Geffroy, Olivier; Lallemand, Elodie; Weiss, Pierre; Guicheux, Jérôme; Vinatier, Claire

    2013-01-01

    Multipotent stromal cell (MSC)-based regenerative strategy has shown promise for the repair of cartilage, an avascular tissue in which cells experience hypoxia. Hypoxia is known to promote the early chondrogenic differentiation of MSC. The aim of our study was therefore to determine whether low oxygen tension could be used to enhance the regenerative potential of MSC for cartilage repair. MSC from rabbit or human adipose stromal cells (ASC) were preconditioned in vitro in control or chondrogenic (ITS and TGF-β) medium and in 21 or 5% O2. Chondrogenic commitment was monitored by measuring COL2A1 and ACAN expression (real-time PCR). Preconditioned rabbit and human ASC were then incorporated into an Si-HPMC hydrogel and injected (i) into rabbit articular cartilage defects for 18 weeks or (ii) subcutaneously into nude mice for five weeks. The newly formed tissue was qualitatively and quantitatively evaluated by cartilage-specific immunohistological staining and scoring. The phenotype of ASC cultured in a monolayer or within Si-HPMC in control or chondrogenic medium and in 21 or 5% O2 was finally evaluated using real-time PCR. 5% O2 increased the in vitro expression of chondrogenic markers in ASC cultured in induction medium. Cells implanted within Si-HPMC hydrogel and preconditioned in chondrogenic medium formed a cartilaginous tissue, regardless of the level of oxygen. In addition, the 3D in vitro culture of ASC within Si-HPMC hydrogel was found to reinforce the pro-chondrogenic effects of the induction medium and 5% O2. These data together indicate that although 5% O2 enhances the in vitro chondrogenic differentiation of ASC, it does not enhance their in vivo chondrogenesis. These results also highlight the in vivo chondrogenic potential of ASC and their potential value in cartilage repair.

  7. Early alterations in vascular contractility associated to changes in fatty acid composition and oxidative stress markers in perivascular adipose tissue.

    PubMed

    Rebolledo, Alejandro; Rebolledo, Oscar R; Marra, Carlos A; García, María E; Roldán Palomo, Ana R; Rimorini, Laura; Gagliardino, Juan J

    2010-10-21

    To test the early effect of fructose-induced changes in fatty acid composition and oxidative stress markers in perivascular adipose tissue (PVAT) upon vascular contractility. Adult male Wistar rats were fed a commercial diet without (CD) or with 10% fructose (FRD) in the drinking water for 3 weeks. We measured plasma metabolic parameters, lipid composition and oxidative stress markers in aortic PVAT. Vascular contractility was measured in aortic rings sequentially, stimulated with serotonin (5-HT) and high K+-induced depolarization using intact and thereafter PVAT-deprived rings. Comparable body weights were recorded in both groups. FRD rats had increased plasma triglyceride and fructosamine levels. Their PVAT had an increased saturated to mono- or poly-unsaturated fatty acid ratio, a significant decrease in total superoxide dismutase and glutathione peroxidase activities and in the total content of glutathione. Conversely, lipid peroxidation (TBARS), nitric oxide content, and gluthathione reductase activity were significantly higher, indicating an increase in oxidative stress. In aortic rings, removal of PVAT increased serotonin-induced contractions, but the effect was significantly lower in rings from FRD rats. This effect was no longer observed when the two contractions were performed in PVAT-deprived rings. PVAT did not affect the contractions triggered by high K+-induced depolarization either in CD or FRD rats. FRD induces multiple metabolic and endocrine systemic alterations which also alter PVAT and the vascular relaxant properties of this tissue. The changes in PVAT would affect its paracrine modulation of vascular function.

  8. Defining the identity of human adipose-derived mesenchymal stem cells.

    PubMed

    Montelatici, Elisa; Baluce, Barbara; Ragni, Enrico; Lavazza, Cristiana; Parazzi, Valentina; Mazzola, Riccardo; Cantarella, Giovanna; Brambilla, Massimiliano; Giordano, Rosaria; Lazzari, Lorenza

    2015-02-01

    Adipose-derived mesenchymal stem cells (ADMSCs) are an ideal population for regenerative medical application. Both the isolation procedure and the culturing conditions are crucial steps, since low yield can limit further cell therapies, especially when minimal adipose tissue harvests are available for cell expansion. To date, a standardized procedure encompassing both isolation sites and expansion methods is missing, thus making the choice of the most appropriate conditions for the preparation of ADMSCs controversial, especially in view of the different applications needed. In this study, we compared the effects of three different commercial media (DMEM, aMEM, and EGM2), routinely used for ADMSCs expansion, and two supplements, FBS and human platelet lysate, recently proven to be an effective alternative to prevent xenogeneic antibody transfer and immune alloresponse in the host. Notably, all the conditions resulted in being safe for ADMSCs isolation and expansion with platelet lysate supplementation giving the highest isolation and proliferation rates, together with a commitment for osteogenic lineage. Then, we proved that the high ADMSC hematopoietic supportive potential is performed through a constant and abundant secretion of both GCSF and SCF. In conclusion, this study further expands the knowledge on ADMSCs, defining their identity definition and offers potential options for in vitro protocols for clinical production, especially related to HSC expansion without use of exogenous cytokines or genetic modifications.

  9. The effect of two novel amino acid-coated magnetic nanoparticles on survival in vascular endothelial cells, bone marrow stromal cells, and macrophages

    NASA Astrophysics Data System (ADS)

    Wu, Qinghua; Meng, Ning; Zhang, Yanru; Han, Lei; Su, Le; Zhao, Jing; Zhang, Shangli; Zhang, Yun; Zhao, Baoxiang; Miao, Junying

    2014-09-01

    Magnetic nanoparticles (MNPs) have been popularly used in many fields. Recently, many kinds of MNPs are modified as new absorbents, which have attracted considerable attention and are promising to be applied in waste water. In our previous study, we synthesized two novel MNPs surface-coated with glycine or lysine, which could efficiently remove many anionic and cationic dyes under severe conditions. It should be considered that MNP residues in water may exert some side effects on human health. In the present study, we evaluated the potential nanotoxicity of MNPs in human endothelial cells, macrophages, and rat bone marrow stromal cells. The results showed that the two kinds of nanoparticles were consistently absorbed into the cell cytoplasm. The concentration of MNPs@Gly that could distinctly decrease survival was 15 μg/ml in human umbilical vascular endothelial cells (HUVECs) or bone marrow stromal cells (BMSCs) and 10 μg/ml in macrophages. While the concentration of MNPs@Lys that obviously reduced viability was 15 μg/ml in HUVECs or macrophages and 50 μg/ml in BMSCs. Furthermore, cell nucleus staining and cell integrity assay indicated that the nanoparticles induced cell apoptosis, but not necrosis even at a high concentration. Altogether, these data suggest that the amino acid-coated magnetic nanoparticles exert relatively high cytotoxicity. By contrast, lysine-coated magnetic nanoparticles are more secure than glycine-coated magnetic nanoparticles.

  10. The skeletal cell-derived molecule sclerostin drives bone marrow adipogenesis.

    PubMed

    Fairfield, Heather; Falank, Carolyne; Harris, Elizabeth; Demambro, Victoria; McDonald, Michelle; Pettitt, Jessica A; Mohanty, Sindhu T; Croucher, Peter; Kramer, Ina; Kneissel, Michaela; Rosen, Clifford J; Reagan, Michaela R

    2018-02-01

    The bone marrow niche is a dynamic and complex microenvironment that can both regulate, and be regulated by the bone matrix. Within the bone marrow (BM), mesenchymal stromal cell (MSC) precursors reside in a multi-potent state and retain the capacity to differentiate down osteoblastic, adipogenic, or chondrogenic lineages in response to numerous biochemical cues. These signals can be altered in various pathological states including, but not limited to, osteoporotic-induced fracture, systemic adiposity, and the presence of bone-homing cancers. Herein we provide evidence that signals from the bone matrix (osteocytes) determine marrow adiposity by regulating adipogenesis in the bone marrow. Specifically, we found that physiologically relevant levels of Sclerostin (SOST), which is a Wnt-inhibitory molecule secreted from bone matrix-embedded osteocytes, can induce adipogenesis in 3T3-L1 cells, mouse ear- and BM-derived MSCs, and human BM-derived MSCs. We demonstrate that the mechanism of SOST induction of adipogenesis is through inhibition of Wnt signaling in pre-adipocytes. We also demonstrate that a decrease of sclerostin in vivo, via both genetic and pharmaceutical methods, significantly decreases bone marrow adipose tissue (BMAT) formation. Overall, this work demonstrates a direct role for SOST in regulating fate determination of BM-adipocyte progenitors. This provides a novel mechanism for which BMAT is governed by the local bone microenvironment, which may prove relevant in the pathogenesis of certain diseases involving marrow adipose. Importantly, with anti-sclerostin therapy at the forefront of osteoporosis treatment and a greater recognition of the role of BMAT in disease, these data are likely to have important clinical implications. © 2017 Wiley Periodicals, Inc.

