Characterization of basic amino acids-conjugated PAMAM dendrimers as gene carriers for human adipose-derived mesenchymal stem cells.

PubMed

Bae, Yoonhee; Lee, Sunray; Green, Eric S; Park, Jung Hyun; Ko, Kyung Soo; Han, Jin; Choi, Joon Sig

2016-03-30

Since mesenchymal stem cells (MSCs) can self-renew and differentiate into multiple cell types, the delivery of genes to this type of cell can be an important tool in the emerging field of tissue regeneration and engineering. However, development of more efficient and safe nonviral vectors for gene delivery to stem cells in particular still remains a great challenge. In this study, we describe a group of nonviral gene delivery vectors, conjugated PAMAM derivatives (PAMAM-H-R, PAMAM-H-K, and PAMAM-H-O), displaying affinity toward human adipose-derived mesenchymal stem cells (AD-MSCs). Transfection efficiency using pDNA encoding for luciferase (Luc) and enhanced green fluorescent protein (EGFP), and cytotoxicity assays were performed in human AD-MSCs. The results show that transfection efficiencies of conjugated PAMAM derivatives are improved significantly compared to native PAMAM dendrimer, and that among PAMAM derivatives, cytotoxicity of PAMAM-H-K and PAMAM-H-O were very low. Also, treatment of human AD-MSCs to polyplex formation in conjugated PAMAM derivatives, their cellular uptake and localization were analyzed by flow cytometry and confocal microscopy. Copyright © 2016 Elsevier B.V. All rights reserved.

Generation and characterization of human iPSC line generated from mesenchymal stem cells derived from adipose tissue.

PubMed

Zapata-Linares, Natalia; Rodriguez, Saray; Mazo, Manuel; Abizanda, Gloria; Andreu, Enrique J; Barajas, Miguel; Prosper, Felipe; Rodriguez-Madoz, Juan R

2016-01-01

In this work, mesenchymal stem cells derived from adipose tissue (ADSCs) were used for the generation of the human-induced pluripotent stem cell line G15.AO. Cell reprogramming was performed using retroviral vectors containing the Yamanaka factors, and the generated G15.AO hiPSC line showed normal karyotype, silencing of the exogenous reprogramming factors, induction of the typical pluripotency-associated markers, alkaline phosphatase enzymatic activity, and in vivo and in vitro differentiation ability to the three germ layers. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

Matrix directed adipogenesis and neurogenesis of mesenchymal stem cells derived from adipose tissue and bone marrow.

PubMed

Lee, Junmin; Abdeen, Amr A; Tang, Xin; Saif, Taher A; Kilian, Kristopher A

2016-09-15

Mesenchymal stem cells (MSCs) can differentiate into multiple lineages through guidance from the biophysical and biochemical properties of the extracellular matrix. In this work we conduct a combinatorial study of matrix properties that influence adipogenesis and neurogenesis including: adhesion proteins, stiffness, and cell geometry, for mesenchymal stem cells derived from adipose tissue (AT-MSCs) and bone marrow (BM-MSCs). We uncover distinct differences in integrin expression, the magnitude of traction stress, and lineage specification to adipocytes and neuron-like cells between cell sources. In the absence of media supplements, adipogenesis in AT-MSCs is not significantly influenced by matrix properties, while the converse is true in BM-MSCs. Both cell types show changes in the expression of neurogenesis markers as matrix cues are varied. When cultured on laminin conjugated microislands of the same adhesive area, BM-MSCs display elevated adipogenesis markers, while AT-MSCs display elevated neurogenesis markers; integrin analysis suggests neurogenesis in AT-MSCs is guided by adhesion through integrin αvβ3. Overall, the properties of the extracellular matrix guides MSC adhesion and lineage specification to different degrees and outcomes, in spite of their similarities in general characteristics. This work will help guide the selection of MSCs and matrix components for applications where high fidelity of differentiation outcome is desired. Mesenchymal stem cells (MSCs) are an attractive cell type for stem cell therapies; however, in order for these cells to be useful in medicine, we need to understand how they respond to the physical and chemical environments of tissue. Here, we explore how two promising sources of MSCs-those derived from bone marrow and from adipose tissue-respond to the compliance and composition of tissue using model extracellular matrices. Our results demonstrate a source-specific propensity to undergo adipogenesis and neurogenesis, and

Immortalization of canine adipose-derived mesenchymal stem cells and their seminiferous tubule transplantation.

PubMed

Fang, Jia; Wei, Yudong; Teng, Xin; Zhao, Shanting; Hua, Jinlian

2018-04-01

Adipose-derived mesenchymal stem cells (ADSCs) are proven to provide good effects in numerous tissue engineering application and other cell-based therapies. However, the difficulty in the proliferation of ADSCs, known as the "Hayflick limit" in vitro, limits their clinical application. Here, we immortalized canine ADSCs (cADSCs) with SV40 gene and transplanted them into busulfan-induced seminiferous tubules of infertile mice. The proliferation of these immortalized cells was improved significantly. Then, cellular differentiation assays showed that the immortalized cADSCs could differentiate into three-germ-layer cells, osteogenesis, chondrogenesis, adipogenesis phenotypes, and primordial germ cell-like cells (PGCLCs). In addition, the immortalized cADSCs can proliferate in the busulfan-induced seminiferous tubules of infertile mice. These findings confirmed that the immortalized cADSCs maintain the criteria of cADSCs. © 2017 Wiley Periodicals, Inc.

GC-Rich Extracellular DNA Induces Oxidative Stress, Double-Strand DNA Breaks, and DNA Damage Response in Human Adipose-Derived Mesenchymal Stem Cells.

PubMed

Kostyuk, Svetlana; Smirnova, Tatiana; Kameneva, Larisa; Porokhovnik, Lev; Speranskij, Anatolij; Ershova, Elizaveta; Stukalov, Sergey; Izevskaya, Vera; Veiko, Natalia

2015-01-01

Cell free DNA (cfDNA) circulates throughout the bloodstream of both healthy people and patients with various diseases. CfDNA is substantially enriched in its GC-content as compared with human genomic DNA. Exposure of haMSCs to GC-DNA induces short-term oxidative stress (determined with H2DCFH-DA) and results in both single- and double-strand DNA breaks (comet assay and γH2AX, foci). As a result in the cells significantly increases the expression of repair genes (BRCA1 (RT-PCR), PCNA (FACS)) and antiapoptotic genes (BCL2 (RT-PCR and FACS), BCL2A1, BCL2L1, BIRC3, and BIRC2 (RT-PCR)). Under the action of GC-DNA the potential of mitochondria was increased. Here we show that GC-rich extracellular DNA stimulates adipocyte differentiation of human adipose-derived mesenchymal stem cells (haMSCs). Exposure to GC-DNA leads to an increase in the level of RNAPPARG2 and LPL (RT-PCR), in the level of fatty acid binding protein FABP4 (FACS analysis) and in the level of fat (Oil Red O). GC-rich fragments in the pool of cfDNA can potentially induce oxidative stress and DNA damage response and affect the direction of mesenchymal stem cells differentiation in human adipose-derived mesenchymal stem cells. Such a response may be one of the causes of obesity or osteoporosis.

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

Adipose-derived mesenchymal stem cells from liposuction and resected fat are feasible sources for regenerative medicine.

PubMed

Schneider, Sandra; Unger, Marina; van Griensven, Martijn; Balmayor, Elizabeth R

2017-05-19

The use of mesenchymal stem cells (MSCs) in research and in regenerative medicine has progressed. Bone marrow as a source has drawbacks because of subsequent morbidities. An easily accessible and valuable source is adipose tissue. This type of tissue contains a high number of MSCs, and obtaining higher quantities of tissue is more feasible. Fat tissue can be harvested using different methods such as liposuction and resection. First, a detailed isolation protocol with complete characterization is described. This also includes highlighting problems and pitfalls. Furthermore, some comparisons of these different harvesting methods exist. However, the later characterization of the cells is conducted poorly in most cases. We performed an in-depth characterization over five passages including an investigation of the effect of freezing and thawing. Characterization was performed using flow cytometry with CD markers, metabolic activity with Alamar Blue, growth potential in between passages, and cytoskeleton staining. Our results show that the cells isolated with distinct isolation methods (solid versus liposuction "liquid") have the same MSC potential. However, the percentage of cells positive for the markers CD73, CD90, and CD105 is initially quite low. The cells isolated from the liquid fat tissue grow faster at higher passages, and significantly more cells display MSC markers. In summary, we show a simple and efficient method to isolate adipose-derived mesenchymal stem cells from different preparations. Liposuctions and resection can be used, whereas liposuction has more growth potential at higher passages.

In vivo cartilage formation using chondrogenic-differentiated human adipose-derived mesenchymal stem cells mixed with fibrin glue.

PubMed

Jung, Sung-No; Rhie, Jong Won; Kwon, Ho; Jun, Young Joon; Seo, Je-Won; Yoo, Gyeol; Oh, Deuk Young; Ahn, Sang Tae; Woo, Jihyoun; Oh, Jieun

2010-03-01

Human adipose-derived mesenchymal stem cells (MSCs) were differentiated into chondrogenic MSCs, and fibrin glue was used together to explore the feasibility of whether cartilages can be generated in vivo by injecting the differentiated cells. Mesenchymal stem cells extracted from human adipose were differentiated into chondrogenic MSCs, and such differentiated cells mixed with fibrin glue were injected subcutaneously into the back of the nude mouse. In addition to visual evaluation of the tissues formed after 4, 8, and 12 weeks, hematoxylin-eosin staining, Masson trichrome staining, measurement of glycosaminoglycan concentration using dimethylmethylene blue, agreecan through reverse transcriptase-polymerase chain reaction, type II collagen, and expression of SOX-9 were verified. Moreover, the results were compared with 2 groups of controls: 1 control group that received only injection of chondrogenic-differentiated MSC and the supporting control group that received only fibrin glue injection. For the experimental group, cartilage-like tissues were formed after 4, 8, and 12 weeks. Formation of cartilage tissues was not observed in any of 4, 8, and 12 weeks of the control group. The supporting control group had only a small structure formation after 4 weeks, but the formed structure was completely decomposed by the 8th and 12th weeks. The range of staining dramatically increased with time at 4, 8, and 12 weeks in Masson trichrome staining. The concentration of glycosaminoglycan also increased with time. The increased level was statistically significant with more than 3 times more after 8 weeks compared with 4 weeks and more than 2 times more after 12 weeks compared with 8 weeks. Also, in reverse transcriptase-polymerase chain reaction at 4, 8, and 12 weeks, all results expressed a cartilage-specific gene called aggrecan, type II collagen, and SOX-9. The study verified that the chondrogenic-differentiated MSCs derived from human adipose tissues with fibrin glue can

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

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

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.

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

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

Mesenchymal Stem Cells From Bone Marrow, Adipose Tissue, and Lung Tissue Differentially Mitigate Lung and Distal Organ Damage in Experimental Acute Respiratory Distress Syndrome.

PubMed

Silva, Johnatas D; Lopes-Pacheco, Miquéias; Paz, Ana H R; Cruz, Fernanda F; Melo, Elga B; de Oliveira, Milena V; Xisto, Débora G; Capelozzi, Vera L; Morales, Marcelo M; Pelosi, Paolo; Cirne-Lima, Elizabeth; Rocco, Patricia R M

2018-02-01

Mesenchymal stem cells-based therapies have shown promising effects in experimental acute respiratory distress syndrome. Different mesenchymal stem cells sources may result in diverse effects in respiratory diseases; however, there is no information regarding the best source of mesenchymal stem cells to treat pulmonary acute respiratory distress syndrome. We tested the hypothesis that mesenchymal stem cells derived from bone marrow, adipose tissue, and lung tissue would lead to different beneficial effects on lung and distal organ damage in experimental pulmonary acute respiratory distress syndrome. Animal study and primary cell culture. Laboratory investigation. Seventy-five Wistar rats. Wistar rats received saline (control) or Escherichia coli lipopolysaccharide (acute respiratory distress syndrome) intratracheally. On day 2, acute respiratory distress syndrome animals were further randomized to receive saline or bone marrow, adipose tissue, or lung tissue mesenchymal stem cells (1 × 10 cells) IV. Lung mechanics, histology, and protein levels of inflammatory mediators and growth factors were analyzed 5 days after mesenchymal stem cells administration. RAW 264.7 cells (a macrophage cell line) were incubated with lipopolysaccharide followed by coculture or not with bone marrow, adipose tissue, and lung tissue mesenchymal stem cells (10 cells/mL medium). Regardless of mesenchymal stem cells source, cells administration improved lung function and reduced alveolar collapse, tissue cellularity, collagen, and elastic fiber content in lung tissue, as well as decreased apoptotic cell counts in liver. Bone marrow and adipose tissue mesenchymal stem cells administration also reduced levels of tumor necrosis factor-α, interleukin-1β, keratinocyte-derived chemokine, transforming growth factor-β, and vascular endothelial growth factor, as well as apoptotic cell counts in lung and kidney, while increasing expression of keratinocyte growth factor in lung tissue

Metformin preconditioned adipose derived mesenchymal stem cells is a better option for the reversal of diabetes upon transplantation.

PubMed

Shree, Nitya; Bhonde, Ramesh R

2016-12-01

Metformin is used worldwide as an insulin sensitizer. Adipose derived mesenchymal stem cells have shown promising results in the reducing hyperglycemia. We examined whether preconditioning of adipose derived mesenchymal stem cells (ASCs) with metformin could have a better therapeutic value for the reversal of type 2 diabetes. We compared the effect of metformin, ASCs and metformin preconditioned ASCs (MetASCs) in high fat diet induced C57BL/6 mice by injecting the cells intramuscularly only once where as metformin was given at a concentration of 300mg per kg body weight orally daily. Fasting glucose was measured every week for 4 weeks. At the end of the study insulin, triglycerides, IL6 and oxidised LDL were evaluated from the serum. Gene expression studies were performed for muscle (GLUT4) and liver tissues (IL6 and PAI1).There was a remarkable decrease in hyperglycemia within two weeks of injection by MetASCs as compared to metformin and ASCs alone. A significant decrement of hyperinsulinemia, triglyceridemia, serum IL6 and oxidised LDL were observed at the end of the study. Gene expression studies for muscle tissue revealed the drastic upregulation of GLUT4 gene levels in the MetASCs group indicating enhanced glucose uptake in muscle. Liver tissue analysed for the genes involved in inflammation viz. IL6 and PAI1 showed significant downregulation in the MetASCs group as compared to the other groups. This is a first report demonstrating the synergistic effect of metformin preconditioning of ASCs leading to reversal of hyperglycemia, hyperinsulinemia and triglyceridemia. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

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.

Isolation, culture and chondrogenic differentiation of canine adipose tissue- and bone marrow-derived mesenchymal stem cells--a comparative study.

PubMed

Reich, Christine M; Raabe, Oksana; Wenisch, Sabine; Bridger, Philip S; Kramer, Martin; Arnhold, Stefan

2012-06-01

In the dog, mesenchymal stem cells (MSCs) have been shown to reside in the bone marrow (bone marrow-derived mesenchymal stem cells: BM-MSCs) as well as in the adipose tissue (adipose tissue-derived stem cells: ADSCs). Potential application fields for these multipotent MSCs in small animal practice are joint diseases as MSCs of both sources have shown to possess chondrogenic differentiation ability. However, it is not clear whether the chondrogenic differentiation potential of cells of these two distinct tissues is truly equal. Therefore, we compared MSCs of both origins in this study in terms of their chondrogenic differentiation ability and suitability for clinical application. BM-MSCs harvested from the femoral neck and ADSCs from intra-abdominal fat tissue were examined for their morphology, population doubling time (PDT) and CD90 surface antigen expression. RT-PCR served to assess expression of pluripotency marker Oct4 and early differentiation marker genes. Chondrogenic differentiation ability was compared and validated using histochemistry, transmission electron microscopy (TEM) and quantitative RT-PCR. Both cell populations presented a highly similar morphology and marker expression in an undifferentiated stage except that freshly isolated ADSCs demonstrated a significantly faster PDT than BM-MSCs. In contrast, BM-MSCs revealed a morphological superior cartilage formation by the production of a more abundant and structured hyaline matrix and higher expression of lineage specific genes under the applied standard differentiation protocol. However, further investigations are necessary in order to find out if chondrogenic differentiation can be improved in canine ADSCs using different protocols and/or supplements.

L-carnitine significantly decreased aging of rat adipose tissue-derived mesenchymal stem cells.

PubMed

Mobarak, Halimeh; Fathi, Ezzatollah; Farahzadi, Raheleh; Zarghami, Nosratollah; Javanmardi, Sara

2017-03-01

Mesenchymal stem cells are undifferentiated cells that have the ability to divide continuously and tissue regeneration potential during the transplantation. Aging and loss of cell survival, is one of the main problems in cell therapy. Since the production of free radicals in the aging process is effective, the use of antioxidant compounds can help in scavenging free radicals and prevent the aging of cells. The aim of this study is evaluate the effects of L-carnitine (LC) on proliferation and aging of rat adipose tissue-derived mesenchymal stem cells (rADSC). rADSCs were isolated from inguinal region of 5 male Rattus rats. Oil red-O, alizarin red-S and toluidine blue staining were performed to evaluate the adipogenic, osteogenic and chondrogenic differentiation of rADSCs, respectively. Flow cytometric analysis was done for investigating the cell surface markers. The methyl thiazol tetrazolium (MTT) method was used to determine the cell proliferation of rADSCs following exposure to different concentrations of LC. rADSCs aging was evaluated by beta-galactosidase staining. The results showed significant proliferation of rADSCs 48 h after treatment with concentrations of 0.2 mM LC. In addition, in the presence of 0.2 mM LC, rADSCs appeared to be growing faster than control group and 0.2 mM LC supplementation could significantly decrease the population doubling time and aging of rADSCs. It seems that LC would be a good antioxidant to improve lifespan of rADSCs due to the decrease in aging.

Diabetic human adipose tissue-derived mesenchymal stem cells fail to differentiate in functional adipocytes.

PubMed

Barbagallo, Ignazio; Li Volti, Giovanni; Galvano, Fabio; Tettamanti, Guido; Pluchinotta, Francesca R; Bergante, Sonia; Vanella, Luca

2017-05-01

Adipose tissue dysfunction represents a hallmark of diabetic patients and is a consequence of the altered homeostasis of this tissue. Mesenchymal stem cells (MSCs) and their differentiation into adipocytes contribute significantly in maintaining the mass and function of adult adipose tissue. The aim of this study was to evaluate the differentiation of MSCs from patients suffering type 2 diabetes (dASC) and how such process results in hyperplasia or rather a stop of adipocyte turnover resulting in hypertrophy of mature adipocytes. Our results showed that gene profile of all adipogenic markers is not expressed in diabetic cells after differentiation indicating that diabetic cells fail to differentiate into adipocytes. Interestingly, delta like 1, peroxisome proliferator-activated receptor alpha, and interleukin 1β were upregulated whereas Sirtuin 1 and insulin receptor substrate 1 gene expression were found downregulated in dASC compared to cells obtained from healthy subjects. Taken together our data indicate that dASC lose their ability to differentiate into mature and functional adipocytes. In conclusion, our in vitro study is the first to suggest that diabetic patients might develop obesity through a hypertrophy of existing mature adipocytes due to failure turnover of adipose tissue. Impact statement In the present manuscript, we evaluated the differentiative potential of mesenchymal stem cells (MSCs) in adipocytes obtained from healthy and diabetic patients. This finding could be of great potential interest for the field of obesity in order to exploit such results to further understand the pathophysiological processes underlying metabolic syndrome. In particular, inflammation in diabetic patients causes a dysfunction in MSCs differentiation and a decrease in adipocytes turnover leading to insulin resistance.

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.

Treatment with adipose derived mesenchymal stem cells and their conditioned media reverse carrageenan induced paw oedema in db/db mice.

PubMed

Shree, Nitya; Venkategowda, Sunil; Venkatranganna, M V; Bhonde, Ramesh R

2017-06-01

Mesenchymal stem cells are known for anti inflammatory and immunomodulatory activities. The aim of our study was to evaluate the effect of human adipose derived mesenchymal stem cells (hADMSCs) and its conditioned media (CM) on carrageenan induced acute inflammation in db/db mice. We injected 5×10 5 ADMSCs or the CM in the inflamed paw. We assessed the paw volume, serum IL6 levels and histopathology of the paw to reveal the anti inflammatory effect. We observed a single injection of hADMSCs or CM could reverse the inflammation within 24h as evidenced by reduction in paw volume, IL6 levels and histological examination. Our result equivocally demonstrates the role of CM in normalising the inflammation better than hADMSCs. This study will pave way for an alternative to anti inflammatory drugs. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

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

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.

Comparative analysis of human UCB and adipose tissue derived mesenchymal stem cells for their differentiation potential into brown and white adipocytes.

PubMed

Rashnonejad, Afrooz; Ercan, Gulinnaz; Gunduz, Cumhur; Akdemir, Ali; Tiftikcioglu, Yigit Ozer

2018-06-01

The differentiation potential of umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) into brown and white adipocytes in comparison to Adipose tissue derived MSCs (AD-MSCs) were investigated in order to characterize their potency for future cell therapies. MSCs were isolated from ten UCB samples and six liposuction materials. MSCs were differentiated into white and brown adipocytes after characterization by flow cytometry. Differentiated adipocytes were stained with Oil Red O and hematoxylin/eosin. The UCP1 protein levels in brown adipocytes were investigated by immunofluoresence and western blot analysis. Cells that expressed mesenchymal stem cells markers (CD34-, CD45-, CD90+ and CD105+) were successfully isolated from UCB and adipose tissue. Oil Red O staining demonstrated that white and brown adipocytes obtained from AD-MSCs showed 85 and 61% of red pixels, while it was 3 and 1.9%, respectively for white and brown adipocytes obtained from UCB-MSCs. Fluorescence microscopy analysis showed strong uncoupling protein 1 (UCP1) signaling in brown adipocytes, especially which were obtained from AD-MSCs. Quantification of UCP1 protein amount showed 4- and 10.64-fold increase in UCP1 contents of brown adipocytes derived from UCB-MSCs and AD-MSCs, respectively in comparison to undifferentiated MSCs (P < 0.004). UCB-MSCs showed only a little differentiation tendency into adipocytes means it is not an appropriate stem cell type to be differentiated into these cell types. In contrast, high differentiation efficiency of AD-MSCs into brown and white adipocytes make it appropriate stem cell type to use in future regenerative medicine of soft tissue disorders or fighting with obesity and its related disorders.