  11. Relations among Adiposity and Insulin Resistance with Flow-Mediated Dilation, Carotid Intima-Media Thickness, and Arterial Stiffness in Children.

    PubMed

    Ryder, Justin R; Dengel, Donald R; Jacobs, David R; Sinaiko, Alan R; Kelly, Aaron S; Steinberger, Julia

    2016-01-01

    To determine the associations of adiposity and insulin resistance with measures of vascular structure and function in children. A cross-sectional study included 252 children (age 15.1 ± 2.4 years; body mass index percentile 68.2 ± 26.5%; Tanner 2-5). Measurements of body fat percentage were obtained with dual-energy X-ray absorptiometry and visceral adipose tissue (VAT) with computed tomography. Insulin resistance was measured with hyperinsulinemic euglycemic clamp. Vascular measurements for endothelial function (brachial artery flow-mediated dilation [FMD]), vascular structure (carotid intima-media thickness [cIMT]), vascular stiffness (carotid incremental elastic modulus), and pulse wave velocity were analyzed by tertiles of adiposity and insulin resistance. Additional analyses with ANCOVA and linear regression were adjusted for Tanner, sex, race, and family relationship; FMD was also adjusted for baseline artery diameter. FMD was positively associated with high adiposity (body mass index, body fat percentage, and VAT) (P < .01 all). Insulin resistance was not associated with FMD. cIMT was significantly, positively related to obesity, VAT, and insulin resistance (P < .05 all). No differences in carotid incremental elastic modulus and pulse wave velocity were observed in relation to adiposity or insulin resistance. The findings suggest that adiposity is associated with higher FMD, and insulin resistance and VAT are associated with higher cIMT in children. Further research is needed to clarify the progression of these relations. Copyright © 2016 Elsevier Inc. All rights reserved.

  12. [Effect of cryotherapy over the expression of vascular endothelial growth factor and pigment epithelium-derived factor].

    PubMed

    Toscano-Garibay, Julia Dolores; Quiroz-Mercado, Hugo; Espitia-Pinzón, Clara; Gil-Carrasco, Félix; Flores-Estrada, José Javier

    2014-01-01

    Cryotherapy is a no invasive technique that uses intense cold to freeze and destroy cancer tissues. There are no descriptions of its effects over the expression of vascular endothelial growth factor and pigment epithelium-derived factor. Experimental study in cryogenic spot were applied in the right sclera of twelve pigs for ten minutes. Other 3 pigs were used as normal controls. Animals were sacrificed at 7, 14 and 21 and the tissues of choriodes and retina were dissected in areas of approximately 1 cm2 surrounding cryogenic spots. Expression levels of vascular endothelial growth factor and pigment epithelium-derived factor were determined analyzed using polymerase chain reaction coupled to reverse-transcription. Vascular endothelial growth factor was significantly downregulated (24%, p< 0.05) seven days post-treatment meanwhile pigment epithelium-derived factor levels increased 44.8% (p< 0.05) as compared to normal controls (untreated). Both vascular endothelial growth factor and pigment epithelium-derived factor levels remain the same until day 14 but returned to basal expression at day 21. This work expose the relation of cryotherapy with the expression of two factors related to angiogenesis. RESULTS showed significant changes on the expression of vascular endothelial growth factor and pigment epithelium-derived factor illustrating that both proteins are regulated in response to cryogenic treatment in relatively short periods (21 days).

  13. Defocused low-energy shock wave activates adipose tissue-derived stem cells in vitro via multiple signaling pathways.

    PubMed

    Xu, Lina; Zhao, Yong; Wang, Muwen; Song, Wei; Li, Bo; Liu, Wei; Jin, Xunbo; Zhang, Haiyang

    2016-12-01

    We found defocused low-energy shock wave (DLSW) could be applied in regenerative medicine by activating mesenchymal stromal cells. However, the possible signaling pathways that participated in this process remain unknown. In the present study, DLSW was applied in cultured rat adipose tissue-derived stem cells (ADSCs) to explore its effect on ADSCs and the activated signaling pathways. After treating with DLSW, the cellular morphology and cytoskeleton of ADSCs were observed. The secretions of ADSCs were detected. The expressions of ADSC surface antigens were analyzed using flow cytometry. The expressions of proliferating cell nuclear antigen and Ki67 were analyzed using western blot. The expression of CXCR2 and the migrations of ADSCs in vitro and in vivo were detected. The phosphorylation of selected signaling pathways with or without inhibitors was also detected. DLSW did not change the morphology and phenotype of ADSCs, and could promote the secretion, proliferation and migration of ADSCs. The phosphorylation levels were significantly higher in mitogen-activated protein kinases (MAPK) pathway, phosphoinositide 3-kinase (PI-3K)/AKT pathway and nuclear factor-kappa B (NF-κB) signaling pathway but not in Janus kinase/signal transducer and activator of transcription (JAK/STAT) pathway. Furthermore, ADSCs were not activated by DLSW after adding the inhibitors of these pathways simultaneously. Our results demonstrated for the first time that DLSW could activate ADSCs through MAPK, PI-3K/AKT and NF-κB signaling pathways. Combination of DLSW and agonists targeting these pathways might improve the efficacy of ADSCs in regenerative medicine in the future. Copyright © 2016 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

  14. Active spice-derived components can inhibit inflammatory responses of adipose tissue in obesity by suppressing inflammatory actions of macrophages and release of monocyte chemoattractant protein-1 from adipocytes.

    PubMed

    Woo, Hae-Mi; Kang, Ji-Hye; Kawada, Teruo; Yoo, Hoon; Sung, Mi-Kyung; Yu, Rina

    2007-02-13

    Inflammation plays a key role in obesity-related pathologies such as cardiovascular disease, type II diabetes, and several types of cancer. Obesity-induced inflammation entails the enhancement of the recruitment of macrophages into adipose tissue and the release of various proinflammatory proteins from fat tissue. Therefore, the modulation of inflammatory responses in obesity may be useful for preventing or ameliorating obesity-related pathologies. Some spice-derived components, which are naturally occurring phytochemicals, elicit antiobesity and antiinflammatory properties. In this study, we investigated whether active spice-derived components can be applied to the suppression of obesity-induced inflammatory responses. Mesenteric adipose tissue was isolated from obese mice fed a high-fat diet and cultured to prepare an adipose tissue-conditioned medium. Raw 264.7 macrophages were treated with the adipose tissue-conditioned medium with or without active spice-derived components (i.e., diallyl disulfide, allyl isothiocyanate, piperine, zingerone and curcumin). Chemotaxis assay was performed to measure the degree of macrophage migration. Macrophage activation was estimated by measuring tumor necrosis factor-alpha (TNF-alpha), nitric oxide, and monocyte chemoattractant protein-1 (MCP-1) concentrations. The active spice-derived components markedly suppressed the migration of macrophages induced by the mesenteric adipose tissue-conditioned medium in a dose-dependent manner. Among the active spice-derived components studied, allyl isothiocyanate, zingerone, and curcumin significantly inhibited the cellular production of proinflammatory mediators such as TNF-alpha and nitric oxide, and significantly inhibited the release of MCP-1 from 3T3-L1 adipocytes. Our findings suggest that the spice-derived components can suppress obesity-induced inflammatory responses by suppressing adipose tissue macrophage accumulation or activation and inhibiting MCP-1 release from adipocytes

  15. Human adipose tissue mesenchymal stromal cells and their extracellular vesicles act differentially on lung mechanics and inflammation in experimental allergic asthma.