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.

Fas-L promotes the stem cell potency of adipose-derived mesenchymal cells.

PubMed

Solodeev, Inna; Meilik, Benjamin; Volovitz, Ilan; Sela, Meirav; Manheim, Sharon; Yarkoni, Shai; Zipori, Dov; Gur, Eyal; Shani, Nir

2018-06-11

Fas-L is a TNF family member known to trigger cell death. It has recently become evident that Fas-L can transduce also non-apoptotic signals. Mesenchymal stem cells (MSCs) are multipotent cells that are derived from various adult tissues. Although MSCs from different tissues display common properties they also display tissue-specific characteristics. Previous works have demonstrated massive apoptosis following Fas-L treatment of bone marrow-derived MSCs both in vitro and following their administration in vivo. We therefore set to examine Fas-L-induced responses in adipose-derived stem cells (ASCs). Human ASCs were isolated from lipoaspirates and their reactivity to Fas-L treatment was examined. ASCs responded to Fas-L by simultaneous apoptosis and proliferation, which yielded a net doubling of cell quantities and a phenotypic shift, including reduced expression of CD105 and increased expression of CD73, in association with increased bone differentiation potential. Treatment of freshly isolated ASCs led to an increase in large colony forming unit fibroblasts, likely produced by early stem cell progenitor cells. Fas-L-induced apoptosis and proliferation signaling were found to be independent as caspase inhibition attenuated Fas-L-induced apoptosis without impacting proliferation, whereas inhibition of PI3K and MEK, but not of JNK, attenuated Fas-L-dependent proliferation, but not apoptosis. Thus, Fas-L signaling in ASCs leads to their expansion and phenotypic shift toward a more potent stem cell state. We speculate that these reactions ensure the survival of ASC progenitor cells encountering Fas-L-enriched environments during tissue damage and inflammation and may also enhance ASC survival following their administration in vivo.

The morphology, proliferation rate, and population doubling time factor of adipose-derived mesenchymal stem cells cultured on to non-aqueous SiO2, TiO2, and hybrid sol-gel-derived oxide coatings.

PubMed

Marycz, Krzysztof; Krzak-Roś, Justyna; Donesz-Sikorska, Anna; Śmieszek, Agnieszka

2014-11-01

In recent years, much attention has been paid to the development of tissue engineering and regenerative medicine, especially when stem cells of various sources are concerned. In addition to the interest in mesenchymal stem cells isolated from bone marrow, recently more consideration has been given to stem cells isolated from adipose tissue (AdMSCs), due to their less invasive method of collection as well as their ease of isolation and culture. However, the development of regenerative medicine requires both the application of biocompatible material and the stem cells to accelerate the regeneration. In this study, we investigated the morphology, proliferation rate index (PRi), and population doubling time factor of adipose-derived mesenchymal stem cells cultured on non-aqueous sol-gel-derived SiO2, TiO2, and SiO2/TiO2 oxide coatings. The results indicated an increase in PRi of AdMSCs when cultured on to titanium dioxide, suggesting its high attractiveness for AdMSCs. In addition, the proper morphology and the shortest doubling time of AdMSCs were observed when cultured on titanium dioxide coating. © 2014 Wiley Periodicals, Inc.

Efficient transduction of equine adipose-derived mesenchymal stem cells by VSV-G pseudotyped lentiviral vectors.

PubMed

Petersen, Gayle F; Hilbert, Bryan; Trope, Gareth; Kalle, Wouter; Strappe, Padraig

2014-12-01

Equine adipose-derived mesenchymal stem cells (EADMSC) provide a unique cell-based approach for treatment of a variety of equine musculoskeletal injuries, via regeneration of diseased or damaged tissue, or the secretion of immunomodulatory molecules. These capabilities can be further enhanced by genetic modification using lentiviral vectors, which provide a safe and efficient method of gene delivery. We investigated the suitability of lentiviral vector technology for gene delivery into EADMSC, using GFP expressing lentiviral vectors pseudotyped with the G glycoprotein from the vesicular stomatitis virus (V-GFP) or, for the first time, the baculovirus gp64 envelope protein (G-GFP). In this study, we produced similarly high titre V-GFP and G-GFP lentiviral vectors. Flow cytometric analysis showed efficient transduction using V-GFP; however G-GFP exhibited a poor ability to transduce EADMSC. Transduction resulted in sustained GFP expression over four passages, with minimal effects on cell viability and doubling time, and an unaltered chondrogenic differentiation potential. Copyright © 2014 Elsevier Ltd. All rights reserved.

Functional characteristics of mesenchymal stem cells derived from the adipose tissue of a patient with achondroplasia.

PubMed

Park, Jeong-Ran; Lee, Hanbyeol; Kim, Chung-Hyo; Hong, Seok-Ho; Ha, Kwon-Soo; Yang, Se-Ran

2016-05-01

Mesenchymal stem cells (MSCs) can be isolated from various tissues including bone marrow, adipose tissue, skin dermis, and umbilical Wharton's jelly as well as injured tissues. MSCs possess the capacity for self-renewal and the potential for differentiation into adipogenic, osteogenic, and chondrogenic lineages. However, the characteristics of MSCs in injured tissues, such as achondroplasia (ACH), are not well known. In this study, we isolated MSCs from human subcutaneous adipose (ACH-SAMSCs) tissue and circumjacent human adipose tissue of the cartilage (ACH-CAMSCs) from a patient with ACH. We then analyzed the characterization of ACH-SAMSCs and ACH-CAMSCs, compared with normal human dermis-derived MSCs (hDMSCs). In flow cytometry analysis, the isolated ACH-MSCs expressed low levels of CD73, CD90, and CD105, compared with hDMSCs. Moreover, both ACH- SAMSCs and ACH-CAMSCs had constitutionally overactive fibroblast growth factor receptor 3 (FGFR3) and exhibited significantly reduced osteogenic differentiation, compared to enhanced adipogenic differentiation. The activity of extracellular signal-regulated kinases 1/2 (ERK1/2) and p38 mitogen-activated protein kinases (p38 MAPK) was increased in ACH-MSCs. In addition, the efficacy of osteogenic differentiation was slightly restored in osteogenic differentiation medium with MAPKs inhibitors. These results suggest that they play essential roles in MSC differentiation toward adipogenesis in ACH pathology. In conclusion, the identification of the characteristics of ACH-MSCs and the favoring of adipogenic differentiation via the FGFR3/MAPK axis might help to elucidate the pathogenic mechanisms relevant to other skeletal diseases and could provide targets for therapeutic interventions.

From Human Mesenchymal Stem Cells to Insulin-Producing Cells: Comparison between Bone Marrow- and Adipose Tissue-Derived Cells.

PubMed

Gabr, Mahmoud M; Zakaria, Mahmoud M; Refaie, Ayman F; Abdel-Rahman, Engy A; Reda, Asmaa M; Ali, Sameh S; Khater, Sherry M; Ashamallah, Sylvia A; Ismail, Amani M; Ismail, Hossam El-Din A; El-Badri, Nagwa; Ghoneim, Mohamed A

2017-01-01

The aim of this study is to compare human bone marrow-derived mesenchymal stem cells (BM-MSCs) and adipose tissue-derived mesenchymal stem cells (AT-MSCs), for their differentiation potentials to form insulin-producing cells. BM-MSCs were obtained during elective orthotopic surgery and AT-MSCs from fatty aspirates during elective cosmetics procedures. Following their expansion, cells were characterized by phenotyping, trilineage differentiation ability, and basal gene expression of pluripotency genes and for their metabolic characteristics. Cells were differentiated according to a Trichostatin-A based protocol. The differentiated cells were evaluated by immunocytochemistry staining for insulin and c-peptide. In addition the expression of relevant pancreatic endocrine genes was determined. The release of insulin and c-peptide in response to a glucose challenge was also quantitated. There were some differences in basal gene expression and metabolic characteristics. After differentiation the proportion of the resulting insulin-producing cells (IPCs), was comparable among both cell sources. Again, there were no differences neither in the levels of gene expression nor in the amounts of insulin and c-peptide release as a function of glucose challenge. The properties, availability, and abundance of AT-MSCs render them well-suited for applications in regenerative medicine. Conclusion . BM-MSCs and AT-MSCs are comparable regarding their differential potential to form IPCs. The availability and properties of AT-MSCs render them well-suited for applications in regenerative medicine.

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

Osteogenic Performance of Donor-Matched Human Adipose and Bone Marrow Mesenchymal Cells Under Dynamic Culture

PubMed Central

Wu, Wei; Le, Andrew V.; Mendez, Julio J.; Chang, Julie; Niklason, Laura E.

2015-01-01

Adipose-derived mesenchymal cells (ACs) and bone marrow-derived mesenchymal cells (BMCs) have been widely used for bone regeneration and can be seeded on a variety of rigid scaffolds. However, to date, a direct comparison of mesenchymal cells (MC) harvested from different tissues from the same donor and cultured in identical osteogenic conditions has not been investigated. Indeed, it is unclear whether marrow-derived or fat-derived MC possess intrinsic differences in bone-forming capabilities, since within-patient comparisons have not been previously done. This study aims at comparing ACs and BMCs from three donors ranging in age from neonatal to adult. Matched cells from each donor were studied in three distinct bioreactor settings, to determine the best method to create a viable osseous engineered construct. Human ACs and BMCs were isolated from each donor, cultured, and seeded on decellularized porcine bone (DCB) constructs. The constructs were then subjected to either static or dynamic (stirring or perfusion) bioreactor culture conditions for 7–21 days. Afterward, the constructs were analyzed for cell adhesion and distribution and osteogenic differentiation. ACs demonstrated higher seeding efficiency than BMCs. However, static and dynamic culture significantly increased BMCs proliferation more than ACs. In all conditions, BMCs demonstrated stronger osteogenic activity as compared with ACs, through higher alkaline phosphatase activity and gene expression for various bony markers. Conversely, ACs expressed more collagen I, which is a nonspecific matrix molecule in most connective tissues. Overall, dynamic bioreactor culture conditions enhanced osteogenic gene expression in both ACs and BMCs. Scaffolds seeded with BMCs in dynamic stirring culture conditions exhibit the greatest osteogenic proliferation and function in vitro, proving that marrow-derived MC have superior bone-forming potential as compared with adipose-derived cells. PMID:25668104

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.

Impact of bacteria and bacterial components on osteogenic and adipogenic differentiation of adipose-derived mesenchymal stem cells

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

Fiedler, Tomas, E-mail: tomas.fiedler@med.uni-rostock.de; Salamon, Achim; Adam, Stefanie

Adult mesenchymal stem cells (MSC) are present in several tissues, e.g. bone marrow, heart muscle, brain and subcutaneous adipose tissue. In invasive infections MSC get in contact with bacteria and bacterial components. Not much is known about how bacterial pathogens interact with MSC and how contact to bacteria influences MSC viability and differentiation potential. In this study we investigated the impact of three different wound infection relevant bacteria, Escherichia coli, Staphylococcus aureus, and Streptococcus pyogenes, and the cell wall components lipopolysaccharide (LPS; Gram-negative bacteria) and lipoteichoic acid (LTA; Gram-positive bacteria) on viability, proliferation, and osteogenic as well as adipogenic differentiationmore » of human adipose tissue-derived mesenchymal stem cells (adMSC). We show that all three tested species were able to attach to and internalize into adMSC. The heat-inactivated Gram-negative E. coli as well as LPS were able to induce proliferation and osteogenic differentiation but reduce adipogenic differentiation of adMSC. Conspicuously, the heat-inactivated Gram-positive species showed the same effects on proliferation and adipogenic differentiation, while its cell wall component LTA exhibited no significant impact on adMSC. Therefore, our data demonstrate that osteogenic and adipogenic differentiation of adMSC is influenced in an oppositional fashion by bacterial antigens and that MSC-governed regeneration is not necessarily reduced under infectious conditions. - Highlights: • Staphylococcus aureus, Streptococcus pyogenes and Escherichia coli bind to and internalize into adMSC. • Heat-inactivated cells of these bacterial species trigger proliferation of adMSC. • Heat-inactivated E. coli and LPS induce osteogenic differentiation of adMSC. • Heat-inactivated E. coli and LPS reduce adipogenic differentiation of adMSC. • LTA does not influence adipogenic or osteogenic differentiation of adMSC.« less

The enhancement of differentiating adipose derived mesenchymal stem cells toward hepatocyte like cells using gelatin cryogel scaffold.

PubMed

Ghaderi Gandomani, Maryam; Sahebghadam Lotfi, Abbas; Kordi Tamandani, Dormohammad; Arjmand, Sareh; Alizadeh, Shaban

2017-09-30

Liver tissue engineering creates a promising methodology for developing functional tissue to restore or improve the function of lost or damaged liver by using appropriate cells and biologically compatible scaffolds. The present paper aims to study the hepatogenic potential of human adipose derived mesenchymal stem cells (hADSCs) on a 3D gelatin scaffold in vitro. For this purpose, mesenchymal stem cells were isolated from human adipose tissue and characterized by flowcytometry analysis and mesodermal lineage differentiation capacity. Then, porous cryogel scaffolds were fabricated by cryogelating the gelatin using glutaraldehyde as the crosslinking agent. The structure of the scaffolds as well as the adhesion and proliferation of the cells were then determined by Scanning Electron Microscopy (SEM) analysis and MTT assay, respectively. The efficiency of hepatic differentiation of hADSCs on 2D and 3D culture systems has been assessed by means of morphological, cytological, molecular and biochemical approaches. Based on the results of flowcytometry, the isolated cells were positive for hMSC specific markers and negative for hematopoietic markers. Further, the multipotency of these cells was confirmed by adipogenic and osteogenic differentiation and the highly porous structure of scaffolds was characterized by SEM images. Biocompatibility was observed in the fabricated gelatin scaffolds and the adhesion and proliferation of hADSCs were promoted without any cytotoxicity effects. In addition, compared to 2D TCPS, the fabricated scaffolds provided more appropriate microenvironment resulting in promoting the differentiation of hADSCs toward hepatocyte-like cells with higher expression of hepatocyte-specific markers and appropriate functional characteristics such as increased levels of urea biosynthesis and glycogen storage. Finally, the created 3D gelatin scaffold could provide an appropriate matrix for hepatogenic differentiation of hADSCs, which could be considered for

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

Adipose-derived mesenchymal stem cells for cartilage tissue engineering: state-of-the-art in in vivo studies.

PubMed

Veronesi, Francesca; Maglio, Melania; Tschon, Matilde; Aldini, Nicolò Nicoli; Fini, Milena

2014-07-01

Several therapeutic approaches have been developed to address hyaline cartilage regeneration, but to date, there is no universal procedure to promote the restoration of mechanical and functional properties of native cartilage, which is one of the most important challenges in orthopedic surgery. For cartilage tissue engineering, adult mesenchymal stem cells (MSCs) are considered as an alternative cell source to chondrocytes. Since little is known about adipose-derived mesenchymal stem cell (ADSC) cartilage regeneration potential, the aim of this review was to give an overview of in vivo studies about the chondrogenic potential and regeneration ability of culture-expanded ADSCs when implanted in heterotopic sites or in osteoarthritic and osteochondral defects. The review compares the different studies in terms of number of implanted cells and animals, cell harvesting sites, in vitro expansion and chondrogenic induction conditions, length of experimental time, defect dimensions, used scaffolds and post-explant analyses of the cartilage regeneration. Despite variability of the in vivo protocols, it seems that good cartilage formation and regeneration were obtained with chondrogenically predifferentiated ADSCs (1 × 10(7) cells for heterotopic cartilage formation and 1 × 10(6) cells/scaffold for cartilage defect regeneration) and polymeric scaffolds, even if many other aspects need to be clarified in future studies. © 2013 Wiley Periodicals, Inc.

Human adipose tissue-derived mesenchymal stem cells inhibit T-cell lymphoma growth in vitro and in vivo.

PubMed

Ahn, Jin-Ok; Chae, Ji-Sang; Coh, Ye-Rin; Jung, Woo-Sung; Lee, Hee-Woo; Shin, Il-Seob; Kang, Sung-Keun; Youn, Hwa-Young

2014-09-01

Human mesenchymal stem cells (hMSCs) are thought to be one of the most reliable stem cell sources for a variety of cell therapies. This study investigated the anti-tumor effect of human adipose tissue-derived mesenchymal stem cells (hAT-MSCs) on EL4 murine T-cell lymphoma in vitro and in vivo. The growth-inhibitory effect of hAT-MSCs on EL4 tumor cells was evaluated using a WST-1 cell proliferation assay. Cell-cycle arrest and apoptosis were investigated by flow cytometry and western blot. To evaluate an anti-tumor effect of hAT-MSCs on T-cell lymphoma in vivo, CM-DiI-labeled hAT-MSCs were circumtumorally injected in tumor-bearing nude mice, and tumor size was measured. hAT-MSCs inhibited T-cell lymphoma growth by altering cell-cycle progression and inducing apoptosis in vitro. hAT-MSCs inhibited tumor growth in tumor-bearing nude mice and prolonged survival time. Immunofluorescence analysis showed that hAT-MSCs migrated to tumor sites. hAT-MSCs suppress the growth of T-cell lymphoma, suggesting a therapeutic option for T-cell lymphoma. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Mesenchymal Stem Cells from Adipose Tissue in Clinical Applications for Dermatological Indications and Skin Aging.

PubMed

Gaur, Meenakshi; Dobke, Marek; Lunyak, Victoria V

2017-01-20

Operating at multiple levels of control, mesenchymal stem cells from adipose tissue (ADSCs) communicate with organ systems to adjust immune response, provide signals for differentiation, migration, enzymatic reactions, and to equilibrate the regenerative demands of balanced tissue homeostasis. The identification of the mechanisms by which ADSCs accomplish these functions for dermatological rejuvenation and wound healing has great potential to identify novel targets for the treatment of disorders and combat aging. Herein, we review new insights into the role of adipose-derived stem cells in the maintenance of dermal and epidermal homeostasis, and recent advances in clinical applications of ADSCs related to dermatology.

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-mesenchymal

Re-using blood products as an alternative supplement in the optimisation of clinical-grade adipose-derived mesenchymal stem cell culture

PubMed Central

Phetfong, J.; Tawonsawatruk, T.; Seenprachawong, K.; Srisarin, A.; Isarankura-Na-Ayudhya, C.

2017-01-01

Objectives Adipose-derived mesenchymal stem cells (ADMSCs) are a promising strategy for orthopaedic applications, particularly in bone repair. Ex vivo expansion of ADMSCs is required to obtain sufficient cell numbers. Xenogenic supplements should be avoided in order to minimise the risk of infections and immunological reactions. Human platelet lysate and human plasma may be an excellent material source for ADMSC expansion. In the present study, use of blood products after their recommended transfusion date to prepare human platelet lysate (HPL) and human plasma (Hplasma) was evaluated for in vitro culture expansion and osteogenesis of ADMSCs. Methods Human ADMSCs were cultured in medium supplemented with HPL, Hplasma and a combination of HPL and Hplasma (HPL+Hplasma). Characteristics of these ADMSCs, including osteogenesis, were evaluated in comparison with those cultured in fetal bovine serum (FBS). Results HPL and HPL+Hplasma had a significantly greater growth-promoting effect than FBS, while Hplasma exhibited a similar growth-promoting effect to that of FBS. ADMSCs cultured in HPL and/or Hplasma generated more colony-forming unit fibroblasts (CFU-F) than those cultured in FBS. After long-term culture, ADMSCs cultured in HPL and/or Hplasma showed reduced cellular senescence, retained typical cell phenotypes, and retained differentiation capacities into osteogenic and adipogenic lineages. Conclusion HPL and Hplasma prepared from blood products after their recommended transfusion date can be used as an alternative and effective source for large-scale ex vivo expansion of ADMSCs. Cite this article: J. Phetfong, T. Tawonsawatruk, K. Seenprachawong, A. Srisarin, C. Isarankura-Na-Ayudhya, A. Supokawej. Re-using blood products as an alternative supplement in the optimisation of clinical-grade adipose-derived mesenchymal stem cell culture. Bone Joint Res 2017;6:414–422. DOI: 10.1302/2046-3758.67.BJR-2016-0342.R1. PMID:28720606

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.

Cholinergic and dopaminergic neuronal differentiation of human adipose tissue derived mesenchymal stem cells.