    PubMed

    de Castro, Ligia Lins; Xisto, Debora Gonçalves; Kitoko, Jamil Zola; Cruz, Fernanda Ferreira; Olsen, Priscilla Christina; Redondo, Patricia Albuquerque Garcia; Ferreira, Tatiana Paula Teixeira; Weiss, Daniel Jay; Martins, Marco Aurélio; Morales, Marcelo Marcos; Rocco, Patricia Rieken Macedo

    2017-06-24

    Asthma is a chronic inflammatory disease that can be difficult to treat due to its complex pathophysiology. Most current drugs focus on controlling the inflammatory process, but are unable to revert the changes of tissue remodeling. Human mesenchymal stromal cells (MSCs) are effective at reducing inflammation and tissue remodeling; nevertheless, no study has evaluated the therapeutic effects of extracellular vesicles (EVs) obtained from human adipose tissue-derived MSCs (AD-MSC) on established airway remodeling in experimental allergic asthma. C57BL/6 female mice were sensitized and challenged with ovalbumin (OVA). Control (CTRL) animals received saline solution using the same protocol. One day after the last challenge, each group received saline, 10 5 human AD-MSCs, or EVs (released by 10 5  AD-MSCs). Seven days after treatment, animals were anesthetized for lung function assessment and subsequently euthanized. Bronchoalveolar lavage fluid (BALF), lungs, thymus, and mediastinal lymph nodes were harvested for analysis of inflammation. Collagen fiber content of airways and lung parenchyma were also evaluated. In OVA animals, AD-MSCs and EVs acted differently on static lung elastance and on BALF regulatory T cells, CD3 + CD4 + T cells, and pro-inflammatory mediators (interleukin [IL]-4, IL-5, IL-13, and eotaxin), but similarly reduced eosinophils in lung tissue, collagen fiber content in airways and lung parenchyma, levels of transforming growth factor-β in lung tissue, and CD3 + CD4 + T cell counts in the thymus. No significant changes were observed in total cell count or percentage of CD3 + CD4 + T cells in the mediastinal lymph nodes. In this immunocompetent mouse model of allergic asthma, human AD-MSCs and EVs effectively reduced eosinophil counts in lung tissue and BALF and modulated airway remodeling, but their effects on T cells differed in lung and thymus. EVs may hold promise for asthma; however, further studies are required to elucidate the different

  16. Biologic and immunomodulatory properties of mesenchymal stromal cells derived from human pancreatic islets

    PubMed Central

    KIM, JAEHYUP; BREUNIG, MELISSA J.; ESCALANTE, LEAH E.; BHATIA, NEEHAR; DENU, RYAN A.; DOLLAR, BRIDGET A.; STEIN, ANDREW P.; HANSON, SUMMER E.; NADERI, NADIA; RADEK, JAMES; HAUGHY, DERMOT; BLOOM, DEBRA D.; ASSADI-PORTER, FARIBA M.; HEMATTI, PEIMAN

    2012-01-01

    Background aims Mesenchymal stromal cells (MSC) have now been shown to reside in numerous tissues throughout the body, including the pancreas. Ex vivo culture-expanded MSC derived from many tissues display important interactions with different types of immune cells in vitro and potentially play a significant role in tissue homeostasis in vivo. In this study, we investigated the biologic and immunomodulatory properties of human pancreatic islet-derived MSC. Methods We culture-expanded MSC from cadaveric human pancreatic islets and characterized them using flow cytometry, differentiation assays and nuclear magnetic resonance-based metabolomics. We also investigated the immunologic properties of pancreatic islet-derived MSC compared with bone marrow (BM) MSC. Results Pancreatic islet and BM-derived MSC expressed the same cell-surface markers by flow cytometry, and both could differentiate into bone, fat and cartilage. Metabolomics analysis of MSC from BM and pancreatic islets also showed a similar set of metabolic markers but quantitative polymerase chain reactions showed that pancreatic islet MSC expressed more interleukin(IL)-1b, IL-6, STAT3 and FGF9 compared with BM MSC, and less IL-10. However, similar to BM MSC, pancreatic islet MSC were able to suppress proliferation of allogeneic T lymphocytes stimulated with anti-CD3 and anti-CD28 antibodies. Conclusions Our in vitro analysis shows pancreatic islet-derived MSC have phenotypic, biologic and immunomodulatory characteristics similar, but not identical, to BM-derived MSC. We propose that pancreatic islet-derived MSC could potentially play an important role in improving the outcome of pancreatic islet transplantation by promoting engraftment and creating a favorable immune environment for long-term survival of islet allografts. PMID:22571381

  17. Differentiation and Application of Induced Pluripotent Stem Cell-Derived Vascular Smooth Muscle Cells.

    PubMed

    Maguire, Eithne Margaret; Xiao, Qingzhong; Xu, Qingbo

    2017-11-01

    Vascular smooth muscle cells (VSMCs) play a role in the development of vascular disease, for example, neointimal formation, arterial aneurysm, and Marfan syndrome caused by genetic mutations in VSMCs, but little is known about the mechanisms of the disease process. Advances in induced pluripotent stem cell technology have now made it possible to derive VSMCs from several different somatic cells using a selection of protocols. As such, researchers have set out to delineate key signaling processes involved in triggering VSMC gene expression to grasp the extent of gene regulatory networks involved in phenotype commitment. This technology has also paved the way for investigations into diseases affecting VSMC behavior and function, which may be treatable once an identifiable culprit molecule or gene has been repaired. Moreover, induced pluripotent stem cell-derived VSMCs are also being considered for their use in tissue-engineered blood vessels as they may prove more beneficial than using autologous vessels. Finally, while several issues remains to be clarified before induced pluripotent stem cell-derived VSMCs can become used in regenerative medicine, they do offer both clinicians and researchers hope for both treating and understanding vascular disease. In this review, we aim to update the recent progress on VSMC generation from stem cells and the underlying molecular mechanisms of VSMC differentiation. We will also explore how the use of induced pluripotent stem cell-derived VSMCs has changed the game for regenerative medicine by offering new therapeutic avenues to clinicians, as well as providing researchers with a new platform for modeling of vascular disease. © 2017 American Heart Association, Inc.

  18. Expansion and delivery of adipose-derived mesenchymal stem cells on three microcarriers for soft tissue regeneration.

    PubMed

    Zhou, Yalei; Yan, Zhiwei; Zhang, Hongmei; Lu, Wei; Liu, Shiyu; Huang, Xinhui; Luo, Hailang; Jin, Yan

    2011-12-01

    Cell/microcarrier combinations can be injected to repair tissue defects, but whether currently available microcarriers can be utilized to repair different tissue defects remains unknown. Here, we compared the suitability of fabricated micronized acellular dermal matrix (MADM), micronized small intestinal submucosa (MSIS), and gelatin microspheres as expansion and delivery scaffolds for adipose-derived mesenchymal stem cells (ADSCs). The results of MTS assay, scanning electron microscopy (SEM), and flow cytometry suggested that the three microcarriers all have good biocompatibility. Quantitative polymerase chain reaction revealed enhanced epidermal growth factor, vascular endothelial growth factor, basal fibroblast growth factor, and transforming growth factor-β expression levels after ADSCs had been cultured on MADM or MSIS for 5 days. After culturing ADSCs on microcarriers in osteogenic medium for 7 days, the expression levels of bone formation-related genes were enhanced. ADSC/microcarrier treatment accelerated wound closure. The ADSC/MADM and ADSC/MSIS combinations retained more of the original implant volume at 1 month postimplantation than ADSC/gelatin microspheres combination in soft-tissue augmentation studies. All implants displayed fibroblast and capillary vessel infiltrations; but ectopic bone formation did not occur, and the calvarial defect repair results were unfavorable. Our study demonstrates the potential utility of these microcarriers not only as a cell-culture substrate but also as a cell-transplantation vehicle for skin regeneration and soft-tissue reconstruction.

  19. A Novel Combination of Homeobox Genes Is Expressed in Mesenchymal Chorionic Stem/Stromal Cells in First Trimester and Term Pregnancies

    PubMed Central

    Liu, Haiying; Murthi, Padma; Qin, Sharon; Kusuma, Gina D.; Borg, Anthony J.; Knöfler, Martin; Haslinger, Peter; Manuelpillai, Ursula; Pertile, Mark D.; Abumaree, Mohamed

    2014-01-01

    Human chorionic mesenchymal stem/stromal cells (CMSCs) derived from the placenta are similar to adult tissue-derived MSCs. The aim of this study was to investigate the role of these cells in normal placental development. Transcription factors, particularly members of the homeobox gene family, play crucial roles in maintaining stem cell proliferation and lineage specification in embryonic tissues. In adult tissues and organs, stem cells proliferate at low levels in their niche until they receive cues from the microenvironment to differentiate. The homeobox genes that are expressed in the CMSC niche in placental tissues have not been identified. We used the novel strategy of laser capture microdissection to isolate the stromal component of first trimester villi and excluded the cytotrophoblast and syncytiotrophoblast layers that comprise the outer layer of the chorionic villi. Microarray analysis was then used to screen for homeobox genes in the microdissected tissue. Candidate homeobox genes were selected for further RNA analysis. Immunohistochemistry of candidate genes in first trimester placental villous stromal tissue revealed homeobox genes Meis1, myeloid ectropic viral integration site 1 homolog 2 (MEIS2), H2.0-like Drosophila (HLX), transforming growth factor β-induced factor (TGIF), and distal-less homeobox 5 (DLX5) were expressed in the vascular niche where CMSCs have been shown to reside. Expression of MEIS2, HLX, TGIF, and DLX5 was also detected in scattered stromal cells. Real-time polymerase chain reaction and immunocytochemistry verified expression of MEIS2, HLX, TGIF, and DLX5 homeobox genes in first trimester and term CMSCs. These data suggest a combination of regulatory homeobox genes is expressed in CMSCs from early placental development to term, which may be required for stem cell proliferation and differentiation. PMID:24692208

  20. Botulinum toxin A improves adipose tissue engraftment by promoting cell proliferation, adipogenesis and angiogenesis

    PubMed Central

    Tang, Qi; Chen, Chang; Wang, Xiaqi; Li, Wei; Zhang, Yan; Wang, Muyao; Jing, Wei; Wang, Hang; Guo, Weihua; Tian, Weidong

    2017-01-01

    Adipose tissue engraftment has become a well-established therapy in plastic and reconstructive surgery used to restore age-related or injury-related soft tissue loss. However, the unpredictable absorption rates limit its further application. Some clinicians have noted that more optimal aesthetic results are achieved when botulinum toxin A (BoNTA) is applied prior to adipose tissue grafting. In the present study, we transplanted allogeneic adipose tissue treated with or without BoNTA in SD rats in vivo. We subsequently evaluated the survival rate (weight, volume, apoptosis and cellular integrity) and revascularization of the adipose tissue. The results revealed that BoNTA improved the long-term weight and volume retention of the graft, and preserved cellular integrity. BoNTA significantly increased the expression levels of CD31 and vascular endothelial growth factor (VEGF), suggesting enhanced vasodilation and endothelial cell proliferation. In vitro, adipose-derived stem cells (ASCs) were isolated, identified and induced to proliferate and differentiate with or without BoNTA. Furthermore, to evaluate the proliferative, adipogenic and angiogenic ability of the ASCs, CCK-8 assay and Oil Red O staining were conducted. Gene and protein expression levels were analyzed by RT-qPCR and western blot analysis. The results revealed that 8×10−2 U/ml BoNTA as the optimal dose increased ASC proliferation and adipogenic differentiation capacity, as well as the expression level of the key cytokine of angiogenesis. On the whole, our findings indicate that BoNTA improves adipose tissue engraftment and promotes ASC regeneration, which could benefit future clinical applications. PMID:28731141

  1. TPL2 (Therapeutic Targeting Tumor Progression Locus-2)/ATF4 (Activating Transcription Factor-4)/SDF1α (Chemokine Stromal Cell-Derived Factor-α) Axis Suppresses Diabetic Retinopathy.

    PubMed

    Lai, De-Wei; Lin, Keng-Hung; Sheu, Wayne Huey-Herng; Lee, Maw-Rong; Chen, Chung-Yu; Lee, Wen-Jane; Hung, Yi-Wen; Shen, Chin-Chang; Chung, Tsung-Ju; Liu, Shing-Hwa; Sheu, Meei-Ling

    2017-09-01

    Diabetic retinopathy is characterized by vasopermeability, vascular leakage, inflammation, blood-retinal barrier breakdown, capillary degeneration, and neovascularization. However, the mechanisms underlying the association between diabetes mellitus and progression retinopathy remain unclear. TPL2 (tumor progression locus 2), a serine-threonine protein kinase, exerts a pathological effect on vascular angiogenesis. This study investigated the role of N ε -(carboxymethyl)lysine, a major advanced glycation end products, and the involved TPL2-related molecular signals in diabetic retinopathy using models of in vitro and in vivo and human samples. Serum N ε -(carboxymethyl)lysine levels and TPL2 kinase activity were significantly increased in clinical patients and experimental animals with diabetic retinopathy. Intravitreal administration of pharmacological blocker or neutralizing antibody inhibited TPL2 and effectively suppressed the pathological characteristics of retinopathy in streptozotocin-induced diabetic animal models. Intravitreal VEGF (vascular endothelial growth factor) neutralization also suppressed the diabetic retinopathy in diabetic animal models. Mechanistic studies in primary human umbilical vein endothelial cells and primary retinal microvascular endothelial cells from streptozotocin-diabetic rats, db/db mice, and samples from patients with diabetic retinopathy revealed a positive parallel correlation between N ε -(carboxymethyl)lysine and the TPL2/chemokine SDF1α (stromal cell-derived factor-α) axis that is dependent on endoplasmic reticulum stress-related molecules, especially ATF4 (activating transcription factor-4). This study demonstrates that inhibiting the N ε -(carboxymethyl)lysine-induced TPL2/ATF4/SDF1α axis can effectively prevent diabetes mellitus-mediated retinal microvascular dysfunction. This signaling axis may include the therapeutic potential for other diseases involving pathological neovascularization or macular edema. © 2017

  2. Decellularized silk fibroin scaffold primed with adipose mesenchymal stromal cells improves wound healing in diabetic mice

    PubMed Central

    2014-01-01

    Introduction Silk fibroin (SF) scaffolds have been shown to be a suitable substrate for tissue engineering and to improve tissue regeneration when cellularized with mesenchymal stromal cells (MSCs). We here demonstrate, for the first time, that electrospun nanofibrous SF patches cellularized with human adipose-derived MSCs (Ad-MSCs-SF), or decellularized (D-Ad-MSCs-SF), are effective in the treatment of skin wounds, improving skin regeneration in db/db diabetic mice. Methods The conformational and structural analyses of SF and D-Ad-MSCs-SF patches were performed by scanning electron microscopy, confocal microscopy, Fourier transform infrared spectroscopy and differential scanning calorimetry. Wounds were performed by a 5 mm punch biopsy tool on the mouse’s back. Ad-MSCs-SF and D-Ad-MSCs-SF patches were transplanted and the efficacy of treatments was assessed by measuring the wound closure area, by histological examination and by gene expression profile. We further investigated the in vitro angiogenic properties of Ad-MSCs-SF and D-Ad-MSCs-SF patches by affecting migration of human umbilical vein endothelial cells (HUVECs), keratinocytes (KCs) and dermal fibroblasts (DFs), through the aortic ring assay and, finally, by evaluating the release of angiogenic factors. Results We found that Ad-MSCs adhere and grow on SF, maintaining their phenotypic mesenchymal profile and differentiation capacity. Conformational and structural analyses on SF and D-Ad-MSCs-SF samples, showed that sterilization, decellularization, freezing and storing did not affect the SF structure. When grafted in wounds of diabetic mice, both Ad-MSCs-SF and D-Ad-MSCs-SF significantly improved tissue regeneration, reducing the wound area respectively by 40% and 35%, within three days, completing the process in around 10 days compared to 15–17 days of controls. RT2 gene profile analysis of the wounds treated with Ad-MSCs-SF and D-Ad-MSCs-SF showed an increment of genes involved in angiogenesis and