PubMed

Marei, Hany El Sayed; El-Gamal, Aya; Althani, Asma; Afifi, Nahla; Abd-Elmaksoud, Ahmed; Farag, Amany; Cenciarelli, Carlo; Thomas, Caceci; Anwarul, Hasan

2018-02-01

Mesenchymal stem cells (MSCs) are multipotent cells that can differentiate into various cell types such as cartilage, bone, and fat cells. Recent studies have shown that induction of MSCs in vitro by growth factors including epidermal growth factor (EGF) and fibroblast growth factor (FGF2) causes them to differentiate into neural like cells. These cultures also express ChAT, a cholinergic marker; and TH, a dopaminergic marker for neural cells. To establish a protocol with maximum differentiation potential, we examined MSCs under three experimental culture conditions using neural induction media containing FGF2, EGF, BMP-9, retinoic acid, and heparin. Adipose-derived MSCs were extracted and expanded in vitro for 3 passages after reaching >80% confluency, for a total duration of 9 days. Cells were then characterized by flow cytometry for CD markers as CD44 positive and CD45 negative. MSCs were then treated with neural induction media and were characterized by morphological changes and Q-PCR. Differentiated MSCs expressed markers for immature and mature neurons; β Tubulin III (TUBB3) and MAP2, respectively, showing the neural potential of these cells to differentiate into functional neurons. Improved protocols for MSCs induction will facilitate and ensure the reproducibility and standard production of MSCs for therapeutic applications in neurodegenerative diseases. © 2017 Wiley Periodicals, Inc.

From Human Mesenchymal Stem Cells to Insulin-Producing Cells: Comparison between Bone Marrow- and Adipose Tissue-Derived Cells

PubMed Central

Abdel-Rahman, Engy A.; Reda, Asmaa M.; Ashamallah, Sylvia A.; Ismail, Amani M.; Ismail, Hossam El-Din A.; El-Badri, Nagwa

2017-01-01

The aim of this study is to compare human bone marrow-derived mesenchymal stem cells (BM-MSCs) and adipose tissue-derived mesenchymal stem cells (AT-MSCs), for their differentiation potentials to form insulin-producing cells. BM-MSCs were obtained during elective orthotopic surgery and AT-MSCs from fatty aspirates during elective cosmetics procedures. Following their expansion, cells were characterized by phenotyping, trilineage differentiation ability, and basal gene expression of pluripotency genes and for their metabolic characteristics. Cells were differentiated according to a Trichostatin-A based protocol. The differentiated cells were evaluated by immunocytochemistry staining for insulin and c-peptide. In addition the expression of relevant pancreatic endocrine genes was determined. The release of insulin and c-peptide in response to a glucose challenge was also quantitated. There were some differences in basal gene expression and metabolic characteristics. After differentiation the proportion of the resulting insulin-producing cells (IPCs), was comparable among both cell sources. Again, there were no differences neither in the levels of gene expression nor in the amounts of insulin and c-peptide release as a function of glucose challenge. The properties, availability, and abundance of AT-MSCs render them well-suited for applications in regenerative medicine. Conclusion. BM-MSCs and AT-MSCs are comparable regarding their differential potential to form IPCs. The availability and properties of AT-MSCs render them well-suited for applications in regenerative medicine. PMID:28584815

Osteogenic differentiation of equine adipose tissue derived mesenchymal stem cells using CaCl2.

PubMed

Elashry, Mohamed I; Baulig, Nadine; Heimann, Manuela; Bernhardt, Caroline; Wenisch, Sabine; Arnhold, Stefan

2018-04-01

Adipose tissue derived mesenchymal stem cells (ASCs) may be used to cure bone defects after osteogenic differentiation. In this study we tried to optimize osteogenic differentiation for equine ASCs using various concentrations of CaCl 2 in comparison to the standard osteogenic protocol. ASCs were isolated from subcutaneous adipose tissue from mixed breed horses. The osteogenic induction protocols were (1) the standard osteogenic medium (OM) composed of dexamethasone, ascorbic acid and β-glycerol phosphate; (2) CaCl 2 based protocol composed of 3, 5 and 7.5mM CaCl 2 . Differentiation and proliferation were evaluated at 7, 10, 14 and 21days post-differentiation induction using the alizarin red staining (ARS) detecting matrix calcification. Semi-quantification of cell protein content, ARS and alkaline phosphatase activity (ALP) were performed using an ELISA reader. Quantification of the transcription level for the common osteogenic markers alkaline phosphatase (ALP) and Osteopontin (OP) was performed using RT-qPCR. In the presence of CaCl 2 , a concentration dependent effect on the osteogenic differentiation capacity was evident by the ARS evaluation and OP gene expression. We provide evidence that 5 and 7mM CaCl 2 enhance the osteogenic differentiation compared to the OM protocol. Although, there was a clear commitment of ASCs to the osteogenic fate in the presence of 5 and 7mM CaCl 2 , cell proliferation was increased compared to OM. We report that an optimized CaCl 2 protocol reliably influences ASCs osteogenesis while conserving the proliferation capacity. Thus, using these protocols provide a platform for using ASCs as a cell source in bone tissue engineering. Copyright © 2017 Elsevier Ltd. All rights reserved.

[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.

Silk fibroin/chitosan thin film promotes osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells.

PubMed

Li, Da-Wei; He, Jin; He, Feng-Li; Liu, Ya-Li; Liu, Yang-Yang; Ye, Ya-Jing; Deng, Xudong; Yin, Da-Chuan

2018-04-01

As a biodegradable polymer thin film, silk fibroin/chitosan composite film overcomes the defects of pure silk fibroin and chitosan films, respectively, and shows remarkable biocompatibility, appropriate hydrophilicity and mechanical properties. Silk fibroin/chitosan thin film can be used not only as metal implant coating for bone injury repair, but also as tissue engineering scaffold for skin, cornea, adipose, and other soft tissue injury repair. However, the biocompatibility of silk fibroin/chitosan thin film for mesenchymal stem cells, a kind of important seed cell of tissue engineering and regenerative medicine, is rarely reported. In this study, silk fibroin/chitosan film was prepared by solvent casting method, and the rat bone marrow-derived mesenchymal stem cells were cultured on the silk fibroin/chitosan thin film. Osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells were induced, respectively. The proliferation ability, osteogenic and adipogenic differentiation abilities of rat bone marrow-derived mesenchymal stem cells were systematically compared between silk fibroin/chitosan thin film and polystyrene tissue culture plates. The results showed that silk fibroin/chitosan thin film not only provided a comparable environment for the growth and proliferation of rat bone marrow-derived mesenchymal stem cells but also promoted their osteogenic and adipogenic differentiation. This work provided information of rat bone marrow-derived mesenchymal stem cells behavior on silk fibroin/chitosan thin film and extended the application of silk fibroin/chitosan thin film. Based on the results, we suggested that the silk fibroin/chitosan thin film could be a promising material for tissue engineering of bone, cartilage, adipose, and skin.

Effect of extracorporeal shock wave on proliferation and differentiation of equine adipose tissue-derived mesenchymal stem cells in vitro

PubMed Central

Raabe, O; Shell, K; Goessl, A; Crispens, C; Delhasse, Y; Eva, A; Scheiner-Bobis, G; Wenisch, S; Arnhold, S

2013-01-01

Mesenchymal stem cells are regarded as common cellular precursors of the musculoskeletal tissue and are responsible for tissue regeneration in the course of musculoskeletal disorders. In equine veterinary medicine extracorporeal shock wave therapy (ESWT) is used to optimize healing processes of bone, tendon and cartilage. Nevertheless, little is known about the effects of the shock waves on cells and tissues. Thus, the aim of this study was to investigate the influence of focused ESWT on the viability, proliferation, and differentiation capacity of adipose tissue-derived mesenchymal stem cells (ASCs) and to explore its effects on gap junctional communication and the activation of signalling cascades associated with cell proliferation and differentiation. ASCs were treated with different pulses of focused ESWT. Treated cells showed increased proliferation and expression of Cx43, as detected by means of qRT-PCR, histological staining, immunocytochemistry and western blot. At the same time, cells responded to ESWT by significant activation (phosphorylation) of Erk1/2, detected in western blots. No significant effects on the differentiation potential of the ASCs were evident. Taken together, the present results show significant effects of shock waves on stem cells in vitro. PMID:23671817

Mesenchymal Stem Cells from Adipose Tissue in Clinical Applications for Dermatological Indications and Skin Aging

PubMed Central

Gaur, Meenakshi; Dobke, Marek; Lunyak, Victoria V.

2017-01-01

Operating at multiple levels of control, mesenchymal stem cells from adipose tissue (ADSCs) communicate with organ systems to adjust immune response, provide signals for differentiation, migration, enzymatic reactions, and to equilibrate the regenerative demands of balanced tissue homeostasis. The identification of the mechanisms by which ADSCs accomplish these functions for dermatological rejuvenation and wound healing has great potential to identify novel targets for the treatment of disorders and combat aging. Herein, we review new insights into the role of adipose-derived stem cells in the maintenance of dermal and epidermal homeostasis, and recent advances in clinical applications of ADSCs related to dermatology. PMID:28117680

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.

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

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.

GC-Rich Extracellular DNA Induces Oxidative Stress, Double-Strand DNA Breaks, and DNA Damage Response in Human Adipose-Derived Mesenchymal Stem Cells

PubMed Central

Smirnova, Tatiana; Kameneva, Larisa; Porokhovnik, Lev; Speranskij, Anatolij; Ershova, Elizaveta; Stukalov, Sergey; Izevskaya, Vera; Veiko, Natalia

2015-01-01

Background. Cell free DNA (cfDNA) circulates throughout the bloodstream of both healthy people and patients with various diseases. CfDNA is substantially enriched in its GC-content as compared with human genomic DNA. Principal Findings. Exposure of haMSCs to GC-DNA induces short-term oxidative stress (determined with H2DCFH-DA) and results in both single- and double-strand DNA breaks (comet assay and γH2AX, foci). As a result in the cells significantly increases the expression of repair genes (BRCA1 (RT-PCR), PCNA (FACS)) and antiapoptotic genes (BCL2 (RT-PCR and FACS), BCL2A1, BCL2L1, BIRC3, and BIRC2 (RT-PCR)). Under the action of GC-DNA the potential of mitochondria was increased. Here we show that GC-rich extracellular DNA stimulates adipocyte differentiation of human adipose-derived mesenchymal stem cells (haMSCs). Exposure to GC-DNA leads to an increase in the level of RNAPPARG2 and LPL (RT-PCR), in the level of fatty acid binding protein FABP4 (FACS analysis) and in the level of fat (Oil Red O). Conclusions. GC-rich fragments in the pool of cfDNA can potentially induce oxidative stress and DNA damage response and affect the direction of mesenchymal stem cells differentiation in human adipose—derived mesenchymal stem cells. Such a response may be one of the causes of obesity or osteoporosis. PMID:26273425

Human Adipose-Derived Mesenchymal Stem Cell-Secreted CXCL1 and CXCL8 Facilitate Breast Tumor Growth By Promoting Angiogenesis.

PubMed

Wang, Yuan; Liu, Junli; Jiang, Qingyuan; Deng, Jie; Xu, Fen; Chen, Xiaolei; Cheng, Fuyi; Zhang, Yujing; Yao, Yunqi; Xia, Zhemin; Xu, Xia; Su, Xiaolan; Huang, Meijuan; Dai, Lei; Yang, Yang; Zhang, Shuang; Yu, Dechao; Zhao, Robert Chunhua; Wei, Yuquan; Deng, Hongxin

2017-09-01

Autologous adipose tissue or adipose tissue with additive adipose-derived mesenchymal stem cells (ADSCs) is used in the breast reconstruction of breast cancer patients who undergo mastectomy. ADSCs play an important role in the angiogenesis and adipogenesis, which make it much better than other materials. However, ADSCs may promote residual tumor cells to proliferate or metastasize, and the mechanism is still not fully understood. In this study, we demonstrated that human ADSCs (hADSCs) could facilitate tumor cells growth after co-injection with MCF7 and ZR-75-30 breast cancer cells (BCCs) by promoting angiogenesis, but hADSCs showed limited effect on the growth of MDA-MB-231 BCCs. Intriguingly, compared with ZR-75-30 tumor cells, MCF7 tumor cells were more potentially promoted by hADSCs in the aspects of angiogenesis and proliferation. Consistent with this, cytokine and angiogenesis array analyses showed that after co-injection with hADSCs, the CXCL1 and CXCL8 concentration were significantly increased in MCF7 tumor, but only moderately increased in ZR-75-30 tumor and did not increase in MDA-MB-231 tumor. Furthermore, we found that CXCL1/8 were mainly derived from hADSCs and could increase the migration and tube formation of human umbilical vein endothelial cells (HUVECs) by signaling via their receptors CXCR1 and CXCR2. A CXCR1/2-specific antagonist (SCH527123) attenuated the angiogenesis and tumor growth in vivo. Our findings suggest that CXCL1/8 secreted by hADSCs could promote breast cancer angiogenesis and therefore provide better understanding of safety concerns regarding the clinical application of hADSCs and suggestion in further novel therapeutic options. Stem Cells 2017;35:2060-2070. © 2017 AlphaMed Press.

Bone morphogenetic protein 9 (BMP9) induces effective bone formation from reversibly immortalized multipotent adipose-derived (iMAD) mesenchymal stem cells.

PubMed

Lu, Shun; Wang, Jing; Ye, Jixing; Zou, Yulong; Zhu, Yunxiao; Wei, Qiang; Wang, Xin; Tang, Shengli; Liu, Hao; Fan, Jiaming; Zhang, Fugui; Farina, Evan M; Mohammed, Maryam M; Song, Dongzhe; Liao, Junyi; Huang, Jiayi; Guo, Dan; Lu, Minpeng; Liu, Feng; Liu, Jianxiang; Li, Li; Ma, Chao; Hu, Xue; Lee, Michael J; Reid, Russell R; Ameer, Guillermo A; Zhou, Dongsheng; He, Tongchuan

2016-01-01

Regenerative medicine and bone tissue engineering using mesenchymal stem cells (MSCs) hold great promise as an effective approach to bone and skeletal reconstruction. While adipose tissue harbors MSC-like progenitors, or multipotent adipose-derived cells (MADs), it is important to identify and characterize potential biological factors that can effectively induce osteogenic differentiation of MADs. To overcome the time-consuming and technically challenging process of isolating and culturing primary MADs, here we establish and characterize the reversibly immortalized mouse multipotent adipose-derived cells (iMADs). The isolated mouse primary inguinal MAD cells are reversibly immortalized via the retrovirus-mediated expression of SV40 T antigen flanked with FRT sites. The iMADs are shown to express most common MSC markers. FLP-mediated removal of SV40 T antigen effectively reduces the proliferative activity and cell survival of iMADs, indicating the immortalization is reversible. Using the highly osteogenic BMP9, we find that the iMADs are highly responsive to BMP9 stimulation, express multiple lineage regulators, and undergo osteogenic differentiation in vitro upon BMP9 stimulation. Furthermore, we demonstrate that BMP9-stimulated iMADs form robust ectopic bone with a thermoresponsive biodegradable scaffold material. Collectively, our results demonstrate that the reversibly immortalized iMADs exhibit the characteristics of multipotent MSCs and are highly responsive to BMP9-induced osteogenic differentiation. Thus, the iMADs should provide a valuable resource for the study of MAD biology, which would ultimately enable us to develop novel and efficacious strategies for MAD-based bone tissue engineering.

Immunophenotypical characterization of canine mesenchymal stem cells from perivisceral and subcutaneous adipose tissue by a species-specific panel of antibodies.

PubMed

Ivanovska, Ana; Grolli, Stefano; Borghetti, Paolo; Ravanetti, Francesca; Conti, Virna; De Angelis, Elena; Macchi, Francesca; Ramoni, Roberto; Martelli, Paolo; Gazza, Ferdinando; Cacchioli, Antonio

2017-10-01

Immunophenotypical characterization of mesenchymal stem cells is fundamental for the design and execution of sound experimental and clinical studies. The scarce availability of species-specific antibodies for canine antigens has hampered the immunophenotypical characterization of canine mesenchymal stem cells (MSC). The aim of this study was to select a panel of species-specific direct antibodies readily useful for canine mesenchymal stem cells characterization. They were isolated from perivisceral and subcutaneous adipose tissue samples collected during regular surgeries from 8 dogs. Single color flow cytometric analysis of mesenchymal stem cells (P3) deriving from subcutaneous and perivisceral adipose tissue with a panel of 7 direct anti-canine antibodies revealed two largely homogenous cell populations with a similar pattern: CD29 + , CD44 + , CD73 + , CD90 + , CD34 - , CD45 - and MHC-II - with no statistically significant differences among them. Antibody reactivity was demonstrated on canine peripheral blood mononuclear cells. The similarities are reinforced by their in vitro cell morphology, trilineage differentiation ability and RT-PCR analysis (CD90 + , CD73 + , CD105 + , CD44 + , CD13 + , CD29 + , Oct-4 + gene and CD31 - and CD45 - expression). Our results report for the first time a comparison between the immunophenotypic profile of canine MSC deriving from perivisceral and subcutaneous adipose tissue. The substantial equivalence between the two populations has practical implication on clinical applications, giving the opportunity to choose the source depending on the patient needs. The results contribute to routine characterization of MSC populations grown in vitro, a mandatory process for the definition of solid and reproducible laboratory and therapeutic procedures. Copyright © 2017 Elsevier Ltd. All rights reserved.

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. �

Extracellular Vesicles from Adipose-Derived Mesenchymal Stem Cells Downregulate Senescence Features in Osteoarthritic Osteoblasts

PubMed Central

Tofiño-Vian, Miguel; Pérez del Caz, María Dolores; Castejón, Miguel Angel

2017-01-01

Osteoarthritis (OA) affects all articular tissues leading to pain and disability. The dysregulation of bone metabolism may contribute to the progression of this condition. Adipose-derived mesenchymal stem cells (ASC) are attractive candidates in the search of novel strategies for OA treatment and exert anti-inflammatory and cytoprotective effects on cartilage. Chronic inflammation in OA is a relevant factor in the development of cellular senescence and joint degradation. In this study, we extend our previous observations of ASC paracrine effects to study the influence of conditioned medium and extracellular vesicles from ASC on senescence induced by inflammatory stress in OA osteoblasts. Our results in cells stimulated with interleukin- (IL-) 1β indicate that conditioned medium, microvesicles, and exosomes from ASC downregulate senescence-associated β-galactosidase activity and the accumulation of γH2AX foci. In addition, they reduced the production of inflammatory mediators, with the highest effect on IL-6 and prostaglandin E2. The control of mitochondrial membrane alterations and oxidative stress may provide a mechanism for the protective effects of ASC in OA osteoblasts. We have also shown that microvesicles and exosomes mediate the paracrine effects of ASC. Our study suggests that correction of abnormal osteoblast metabolism by ASC products may contribute to their protective effects. PMID:29230269

Extracellular Vesicles from Adipose-Derived Mesenchymal Stem Cells Downregulate Senescence Features in Osteoarthritic Osteoblasts.

PubMed

Tofiño-Vian, Miguel; Guillén, Maria Isabel; Pérez Del Caz, María Dolores; Castejón, Miguel Angel; Alcaraz, Maria José

2017-01-01

Osteoarthritis (OA) affects all articular tissues leading to pain and disability. The dysregulation of bone metabolism may contribute to the progression of this condition. Adipose-derived mesenchymal stem cells (ASC) are attractive candidates in the search of novel strategies for OA treatment and exert anti-inflammatory and cytoprotective effects on cartilage. Chronic inflammation in OA is a relevant factor in the development of cellular senescence and joint degradation. In this study, we extend our previous observations of ASC paracrine effects to study the influence of conditioned medium and extracellular vesicles from ASC on senescence induced by inflammatory stress in OA osteoblasts. Our results in cells stimulated with interleukin- (IL-) 1 β indicate that conditioned medium, microvesicles, and exosomes from ASC downregulate senescence-associated β -galactosidase activity and the accumulation of γ H2AX foci. In addition, they reduced the production of inflammatory mediators, with the highest effect on IL-6 and prostaglandin E 2 . The control of mitochondrial membrane alterations and oxidative stress may provide a mechanism for the protective effects of ASC in OA osteoblasts. We have also shown that microvesicles and exosomes mediate the paracrine effects of ASC. Our study suggests that correction of abnormal osteoblast metabolism by ASC products may contribute to their protective effects.

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

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.

Lysophosphatidic acid-induced ADAM12 expression mediates human adipose tissue-derived mesenchymal stem cell-stimulated tumor growth.

PubMed

Do, Eun Kyoung; Kim, Young Mi; Heo, Soon Chul; Kwon, Yang Woo; Shin, Sang Hun; Suh, Dong-Soo; Kim, Ki-Hyung; Yoon, Man-Soo; Kim, Jae Ho

2012-11-01

Lysophosphatidic acid (LPA) is involved in mesenchymal stem cell-stimulated tumor growth in vivo. However, the molecular mechanism by which mesenchymal stem cells promote tumorigenesis remains elusive. In the present study, we demonstrate that conditioned medium from A549 human lung adenocarcinoma cells (A549 CM) induced the expression of ADAM12, a disintegrin and metalloproteases family member, in human adipose tissue-derived mesenchymal stem cells (hASCs). A549 CM-stimulated ADAM12 expression was abrogated by pretreatment of hASCs with the LPA receptor 1 inhibitor Ki16425 or by small interfering RNA-mediated silencing of LPA receptor 1, suggesting a key role for the LPA-LPA receptor 1 signaling axis in A549 CM-stimulated ADAM12 expression. Silencing of ADAM12 expression using small interfering RNA or short hairpin RNA abrogated LPA-induced expression of both α-smooth muscle actin, a marker of carcinoma-associated fibroblasts, and ADAM12 in hASCs. Using a xenograft transplantation model of A549 cells, we demonstrated that silencing of ADAM12 inhibited the hASC-stimulated in vivo growth of A549 xenograft tumors and the differentiation of transplanted hASCs to α-smooth muscle actin-positive carcinoma-associated fibroblasts. LPA-conditioned medium from hASCs induced the adhesion of A549 cells and silencing of ADAM12 inhibited LPA-induced expression of extracellular matrix proteins, periostin and βig-h3, in hASCs and LPA-conditioned medium-stimulated adhesion of A549 cells. These results suggest a pivotal role for LPA-stimulated ADAM12 expression in tumor growth and the differentiation of hASCs to carcinoma-associated fibroblasts expressing α-smooth muscle actin, periostin, and βig-h3. Copyright © 2012 Elsevier Ltd. All rights reserved.