  3. Development of a 3D bone marrow adipose tissue model.

    PubMed

    Fairfield, Heather; Falank, Carolyne; Farrell, Mariah; Vary, Calvin; Boucher, Joshua M; Driscoll, Heather; Liaw, Lucy; Rosen, Clifford J; Reagan, Michaela R

    2018-01-26

    Over the past twenty years, evidence has accumulated that biochemically and spatially defined networks of extracellular matrix, cellular components, and interactions dictate cellular differentiation, proliferation, and function in a variety of tissue and diseases. Modeling in vivo systems in vitro has been undeniably necessary, but when simplified 2D conditions rather than 3D in vitro models are used, the reliability and usefulness of the data derived from these models decreases. Thus, there is a pressing need to develop and validate reliable in vitro models to reproduce specific tissue-like structures and mimic functions and responses of cells in a more realistic manner for both drug screening/disease modeling and tissue regeneration applications. In adipose biology and cancer research, these models serve as physiologically relevant 3D platforms to bridge the divide between 2D cultures and in vivo models, bringing about more reliable and translationally useful data to accelerate benchtop to bedside research. Currently, no model has been developed for bone marrow adipose tissue (BMAT), a novel adipose depot that has previously been overlooked as "filler tissue" but has more recently been recognized as endocrine-signaling and systemically relevant. Herein we describe the development of the first 3D, BMAT model derived from either human or mouse bone marrow (BM) mesenchymal stromal cells (MSCs). We found that BMAT models can be stably cultured for at least 3 months in vitro, and that myeloma cells (5TGM1, OPM2 and MM1S cells) can be cultured on these for at least 2 weeks. Upon tumor cell co-culture, delipidation occurred in BMAT adipocytes, suggesting a bidirectional relationship between these two important cell types in the malignant BM niche. Overall, our studies suggest that 3D BMAT represents a "healthier," more realistic tissue model that may be useful for elucidating the effects of MAT on tumor cells, and tumor cells on MAT, to identify novel therapeutic

  4. miR-21 modulates tumor outgrowth induced by human adipose tissue-derived mesenchymal stem cells in vivo

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shin, Keun Koo; Lee, Ae Lim; Kim, Jee Young

    2012-06-15

    Highlights: Black-Right-Pointing-Pointer miR-21 modulates hADSC-induced increase of tumor growth. Black-Right-Pointing-Pointer The action is mostly mediated by the modulation of TGF-{beta} signaling. Black-Right-Pointing-Pointer Inhibition of miR-21 enhances the blood flow recovery in hindlimb ischemia. -- Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in clinical situations, due principally to their potential use in regenerative medicine and tissue engineering applications. However, the therapeutic application of MSCs remains limited, unless the favorable effects of MSCs on tumor growth in vivo, and the long-term safety of the clinical applications of MSCs, can be more thoroughly understood. In this study, wemore » determined whether microRNAs can modulate MSC-induced tumor outgrowth in BALB/c nude mice. Overexpression of miR-21 in human adipose-derived stem cells (hADSCs) inhibited hADSC-induced tumor growth, and inhibition of miR-21 increased it. Downregulation of transforming growth factor beta receptor II (TGFBR2), but not of signal transducer and activator of transcription 3, in hADSCs showed effects similar to those of miR-21 overexpression. Downregulation of TGFBR2 and overexpression of miR21 decreased tumor vascularity. Inhibition of miR-21 and the addition of TGF-{beta} increased the levels of vascular endothelial growth factor and interleukin-6 in hADSCs. Transplantation of miR-21 inhibitor-transfected hADSCs increased blood flow recovery in a hind limb ischemia model of nude mice, compared with transplantation of control oligo-transfected cells. These findings indicate that MSCs might favor tumor growth in vivo. Thus, it is necessary to study the long-term safety of this technique before MSCs can be used as therapeutic tools in regenerative medicine and tissue engineering.« less

  5. VEGF and VEGFB Play Balancing Roles in Adipose Differentiation, Gene Expression, and Function.

    PubMed

    Jin, Honghong; Li, Dan; Wang, Xutong; Jia, Jia; Chen, Yang; Yao, Yapeng; Zhao, Chunlan; Lu, Xiaodan; Zhang, Shujie; Togo, Jacques; Ji, Yan; Zhang, Luqing; Feng, Xuechao; Zheng, Yaowu

    2018-05-01

    Obesity is the result of abnormal adipose development and energy metabolism. Using vascular endothelial growth factor (VEGF) B-knockout and inducible VEGF downregulation mouse models, we have shown that VEGFB inactivation caused expansion of white adipose, whitening of brown adipose, an increase in fat accumulation, and a reduction in energy consumption. At the same time, expression of the white adipose-associated genes was increased and brown adipose-associated genes decreased. VEGF repression, in contrast, induced brown adipose expansion and brown adipocyte development in white adipose, increased energy expenditure, upregulated brown adipose-associated genes, and downregulated white adipose-associated genes. When VEGFB-knockout and VEGF-repressed mice are crossed together, VEGF and VEGFB can counteractively regulate large numbers of genes and efficiently reverse each other's roles. These genes, under counteractive VEGF and VEGFB regulations, include transcription factors, adhesion molecules, and metabolic enzymes. This balancing role is confirmed by morphologic and functional changes. This study reports that VEGF and VEGFB counteractively regulate adipose development and function in energy metabolism.

  6. Isolation and characterization of equine endometrial mesenchymal stromal cells.

    PubMed

    Rink, B Elisabeth; Amilon, Karin R; Esteves, Cristina L; French, Hilari M; Watson, Elaine; Aurich, Christine; Donadeu, F Xavier

    2017-07-12

    Equine mesenchymal stromal/stem cells (MSCs) are most commonly harvested from bone marrow (BM) or adipose tissue, requiring the use of surgical procedures. By contrast, the uterus can be accessed nonsurgically, and may provide a more readily available cell source. While human endometrium is known to harbor mesenchymal precursor cells, MSCs have not been identified in equine endometrium. This study reports the isolation, culture, and characterization of MSCs from equine endometrium. The presence of MSC and pericyte markers in endometrial sections was determined using immunohistochemistry. Stromal cells were harvested and cultured after separation of epithelial cells from endometrial fragments using Mucin-1-bound beads. For comparison, MSCs were also harvested from BM. The expression of surface markers in endometrial and BM-derived MSCs was characterized using flow cytometry and quantitative polymerase chain reaction. MSCs were differentiated in vitro into adipogenic, chondrogenic, osteogenic, and smooth muscle lineages. Typical markers of MSCs (CD29, CD44, CD90, and CD105) and pericytes (NG2 and CD146) were localized in the equine endometrium. Both endometrial and BM MSCs grew clonally and robustly expressed MSC and pericyte markers in culture while showing greatly reduced or negligible expression of hematopoietic markers (CD45, CD34) and MHC-II. Additionally, both endometrial and BM MSCs differentiated into adipogenic, osteogenic, and chondrogenic lineages in vitro, and endometrial MSCs had a distinct ability to undergo smooth muscle differentiation. We have demonstrated for the first time the presence of cells in equine endometrium that fulfill the definition of MSCs. The equine endometrium may provide an alternative, easily accessible source of MSCs, not only for therapeutic regeneration of the uterus, but also for other tissues where MSCs from other sources are currently being used therapeutically.

  7. Whole transcriptome profiling of patient-derived xenograft models as a tool to identify both tumor and stromal specific biomarkers.

    PubMed

    Bradford, James R; Wappett, Mark; Beran, Garry; Logie, Armelle; Delpuech, Oona; Brown, Henry; Boros, Joanna; Camp, Nicola J; McEwen, Robert; Mazzola, Anne Marie; D'Cruz, Celina; Barry, Simon T

    2016-04-12

    The tumor microenvironment is emerging as a key regulator of cancer growth and progression, however the exact mechanisms of interaction with the tumor are poorly understood. Whilst the majority of genomic profiling efforts thus far have focused on the tumor, here we investigate RNA-Seq as a hypothesis-free tool to generate independent tumor and stromal biomarkers, and explore tumor-stroma interactions by exploiting the human-murine compartment specificity of patient-derived xenografts (PDX).Across a pan-cancer cohort of 79 PDX models, we determine that mouse stroma can be separated into distinct clusters, each corresponding to a specific stromal cell type. This implies heterogeneous recruitment of mouse stroma to the xenograft independent of tumor type. We then generate cross-species expression networks to recapitulate a known association between tumor epithelial cells and fibroblast activation, and propose a potentially novel relationship between two hypoxia-associated genes, human MIF and mouse Ddx6. Assessment of disease subtype also reveals MMP12 as a putative stromal marker of triple-negative breast cancer. Finally, we establish that our ability to dissect recruited stroma from trans-differentiated tumor cells is crucial to identifying stem-like poor-prognosis signatures in the tumor compartment.In conclusion, RNA-Seq is a powerful, cost-effective solution to global analysis of human tumor and mouse stroma simultaneously, providing new insights into mouse stromal heterogeneity and compartment-specific disease markers that are otherwise overlooked by alternative technologies. The study represents the first comprehensive analysis of its kind across multiple PDX models, and supports adoption of the approach in pre-clinical drug efficacy studies, and compartment-specific biomarker discovery.

  8. CASTIN: a system for comprehensive analysis of cancer-stromal interactome.

    PubMed

    Komura, Daisuke; Isagawa, Takayuki; Kishi, Kazuki; Suzuki, Ryohei; Sato, Reiko; Tanaka, Mariko; Katoh, Hiroto; Yamamoto, Shogo; Tatsuno, Kenji; Fukayama, Masashi; Aburatani, Hiroyuki; Ishikawa, Shumpei

    2016-11-09

    Cancer microenvironment plays a vital role in cancer development and progression, and cancer-stromal interactions have been recognized as important targets for cancer therapy. However, identifying relevant and druggable cancer-stromal interactions is challenging due to the lack of quantitative methods to analyze whole cancer-stromal interactome. We present CASTIN (CAncer-STromal INteractome analysis), a novel framework for the evaluation of cancer-stromal interactome from RNA-Seq data using cancer xenograft models. For each ligand-receptor interaction which is derived from curated protein-protein interaction database, CASTIN summarizes gene expression profiles of cancer and stroma into three evaluation indices. These indices provide quantitative evaluation and comprehensive visualization of interactome, and thus enable to identify critical cancer-microenvironment interactions, which would be potential drug targets. We applied CASTIN to the dataset of pancreas ductal adenocarcinoma, and successfully characterized the individual cancer in terms of cancer-stromal relationships, and identified both well-known and less-characterized druggable interactions. CASTIN provides comprehensive view of cancer-stromal interactome and is useful to identify critical interactions which may serve as potential drug targets in cancer-microenvironment. CASTIN is available at: http://github.com/tmd-gpat/CASTIN .

  9. Supercritical carbon dioxide extracted extracellular matrix material from adipose tissue.

    PubMed

    Wang, Jun Kit; Luo, Baiwen; Guneta, Vipra; Li, Liang; Foo, Selin Ee Min; Dai, Yun; Tan, Timothy Thatt Yang; Tan, Nguan Soon; Choong, Cleo; Wong, Marcus Thien Chong

    2017-06-01

    Adipose tissue is a rich source of extracellular matrix (ECM) material that can be isolated by delipidating and decellularizing the tissue. However, the current delipidation and decellularization methods either involve tedious and lengthy processes or require toxic chemicals, which may result in the elimination of vital proteins and growth factors found in the ECM. Hence, an alternative delipidation and decellularization method for adipose tissue was developed using supercritical carbon dioxide (SC-CO 2 ) that eliminates the need of any harsh chemicals and also reduces the amount of processing time required. The resultant SC-CO 2 -treated ECM material showed an absence of nuclear content but the preservation of key proteins such as collagen Type I, collagen Type III, collagen Type IV, elastin, fibronectin and laminin. In addition, other biological factors such as glycosaminoglycans (GAGs) and growth factors such as basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) were also retained. Subsequently, the resulting SC-CO 2 -treated ECM material was used as a bioactive coating on tissue culture plastic (TCP). Four different cell types including adipose tissue-derived mesenchymal stem cells (ASCs), human umbilical vein endothelial cells (HUVECs), immortalized human keratinocyte (HaCaT) cells and human monocytic leukemia cells (THP-1) were used in this study to show that the SC-CO 2 -treated ECM coating can be potentially used for various biomedical applications. The SC-CO 2 -treated ECM material showed improved cell-material interactions for all cell types tested. In addition, in vitro scratch wound assay using HaCaT cells showed that the presence of SC-CO 2 -treated ECM material enhanced keratinocyte migration whilst the in vitro cellular studies using THP-1-derived macrophages showed that the SC-CO 2 -treated ECM material did not evoke pro-inflammatory responses from the THP-1-derived macrophages. Overall, this study shows the efficacy

  10. Induction of angiogenesis in tissue-engineered scaffolds designed for bone repair: a combined gene therapy-cell transplantation approach.

    PubMed

    Jabbarzadeh, Ehsan; Starnes, Trevor; Khan, Yusuf M; Jiang, Tao; Wirtel, Anthony J; Deng, Meng; Lv, Qing; Nair, Lakshmi S; Doty, Steven B; Laurencin, Cato T

    2008-08-12

    One of the fundamental principles underlying tissue engineering approaches is that newly formed tissue must maintain sufficient vascularization to support its growth. Efforts to induce vascular growth into tissue-engineered scaffolds have recently been dedicated to developing novel strategies to deliver specific biological factors that direct the recruitment of endothelial cell (EC) progenitors and their differentiation. The challenge, however, lies in orchestration of the cells, appropriate biological factors, and optimal factor doses. This study reports an approach as a step forward to resolving this dilemma by combining an ex vivo gene transfer strategy and EC transplantation. The utility of this approach was evaluated by using 3D poly(lactide-co-glycolide) (PLAGA) sintered microsphere scaffolds for bone tissue engineering applications. Our goal was achieved by isolation and transfection of adipose-derived stromal cells (ADSCs) with adenovirus encoding the cDNA of VEGF. We demonstrated that the combination of VEGF releasing ADSCs and ECs results in marked vascular growth within PLAGA scaffolds. We thereby delineate the potential of ADSCs to promote vascular growth into biomaterials.

  11. Induction of angiogenesis in tissue-engineered scaffolds designed for bone repair: A combined gene therapy–cell transplantation approach

    PubMed Central

    Jabbarzadeh, Ehsan; Starnes, Trevor; Khan, Yusuf M.; Jiang, Tao; Wirtel, Anthony J.; Deng, Meng; Lv, Qing; Nair, Lakshmi S.; Doty, Steven B.; Laurencin, Cato T.

    2008-01-01

    One of the fundamental principles underlying tissue engineering approaches is that newly formed tissue must maintain sufficient vascularization to support its growth. Efforts to induce vascular growth into tissue-engineered scaffolds have recently been dedicated to developing novel strategies to deliver specific biological factors that direct the recruitment of endothelial cell (EC) progenitors and their differentiation. The challenge, however, lies in orchestration of the cells, appropriate biological factors, and optimal factor doses. This study reports an approach as a step forward to resolving this dilemma by combining an ex vivo gene transfer strategy and EC transplantation. The utility of this approach was evaluated by using 3D poly(lactide-co-glycolide) (PLAGA) sintered microsphere scaffolds for bone tissue engineering applications. Our goal was achieved by isolation and transfection of adipose-derived stromal cells (ADSCs) with adenovirus encoding the cDNA of VEGF. We demonstrated that the combination of VEGF releasing ADSCs and ECs results in marked vascular growth within PLAGA scaffolds. We thereby delineate the potential of ADSCs to promote vascular growth into biomaterials. PMID:18678895

  12. Early alterations in vascular contractility associated to changes in fatty acid composition and oxidative stress markers in perivascular adipose tissue