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.

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.

Engineering Adipose-like Tissue in vitro and in vivo Utilizing Human Bone Marrow and Adipose-derived Mesenchymal Stem Cells with Silk Fibroin 3D Scaffolds

PubMed Central

Mauney, Joshua R; Nguyen, Trang; Gillen, Kelly; Kirker-Head, Carl; Gimble, Jeffrey M.; Kaplan, David L.

2009-01-01

Biomaterials derived from silk fibrion prepared by aqueous (AB) and organic (HFIP) solvent based processes, along with collagen (COL) and poly-lactic acid (PLA) based scaffolds were studied in vitro and in vivo for their utility in adipose tissue engineering strategies. For in vitro studies, human bone marrow and adipose-derived mesenchymal stem cells (hMSCs and hASCs) were seeded on the various biomaterials and cultured for 21 days in the presence of adipogenic stimulants (AD) or maintained as noninduced controls. Alamar Blue analysis revealed each biomaterial supported initial attachment of hMSCs and hASCs to similar levels for all matrices except COL in which higher levels were observed. hASCs and hMSCs cultured on all biomaterials in the presence of AD showed significant upregulation of adipogenic mRNA transcript levels (LPL, GLUT4, FABP4, PPARγ, adipsin, ACS) to similar extents when compared to noninduced controls. Similarly Oil-Red O analysis of hASC or hMSC-seeded scaffolds displayed substantial amounts of lipid accumulating adipocytes following cultivation with AD. The data revealed AB and HFIP scaffolds supported similar extents of lipid accumulating cells while PLA and COL scaffolds qualitatively displayed lower and higher extents by comparison, respectively. Following a 4 week implantation period in a rat muscle pouch defect model, both AB and HFIP scaffolds supported in vivo adipogenesis either alone or seeded with hASCs or hMSCs as assessed by Oil-Red O analysis, however the presence of exogenous cell sources substantially increased the extent and frequency of adipogenesis observed. In contrast, COL and PLA scaffolds underwent rapid scaffold degradation and were irretrievable following the implantation period. The results suggest that macroporous 3D AB and HFIP silk fibroin scaffolds offer an important platform for cell-based adipose tissue engineering applications, and in particular, provide longer-term structural integrity to promote the maintenance

Long Noncoding RNAs: New Players in the Osteogenic Differentiation of Bone Marrow- and Adipose-Derived Mesenchymal Stem Cells.

PubMed

Yang, Qiaolin; Jia, Lingfei; Li, Xiaobei; Guo, Runzhi; Huang, Yiping; Zheng, Yunfei; Li, Weiran

2018-06-01

Mesenchymal stem cells (MSCs) are an important population of multipotent stem cells that differentiate into multiple lineages and display great potential in bone regeneration and repair. Although the role of protein-coding genes in the osteogenic differentiation of MSCs has been extensively studied, the functions of noncoding RNAs in the osteogenic differentiation of MSCs are unclear. The recent application of next-generation sequencing to MSC transcriptomes has revealed that long noncoding RNAs (lncRNAs) are associated with the osteogenic differentiation of MSCs. LncRNAs are a class of non-coding transcripts of more than 200 nucleotides in length. Noncoding RNAs are thought to play a key role in osteoblast differentiation through various regulatory mechanisms including chromatin modification, transcription factor binding, competent endogenous mechanism, and other post-transcriptional mechanisms. Here, we review the roles of lncRNAs in the osteogenic differentiation of bone marrow- and adipose-derived stem cells and provide a theoretical foundation for future research.

Human adipose-derived mesenchymal stem cells for osteoarthritis: a pilot study with long-term follow-up and repeated injections.

PubMed

Song, Yang; Du, Hui; Dai, Chengxiang; Zhang, Li; Li, Suke; Hunter, David J; Lu, Liangjing; Bao, Chunde

2018-04-01

This study aimed to evaluate the safety and therapeutic potential of autologous human adipose-derived mesenchymal stem cells (haMSCs) in patients with osteoarthritis. Safety and efficacy of haMSCs were preclinically assessed in vitro and in BALB/c-nu nude mice. 18 patients were enrolled and divided into three dose groups: the low-dose, mid-dose and high-dose group (1 × 10 7 , 2 × 10 7 and 5 × 10 7 cells, respectively), provided three injections and followed up for 96 weeks. The preclinical study established the safety and efficacy of haMSCs. Intra-articular injections of haMSCs were safe and improved pain, function and cartilage volume of the knee joint, rendering them a promising novel treatment for knee osteoarthritis. The dosage of 5 × 10 7 haMSCs exhibited the highest improvement (ClinicalTrials.gov Identifier: NCT01809769).

Obesity Determines the Immunophenotypic Profile and Functional Characteristics of Human Mesenchymal Stem Cells From Adipose Tissue

PubMed Central

Pachón-Peña, Gisela; Serena, Carolina; Ejarque, Miriam; Petriz, Jordi; Duran, Xevi; Oliva-Olivera, W.; Simó, Rafael; Tinahones, Francisco J.

2016-01-01

Adipose tissue is a major source of mesenchymal stem cells (MSCs), which possess a variety of properties that make them ideal candidates for regenerative and immunomodulatory therapies. Here, we compared the immunophenotypic profile of human adipose-derived stem cells (hASCs) from lean and obese individuals, and explored its relationship with the apparent altered plasticity of hASCs. We also hypothesized that persistent hypoxia treatment of cultured hASCs may be necessary but not sufficient to drive significant changes in mature adipocytes. hASCs were obtained from subcutaneous adipose tissue of healthy, adult, female donors undergoing abdominal plastic surgery: lean (n = 8; body mass index [BMI]: 23 ± 1 kg/m2) and obese (n = 8; BMI: 35 ± 5 kg/m2). Cell surface marker expression, proliferation and migration capacity, and adipogenic differentiation potential of cultured hASCs at two different oxygen conditions were studied. Compared with lean-derived hASCs, obese-derived hASCs demonstrated increased proliferation and migration capacity but decreased lipid droplet accumulation, correlating with a higher expression of human leukocyte antigen (HLA)-II and cluster of differentiation (CD) 106 and lower expression of CD29. Of interest, adipogenic differentiation modified CD106, CD49b, HLA-ABC surface protein expression, which was dependent on the donor’s BMI. Additionally, low oxygen tension increased proliferation and migration of lean but not obese hASCs, which correlated with an altered CD36 and CD49b immunophenotypic profile. In summary, the differences observed in proliferation, migration, and differentiation capacity in obese hASCs occurred in parallel with changes in cell surface markers, both under basal conditions and during differentiation. Therefore, obesity is an important determinant of stem cell function independent of oxygen tension. Significance The obesity-related hypoxic environment may have latent effects on human adipose tissue-derived mesenchymal

Zinc Promotes Adipose-Derived Mesenchymal Stem Cell Proliferation and Differentiation towards a Neuronal Fate.

PubMed

Moon, Mi-Young; Kim, Hyun Jung; Choi, Bo Young; Sohn, Min; Chung, Tae Nyoung; Suh, Sang Won

2018-01-01

Zinc is an essential element required for cell division, migration, and proliferation. Under zinc-deficient conditions, proliferation and differentiation of neural progenitors are significantly impaired. Adipose-derived mesenchymal stem cells (AD-MSCs) are multipotent stem cells that can differentiate into neurons. The aim of this study was to evaluate the effect of zinc on AD-MSC proliferation and differentiation. We initially examined the effect of zinc on stem cell proliferation at the undifferentiated stage. AD-MSCs showed high proliferation rates on day 6 in 30  μ M and 100  μ M of ZnCl 2 . Zinc chelation inhibited AD-MSC proliferation via downregulation of ERK1/2 activity. We then assessed whether zinc was involved in cell migration and neurite outgrowth during differentiation. After three days of neuronal differentiation, TUJ-1-positive cells were observed, implying that AD-MSCs had differentiated into early neuron or neuron-like cells. Neurite outgrowth was increased in the zinc-treated group, while the CaEDTA-treated group showed diminished, shrunken neurites. Furthermore, we showed that zinc promoted neurite outgrowth via the inactivation of RhoA and led to the induction of neuronal gene expression (MAP2 and nestin) in differentiated stem cells. Taken together, zinc promoted AD-MSC proliferation and affected neuronal differentiation, mainly by increasing neurite outgrowth.

Zinc Promotes Adipose-Derived Mesenchymal Stem Cell Proliferation and Differentiation towards a Neuronal Fate

PubMed Central

Moon, Mi-Young; Kim, Hyun Jung; Choi, Bo Young; Sohn, Min

2018-01-01

Zinc is an essential element required for cell division, migration, and proliferation. Under zinc-deficient conditions, proliferation and differentiation of neural progenitors are significantly impaired. Adipose-derived mesenchymal stem cells (AD-MSCs) are multipotent stem cells that can differentiate into neurons. The aim of this study was to evaluate the effect of zinc on AD-MSC proliferation and differentiation. We initially examined the effect of zinc on stem cell proliferation at the undifferentiated stage. AD-MSCs showed high proliferation rates on day 6 in 30 μM and 100 μM of ZnCl2. Zinc chelation inhibited AD-MSC proliferation via downregulation of ERK1/2 activity. We then assessed whether zinc was involved in cell migration and neurite outgrowth during differentiation. After three days of neuronal differentiation, TUJ-1-positive cells were observed, implying that AD-MSCs had differentiated into early neuron or neuron-like cells. Neurite outgrowth was increased in the zinc-treated group, while the CaEDTA-treated group showed diminished, shrunken neurites. Furthermore, we showed that zinc promoted neurite outgrowth via the inactivation of RhoA and led to the induction of neuronal gene expression (MAP2 and nestin) in differentiated stem cells. Taken together, zinc promoted AD-MSC proliferation and affected neuronal differentiation, mainly by increasing neurite outgrowth. PMID:29765417

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.

Intermittent hydrostatic pressure enhances growth factor-induced chondroinduction of human adipose-derived mesenchymal stem cells.

PubMed

Safshekan, Farzaneh; Tafazzoli-Shadpour, Mohammad; Shokrgozar, Mohammad Ali; Haghighipour, Nooshin; Mahdian, Reza; Hemmati, Alireza

2012-12-01

Hydrostatic pressure (HP) plays an essential role in regulating function of chondrocytes and chondrogenic differentiation. The objective of this study was to examine effects of intermittent HP on chondrogenic differentiation of human adipose-derived mesenchymal stem cells (hASCs) in the presence or absence of chemical chondrogenic medium. Cells were isolated from abdominal fat tissue and confirmed for expression of ASC surface proteins and differentiation potential. Passage 3 pellets were treated with chemical (growth factor), mechanical (HP of 5 MPa and 0.5 Hz with duration of 4 h/day for 7 consecutive days), and combined chemical-mechanical stimuli. Using real-time polymerase chain reaction, the expression of Sox9, collagen II, and aggrecan as three major chondrogenic markers were quantified among three experimental groups and compared to those of stem cells and human cartilage tissue. In comparison to the chemical and mechanical groups, the chemical-mechanical group showed the highest expression for all three chondrogenic genes close to that of cartilage tissue. Results show the beneficial role of intermittent HP on chondrogenic differentiation of hASCs, and that this loading regime in combination with chondrogenic medium can be used in cartilage tissue engineering. © 2012, Copyright the Authors. Artificial Organs © 2012, International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.

Functional Roles of Pattern Recognition Receptors That Recognize Virus Nucleic Acids in Human Adipose-Derived Mesenchymal Stem Cells

PubMed Central

Wang, Fangchao; Yang, Can; Liu, Guoyan; Song, Xiangfeng

2016-01-01

Human adipose-derived mesenchymal stem cells (hAD-MSCs) are mesenchymal stem cells with the capability to modulate immune responses. Evidence showing that hAD-MSCs could mediate innate immune responses through pattern recognition receptors (PRRs) is increasing. However, the roles of PRRs in regulating the innate sensing of virus nucleic acids (RNA and DNA) in hAD-MSCs have not yet been investigated. This study focused on the abundant expression of PRRs, including Toll-like receptor 3 (TLR3) and retinoic acid-inducible gene I (RIG-I), which recognize viral RNA, and gamma-interferon inducible protein 16 (IFI16), which recognizes viral DNA in hAD-MSCs. Poly(I:C), a synthetic dsRNA analogy, activated TLR3 and RIG-I and induced the expression of type I interferons (IFN-α/β) and antivirus proteins, including IFN-stimulating gene 15, 2′5′-oligoadenylate synthetase, and Mx GTPase 1 in hAD-MSCs, which were attenuated by the knockdown of each TLR3 or RIG-I. Synthetic herpes simplex viral DNA (HSV60) activated IFI16 and induced the expression of IFN-α/β and antivirus proteins in hAD-MSCs, which were inhibited by the knockdown of IFI16. Both poly(I:C) and HSV60 induced the expression of IFN-α/β through the phosphorylation of IFN-regulatory factor 3. All these results indicated that PRRs recognizing virus nucleic acids were expressed and can mediate antivirus responses in hAD-MSCs. PMID:28105439

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.

Human Adipose-derived Mesenchymal Stem Cells Attenuate Early Stage of Bleomycin Induced Pulmonary Fibrosis: Comparison with Pirfenidone

PubMed Central

Reddy, Manoj; Fonseca, Lyle; Gowda, Shashank; Chougule, Basavraj; Hari, Aarya; Totey, Satish

2016-01-01

Background and Objectives Idiopathic pulmonary fibrosis (IPF) is a progressive, irreversible, invariably fatal fibrotic lung disease with no lasting option for therapy. Mesenchymal stem cells (MSCs) could be a promising modality for the treatment of IPF. Aim of the study was to investigate improvement in survivability and anti-fibrotic efficacy of human adipose-derived mesenchymal stem cells (AD-MSCs) in comparison with pirfenidone in the bleomycin-induced pulmonary fibrosis model. Methods Human AD-MSCs were administered intravenously on day 3, 6 and 9 after an intra-tracheal challenge with bleomycin, whereas, pirfenidone was given orally in drinking water at the rate of 100 mg/kg body weight three times a day daily from day 3 onward. AD-MSCs were labelled with PKH-67 before administration to detect engraftment. Disease severity and improvement was assessed and compared between sham control and vehicle control groups using Kaplan-Meier survival analysis, biochemical and molecular analysis, histopathology and high resolution computed tomography (HRCT) parameters at the end of study. Results Results demonstrated that AD-MSCs significantly increase survivability; reduce organ weight and collagen deposition better than pirfenidone group. Histological analyses and HRCT of the lung revealed that AD-MSCs afforded protection against bleomycin induced fibrosis and protect architecture of the lung. Gene expression analysis revealed that AD-MSCs potently suppressed pro-fibrotic genes induced by bleomycin. More importantly, AD-MSCs were found to inhibit pro-inflammatory related transcripts. Conclusions Our results provided direct evidence that AD-MSC-mediated immunomodulation and anti-fibrotic effect in the lungs resulted in marked protection in pulmonary fibrosis, but at an early stage of disease. PMID:27871152

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.

Platelet-Rich Plasma Favors Proliferation of Canine Adipose-Derived Mesenchymal Stem Cells in Methacrylate-Endcapped Caprolactone Porous Scaffold Niches

PubMed Central

Rodríguez-Jiménez, Francisco Javier; Valdes-Sánchez, Teresa; Carrillo, José M.; Rubio, Mónica; Monleon-Prades, Manuel; García-Cruz, Dunia Mercedes; García, Montserrat; Cugat, Ramón; Moreno-Manzano, Victoria

2012-01-01

Osteoarticular pathologies very often require an implementation therapy to favor regeneration processes of bone, cartilage and/or tendons. Clinical approaches performed on osteoarticular complications in dogs constitute an ideal model for human clinical translational applications. The adipose-derived mesenchymal stem cells (ASCs) have already been used to accelerate and facilitate the regenerative process. ASCs can be maintained in vitro and they can be differentiated to osteocytes or chondrocytes offering a good tool for cell replacement therapies in human and veterinary medicine. Although ACSs can be easily obtained from adipose tissue, the amplification process is usually performed by a time consuming process of successive passages. In this work, we use canine ASCs obtained by using a Bioreactor device under GMP cell culture conditions that produces a minimum of 30 million cells within 2 weeks. This method provides a rapid and aseptic method for production of sufficient stem cells with potential further use in clinical applications. We show that plasma rich in growth factors (PRGF) treatment positively contributes to viability and proliferation of canine ASCs into caprolactone 2-(methacryloyloxy) ethyl ester (CLMA) scaffolds. This biomaterial does not need additional modifications for cASCs attachment and proliferation. Here we propose a framework based on a combination of approaches that may contribute to increase the therapeutical capability of stem cells by the use of PRGF and compatible biomaterials for bone and connective tissue regeneration. PMID:24955632

The Role of Magnesium Ion Substituted Biphasic Calcium Phosphate Spherical Micro-Scaffolds in Osteogenic Differentiation of Human Adipose Tissue-Derived Mesenchymal Stem Cells.

PubMed

Kim, Dong-Hyun; Shin, Keun-Koo; Jung, Jin Sup; Chun, Ho Hwan; Park, Seong Soo; Lee, Jong Kook; Park, Hong-Chae; Yoon, Seog-Young

2015-08-01

This study was investigated the role of magnesium (Mg2+) ion substituted biphasic calcium phosphate (Mg-BCP) spherical micro-scaffolds in osteogenic differentiation of human adipose tissue-derived mesenchymal stem cells (hAT-MSCs). Mg-BCP micro-scaffolds with spherical morphology were successfully prepared using in situ co-precipitation and spray drying atomization process. The in vitro cell proliferation and differentiation of hAT-MSCs were determined up to day 14. After in vitro biological tests, Mg-BCP micro-scaffolds with hAT-MSCs showed more enhanced osteogenicity than pure hAT-MSCs as control group by unique biodegradation of TCP phase and influence of substituted Mg2+ ion in biphasic nanostructure. Therefore, these results suggest that Mg-BCP micro-scaffolds promote osteogenic differentiation of hAT-MSCs.

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.

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

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.

Characterization of glycol chitosan grafted with low molecular weight polyethylenimine as a gene carrier for human adipose-derived mesenchymal stem cells.

PubMed

Bae, Yoonhee; Lee, Young Hwa; Lee, Sunray; Han, Jin; Ko, Kyung Soo; Choi, Joon Sig

2016-11-20

Mesenchymal stem cells (MSCs) have a great capacity for self-renewal while still maintaining their multipotency, and can differentiate into a variety of cell types. The delivery of genes to a site of injury is a current and interesting field of gene therapy. In the present study, we describe a nonviral gene delivery carrier, glycol chitosan-methyl acrylate-polyethylenimine (GMP) polymer targeted towards human adipose-derived mesenchymal stem cells (AD-MSCs). Transfection efficiency, using luciferase (Luc) and a pDNA encoding enhanced green fluorescent protein (EGFP), along with cytotoxicity assays, were performed in human AD-MSCs. The results show that the transfection efficiency of the GMP polymer was similar to that of PEI25kD, and the cytotoxicity was lower. Moreover, human AD-MSCs were treated with the GMP polymer/pDNA polyplex and its cellular uptake and distribution were analyzed by flow cytometry and confocal microscopy. Furthermore, we performed endosomal escape analysis using LysoTracker Red, and found that the conjugated GMP polymer could escape from the endosome to the cytosol. Human AD-MSCs treated with the GMP polymer maintained their potential for osteogenic differentiation and phenotypic expression of human AD-MSCs based on flow cytometry analysis. The present study demonstrates that the GMP polymer can be used as a potential targeted-delivery carrier for effective gene delivery. Copyright © 2016 Elsevier Ltd. All rights reserved.

Lower Oncogenic Potential of Human Mesenchymal Stem Cells Derived from Cord Blood Compared to Induced Pluripotent Stem Cells

PubMed Central

Foroutan, T.; Najmi, M.; Kazemi, N.; Hasanlou, M.; Pedram, A.