    PubMed Central

    2010-01-01

    Aim To test the early effect of fructose-induced changes in fatty acid composition and oxidative stress markers in perivascular adipose tissue (PVAT) upon vascular contractility. Methods Adult male Wistar rats were fed a commercial diet without (CD) or with 10% fructose (FRD) in the drinking water for 3 weeks. We measured plasma metabolic parameters, lipid composition and oxidative stress markers in aortic PVAT. Vascular contractility was measured in aortic rings sequentially, stimulated with serotonin (5-HT) and high K+-induced depolarization using intact and thereafter PVAT-deprived rings. Results Comparable body weights were recorded in both groups. FRD rats had increased plasma triglyceride and fructosamine levels. Their PVAT had an increased saturated to mono- or poly-unsaturated fatty acid ratio, a significant decrease in total superoxide dismutase and glutathione peroxidase activities and in the total content of glutathione. Conversely, lipid peroxidation (TBARS), nitric oxide content, and gluthathione reductase activity were significantly higher, indicating an increase in oxidative stress. In aortic rings, removal of PVAT increased serotonin-induced contractions, but the effect was significantly lower in rings from FRD rats. This effect was no longer observed when the two contractions were performed in PVAT-deprived rings. PVAT did not affect the contractions triggered by high K+-induced depolarization either in CD or FRD rats. Conclusions FRD induces multiple metabolic and endocrine systemic alterations which also alter PVAT and the vascular relaxant properties of this tissue. The changes in PVAT would affect its paracrine modulation of vascular function. PMID:20964827

  13. Identification of suitable reference genes for quantitative gene expression analysis in rat adipose stromal cells induced to trilineage differentiation.

    PubMed

    Santos, Bruno Paiva Dos; da Costa Diesel, Luciana Fraga; da Silva Meirelles, Lindolfo; Nardi, Nance Beyer; Camassola, Melissa

    2016-12-15

    This study was designed to (i) identify stable reference genes for the analysis of gene expression during in vitro differentiation of rat adipose stromal cells (rASCs), (ii) recommend stable genes for individual treatment conditions, and (iii) validate these genes by comparison with normalization results from stable and unstable reference genes. On the basis of a literature review, eight genes were selected: Actb, B2m, Hprt1, Ppia, Rplp0, Rpl13a, Rpl5, and Ywhaz. Genes were ranked according to their stability under different culture conditions as assessed using GenNorm, NormFinder, and RefFinder algorithms. Although the employed algorithms returned different rankings, the most frequently top-ranked genes were: B2m and/or Ppia for all 28day treatments (ALL28); Ppia and Hprt1 (adipogenic differentiation; A28), B2m (chondrogenic differentiation; C28), Rpl5 (controls maintained in complete culture medium; CCM), Rplp0 (osteogenic differentiation for 3days; O3), Rpl13a and Actb (osteogenic differentiation for 7days; O7), Rplp0 and Ppia (osteogenic differentiation for 14days; O14), Hprt1 and Ppia (osteogenic differentiation for 28days; O28), as well as Actb (all osteogenesis time points combined; ALLOSTEO). The obtained results indicate that the performance of reference genes depends on the differentiation protocol and on the analysis time, thus providing valuable information for the design of RT-PCR experiments. Copyright © 2016. Published by Elsevier B.V.

  14. Comparison of the paracrine activity of mesenchymal stem cells derived from human umbilical cord, amniotic membrane and adipose tissue.

    PubMed

    Dabrowski, Filip A; Burdzinska, Anna; Kulesza, Agnieszka; Sladowska, Anna; Zolocinska, Aleksandra; Gala, Kamila; Paczek, Leszek; Wielgos, Miroslaw

    2017-11-01

    The study was conducted to investigate secretory activity and define the paracrine potential of mesenchymal stem cells from human umbilical cord and amniotic membrane (UC-MSCs and AM-MSCs, respectively). UC-MSCs (n = 6) were obtained from tissue explants using an adherent method after two weeks of incubation. AM-MSCs (n = 6) were obtained by digestion with tripsin and collagenase. MSC phenotype was confirmed in vitro by performing flow cytometry, differentiation assays and vimentin staining. Supernatants were collected after 48 h culturing in serum-free conditions and the following concentrations were determined: epidermal growth factor (EGF), interleukin (IL)-6, IL-10, tumor necrosis factor-α, transforming growth factor-β (TGF-β), vascular endothelial growth factor-α (VEGF-α) and metalloproteinase (MMP) 1, 8 and 13, using multiplex supernatant cytokine assay. Data were compared with adipose tissue derived MSCs (AD-MSCs, n = 6). Both UC-MSC and AM-MSC populations were positively identified as MSCs by flow cytometry and differentiation potential into bone, cartilage and adipose tissue. Using a multiple cytokine detection assay, we proved that both UC-MSCs and AM-MSCs show high secretive capacity. However, the secretion profile differed between cells from various sources. UC-MSCs showed significantly higher production of TGF-β and lower production of VEGF-α, compared to AD-MSCs (P = 0.004) and AM-MSCs (P = 0.039) and lower levels of EGF (P = 0005). AM-MSCs showed significantly lower levels of MMP-8 than UC-MSCs (P = 0.024); however, there was no difference in levels of released cytokines compared to AD-MSCs. AM-MSCs show similar IL production as AD-MSCs, while UC-MSCs have a significantly different profile, which suggests diverse biological potential of both cell types for immunomodulative and regenerative therapy. © 2017 Japan Society of Obstetrics and Gynecology.

  15. Rhodamine bound maghemite as a long-term dual imaging nanoprobe of adipose tissue-derived mesenchymal stromal cells.

    PubMed

    Cmiel, Vratislav; Skopalik, Josef; Polakova, Katerina; Solar, Jan; Havrdova, Marketa; Milde, David; Justan, Ivan; Magro, Massimiliano; Starcuk, Zenon; Provaznik, Ivo

    2017-07-01

    In the last few years, magnetically labeled cells have been intensively explored, and non-invasive cell tracking and magnetic manipulation methods have been tested in preclinical studies focused on cell transplantation. For clinical applications, it is desirable to know the intracellular pathway of nanoparticles, which can predict their biocompatibility with cells and the long-term imaging properties of labeled cells. Here, we quantified labeling efficiency, localization, and fluorescence properties of Rhodamine derivatized superparamagnetic maghemite nanoparticles (SAMN-R) in mesenchymal stromal cells (MSC). We investigated the stability of SAMN-R in the intracellular space during a long culture (20 days). Analyses were based on advanced confocal microscopy accompanied by atomic absorption spectroscopy (AAS) and magnetic resonance imaging. SAMN-R displayed excellent cellular uptake (24 h of labeling), and no toxicity of SAMN-R labeling was found. 83% of SAMN-R nanoparticles were localized in lysosomes, only 4.8% were found in mitochondria, and no particles were localized in the nucleus. On the basis of the MSC fluorescence measurement every 6 days, we also quantified the continual decrease of SAMN-R fluorescence in the average single MSC during 18 days. An additional set of analyses showed that the intracellular SAMN-R signal decrease was minimally caused by fluorophore degradation or nanoparticles extraction from the cells, main reason is a cell division. The fluorescence of SAMN-R nanoparticles within the cells was detectable minimally for 20 days. These observations indicate that SAMN-R nanoparticles have a potential for application in transplantation medicine.

  16. γδ T cells producing interleukin-17A regulate adipose regulatory T cell homeostasis and thermogenesis.

    PubMed

    Kohlgruber, Ayano C; Gal-Oz, Shani T; LaMarche, Nelson M; Shimazaki, Moto; Duquette, Danielle; Nguyen, Hung N; Mina, Amir I; Paras, Tyler; Tavakkoli, Ali; von Andrian, Ulrich; Banks, Alexander S; Shay, Tal; Brenner, Michael B; Lynch, Lydia

    2018-05-01

    γδ T cells are situated at barrier sites and guard the body from infection and damage. However, little is known about their roles outside of host defense in nonbarrier tissues. Here, we characterize a highly enriched tissue-resident population of γδ T cells in adipose tissue that regulate age-dependent regulatory T cell (T reg ) expansion and control core body temperature in response to environmental fluctuations. Mechanistically, innate PLZF + γδ T cells produced tumor necrosis factor and interleukin (IL) 17 A and determined PDGFRα + and Pdpn + stromal-cell production of IL-33 in adipose tissue. Mice lacking γδ T cells or IL-17A exhibited decreases in both ST2 + T reg cells and IL-33 abundance in visceral adipose tissue. Remarkably, these mice also lacked the ability to regulate core body temperature at thermoneutrality and after cold challenge. Together, these findings uncover important physiological roles for resident γδ T cells in adipose tissue immune homeostasis and body-temperature control.