2015-01-01

Background: In regenerative medicine, use of each of the mesenchymal stem cells derived from bone marrow, cord blood, and adipose tissue, has several cons and pros. Mesenchymal stem cells derived from cord blood have been considered the best source for precursor transplantation. Direct reprogramming of a somatic cell into induced pluripotent stem cells by over-expression of 6 transcription factors Oct4, Sox2, Klf4, lin28, Nanog, and c-Myc has great potential for regenerative medicine, eliminating the ethical issues of embryonic stem cells and the rejection problems of using non-autologous cells. Objective: To compare reprogramming and pluripotent markers OCT4, Sox-2, c-Myc, Klf4, Nanog, and lin28 in mesenchymal stem cells derived from cord blood and induced pluripotent stem cells. Methods: We analyzed the expression level of OCT4, Sox-2, c-Myc, Klf4, Nanog and lin28 genes in human mesenchymal stem cells derived from cord blood and induced pluripotent stem cells by cell culture and RT-PCR. Results: The expression level of pluripotent genes OCT4 and Sox-2, Nanog and lin28 in mesenchymal stem cells derived from cord blood were significantly higher than those in induced pluripotent stem cells. In contrast to OCT-4A and Sox-2, Nanog and lin28, the expression level of oncogenic factors c-Myc and Klf4 were significantly higher in induced pluripotent stem cells than in mesenchymal stem cells derived from cord blood. Conclusion: It could be concluded that mesenchymal stem cells derived from human cord blood have lower oncogenic potential compared to induced pluripotent stem cells. PMID:26306155

Impact of adipose tissue or umbilical cord derived mesenchymal stem cells on the immunogenicity of human cord blood derived endothelial progenitor cells

PubMed Central

Tan, Kefang; Zheng, Ke; Li, Daiye; Lu, Haiyuan; Wang, Siqi; Sun, Xuan

2017-01-01

The application of autologous endothelial progenitor cell (EPC) transplantation is a promising approach in therapeutic cardiovascular diseases and ischemic diseases. In this study, we compared the immunogenicity of EPCs, adipose tissue (AD)-derived mesenchymal stem cells (MSCs) and umbilical cord (UC)-derived MSCs by flow cytometry and the mixed lymphocyte reaction. The impact of AD-MSCs and UC-MSCs on the immunogenicity of EPCs was analyzed by the mixed lymphocyte reaction and cytokine secretion in vitro and was further tested by allogenic peripheral blood mononuclear cell (PBMC) induced immuno-rejection on a cell/matrigel graft in an SCID mouse model. EPCs and AD-MSCs express higher levels of MHC class I than UC-MSCs. All three kinds of cells are negative for MHC class II. UC-MSCs also express lower levels of IFN-γ receptor mRNA when compared with EPCs and AD-MSCs. EPCs can stimulate higher rates of proliferation of lymphocytes than AD-MSCs and UC-MSCs. Furthermore, AD-MSCs and UC-MSCs can modulate immune response and inhibit lymphocyte proliferation induced by EPCs, mainly through inhibition of the proliferation of CD8+ T cells. Compared with UC-MSCs, AD-MSCs can significantly improve vessel formation and maintain the integrity of neovascular structure in an EPC+MSC/matrigel graft in SCID mice, especially under allo-PBMC induced immuno-rejection. In conclusion, our study shows that AD-MSC is a powerful candidate to minimize immunological rejection and improve vessel formation in EPC transplantation treatment. PMID:28562647

Generation of Two Biological Wound Dressings as a Potential Delivery System of Human Adipose-Derived Mesenchymal Stem Cells

PubMed Central

Brena-Molina, Ana; Martínez-López, Valentín; Melgarejo-Ramírez, Yaaziel; Tamay de Dios, Lenin; Gómez-García, Ricardo; Reyes-Frías, Ma. de Lourdes; Rodríguez-Rodríguez, Lourdes; Garciadiego-Cázares, David; Lugo-Martínez, Haydée; Ibarra, Clemente

2015-01-01

Human adipose-derived mesenchymal stem cells (hADMSCs) are believed to be potential key factors for starting the regenerative process after tissue injury. However, an efficient method of delivering these regenerative cells to an external wound site is still lacking. Human amnion and pig skin have long been used as skin wound dressings for the treatment of burns and other skin lesions. Herein, we present the generation of two constructs using these two biomaterials as effective scaffolds for the culture of hADMSCs. It was found that hADMSCs seeded onto radiosterilized human amnion and pig skin are viable and proliferate. These cells are able to migrate over these scaffolds as demonstrated by using time-lapse microscopy. In addition, the scaffolds induce hADMSCs to secrete interleukin-10, an important negative regulator of inflammation, and interleukin-1β, a proinflammatory protein. The interplay between these two proteins has been proven to be vital for a balanced restoration of all necessary tissues. Thus, radiosterilized human amnion and pig skin are likely suitable scaffolds for delivery of hADMSCs transplants that could promote tissue regeneration in skin injuries like patients with burn injuries. PMID:26418201

[Proliferation and osteogenic differentiation of mesenchymal stem cells in hydrogels of human blood plasma].

PubMed

Linero, Itali M; Doncel, Adriana; Chaparro, Orlando

2014-01-01

The use of mesenchymal stem cells in clinical practice has increased considerably in the last decade because they play a supporting role in the processes of tissue repair and regeneration, becoming the main tool of cell therapy for the treatment of diseases functionally affecting bone and cartilage tissue . To evaluate in vitro the proliferative and osteogenic differentiation ability of mesenchymal stem cells derived from human adipose tissue in a blood plasma hydrogel. Mesenchymal stem cells were obtained from human adipose tissue explants and characterized by flow cytometry. Their multipotentiality was demonstrated by their ability to differentiate to adipogenic and osteogenic lineages. Cell proliferation and osteogenic differentiation ability of the cells cultured in blood plasma hydrogels were also evaluated. Mesenchymal stem cells derived from human adipose tissue growing in human blood plasma hydrogels showed a pattern of proliferation similar to that of the cells cultured in monolayer and also maintained their ability to differentiate to osteogenic lineage. Human blood plasma hydrogels are a suitable support for proliferation and osteogenic differentiation of mesenchymal stem cells derived from human adipose tissue and provides a substrate that is autologous, biocompatible, reabsorbable, easy to use, potentially injectable and economic, which could be used as a successful strategy for the management and clinical application of cell therapy in regenerative medicine.

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.

The Role of Cellular Proliferation in Adipogenic Differentiation of Human Adipose Tissue-Derived Mesenchymal Stem Cells.

PubMed

Marquez, Maribel P; Alencastro, Frances; Madrigal, Alma; Jimenez, Jossue Loya; Blanco, Giselle; Gureghian, Alex; Keagy, Laura; Lee, Cecilia; Liu, Robert; Tan, Lun; Deignan, Kristen; Armstrong, Brian; Zhao, Yuanxiang

2017-11-01

Mitotic clonal expansion has been suggested as a prerequisite for adipogenesis in murine preadipocytes, but the precise role of cell proliferation during human adipogenesis is unclear. Using adipose tissue-derived human mesenchymal stem cells as an in vitro cell model for adipogenic study, a group of cell cycle regulators, including Cdk1 and CCND1, were found to be downregulated as early as 24 h after adipogenic initiation and consistently, cell proliferation activity was restricted to the first 48 h of adipogenic induction. Cell proliferation was either further inhibited using siRNAs targeting cell cycle genes or enhanced by supplementing exogenous growth factor, basic fibroblast growth factor (bFGF), at specific time intervals during adipogenesis. Expression knockdown of Cdk1 at the initiation of adipogenic induction resulted in significantly increased adipocytes, even though total number of cells was significantly reduced compared to siControl-treated cells. bFGF stimulated proliferation throughout adipogenic differentiation, but exerted differential effect on adipogenic outcome at different phases, promoting adipogenesis during mitotic phase (first 48 h), but significantly inhibiting adipogenesis during adipogenic commitment phase (days 3-6). Our results demonstrate that cellular proliferation is counteractive to adipogenic commitment in human adipogenesis. However, cellular proliferation stimulation can be beneficial for adipogenesis during the mitotic phase by increasing the population of cells capable of committing to adipocytes before adipogenic commitment.

Improved viability and activity of neutrophils differentiated from HL-60 cells by co-culture with adipose tissue-derived mesenchymal stem cells.

PubMed

Park, Yoon Shin; Lim, Goh-Woon; Cho, Kyung-Ah; Woo, So-Youn; Shin, Meeyoung; Yoo, Eun-Sun; Chan Ra, Jeong; Ryu, Kyung-Ha

2012-06-22

Neutropenia is a principal complication of cancer treatment. We investigated the supportive effect of adipose tissue-derived mesenchymal stem cells (AD-MSCs) on the viability and function of neutrophils. Neutrophils were derived from HL-60 cells by dimethylformamide stimulation and cultured with or without AD-MSCs under serum-starved conditions to evaluate neutrophil survival, proliferation, and function. Serum starvation resulted in the apoptosis of neutrophils and decreased cell survival. The co-culture of neutrophils and AD-MSCs resulted in cell survival and inhibited neutrophil apoptosis under serum-starved conditions. The survival rate of neutrophils was prolonged up to 72 h, and the expression levels of interferon (IFN)-α, granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor, and transforming growth factor (TGF)-β in AD-MSCs were increased after co-culture with neutrophils. AD-MSCs promoted the viability of neutrophils by inhibiting apoptosis as well as enhancing respiratory burst, which could potentially be mediated by the increased expression of IFN-α, G-CSF, and TGF-β. Thus, we conclude that the use of AD-MSCs may be a promising cell-based therapy for increasing immunity by accelerating neutrophil function. Copyright © 2012 Elsevier Inc. All rights reserved.

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

The interaction of adipose-derived human mesenchymal stem cells and polyether ether ketone.

PubMed

Wang, Weiwei; Kratz, Karl; Behl, Marc; Yan, Wan; Liu, Yue; Xu, Xun; Baudis, Stefan; Li, Zhengdong; Kurtz, Andreas; Lendlein, Andreas; Ma, Nan

2015-01-01

Polyether ether ketone (PEEK) as a high-performance, thermoplastic implant material entered the field of medical applications due to its structural function and commercial availability. In bone tissue engineering, the combination of mesenchymal stem cells (MSCs) with PEEK implants may accelerate the bone formation and promote the osseointegration between the implant and the adjacent bone tissue. In this concept the question how PEEK influences the behaviour and functions of MSCs is of great interest. Here the cellular response of human adipose-derived MSCs to PEEK was evaluated and compared to tissue culture plate (TCP) as the reference material. Viability and morphology of cells were not altered when cultured on the PEEK film. The cells on PEEK presented a high proliferation activity in spite of a relatively lower initial cell adhesion rate. There was no significant difference on cell apoptosis and senescence between the cells on PEEK and TCP. The inflammatory cytokines and VEGF secreted by the cells on these two surfaces were at similar levels. The cells on PEEK showed up-regulated BMP2 and down-regulated BMP4 and BMP6 gene expression, whereas no conspicuous differences were observed in the committed osteoblast markers (BGLAP, COL1A1 and Runx2). With osteoinduction the cells on PEEK and TCP exhibited a similar osteogenic differentiation potential. Our results demonstrate the biofunctionality of PEEK for human MSC cultivation and differentiation. Its clinical benefits in bone tissue engineering may be achieved by combining MSCs with PEEK implants. These data may also provide useful information for further modification of PEEK with chemical or physical methods to regulate the cellular processes of MSCs and to consequently improve the efficacy of MSC-PEEK based therapies.

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

Adipose-derived mesenchymal stem cells reduce MMP-1 expression in UV-irradiated human dermal fibroblasts: therapeutic potential in skin wrinkling.

PubMed

Son, Woo-Chan; Yun, Jun-Won; Kim, Bae-Hwan

2015-01-01

Adipose-derived mesenchymal stem cells (AdMSCs) have been reported to have therapeutic benefit in skin. The aim of this study was to examine the effects of AdMSCs in UV-irradiated human dermal fibroblasts (HDFs) for therapeutic potential in skin wrinkling. UV irradiation, a model naturally mimic skin wrinkle formation, is known to increase matrix metalloproteinase-1 (MMP-1), making MMP-1 a target for skin photoaging. Our findings identified that AdMSCs reduce MMP-1 level in UV-irradiated HDFs and increase type 1 procollagen in HDFs. A dose-dependent increase in type 1 procollagen was confirmed by AdMSC-conditioned medium. Importantly, our current findings showing the effects of AdMSCs on the induction of MMP-1 in UV-radiated HDFs and the expression of collagen in HDFs can provide an evidence of relationship between MMP-1 and procollagen production for the protection against wrinkle formation. Collectively, AdMSCs may contribute to anti-wrinkle effects in skin but further experiments are needed to identify the mechanism.

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.

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.

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.

GAPDH, β-actin and β2-microglobulin, as three common reference genes, are not reliable for gene expression studies in equine adipose- and marrow-derived mesenchymal stem cells.

PubMed

Nazari, Fatemeh; Parham, Abbas; Maleki, Adham Fani

2015-01-01

Quantitative real time reverse transcription PCR (qRT-PCR) is one of the most important techniques for gene-expression analysis in molecular based studies. Selecting a proper internal control gene for normalizing data is a crucial step in gene expression analysis via this method. The expression levels of reference genes should be remained constant among cells in different tissues. However, it seems that the location of cells in different tissues might influence their expression. The purpose of this study was to determine whether the source of mesenchymal stem cells (MSCs) has any effect on expression level of three common reference genes (GAPDH, β-actin and β2-microglobulin) in equine marrow- and adipose- derived undifferentiated MSCs and consequently their reliability for comparative qRT-PCR. Adipose tissue (AT) and bone marrow (BM) samples were harvested from 3 mares. MSCs were isolated and cultured until passage 3 (P3). Total RNA of P3 cells was extracted for cDNA synthesis. The generated cDNAs were analyzed by quantitative real-time PCR. The PCR reactions were ended with a melting curve analysis to verify the specificity of amplicon. The expression levels of GAPDH were significantly different between AT- and BM- derived MSCs (p < 0.05). Differences in expression level of β-actin (P < 0.001) and B2M (P < 0.006.) between MSCs derived from AT and BM were substantially higher than GAPDH. In addition, the fold change in expression levels of GAPDH, β-actin and B2M in AT-derived MSCs compared to BM-derived MSCs were 2.38, 6.76 and 7.76, respectively. This study demonstrated that GAPDH and especially β-actin and B2M express in different levels in equine AT- and BM- derived MSCs. Thus they cannot be considered as reliable reference genes for comparative quantitative gene expression analysis in MSCs derived from equine bone marrow and adipose tissue.

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.

Expression of a functional epidermal growth factor receptor on human adipose-derived mesenchymal stem cells and its signaling mechanism.

PubMed

Baer, Patrick C; Schubert, Ralf; Bereiter-Hahn, Jürgen; Plösser, Michaela; Geiger, Helmut

2009-05-01

Adult stem cells act as a pluripotent source of regenerative cells during tissue injury. Despite expanded research in stem cell biology, understanding how growth and migration of adipose-derived adult mesenchymal stem cells (ASC) are governed by interactions with growth factors is very limited. One important property of ASC is the presence of the epidermal growth factor (EGF) receptor and the cellular response to soluble EGF. Expression of the EGF receptor was proven by PCR and Western blotting. Signal transduction was analyzed by Western blotting and PhosFlow assay. EGF caused robust phosphorylation of SHC and ERK1/2, which could be inhibited by EGF receptor antagonist AG1478 and MEK inhibitor PD98059. ASC proliferation was determined by MTT assay. Stem cell migration was analyzed in a modified Boyden chamber. Incubation with EGF led to cell proliferation and induced cell migration, but did not change the undifferentiated state of the cells. In the kidney, injured renal tubular cells express high amounts of EGF. Therefore, our results may highlight a mechanism underlying renal regeneration. Thus, future in vivo studies that focus on the effects of EGF on recruitment of ASC to sites of injury are necessary.

Exosomes From Adipose-derived Mesenchymal Stem Cells Protect the Myocardium Against Ischemia/Reperfusion Injury Through Wnt/β-Catenin Signaling Pathway.

PubMed

Cui, Xiaojun; He, Zhangyou; Liang, Zihao; Chen, Zhenyi; Wang, Haifeng; Zhang, Jiankai

2017-10-01

Mesenchymal stem cells (MSCs) and their secreted exosomes exert a cardioprotective role in jeopardized myocardium. However, the specific effects and underlying mechanisms of exosomes derived from adipose-derived MSCs (ADMSCs) on myocardial ischemia/reperfusion (I/R) injury remain largely unclear. In this study, ADMSC-derived exosomes (ADMSCs-ex) were administrated into the rats subjected to I/R injury and H9c2 cells exposed to hypoxia/reoxygenation (H/R). Consequently, administration of ADMSCs-ex significantly reduced I/R-induced myocardial infarction, accompanied with a decrease in serum levels of creatine kinase-myocardial band, lactate dehydrogenase, and cardiac troponin I (cTnI). Simultaneously, ADMSCs-ex dramatically antagonized I/R-induced myocardial apoptosis, along with the upregulation of Bcl-2 and downregulation of Bax, and inhibition of Caspase 3 activity in rat myocardium. Similarly, ADMSCs-ex significantly reduced cell apoptosis and the expression of Bax, but markedly increased cell viability and the expression of Bcl-2 and Cyclin D1 under H/R. Furthermore, ADMSCs-ex observably induced the activation of Wnt/β-catenin signaling by attenuating I/R- and H/R-induced inhibition of Wnt3a, p-GSK-3β (Ser9), and β-catenin expression. Importantly, treatment with Wnt/β-catenin inhibitor XAV939 partly neutralized ADMSC-ex-induced antiapoptotic and prosurvival effects in H9c2 cells. In conclusion, we confirmed that ADMSCs-ex protect ischemic myocardium from I/R injury through the activation of Wnt/β-catenin signaling pathway.

Adipose-derived mesenchymal stem cell-derived exosomes alleviate overwhelming systemic inflammatory reaction and organ damage and improve outcome in rat sepsis syndrome

PubMed Central

Chang, Chia-Lo; Sung, Pei-Hsun; Chen, Kuan-Hung; Shao, Pei-Lin; Yang, Chih-Chao; Cheng, Ben-Chung; Lin, Kun-Chen; Chen, Chih-Hung; Chai, Han-Tan; Chang, Hsueh-Wen; Yip, Hon-Kan; Chen, Hong-Hwa

2018-01-01

This study tested the hypothesis that healthy adipose-derived mesenchymal stem cell (ADMSC)-derived exosomes (HMSCEXO) and apoptotic (A) (induced by 12 h hypoxia/12 h starvation)-ADMSC-derived exosomes (AMSCEXO) were comparably effective at alleviating sepsis syndrome [SS; induced by cecal-ligation and puncture (CLP)]-induced systemic inflammation and reduced organ damage and unfavorable outcomes in rats. SD rats were divided into sham control (SC), SS only, SS + HMSCEXO (100 µg intravenous administration 3 h after CLP), and AMSCEXO. By day 5 after CLP procedure, the mortality rate was significantly higher in SS than in SC and HMSCEXO (all P < 0.01), but it showed no significant different between SC and HMSCEXO, between AMSCEXO and HMSCEXO or between SS and AMSCEXO (P > 0.05). The levels of inflammatory mediators in circulation (CD11b/c/Ly6G/MIF), bronchioalveolar lavage (CD11b/c/Ly6G) and abdominal ascites (CD11b/c/CD14/Ly6G/MIF) were highest in SS, lowest in SC and significantly higher in AMSCEXO than in HMSCEXO (all P < 0.001). The circulating/splenic levels of immune cells (CD34+/CD4+/CD3+/CD8+) were expressed in an identical pattern whereas the T-reg+ cells exhibited an opposite pattern of inflammation among the groups (all P < 0.001). The protein expressions of inflammation (MMP-9/MIF/TNF-α/NF-κB/IL-1β) and oxidative stress (NOX-1/NOX-2/oxidized protein), and cellular expressions (CD14+/CD68+) in lung/kidney parenchyma exhibited an identical pattern of inflammatory mediators (all P < 0.001). The kidney/lung injury scores displayed an identical pattern of inflammatory mediators among the groups (all P < 0.001). In conclusion, HMSCEXO might be superior to AMSCEXO for improving survival and suppressing the inflammatory reactions in rats after SS. PMID:29736200

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.

Priming Adipose-Derived Mesenchymal Stem Cells with Hyaluronan Alters Growth Kinetics and Increases Attachment to Articular Cartilage

PubMed Central

Succar, Peter; Medynskyj, Michael; Breen, Edmond J.; Batterham, Tony; Molloy, Mark P.; Herbert, Benjamin R.

2016-01-01

Background. Biological therapeutics such as adipose-derived mesenchymal stem cell (MSC) therapy are gaining acceptance for knee-osteoarthritis (OA) treatment. Reports of OA-patients show reductions in cartilage defects and regeneration of hyaline-like-cartilage with MSC-therapy. Suspending MSCs in hyaluronan commonly occurs in animals and humans, usually without supporting data. Objective. To elucidate the effects of different concentrations of hyaluronan on MSC growth kinetics. Methods. Using a range of hyaluronan concentrations, we measured MSC adherence and proliferation on culture plastic surfaces and a novel cartilage-adhesion assay. We employed time-course and dispersion imaging to assess MSC binding to cartilage. Cytokine profiling was also conducted on the MSC-secretome. Results. Hyaluronan had dose-dependent effects on growth kinetics of MSCs at concentrations of entanglement point (1 mg/mL). At higher concentrations, viscosity effects outweighed benefits of additional hyaluronan. The cartilage-adhesion assay highlighted for the first time that hyaluronan-primed MSCs increased cell attachment to cartilage whilst the presence of hyaluronan did not. Our time-course suggested patients undergoing MSC-therapy for OA could benefit from joint-immobilisation for up to 8 hours. Hyaluronan also greatly affected dispersion of MSCs on cartilage. Conclusion. Our results should be considered in future trials with MSC-therapy using hyaluronan as a vehicle, for the treatment of OA. PMID:26981136

RNA-Sequencing Gene Expression Profiling of Orbital Adipose-Derived Stem Cell Population Implicate HOX Genes and WNT Signaling Dysregulation in the Pathogenesis of Thyroid-Associated Orbitopathy.

PubMed

Tao, Wensi; Ayala-Haedo, Juan A; Field, Matthew G; Pelaez, Daniel; Wester, Sara T

2017-12-01

The purpose of this study was to characterize the intrinsic cellular properties of orbital adipose-derived stem cells (OASC) from patients with thyroid-associated orbitopathy (TAO) and healthy controls. Orbital adipose tissue was collected from a total of nine patients: four controls and five patients with TAO. Isolated OASC were characterized with mesenchymal stem cell-specific markers. Orbital adipose-derived stem cells were differentiated into three lineages: chondrocytes, osteocytes, and adipocytes. Reverse transcription PCR of genes involved in the adipogenesis, chondrogenesis, and osteogenesis pathways were selected to assay the differentiation capacities. RNA sequencing analysis (RNA-seq) was performed and results were compared to assess for differences in gene expression between TAO and controls. Selected top-ranked results were confirmed by RT-PCR. Orbital adipose-derived stem cells isolated from orbital fat expressed high levels of mesenchymal stem cell markers, but low levels of the pluripotent stem cell markers. Orbital adipose-derived stem cells isolated from TAO patients exhibited an increase in adipogenesis, and a decrease in chondrogenesis and osteogenesis. RNA-seq disclosed 54 differentially expressed genes. In TAO OASC, expression of early neural crest progenitor marker (WNT signaling, ZIC genes and MSX2) was lost. Meanwhile, ectopic expression of HOXB2 and HOXB3 was found in the OASC from TAO. Our results suggest that there are intrinsic genetic and cellular differences in the OASC populations derived from TAO patients. The upregulation in adipogenesis in OASC of TAO may be is consistent with the clinical phenotype. Downregulation of early neural crest markers and ectopic expression of HOXB2 and HOXB3 in TAO OASC demonstrate dysregulation of developmental and tissue patterning pathways.

RNA-Sequencing Gene Expression Profiling of Orbital Adipose-Derived Stem Cell Population Implicate HOX Genes and WNT Signaling Dysregulation in the Pathogenesis of Thyroid-Associated Orbitopathy

PubMed Central

Tao, Wensi; Ayala-Haedo, Juan A.; Field, Matthew G.; Pelaez, Daniel; Wester, Sara T.

2017-01-01

Purpose The purpose of this study was to characterize the intrinsic cellular properties of orbital adipose-derived stem cells (OASC) from patients with thyroid-associated orbitopathy (TAO) and healthy controls. Methods Orbital adipose tissue was collected from a total of nine patients: four controls and five patients with TAO. Isolated OASC were characterized with mesenchymal stem cell–specific markers. Orbital adipose-derived stem cells were differentiated into three lineages: chondrocytes, osteocytes, and adipocytes. Reverse transcription PCR of genes involved in the adipogenesis, chondrogenesis, and osteogenesis pathways were selected to assay the differentiation capacities. RNA sequencing analysis (RNA-seq) was performed and results were compared to assess for differences in gene expression between TAO and controls. Selected top-ranked results were confirmed by RT-PCR. Results Orbital adipose-derived stem cells isolated from orbital fat expressed high levels of mesenchymal stem cell markers, but low levels of the pluripotent stem cell markers. Orbital adipose-derived stem cells isolated from TAO patients exhibited an increase in adipogenesis, and a decrease in chondrogenesis and osteogenesis. RNA-seq disclosed 54 differentially expressed genes. In TAO OASC, expression of early neural crest progenitor marker (WNT signaling, ZIC genes and MSX2) was lost. Meanwhile, ectopic expression of HOXB2 and HOXB3 was found in the OASC from TAO. Conclusion Our results suggest that there are intrinsic genetic and cellular differences in the OASC populations derived from TAO patients. The upregulation in adipogenesis in OASC of TAO may be is consistent with the clinical phenotype. Downregulation of early neural crest markers and ectopic expression of HOXB2 and HOXB3 in TAO OASC demonstrate dysregulation of developmental and tissue patterning pathways. PMID:29214313

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

Chip-based comparison of the osteogenesis of human bone marrow- and adipose tissue-derived mesenchymal stem cells under mechanical stimulation.

PubMed

Park, Sang-Hyug; Sim, Woo Young; Min, Byoung-Hyun; Yang, Sang Sik; Khademhosseini, Ali; Kaplan, David L

2012-01-01

Adipose tissue-derived stem cells (ASCs) are considered as an attractive stem cell source for tissue engineering and regenerative medicine. We compared human bone marrow-derived mesenchymal stem cells (hMSCs) and hASCs under dynamic hydraulic compression to evaluate and compare osteogenic abilities. A novel micro cell chip integrated with microvalves and microscale cell culture chambers separated from an air-pressure chamber was developed using microfabrication technology. The microscale chip enables the culture of two types of stem cells concurrently, where each is loaded into cell culture chambers and dynamic compressive stimulation is applied to the cells uniformly. Dynamic hydraulic compression (1 Hz, 1 psi) increased the production of osteogenic matrix components (bone sialoprotein, oateopontin, type I collagen) and integrin (CD11b and CD31) expression from both stem cell sources. Alkaline phosphatase and Alrizarin red staining were evident in the stimulated hMSCs, while the stimulated hASCs did not show significant increases in staining under the same stimulation conditions. Upon application of mechanical stimulus to the two types of stem cells, integrin (β1) and osteogenic gene markers were upregulated from both cell types. In conclusion, stimulated hMSCs and hASCs showed increased osteogenic gene expression compared to non-stimulated groups. The hMSCs were more sensitive to mechanical stimulation and more effective towards osteogenic differentiation than the hASCs under these modes of mechanical stimulation.

Myogenic potential of mesenchymal stem cells isolated from porcine adipose tissue.

PubMed

Milner, Derek J; Bionaz, Massimo; Monaco, Elisa; Cameron, Jo Ann; Wheeler, Matthew B

2018-06-01

Advances in stem cell biology and materials science have provided a basis for developing tissue engineering methods to repair muscle injury. Among stem cell populations with potential to aid muscle repair, adipose-derived mesenchymal stem cells (ASC) hold great promise. To evaluate the possibility of using porcine ASC for muscle regeneration studies, we co-cultured porcine ASC with murine C 2 C 12 myoblasts. These experiments demonstrated that porcine ASC display significant myogenic potential. Co-culture of ASC expressing green fluorescent protein (GFP) with C 2 C 12 cells resulted in GFP + myotube formation, indicating fusion of ASC with myoblasts to form myotubes. The presence of porcine lamin A/C positive nuclei in myotubes and RTqPCR analysis of porcine myogenin and desmin expression confirmed that myotube nuclei derived from ASC contribute to muscle gene expression. Co-culturing GFP + ASC with porcine satellite cells demonstrated enhanced myogenic capability of ASC, as the percentage of labeled myotubes increased compared to mouse co-cultures. Enhancing myogenic potential of ASC through soluble factor treatment or expansion of ASC with innate myogenic capacity should allow for their therapeutic use to regenerate muscle tissue lost to disease or injury.

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

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

Effects of Ionizing Radiation on Human Adipose Derived Mesenchymal Stem Cells and their Differentiation towards the Osteoblastic Lineage

NASA Astrophysics Data System (ADS)

Konda, Bikash; Baumstark-Khan, Christa; Hellweg, Christine; Reitz, Guenther; Lau, Patrick

Radiation exposure and musculoskeletal disuse are among the major challenges during space missions. Astronauts face the problem to lose bone calcium due to uncoupling of bone formation and resorption. Bone forming osteoblasts can be derived from the undifferentiated mesenchymal stem cell compartment (MSC). In this study, the ability of human adipose tissue derived stem cells (ATSC) to differentiate into the osteoblastic lineage was examined after radiation exposure in presence of medium supplementation with osteogenic additives (ß-glycerophosphate, ascorbic acid and dexamethasone). The SAOS-2 cell line (human osteosarcoma cell line) was used as control for osteoblastic differentiation. Changes in cellular morphology, cell cycle progression, as well as cellular radiation sensitivity were characterized after ionizing radiation exposure with X-rays and heavy ions (Ti). Rapidly proliferating SAOS-2 cells are less radiation-sensitive than slowly proliferating ATSC cells after X-ray (CFA: dose effect curves show D0 values of 1 Gy and 0.75 Gy for SAOS-2 and ATSC, respectively) exposure. Heavy ion (Ti) exposure resulted in a greater extent of cells accumulating in the G2/M phase of the cell cycle in a dose-dependent manner when compared to X-ray exposure. Differentiation of cells towards the osteoblastic lineage was quantified by hydroxyapatite (HA) deposition using Lonza OsteoImageTM mineralization assay. The deposition of HA after X- and Ti-irradiation for highly proliferating SAOS-2 cells showed a dose-dependent time delay while slowly proliferating ATSC showed no effect from radiation exposure. More detailed investigation is required to reveal the radiation dependent mechanism of bone loss in astronauts.

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

Trophic effects of adipose-tissue-derived and bone-marrow-derived mesenchymal stem cells enhance cartilage generation by chondrocytes in co-culture

PubMed Central

Pleumeekers, M. M.; Nimeskern, L.; Koevoet, J. L. M.; Karperien, M.; Stok, K. S.; van Osch, G. J. V. M.

2018-01-01

Aims Combining mesenchymal stem cells (MSCs) and chondrocytes has great potential for cell-based cartilage repair. However, there is much debate regarding the mechanisms behind this concept. We aimed to clarify the mechanisms that lead to chondrogenesis (chondrocyte driven MSC-differentiation versus MSC driven chondroinduction) and whether their effect was dependent on MSC-origin. Therefore, chondrogenesis of human adipose-tissue-derived MSCs (hAMSCs) and bone-marrow-derived MSCs (hBMSCs) combined with bovine articular chondrocytes (bACs) was compared. Methods hAMSCs or hBMSCs were combined with bACs in alginate and cultured in vitro or implanted subcutaneously in mice. Cartilage formation was evaluated with biochemical, histological and biomechanical analyses. To further investigate the interactions between bACs and hMSCs, (1) co-culture, (2) pellet, (3) Transwell® and (4) conditioned media studies were conducted. Results The presence of hMSCs–either hAMSCs or hBMSCs—increased chondrogenesis in culture; deposition of GAG was most evidently enhanced in hBMSC/bACs. This effect was similar when hMSCs and bAC were combined in pellet culture, in alginate culture or when conditioned media of hMSCs were used on bAC. Species-specific gene-expression analyses demonstrated that aggrecan was expressed by bACs only, indicating a predominantly trophic role for hMSCs. Collagen-10-gene expression of bACs was not affected by hBMSCs, but slightly enhanced by hAMSCs. After in-vivo implantation, hAMSC/bACs and hBMSC/bACs had similar cartilage matrix production, both appeared stable and did not calcify. Conclusions This study demonstrates that replacing 80% of bACs by either hAMSCs or hBMSCs does not influence cartilage matrix production or stability. The remaining chondrocytes produce more matrix due to trophic factors produced by hMSCs. PMID:29489829

Trophic effects of adipose-tissue-derived and bone-marrow-derived mesenchymal stem cells enhance cartilage generation by chondrocytes in co-culture.

PubMed

Pleumeekers, M M; Nimeskern, L; Koevoet, J L M; Karperien, M; Stok, K S; van Osch, G J V M

2018-01-01

Combining mesenchymal stem cells (MSCs) and chondrocytes has great potential for cell-based cartilage repair. However, there is much debate regarding the mechanisms behind this concept. We aimed to clarify the mechanisms that lead to chondrogenesis (chondrocyte driven MSC-differentiation versus MSC driven chondroinduction) and whether their effect was dependent on MSC-origin. Therefore, chondrogenesis of human adipose-tissue-derived MSCs (hAMSCs) and bone-marrow-derived MSCs (hBMSCs) combined with bovine articular chondrocytes (bACs) was compared. hAMSCs or hBMSCs were combined with bACs in alginate and cultured in vitro or implanted subcutaneously in mice. Cartilage formation was evaluated with biochemical, histological and biomechanical analyses. To further investigate the interactions between bACs and hMSCs, (1) co-culture, (2) pellet, (3) Transwell® and (4) conditioned media studies were conducted. The presence of hMSCs-either hAMSCs or hBMSCs-increased chondrogenesis in culture; deposition of GAG was most evidently enhanced in hBMSC/bACs. This effect was similar when hMSCs and bAC were combined in pellet culture, in alginate culture or when conditioned media of hMSCs were used on bAC. Species-specific gene-expression analyses demonstrated that aggrecan was expressed by bACs only, indicating a predominantly trophic role for hMSCs. Collagen-10-gene expression of bACs was not affected by hBMSCs, but slightly enhanced by hAMSCs. After in-vivo implantation, hAMSC/bACs and hBMSC/bACs had similar cartilage matrix production, both appeared stable and did not calcify. This study demonstrates that replacing 80% of bACs by either hAMSCs or hBMSCs does not influence cartilage matrix production or stability. The remaining chondrocytes produce more matrix due to trophic factors produced by hMSCs.

Cell culture density affects the stemness gene expression of adipose tissue-derived mesenchymal stem cells.

PubMed

Kim, Dae Seong; Lee, Myoung Woo; Lee, Tae-Hee; Sung, Ki Woong; Koo, Hong Hoe; Yoo, Keon Hee

2017-03-01

The results of clinical trials using mesenchymal stem cells (MSCs) are controversial due to the heterogeneity of human MSCs and differences in culture conditions. In this regard, it is important to identify gene expression patterns according to culture conditions, and to determine how the cells are expanded and when they should be clinically used. In the current study, stemness gene expression was investigated in adipose tissue-derived MSCs (AT-MSCs) harvested following culture at different densities. AT-MSCs were plated at a density of 200 or 5,000 cells/cm 2 . After 7 days of culture, stemness gene expression was examined by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. The proliferation rate of AT-MSCs harvested at a low density (~50% confluent) was higher than that of AT-MSCs harvested at a high density (~90% confluent). Although there were differences in the expression levels of stemness gene, such as octamer-binding transcription factor 4, nanog homeobox ( Nanog ), SRY-box 2, Kruppel like factor 4, v-myc avian myelocytomatosis viral oncogene homolog ( c-Myc ), and lin-28 homolog A, in the AT-MSCs obtained from different donors, RT-qPCR analysis demonstrated differential gene expression patterns according to the cell culture density. Expression levels of stemness genes, particularly Nanog and c-Myc , were upregulated in AT-MSCs harvested at a low density (~50% confluent) in comparison to AT-MSCs from the same donor harvested at a high density (~90% confluent). These results imply that culture conditions, such as the cell density at harvesting, modulate the stemness gene expression and proliferation of MSCs.

Platelet-Rich Plasma and Adipose-Derived Mesenchymal Stem Cells for Regenerative Medicine-Associated Treatments in Bottlenose Dolphins (Tursiops truncatus)

PubMed Central

Griffeth, Richard J.; García-Párraga, Daniel; Mellado-López, Maravillas; Crespo-Picazo, Jose Luis; Soriano-Navarro, Mario; Martinez-Romero, Alicia; Moreno-Manzano, Victoria

2014-01-01

Dolphins exhibit an extraordinary capacity to heal deep soft tissue injuries. Nevertheless, accelerated wound healing in wild or captive dolphins would minimize infection and other side effects associated with open wounds in marine animals. Here, we propose the use of a biological-based therapy for wound healing in dolphins by the application of platelet-rich plasma (PRP). Blood samples were collected from 9 different dolphins and a specific and simple protocol which concentrates platelets greater than two times that of whole blood was developed. As opposed to a commonly employed human protocol for PRP preparation, a single centrifugation for 3 minutes at 900 rpm resulted in the best condition for the concentration of dolphin platelets. By FACS analysis, dolphin platelets showed reactivity to platelet cell-surface marker CD41. Analysis by electron microscopy revealed that dolphin platelets were larger in size than human platelets. These findings may explain the need to reduce the duration and speed of centrifugation of whole blood from dolphins to obtain a 2-fold increase and maintain proper morphology of the platelets. For the first time, levels of several growth factors from activated dolphin platelets were quantified. Compared to humans, concentrations of PDGF-BB were not different, while TGFβ and VEGF-A were significantly lower in dolphins. Additionally, adipose tissue was obtained from cadaveric dolphins found along the Spanish Mediterranean coast, and adipose-derived mesenchymal stem cells (ASCs) were successfully isolated, amplified, and characterized. When dolphin ASCs were treated with 2.5 or 5% dolphin PRP they exhibited significant increased proliferation and improved phagocytotic activity, indicating that in culture, PRP may improve the regenerative capacity of ASCs. Taken together, we show an effective and well-defined protocol for efficient PRP isolation. This protocol alone or in combination with ASCs, may constitute the basis of a biological

Platelet-rich plasma and adipose-derived mesenchymal stem cells for regenerative medicine-associated treatments in bottlenose dolphins (Tursiops truncatus).

PubMed

Griffeth, Richard J; García-Párraga, Daniel; Mellado-López, Maravillas; Crespo-Picazo, Jose Luis; Soriano-Navarro, Mario; Martinez-Romero, Alicia; Moreno-Manzano, Victoria

2014-01-01

Dolphins exhibit an extraordinary capacity to heal deep soft tissue injuries. Nevertheless, accelerated wound healing in wild or captive dolphins would minimize infection and other side effects associated with open wounds in marine animals. Here, we propose the use of a biological-based therapy for wound healing in dolphins by the application of platelet-rich plasma (PRP). Blood samples were collected from 9 different dolphins and a specific and simple protocol which concentrates platelets greater than two times that of whole blood was developed. As opposed to a commonly employed human protocol for PRP preparation, a single centrifugation for 3 minutes at 900 rpm resulted in the best condition for the concentration of dolphin platelets. By FACS analysis, dolphin platelets showed reactivity to platelet cell-surface marker CD41. Analysis by electron microscopy revealed that dolphin platelets were larger in size than human platelets. These findings may explain the need to reduce the duration and speed of centrifugation of whole blood from dolphins to obtain a 2-fold increase and maintain proper morphology of the platelets. For the first time, levels of several growth factors from activated dolphin platelets were quantified. Compared to humans, concentrations of PDGF-BB were not different, while TGFβ and VEGF-A were significantly lower in dolphins. Additionally, adipose tissue was obtained from cadaveric dolphins found along the Spanish Mediterranean coast, and adipose-derived mesenchymal stem cells (ASCs) were successfully isolated, amplified, and characterized. When dolphin ASCs were treated with 2.5 or 5% dolphin PRP they exhibited significant increased proliferation and improved phagocytotic activity, indicating that in culture, PRP may improve the regenerative capacity of ASCs. Taken together, we show an effective and well-defined protocol for efficient PRP isolation. This protocol alone or in combination with ASCs, may constitute the basis of a biological

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.

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

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.

Role of MyD88 in TLR agonist-induced functional alterations of human adipose tissue-derived mesenchymal stem cells.

PubMed

Yu, Sungsook; Cho, Hyun Hwa; Joo, Hye Joon; Bae, Yong Chan; Jung, Jin Sup

2008-10-01

Toll-like receptors (TLRs) sense microorganism components and are critical host mediators of inflammation during infection. Recently, TLRs have been reported to be involved in cell proliferation and differentiation. We previously reported that TLR agonists might affect proliferation and differentiation of human adipose tissue-derived mesenchymal stem cells (hASCs). In this study, we sought to determine whether TLR signaling is dependent on MyD88 in hASCs. The hASCs were downregulated using LV-GFP-miR-MyD88, a lentiviral construct inserted siRNA against human MyD88 that significantly inhibited cell proliferation. MyD88 downregulation reduced NF-kappaB activation and enhancement of osteogenic differentiation induced by peptidoglycan (PGN) more significantly than that induced by lipopolysaccharide (LPS). Although LPS- and PGN-induced cytokine secretions were decreased greatly by MyD88 downregulation, IFN-gamma-induced protein-10 (IP10) and IFNbeta expression were enhanced by LPS irrespective of the downregulation of MyD88. These results suggest that TLR signaling is mediated via MyD88-independent pathways as well as MyD88-dependent pathways in hASCs and that MyD88 contributes to the regulation of cell proliferation and differentiation in hASCs.

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

High-resolution molecular validation of self-renewal and spontaneous differentiation in adipose-tissue derived human mesenchymal stem cells cultured in human platelet lysate

PubMed Central

Dudakovic, Amel Dudakovic; Camilleri, Emily; Riester, Scott M.; Lewallen, Eric A.; Kvasha, Sergiy; Chen, Xiaoyue; Radel, Darcie J.; Anderson, Jarett M.; Nair, Asha A.; Evans, Jared M.; Krych, Aaron J.; Smith, Jay; Deyle, David R.; Stein, Janet L.; Stein, Gary S.; Im, Hee-Jeong; Cool, Simon M.; Westendorf, Jennifer J.; Kakar, Sanjeev; Dietz, Allan B.; van Wijnen, Andre J.

2014-01-01

Improving the effectiveness of adipose-tissue derived human mesenchymal stromal/stem cells (AMSCs) for skeletal therapies requires a detailed characterization of mechanisms supporting cell proliferation and multi-potency. We investigated the molecular phenotype of AMSCs that were either actively proliferating in platelet lysate or in a basal non-proliferative state. Flow cytometry combined with high-throughput RNA sequencing (RNASeq) and RT-qPCR analyses validate that AMSCs express classic mesenchymal cell surface markers (e.g., CD44, CD73/NT5E, CD90/THY1 and CD105/ENG). Expression of CD90 is selectively elevated at confluence. Self-renewing AMSCs express a standard cell cycle program that successively mediates DNA replication, chromatin packaging, cyto-architectural enlargement and mitotic division. Confluent AMSCs preferentially express genes involved in extracellular matrix (ECM) formation and cellular communication. For example, cell cycle-related biomarkers (e.g., cyclins E2 and B2, transcription factor E2F1) and histone-related genes (e.g., H4, HINFP, NPAT) are elevated in proliferating AMSCs, while ECM genes are strongly upregulated (>10 fold) in quiescent AMSCs. AMSCs also express pluripotency genes (e.g., POU5F1, NANOG, KLF4) and early mesenchymal markers (e.g., NES, ACTA2) consistent with their multipotent phenotype. Strikingly, AMSCs modulate expression of WNT signaling components and switch production of WNT ligands (from WNT5A/WNT5B/WNT7B to WNT2/WNT2B), while up-regulating WNT-related genes (WISP2, SFRP2 and SFRP4). Furthermore, post-proliferative AMSCs spontaneously express fibroblastic, osteogenic, chondrogenic and adipogenic biomarkers when maintained in confluent cultures. Our findings validate the biological properties of self-renewing and multi-potent AMSCs by providing high-resolution quality control data that support their clinical versatility. PMID:24905804

Ultrasound-Assisted Liposuction Does Not Compromise the Regenerative Potential of Adipose-Derived Stem Cells

PubMed Central

Duscher, Dominik; Atashroo, David; Maan, Zeshaan N.; Luan, Anna; Brett, Elizabeth A.; Barrera, Janos; Khong, Sacha M.; Zielins, Elizabeth R.; Whittam, Alexander J.; Hu, Michael S.; Walmsley, Graham G.; Pollhammer, Michael S.; Schmidt, Manfred; Schilling, Arndt F.; Machens, Hans-Günther; Huemer, Georg M.; Wan, Derrick C.; Longaker, Michael T.