  17. Heterogeneity of proangiogenic features in mesenchymal stem cells derived from bone marrow, adipose tissue, umbilical cord, and placenta.

    PubMed

    Du, Wen Jing; Chi, Ying; Yang, Zhou Xin; Li, Zong Jin; Cui, Jun Jie; Song, Bao Quan; Li, Xue; Yang, Shao Guang; Han, Zhi Bo; Han, Zhong Chao

    2016-11-10

    Mesenchymal stem cells (MSCs) have been widely proven effective for therapeutic angiogenesis in ischemia animal models as well as clinical vascular diseases. Because of the invasive method, limited resources, and aging problems of adult tissue-derived MSCs, more perinatal tissue-derived MSCs have been isolated and studied as promising substitutable MSCs for cell transplantation. However, fewer studies have comparatively studied the angiogenic efficacy of MSCs derived from different tissues sources. Here, we evaluated whether the in-situ environment would affect the angiogenic potential of MSCs. We harvested MSCs from adult bone marrow (BMSCs), adipose tissue (AMSCs), perinatal umbilical cord (UMSCs), and placental chorionic villi (PMSCs), and studied their "MSC identity" by flow cytometry and in-vitro trilineage differentiation assay. Then we comparatively studied their endothelial differentiation capabilities and paracrine actions side by side in vitro. Our data showed that UMSCs and PMSCs fitted well with the minimum standard of MSCs as well as BMSCs and AMSCs. Interestingly, we found that MSCs regardless of their tissue origins could develop similar endothelial-relevant functions in vitro, including producing eNOS and uptaking ac-LDL during endothelial differentiation in spite of their feeble expression of endothelial-related genes and proteins. Additionally, we surprisingly found that BMSCs and PMSCs could directly form tubular structures in vitro on Matrigel and their conditioned medium showed significant proangiogenic bioactivities on endothelial cells in vitro compared with those of AMSCs and UMSCs. Besides, several angiogenic genes were upregulated in BMSCs and PMSCs in comparison with AMSCs and UMSCs. Moreover, enzyme-linked immunosorbent assay further confirmed that BMSCs secreted much more VEGF, and PMSCs secreted much more HGF and PGE2. Our study demonstrated the heterogeneous proangiogenic properties of MSCs derived from different tissue origins, and

  18. Rehabilitation program after mesenchymal stromal cell transplantation augmented by vascularized bone grafts for idiopathic osteonecrosis of the femoral head: a preliminary study.

    PubMed

    Aoyama, Tomoki; Fujita, Yasuko; Madoba, Katsuyuki; Nankaku, Manabu; Yamada, Minoru; Tomita, Motoko; Goto, Koji; Ikeguchi, Ryosuke; Kakinoki, Ryosuke; Matsuda, Shuichi; Nakamura, Takashi; Toguchida, Junya

    2015-03-01

    To determine the feasibility and safety of implementing a 12-week rehabilitation program after mesenchymal stromal cell (MSC) transplantation augmented by vascularized bone grafting for idiopathic osteonecrosis (ION) of the femoral head. A prospective case series. University clinical research laboratory. Participants (N=10) with ION who received MSC transplantation augmented by vascularized bone grafting. A 12-week exercise program, which included range-of-motion (ROM) exercises, muscle-strengthening exercises, and aerobic training. Measures of ROM, muscle strength, Timed Up and Go test, and Medical Outcomes Study 36-Item Short-Form Health Survey (SF-36) were collected before surgery and again at 6 and 12 months after surgery. All participants completed the 12-week program. External rotation ROM as well as extensor and abductor muscle strength significantly improved 6 months after treatment compared with that before treatment (P<.05). Significant improvements were also seen in physical function, role physical, and bodily pain subgroup scores of the SF-36 (P<.05). No serious adverse events occurred. This study demonstrates the feasibility and safety of a multiplex rehabilitation program after MSC transplantation and provides support for further study on the benefits of rehabilitation programs in regenerative medicine. Copyright © 2015 American Congress of Rehabilitation Medicine. Published by Elsevier Inc. All rights reserved.

  19. Serum Progranulin Concentrations May Be Associated With Macrophage Infiltration Into Omental Adipose Tissue

    PubMed Central

    Youn, Byung-Soo; Bang, Sa-Ik; Klöting, Nora; Park, Ji Woo; Lee, Namseok; Oh, Ji-Eun; Pi, Kyung-Bae; Lee, Tae Hee; Ruschke, Karen; Fasshauer, Mathias; Stumvoll, Michael; Blüher, Matthias

    2009-01-01

    OBJECTIVE—Progranulin is an important molecule in inflammatory response. Chronic inflammation is frequently associated with central obesity and associated disturbances; however, the role of circulating progranulin in human obesity, type 2 diabetes, and dyslipidemia is unknown. RESEARCH DESIGN AND METHODS—For the measurement of progranulin serum concentrations, we developed an enzyme-linked immunosorbent assay (ELISA). Using this ELISA, we assessed circulating progranulin in a cross-sectional study of 209 subjects with a wide range of obesity, body fat distribution, insulin sensitivity, and glucose tolerance and in 60 individuals with normal (NGT) or impaired (IGT) glucose tolerance or type 2 diabetes before and after a 4-week physical training program. Progranulin mRNA and protein expression was measured in paired samples of omental and subcutaneous adipose tissue (adipocytes and cells of the stromal vascular fraction) from 55 lean or obese individuals. Measurement of Erk activation and chemotactic activity induced by progranulin in vitro was performed using THP-1–based cell migration assays. RESULTS—Progranulin serum concentrations were significantly higher in individuals with type 2 diabetes compared with NGT and in obese subjects with predominant visceral fat accumulation. Circulating progranulin significantly correlates with BMI, macrophage infiltration in omental adipose tissue, C-reactive protein (CRP) serum concentrations, A1C values, and total cholesterol. Multivariable linear regression analyses revealed CRP levels as the strongest independent predictor of circulating progranulin. The extent of in vitro progranulin-mediated chemotaxis is similar to that of monocyte chemoattractant protein-1 but independent of Gα. Moreover, in type 2 diabetes, but not in IGT and NGT individuals, physical training for 4 weeks resulted in significantly decreased circulating progranulin levels. CONCLUSIONS—Elevated progranulin serum concentrations are associated

  20. Mesenchymal stromal cells improve human islet function through released products and extracellular matrix.

    PubMed

    Arzouni, Ahmed A; Vargas-Seymour, Andreia; Rackham, Chloe L; Dhadda, Paramjeet; Huang, Guo-Cai; Choudhary, Pratik; Nardi, Nance; King, Aileen J F; Jones, Peter M

    2017-12-01

    The aims of the present study were (i) to determine whether the reported beneficial effects of mesenchymal stromal cells (MSCs) on mouse islet function extend to clinically relevant human tissues (islets and MSCs), enabling translation into improved protocols for clinical human islet transplantation; and (ii) to identify possible mechanisms through which human MSCs influence human islet function. Human islets were co-cultured with human adipose tissue-derived MSCs (hASCs) or pre-treated with its products - extracellular matrix (ECM) and annexin A1 (ANXA1). Mouse islets were pre-treated with mouse MSC-derived ECM. Islet insulin secretory function was assessed in vitro by radioimmunoassay. Quantitative RT-PCR was used to screen human adipMSCs for potential ligands of human islet G-protein-coupled receptors. We show that co-culture with hASCs improves human islet secretory function in vitro , as measured by glucose-stimulated insulin secretion, confirming previous reports using rodent tissues. Furthermore, we demonstrate that these beneficial effects on islet function can be partly attributed to the MSC-derived products ECM and ANXA1. Our results suggest that hASCs have the potential to improve the quality of human islets isolated for transplantation therapy of Type 1 diabetes. Furthermore, it may be possible to achieve improvements in human islet quality in a cell-free culture system by using the MSC-derived products ANXA1 and ECM. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.