2016-01-01

Human mesenchymal stem cells (MSCs) have recently become a focus of regenerative medicine, both for their multilineage differentiation capacity and their excretion of proregenerative cytokines. Adipose-derived mesenchymal stem cells (ASCs) are of particular interest because of their abundance in fat tissue and the ease of harvest via liposuction. However, little is known about the impact of different liposuction methods on the functionality of ASCs. Here we evaluate the regenerative abilities of ASCs harvested via a third-generation ultrasound-assisted liposuction (UAL) device versus ASCs obtained via standard suction-assisted lipoaspiration (SAL). Lipoaspirates were sorted using fluorescent assisted cell sorting based on an established surface-marker profile (CD34+/CD31−/CD45−), to obtain viable ASCs. Yield and viability were compared and the differentiation capacities of the ASCs were assessed. Finally, the regenerative potential of ASCs was examined using an in vivo model of tissue regeneration. UAL- and SAL-derived samples demonstrated equivalent ASC yield and viability, and UAL ASCs were not impaired in their osteogenic, adipogenic, or chondrogenic differentiation capacity. Equally, quantitative real-time polymerase chain reaction showed comparable expression of most osteogenic, adipogenic, and key regenerative genes between both ASC groups. Cutaneous regeneration and neovascularization were significantly enhanced in mice treated with ASCs obtained by either UAL or SAL compared with controls, but there were no significant differences in healing between cell-therapy groups. We conclude that UAL is a successful method of obtaining fully functional ASCs for regenerative medicine purposes. Cells harvested with this alternative approach to liposuction are suitable for cell therapy and tissue engineering applications. Significance Adipose-derived mesenchymal stem cells (ASCs) are an appealing source of therapeutic progenitor cells because of their multipotency

Ultrasound-Assisted Liposuction Does Not Compromise the Regenerative Potential of Adipose-Derived Stem Cells.

PubMed

Duscher, Dominik; Atashroo, David; Maan, Zeshaan N; Luan, Anna; Brett, Elizabeth A; Barrera, Janos; Khong, Sacha M; Zielins, Elizabeth R; Whittam, Alexander J; Hu, Michael S; Walmsley, Graham G; Pollhammer, Michael S; Schmidt, Manfred; Schilling, Arndt F; Machens, Hans-Günther; Huemer, Georg M; Wan, Derrick C; Longaker, Michael T; Gurtner, Geoffrey C

2016-02-01

Human mesenchymal stem cells (MSCs) have recently become a focus of regenerative medicine, both for their multilineage differentiation capacity and their excretion of proregenerative cytokines. Adipose-derived mesenchymal stem cells (ASCs) are of particular interest because of their abundance in fat tissue and the ease of harvest via liposuction. However, little is known about the impact of different liposuction methods on the functionality of ASCs. Here we evaluate the regenerative abilities of ASCs harvested via a third-generation ultrasound-assisted liposuction (UAL) device versus ASCs obtained via standard suction-assisted lipoaspiration (SAL). Lipoaspirates were sorted using fluorescent assisted cell sorting based on an established surface-marker profile (CD34+/CD31-/CD45-), to obtain viable ASCs. Yield and viability were compared and the differentiation capacities of the ASCs were assessed. Finally, the regenerative potential of ASCs was examined using an in vivo model of tissue regeneration. UAL- and SAL-derived samples demonstrated equivalent ASC yield and viability, and UAL ASCs were not impaired in their osteogenic, adipogenic, or chondrogenic differentiation capacity. Equally, quantitative real-time polymerase chain reaction showed comparable expression of most osteogenic, adipogenic, and key regenerative genes between both ASC groups. Cutaneous regeneration and neovascularization were significantly enhanced in mice treated with ASCs obtained by either UAL or SAL compared with controls, but there were no significant differences in healing between cell-therapy groups. We conclude that UAL is a successful method of obtaining fully functional ASCs for regenerative medicine purposes. Cells harvested with this alternative approach to liposuction are suitable for cell therapy and tissue engineering applications. Significance: Adipose-derived mesenchymal stem cells (ASCs) are an appealing source of therapeutic progenitor cells because of their multipotency

Application of a novel sorting system for equine mesenchymal stem cells (MSCs)

PubMed Central

Radtke, Catherine L.; Nino-Fong, Rodolfo; Esparza Gonzalez, Blanca P.; McDuffee, Laurie A.

2014-01-01

The objective of this study was to validate non-equilibrium gravitational field-flow fractionation (GrFFF), an immunotag-less method of sorting mesenchymal stem cells (MSCs) into subpopulations, for use with MSCs derived from equine muscle tissue, periosteal tissue, bone marrow, and adipose tissue. Cells were collected from 6 young, adult horses, postmortem. Cells were isolated from left semitendinosus muscle tissue, periosteal tissue from the distomedial aspect of the right tibia, bone marrow aspirates from the fourth and fifth sternebrae, and left supragluteal subcutaneous adipose tissue. Aliquots of 800 × 103 MSCs from each tissue source were separated and injected into a ribbon-like capillary device by continuous flow (GrFFF proprietary system). Cells were sorted into 6 fractions and absorbencies [optical density (OD)] were read. Six fractions from each of the 6 aliquots were then combined to provide pooled fractions that had adequate cell numbers to seed at equal concentrations into assays. Equine muscle tissue-derived, periosteal tissue-derived, bone marrow-derived, and adipose tissue-derived mesenchymal stem cells were consistently sorted into 6 fractions that remained viable for use in further assays. Fraction 1 had more cuboidal morphology in culture when compared to the other fractions. Statistical analysis of the fraction absorbencies (OD) revealed a P-value of < 0.05 when fractions 2 and 3 were compared to fractions 1, 4, 5, and 6. It was concluded that non-equilibrium GrFFF is a valid method for sorting equine muscle tissue-derived, periosteal tissue-derived, bone marrow-derived, and adipose tissue-derived mesenchymal stem cells into subpopulations that remain viable, thus securing its potential for use in equine stem cell applications and veterinary medicine. PMID:25355998

Matrix-directed differentiation of human adipose-derived mesenchymal stem cells to dermal-like fibroblasts that produce extracellular matrix.

PubMed

Sivan, Unnikrishnan; Jayakumar, K; Krishnan, Lissy K

2016-10-01

Commercially available skin substitutes lack essential non-immune cells for adequate tissue regeneration of non-healing wounds. A tissue-engineered, patient-specific, dermal substitute could be an attractive option for regenerating chronic wounds, for which adipose-derived mesenchymal stem cells (ADMSCs) could become an autologous source. However, ADMSCs are multipotent in nature and may differentiate into adipocytes, osteocytes and chondrocytes in vitro, and may develop into undesirable tissues upon transplantation. Therefore, ADMSCs committed to the fibroblast lineage could be a better option for in vitro or in vivo skin tissue engineering. The objective of this study was to standardize in vitro culture conditions for ADMSCs differentiation into dermal-like fibroblasts which can synthesize extracellular matrix (ECM) proteins. Biomimetic matrix composite, deposited on tissue culture polystyrene (TCPS), and differentiation medium (DM), supplemented with fibroblast-conditioned medium and growth factors, were used as a fibroblast-specific niche (FSN) for cell culture. For controls, ADMSCs were cultured on bare TCPS with either DM or basal medium (BM). Culture of ADMSCs on FSN upregulated the expression of differentiation markers such as fibroblast-specific protein-1 (FSP-1) and a panel of ECM molecules specific to the dermis, such as fibrillin-1, collagen I, collagen IV and elastin. Immunostaining showed the deposition of dermal-specific ECM, which was significantly higher in FSN compared to control. Fibroblasts derived from ADMSCs can synthesize elastin, which is an added advantage for successful skin tissue engineering as compared to fibroblasts from skin biopsy. To obtain rapid differentiation of ADMSCs to dermal-like fibroblasts for regenerative medicine, a matrix-directed differentiation strategy may be employed. Copyright © 2014 John Wiley & Sons, Ltd. Copyright © 2014 John Wiley & Sons, Ltd.

Chip-Based Comparison of the Osteogenesis of Human Bone Marrow- and Adipose Tissue-Derived Mesenchymal Stem Cells under Mechanical Stimulation

PubMed Central

Min, Byoung-Hyun; Yang, Sang Sik; Khademhosseini, Ali; Kaplan, David L.

2012-01-01

Adipose tissue-derived stem cells (ASCs) are considered as an attractive stem cell source for tissue engineering and regenerative medicine. We compared human bone marrow-derived mesenchymal stem cells (hMSCs) and hASCs under dynamic hydraulic compression to evaluate and compare osteogenic abilities. A novel micro cell chip integrated with microvalves and microscale cell culture chambers separated from an air-pressure chamber was developed using microfabrication technology. The microscale chip enables the culture of two types of stem cells concurrently, where each is loaded into cell culture chambers and dynamic compressive stimulation is applied to the cells uniformly. Dynamic hydraulic compression (1 Hz, 1 psi) increased the production of osteogenic matrix components (bone sialoprotein, oateopontin, type I collagen) and integrin (CD11b and CD31) expression from both stem cell sources. Alkaline phosphatase and Alrizarin red staining were evident in the stimulated hMSCs, while the stimulated hASCs did not show significant increases in staining under the same stimulation conditions. Upon application of mechanical stimulus to the two types of stem cells, integrin (β1) and osteogenic gene markers were upregulated from both cell types. In conclusion, stimulated hMSCs and hASCs showed increased osteogenic gene expression compared to non-stimulated groups. The hMSCs were more sensitive to mechanical stimulation and more effective towards osteogenic differentiation than the hASCs under these modes of mechanical stimulation. PMID:23029565

Icaritin, a novel plant-derived osteoinductive agent, enhances the osteogenic differentiation of human bone marrow- and human adipose tissue-derived mesenchymal stem cells.

PubMed

Wu, Tao; Shu, Tao; Kang, Le; Wu, Jinhui; Xing, Jianzhou; Lu, Zhiqin; Chen, Shuxiang; Lv, Jun

2017-04-01

For the treatment of diseases affecting bones using bone regenerative medicine, there is an urgent need to develop safe, inexpensive drugs that can strongly induce bone formation. In the present study, we systematically investigated the effects of icaritin, a metabolic product of icariin, on the osteogenic differentiation of human bone marrow‑derived mesenchymal stem cells (hBMSCs) and human adipose tissue‑derived stem cells (hADSCs) in vitro. After treatment with icaritin at concentrations of 10‑8-10‑5 M, hBMSCs and hADSCs were examined for alkaline phosphatase activity, osteocalcin (OC) secretion, matrix mineralization and expression levels of bone‑related mRNA and proteins. Data showed that icaritin at concentrations 10‑7-10‑5 M significantly increased alkaline phosphatase activity, OC secretion at different time points, and calcium deposition at day 21. In addition, icaritin upregulated the mRNA expression of genes for bone morphogenetic proteins (BMP‑2, ‑4 and ‑7), bone transcription factors (Runx2 and Dlx5) and bone matrix proteins (ALP, OC and Col‑1). Moreover, icaritin increased the protein levels of BMPs, Runx2 and OC, as detected by western blot analysis. These findings suggest that icaritin enhances the osteogenic differentiation of hBMSCS and hADSCs. Icaritin exerts its potent osteogenic effect possibly by directly stimulating the production of BMPs. Although the osteogenic activity of icaritin in vitro was inferior to that of rhBMP‑2, icaritin displayed better results than icariin. Moreover, the low cost, simple extraction procedure, and an abundance of icaritin make it appealing as a bone regenerative medicine.

New Therapy of Skin Repair Combining Adipose-Derived Mesenchymal Stem Cells with Sodium Carboxymethylcellulose Scaffold in a Pre-Clinical Rat Model

PubMed Central

Rodrigues, Cristiano; de Assis, Adriano M.; Moura, Dinara J.; Halmenschlager, Graziele; Saffi, Jenifer; Xavier, Léder Leal; da Cruz Fernandes, Marilda; Wink, Márcia Rosângela

2014-01-01

Lesions with great loss of skin and extensive burns are usually treated with heterologous skin grafts, which may lead rejection. Cell therapy with mesenchymal stem cells is arising as a new proposal to accelerate the healing process. We tested a new therapy consisting of sodium carboxymethylcellulose (CMC) as a biomaterial, in combination with adipose-derived stem cells (ADSCs), to treat skin lesions in an in vivo rat model. This biomaterial did not affect membrane viability and induced a small and transient genotoxicity, only at the highest concentration tested (40 mg/mL). In a rat wound model, CMC at 10 mg/mL associated with ADSCs increased the rate of cell proliferation of the granulation tissue and epithelium thickness when compared to untreated lesions (Sham), but did not increase collagen fibers nor alter the overall speed of wound closure. Taken together, the results show that the CMC is capable to allow the growth of ADSCs and is safe for this biological application up to the concentration of 20 mg/mL. These findings suggest that CMC is a promising biomaterial to be used in cell therapy. PMID:24788779

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.

Hypoxia-cultured human adipose-derived mesenchymal stem cells are non-oncogenic and have enhanced viability, motility, and tropism to brain cancer

PubMed Central

Feng, Y; Zhu, M; Dangelmajer, S; Lee, Y M; Wijesekera, O; Castellanos, C X; Denduluri, A; Chaichana, K L; Li, Q; Zhang, H; Levchenko, A; Guerrero-Cazares, H; Quiñones-Hinojosa, A

2014-01-01

Adult human adipose-derived mesenchymal stem cells (hAMSCs) are multipotent cells, which are abundant, easily collected, and bypass the ethical concerns that plague embryonic stem cells. Their utility and accessibility have led to the rapid development of clinical investigations to explore their autologous and allogeneic cellular-based regenerative potential, tissue preservation capabilities, anti-inflammatory properties, and anticancer properties, among others. hAMSCs are typically cultured under ambient conditions with 21% oxygen. However, physiologically, hAMSCs exist in an environment of much lower oxygen tension. Furthermore, hAMSCs cultured in standard conditions have shown limited proliferative and migratory capabilities, as well as limited viability. This study investigated the effects hypoxic culture conditions have on primary intraoperatively derived hAMSCs. hAMSCs cultured under hypoxia (hAMSCs-H) remained multipotent, capable of differentiation into osteogenic, chondrogenic, and adipogenic lineages. In addition, hAMSCs-H grew faster and exhibited less cell death. Furthermore, hAMSCs-H had greater motility than normoxia-cultured hAMSCs and exhibited greater homing ability to glioblastoma (GBM) derived from brain tumor-initiating cells from our patients in vitro and in vivo. Importantly, hAMSCs-H did not transform into tumor-associated fibroblasts in vitro and were not tumorigenic in vivo. Rather, hAMSCs-H promoted the differentiation of brain cancer cells in vitro and in vivo. These findings suggest an alternative culturing technique that can enhance the function of hAMSCs, which may be necessary for their use in the treatment of various pathologies including stroke, myocardial infarction, amyotrophic lateral sclerosis, and GBM. PMID:25501828

A fat option for the pig: Hepatocytic differentiated mesenchymal stem cells for translational research

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

Brückner, Sandra, E-mail: sandra.brueckner@medizin.uni-leipzig.de; Tautenhahn, Hans-Michael, E-mail: hans-michael.tautenhahn@medizin.uni-leipzig.de; TRM, Translational Centre for Regenerative Medicine, Philipp-Rosenthal-Str. 55, Leipzig D-04103

Study background: Extended liver resection is the only curative treatment option of liver cancer. Yet, the residual liver may not accomplish the high metabolic and regenerative capacity needed, which frequently leads to acute liver failure. Because of their anti-inflammatory and -apoptotic as well as pro-proliferative features, mesenchymal stem cells differentiated into hepatocyte-like cells might provide functional and regenerative compensation. Clinical translation of basic research requires pre-clinical approval in large animals. Therefore, we characterized porcine mesenchymal stem cells (MSC) from adipose tissue and bone marrow and their hepatocyte differentiation potential for future assessment of functional liver support after surgical intervention inmore » the pig model. Methods: Mesenchymal surface antigens and multi-lineage differentiation potential of porcine MSC isolated by collagenase digestion either from bone marrow or adipose tissue (subcutaneous/visceral) were assessed by flow cytometry. Morphology and functional properties (urea-, glycogen synthesis and cytochrome P450 activity) were determined during culture under differentiation conditions and compared with primary porcine hepatocytes. Results: MSC from porcine adipose tissue and from bone marrow express the typical mesenchymal markers CD44, CD29, CD90 and CD105 but not haematopoietic markers. MSC from both sources displayed differentiation into the osteogenic as well as adipogenic lineage. After hepatocyte differentiation, expression of CD105 decreased significantly and cells adopted the typical polygonal morphology of hepatocytes. Glycogen storage was comparable in adipose tissue- and bone marrow-derived cells. Urea synthesis was about 35% lower in visceral than in subcutaneous adipose tissue-derived MSC. Cytochrome P450 activity increased significantly during differentiation and was twice as high in hepatocyte-like cells generated from bone marrow as from adipose tissue. Conclusion: The

Naringin protects human adipose-derived mesenchymal stem cells against hydrogen peroxide-induced inhibition of osteogenic differentiation.

PubMed

Wang, Lei; Zhang, Yu-Ge; Wang, Xiu-Mei; Ma, Long-Fei; Zhang, Yuan-Min

2015-12-05

Extensive evidence indicates that oxidative stress plays a pivotal role in the development of osteoporosis. We show that naringin, a natural antioxidant and anti-inflammatory compound, effectively protects human adipose-derived mesenchymal stem cells (hADMSCs) against hydrogen peroxide (H2O2)-induced inhibition of osteogenic differentiation. Naringin increased viability of hAMDSCs and attenuated H2O2-induced cytotoxicity. Naringin also reversed H2O2-induced oxidative stress. Oxidative stress induced by H2O2 inhibits osteogenic differentiation by decreasing alkaline phosphatase (ALP) activity, calcium content and mRNA expression levels of osteogenesis marker genes RUNX2 and OSX in hADMSCs. However, addition of naringin leads to a significant recovery, suggesting the protective effects of naringin against H2O2-induced inhibition of osteogenic differentiation. Furthermore, the H2O2-induced decrease of protein expressions of β-catenin and clyclin D1, two important transcriptional regulators of Wnt-signaling, was successfully rescued by naringin treatment. Also, in the presence of Wnt inhibitor DKK-1, naringin is no longer effective in stimulating ALP activity, increasing calcium content and mRNA expression levels of RUNX2 and OSX in H2O2-exposed hADMSCs. These data clearly demonstrates that naringin protects hADMSCs against oxidative stress-induced inhibition of osteogenic differentiation, which may involve Wnt signaling pathway. Our work suggests that naringin may be a useful addition to the treatment armamentarium for osteoporosis and activation of Wnt signaling may represent attractive therapeutic strategy for the treatment of degenerative disease of bone tissue. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

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.

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

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

The therapeutic potential of human adipose-derived mesenchymal stem cells producing CXCL10 in a mouse melanoma lung metastasis model.

PubMed

Mirzaei, Hamed; Salehi, Hossein; Oskuee, Reza Kazemi; Mohammadpour, Ali; Mirzaei, Hamid Reza; Sharifi, Mohammad Reza; Salarinia, Reza; Darani, Hossein Yousofi; Mokhtari, Mojgan; Masoudifar, Aria; Sahebkar, Amirhossein; Salehi, Rasoul; Jaafari, Mahmoud Reza

2018-04-10

Interferon γ-induced protein 10 kDa (IP-10) is a potent chemoattractant and has been suggested to enhance antitumor activity and mediate tumor regression through multiple mechanisms of action. Multiple lines of evidence have indicated that genetically-modified adult stem cells represent a potential source for cell-based cancer therapy. In the current study, we assessed therapeutic potential of human adipose derived mesenchymal stem cells (hADSC) genetically-modified to express IP-10 for the treatment of lung metastasis in an immunocompetent mouse model of metastatic melanoma. A Piggybac vector encoding IP-10 was employed to transfect hADSC ex vivo. Expression and bioactivity of the transgenic protein from hADSCs expressing IP-10 were confirmed prior to in vivo studies. Our results indicated that hADSCs expressing IP-10 could inhibit the growth of B16F10 melanoma cells and significantly prolonged survival. Immunohistochemistry analysis, TUNEL assay and western blot analysis indicated that hADSCs expressing IP-10 inhibited tumor cell growth, hindered tumor infiltration of Tregs, restricted angiogenesis and significantly prolonged survival. In conclusion, our results demonstrated that targeting metastatic tumor sites by hADSC expressing IP-10 could reduce melanoma tumor growth and lung metastasis. Copyright © 2018 Elsevier B.V. All rights reserved.

Adipose tissue stem cells in regenerative medicine

PubMed Central

Miana, Vanesa Verónica; González, Elio A Prieto

2018-01-01

Adipose tissue-derived stem cells (ADSCs) are mesenchymal cells with the capacity for self-renewal and multipotential differentiation. This multipotentiality allows them to become adipocytes, chondrocytes, myocytes, osteoblasts and neurocytes among other cell lineages. Stem cells and, in particular, adipose tissue-derived cells, play a key role in reconstructive or tissue engineering medicine as they have already proven effective in developing new treatments. The purpose of this work is to review the applications of ADSCs in various areas of regenerative medicine, as well as some of the risks associated with treatment with ADSCs in neoplastic disease. PMID:29662535

[Wnt/β-catenin pathway involved in the regulation of rat mesangial cell proliferation by adipose-derived mesenchymal stem cells].

PubMed

Li, Zhi; Zhang, Mengying; Li, Xueqin; Lu, Jinming; Xu, Liang

2016-11-01

Objective To investigate the effect of adipose-derived mesenchymal stem cells (ADSCs) on glomerular mesangial cell proliferation via Wnt/β-catenin pathway. Methods The rat glomerular mesangial cells (HBZY-1) were incubated in conditioned ADSC medium. Cell cycle was analyzed with flow cytometry; the proliferation rate of HBZY-1 and the expression levels of relative genes and proteins of Wnt signaling pathway were measured using RNA interference, quantitative real-time PCR and Western blotting, respectively. Results HBZY-1 proliferation was significantly inhibited under the action of conditioned ADSC medium, whereas dickkopf WNT signaling pathway inhibitor 1 (DKK1) mRNA level was up-regulated. Fibronectin and TGF-β1 mRNA expression as well as β-catenin and Bcl-2 protein levels of HBZY-1 were significantly down-regulated. DKK1 gene expression level in ADSCs was significantly higher than that of HBZY-1. After RNA interference, DKK1 expression level in ADSCs was markedly inhibited, yet the β-catenin protein level was notably elevated. The β-catenin and Bcl-2 protein levels of HBZY-1 were also significantly raised in HBZY-1 after cultured with conditioned medium containing ADSCs treated with RNA interference. Conclusion Wnt/β-catenin may be a potential signaling pathway involved in the regulative effect of ADSCs on glomerular mesangial cell proliferation.

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

Generation of highly purified neural stem cells from human adipose-derived mesenchymal stem cells by Sox1 activation.

PubMed

Feng, Nianhua; Han, Qin; Li, Jing; Wang, Shihua; Li, Hongling; Yao, Xinglei; Zhao, Robert Chunhua

2014-03-01

Neural stem cells (NSCs) are ideal candidates in stem cell-based therapy for neurodegenerative diseases. However, it is unfeasible to get enough quantity of NSCs for clinical application. Generation of NSCs from human adipose-derived mesenchymal stem cells (hAD-MSCs) will provide a solution to this problem. Currently, the differentiation of hAD-MSCs into highly purified NSCs with biological functions is rarely reported. In our study, we established a three-step NSC-inducing protocol, in which hAD-MSCs were induced to generate NSCs with high purity after sequentially cultured in the pre-inducing medium (Step1), the N2B27 medium (Step2), and the N2B27 medium supplement with basic fibroblast growth factor and epidermal growth factor (Step3). These hAD-MSC-derived NSCs (adNSCs) can form neurospheres and highly express Sox1, Pax6, Nestin, and Vimentin; the proportion was 96.1% ± 1.3%, 96.8% ± 1.7%, 96.2% ± 1.3%, and 97.2% ± 2.5%, respectively, as detected by flow cytometry. These adNSCs can further differentiate into astrocytes, oligodendrocytes, and functional neurons, which were able to generate tetrodotoxin-sensitive sodium current. Additionally, we found that the neural differentiation of hAD-MSCs were significantly suppressed by Sox1 interference, and what's more, Step1 was a key step for the following induction, probably because it was associated with the initiation and nuclear translocation of Sox1, an important transcriptional factor for neural development. Finally, we observed that bone morphogenetic protein signal was inhibited, and Wnt/β-catenin signal was activated during inducing process, and both signals were related with Sox1 expression. In conclusion, we successfully established a three-step inducing protocol to derive NSCs from hAD-MSCs with high purity by Sox1 activation. These findings might enable to acquire enough autologous transplantable NSCs for the therapy of neurodegenerative diseases in clinic.

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.

Does an Injection of Adipose-Derived Mesenchymal Stem Cells Loaded in Fibrin Glue Influence Rotator Cuff Repair Outcomes? A Clinical and Magnetic Resonance Imaging Study.

PubMed

Kim, Yong Sang; Sung, Chang Hun; Chung, Sung Hoon; Kwak, Sang Joon; Koh, Yong Gon

2017-07-01

The mesenchymal stem cell (MSC)-based tissue engineering approach has been developed to improve the treatment of rotator cuff tears. Hypothesis/Purpose: The purpose was to determine the effect of an injection of adipose-derived MSCs loaded in fibrin glue during arthroscopic rotator cuff repair on clinical outcomes and to evaluate its effect on structural integrity using magnetic resonance imaging (MRI). The hypothesis was that the application of adipose-derived MSCs would improve outcomes after the surgical repair of a rotator cuff tear. Cohort study; Level of evidence, 3. Among 182 patients treated with arthroscopic surgery for a rotator cuff tear, 35 patients treated with arthroscopic rotator cuff repair alone (conventional group) were matched with 35 patients who underwent arthroscopic rotator cuff repair with an injection of adipose-derived MSCs loaded in fibrin glue (injection group) based on sex, age, and lesion size. Outcomes were assessed with respect to the visual analog scale (VAS) for pain, range of motion (ROM) (including forward flexion, external rotation at the side, and internal rotation at the back), and functional measures of the Constant score and University of California, Los Angeles (UCLA) shoulder rating scale. Repaired tendon structural integrity was assessed by using MRI at a minimum of 12 months after surgery, and the mean clinical follow-up was 28.8 ± 4.2 months in the conventional group and 28.3 ± 3.8 months in the injection group. The mean VAS score at rest and during motion improved significantly in both groups after surgery. However, there were no significant differences between the groups at the final follow-up ( P = .256 and .776, respectively). Compared with preoperative measurements, forward flexion and external rotation at the side significantly improved at the final follow-up in both groups (all P < .05). However, no significant improvements in internal rotation at the back were observed in either group ( P = .625 and .834 for

Local injections of adipose-derived mesenchymal stem cells modulate inflammation and increase angiogenesis ameliorating the dystrophic phenotype in dystrophin-deficient skeletal muscle.

PubMed

Pinheiro, Carlos Hermano da Justa; de Queiroz, Jean César Farias; Guimarães-Ferreira, Lucas; Vitzel, Kaio Fernando; Nachbar, Renato Tadeu; de Sousa, Luís Gustavo Oliveira; de Souza, Alcione Lescano; Nunes, Maria Tereza; Curi, Rui

2012-06-01

The effects of adipose-derived mesenchymal stem cells (ADMSC) transplantation on degeneration, regeneration and skeletal muscle function were investigated in dystrophin-deficient mice (24-week-old). ADMSC transplantation improved muscle strength and, resistance to fatigue. An increase in fiber cross-sectional area and in the number of fibers with centralized nuclei and augment of myogenin content were observed. In ADMSC-treated muscles a decrease in muscle content of TNF-α, IL-6 and oxidative stress measured by Amplex(®) reagent were observed. The level of TGF-β1 was lowered whereas that of VEGF, IL-10 and IL-4 were increased by ADMSC treatment. An increase in markers of macrophage M1 (CD11 and F4-80) and a decrease in T lymphocyte marker (CD3) and arginase-1 were also observed in ADMSCs-treated dystrophic muscle. No change was observed in iNOS expression. Increased phosphorylation of Akt, p70S6k and 4E-BP1 was found in dystrophic muscles treated with ADMSC. These results suggest that ADMSC transplantation modulates inflammation and improves muscle tissue regeneration, ameliorating the dystrophic phenotype in dystrophin-deficient mice.

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.

Adipose, Bone Marrow and Synovial Joint-Derived Mesenchymal Stem Cells for Cartilage Repair

PubMed Central

Fellows, Christopher R.; Matta, Csaba; Zakany, Roza; Khan, Ilyas M.; Mobasheri, Ali

2016-01-01

Current cell-based repair strategies have proven unsuccessful for treating cartilage defects and osteoarthritic lesions, consequently advances in innovative therapeutics are required and mesenchymal stem cell-based (MSC) therapies are an expanding area of investigation. MSCs are capable of differentiating into multiple cell lineages and exerting paracrine effects. Due to their easy isolation, expansion, and low immunogenicity, MSCs are an attractive option for regenerative medicine for joint repair. Recent studies have identified several MSC tissue reservoirs including in adipose tissue, bone marrow, cartilage, periosteum, and muscle. MSCs isolated from these discrete tissue niches exhibit distinct biological activities, and have enhanced regenerative potentials for different tissue types. Each MSC type has advantages and disadvantages for cartilage repair and their use in a clinical setting is a balance between expediency and effectiveness. In this review we explore the challenges associated with cartilage repair and regeneration using MSC-based cell therapies and provide an overview of phenotype, biological activities, and functional properties for each MSC population. This paper also specifically explores the therapeutic potential of each type of MSC, particularly focusing on which cells are capable of producing stratified hyaline-like articular cartilage regeneration. Finally we highlight areas for future investigation. Given that patients present with a variety of problems it is unlikely that cartilage regeneration will be a simple “one size fits all,” but more likely an array of solutions that need to be applied systematically to achieve regeneration of a biomechanically competent repair tissue. PMID:28066501

Use of Adipose-Derived Mesenchymal Stem Cells to Accelerate Neovascularization in Interpolation Flaps.

PubMed

Izmirli, Hakki Hayrettin; Alagoz, Murat Sahin; Gercek, Huseyin; Eren, Guler Gamze; Yucel, Ergin; Subasi, Cansu; Isgoren, Serkan; Muezzinoglu, Bahar; Karaoz, Erdal

2016-01-01

Interpolation flaps are commonly used in plastic surgery to cover wide and deep defects. The need to, wait for 2 to 3 weeks until the division of the pedicle still, however, poses a serious challenge, not only extending treatment and hospital stay, but also increasing hospital expenses. To solve this problem, we have aimed to use the angiogenic potential of stem cells to selectively accelerate neovascularization with a view to increasing the viability of interpolation flaps and achieving early pedicle removal. A total of 32 rats were allocated to 2 groups as control (N = 16) and experiment (N = 16). The cranial flaps 6 × 5 cm in size located on the back of the rats were raised. Then, a total suspension containing 3 × 10(6) adipose-derived mesenchymal stem cells (ADSC) tagged with a green fluorescent protein (GFP) was injected diffusely into the distal part of the flap, receiving bed, and wound edges. In the control group, only a medium solution was injected into the same sites. After covering the 3 × 5 cm region in the proximal part of the area where the flap was removed, the distal part of the flap was adapted to the uncovered distal area. The pedicles of 4 rats in each group were divided on postoperative days 5, 8, 11, and 14. The areas were photographed 7 days after the pedicles were released. The photographs were processed using Adobe Acrobat 9 Pro software (San Jose, CA) to measure the flap survival area in millimeters and to compare groups. Seven days after the flap pedicle was divided, the rats were injected with 250 mCi Tc-99 mm (methoxy-isobutyl-isonitrie) from the penile vein, and scintigraphic images were obtained. The images obtained from each group were subjected to a numerical evaluation, which was then used in the comparison between groups. The flaps were then examined by histology to numerically compare the number of newly formed vessels. Neovascularization was also assessed by microangiography. In addition, radiographic images were obtained by

The Potential of GMP-Compliant Platelet Lysate to Induce a Permissive State for Cardiovascular Transdifferentiation in Human Mediastinal Adipose Tissue-Derived Mesenchymal Stem Cells

PubMed Central

Bordin, Antonella; Ponti, Donatella; Iudicone, Paola; Rendina, Erino Angelo; Calogero, Antonella; Pierelli, Luca; Ibrahim, Mohsen; De Falco, Elena

2015-01-01

Human adipose tissue-derived mesenchymal stem cells (ADMSCs) are considered eligible candidates for cardiovascular stem cell therapy applications due to their cardiac transdifferentiation potential and immunotolerance. Over the years, the in vitro culture of ADMSCs by platelet lysate (PL), a hemoderivate containing numerous growth factors and cytokines derived from platelet pools, has allowed achieving a safe and reproducible methodology to obtain high cell yield prior to clinical administration. Nevertheless, the biological properties of PL are still to be fully elucidated. In this brief report we show the potential ability of PL to induce a permissive state of cardiac-like transdifferentiation and to cause epigenetic modifications. RTPCR results indicate an upregulation of Cx43, SMA, c-kit, and Thy-1 confirmed by immunofluorescence staining, compared to standard cultures with foetal bovine serum. Moreover, PL-cultured ADMSCs exhibit a remarkable increase of both acetylated histones 3 and 4, with a patient-dependent time trend, and methylation at lysine 9 on histone 3 preceding the acetylation. Expression levels of p300 and SIRT-1, two major regulators of histone 3, are also upregulated after treatment with PL. In conclusion, PL could unravel novel biological properties beyond its routine employment in noncardiac applications, providing new insights into the plasticity of human ADMSCs. PMID:26495284

The potential of GMP-compliant platelet lysate to induce a permissive state for cardiovascular transdifferentiation in human mediastinal adipose tissue-derived mesenchymal stem cells.

PubMed

Siciliano, Camilla; Chimenti, Isotta; Bordin, Antonella; Ponti, Donatella; Iudicone, Paola; Peruzzi, Mariangela; Rendina, Erino Angelo; Calogero, Antonella; Pierelli, Luca; Ibrahim, Mohsen; De Falco, Elena

2015-01-01

Human adipose tissue-derived mesenchymal stem cells (ADMSCs) are considered eligible candidates for cardiovascular stem cell therapy applications due to their cardiac transdifferentiation potential and immunotolerance. Over the years, the in vitro culture of ADMSCs by platelet lysate (PL), a hemoderivate containing numerous growth factors and cytokines derived from platelet pools, has allowed achieving a safe and reproducible methodology to obtain high cell yield prior to clinical administration. Nevertheless, the biological properties of PL are still to be fully elucidated. In this brief report we show the potential ability of PL to induce a permissive state of cardiac-like transdifferentiation and to cause epigenetic modifications. RTPCR results indicate an upregulation of Cx43, SMA, c-kit, and Thy-1 confirmed by immunofluorescence staining, compared to standard cultures with foetal bovine serum. Moreover, PL-cultured ADMSCs exhibit a remarkable increase of both acetylated histones 3 and 4, with a patient-dependent time trend, and methylation at lysine 9 on histone 3 preceding the acetylation. Expression levels of p300 and SIRT-1, two major regulators of histone 3, are also upregulated after treatment with PL. In conclusion, PL could unravel novel biological properties beyond its routine employment in noncardiac applications, providing new insights into the plasticity of human ADMSCs.

Melatonin treatment further improves adipose-derived mesenchymal stem cell therapy for acute interstitial cystitis in rat.

PubMed

Chen, Yen-Ta; Chiang, Hsin-Ju; Chen, Chih-Hung; Sung, Pei-Hsun; Lee, Fan-Yen; Tsai, Tzu-Hsien; Chang, Chia-Lo; Chen, Hong-Hwa; Sun, Cheuk-Kwan; Leu, Steve; Chang, Hsueh-Wen; Yang, Chih-Chao; Yip, Hon-Kan

2014-10-01

This study tests the hypothesis that combined melatonin and adipose-derived mesenchymal stem cell (ADMSC, 1.2 × 10(6) given intravenously) treatment offer superior protection against cyclophosphamide (CYP 150 mg/kg)-induced acute interstitial cystitis (AIC) in rats. Male adult Sprague-Dawley rats were treated as follows: sham controls, AIC alone, AIC + melatonin, AIC + ADMSC, and AIC + melatonin +ADMSC. When melatonin was used, it was given as follows: 20 mg/kg at 30 min after CYP and 50 mg/kg at 6 and 18 hr after CYP. Twenty-four-hour urine volume, urine albumin level, and severity of hematuria were highest in AIC rats and lowest in the controls; likewise urine volume was higher in AIC + melatonin rats than in AIC + ADMSC and AIC + melatonin + ADMSC treated rats; in all cases, P < 0.001. The numbers of CD14+, CD74+, CD68+, MIP+, Cox-2+, substance P+, cells and protein expression of IL-6, IL-12, RANTES, TNF-α, NF-κB, MMP-9, iNOS (i.e. inflammatory biomarkers), glycosaminoglycan level, expression of oxidized protein, and protein expression of reactive oxygen species (NOX-1, NOX-2, NOX-4) in the bladder tissue exhibited an identical pattern compared with that of hematuria among the five groups (all P < 0.0001). The integrity of epithelial layer and area of collagen deposition displayed an opposite pattern compared to that of hematuria among all groups (P < 0.0001). The cellular expressions of antioxidants (GR, GPx, HO-1, NQO 1) showed a significant progressive increase form controls to AIC + melatonin + ADMSC (all P < 0.0001). Combined regimen of melatonin and ADMSC was superior to either alone in protecting against CYP-induced AIC. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Thermally labile components of aqueous humor potently induce osteogenic potential in adipose-derived mesenchymal stem cells

PubMed Central

Morgan, Joshua T.; Kwon, Heung Sun; Wood, Joshua A.; Borjesson, Dori L.; Tomarev, Stanislav I.; Murphy, Christopher J.; Russell, Paul

2015-01-01

Adipose-derived mesenchymal stem cells (ASCs) hold promise for use in cell-based therapies. Their intrinsic anti-inflammatory properties are potentially useful for treatments of inflammatory conditions such as uveitis, while their ability to differentiate along multiple cell lineages suggests use in regenerating damaged or degenerated tissue. However, how ASCs will respond to the intraocular environment is poorly studied. We have recently reported that aqueous humor (AH), the fluid that nourishes the anterior segment of the eye, potently increases alkaline phosphatase (ALP) activity of ASCs, indicating osteogenic differentiation. Here, we expand on our previous findings to better define the nature of this response. To this end, we cultured ASCs in the presence of 0, 5, 10, and 20% AH and assayed them for ALP activity. We found ALP activity correlates with increasing AH concentrations from 5 to 20%, and that longer treatments result in increased ALP activity. By using serum free media and pretreating AH with dextran-coated charcoal, we found that serum and charcoal-adsorbable AH components augment but are not required for this response. Further, by heat-treating the AH, we established that thermally labile components are required for the osteogenic response. Finally, we showed myocilin, a protein present in AH, could induce ALP activity in ASCs. However, this was to a lesser extent than untreated 5% AH, and myocilin could only partially rescue the effect after heat treatment, documenting there were additional thermally labile constituents of AH involved in the osteogenic response. Our work adds to the understanding of the induction of ALP in ASCs following exposure to AH, providing important insight in how ASCs will be influenced by the ocular environment. In conclusion, increased osteogenic potential upon exposure to AH represents a potential challenge to developing ASC cell-based therapies directed at the eye. PMID:25720657

Novel daidzein analogs enhance osteogenic activity of bone marrow-derived mesenchymal stem cells and adipose-derived stromal/stem cells through estrogen receptor dependent and independent mechanisms

USDA-ARS?s Scientific Manuscript database

Osteoporosis is a disease characterized by low bone mineral density (BMD) and increased risk of fractures. Studies have demonstrated the use of phytoestrogens, or plant-derived estrogens, such as genistein anddaidzein, to effectively increase osteogenic activity of bone marrow-derived mesenchymal s...

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.

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.

Labeling and Imaging Mesenchymal Stem Cells with Quantum Dots

EPA Science Inventory

Mesenchymal stem cells (MSCs) are multipotent cells with the potential to differentiate into bone, cartilage, adipose and muscle cells. Adult derived MSCs are being actively investigated because of their potential to be utilized for therapeutic cell-based transplantation. Methods...

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

Human and feline adipose-derived mesenchymal stem cells have comparable phenotype, immunomodulatory functions, and transcriptome.

PubMed

Clark, Kaitlin C; Fierro, Fernando A; Ko, Emily Mills; Walker, Naomi J; Arzi, Boaz; Tepper, Clifford G; Dahlenburg, Heather; Cicchetto, Andrew; Kol, Amir; Marsh, Lyndsey; Murphy, William J; Fazel, Nasim; Borjesson, Dori L

2017-03-20

Adipose-derived mesenchymal stem cells (ASCs) are a promising cell therapy to treat inflammatory and immune-mediated diseases. Development of appropriate pre-clinical animal models is critical to determine safety and attain early efficacy data for the most promising therapeutic candidates. Naturally occurring diseases in cats already serve as valuable models to inform human clinical trials in oncologic, cardiovascular, and genetic diseases. The objective of this study was to complete a comprehensive side-by-side comparison of human and feline ASCs, with an emphasis on their immunomodulatory capacity and transcriptome. Human and feline ASCs were evaluated for phenotype, immunomodulatory profile, and transcriptome. Additionally, transwells were used to determine the role of cell-cell contact in ASC-mediated inhibition of lymphocyte proliferation in both humans and cats. Similar to human ASCs, feline ASCs were highly proliferative at low passages and fit the minimal criteria of multipotent stem cells including a compatible surface protein phenotype, osteogenic capacity, and normal karyotype. Like ASCs from all species, feline ASCs inhibited mitogen-activated lymphocyte proliferation in vitro, with or without direct ASC-lymphocyte contact. Feline ASCs mimic human ASCs in their mediator secretion pattern, including prostaglandin E2, indoleamine 2,3 dioxygenase, transforming growth factor beta, and interleukin-6, all augmented by interferon gamma secretion by lymphocytes. The transcriptome of three unactivated feline ASC lines were highly similar. Functional analysis of the most highly expressed genes highlighted processes including: 1) the regulation of apoptosis; 2) cell adhesion; 3) response to oxidative stress; and 4) regulation of cell differentiation. Finally, feline ASCs had a similar gene expression profile to noninduced human ASCs. Findings suggest that feline ASCs modulate lymphocyte proliferation using soluble mediators that mirror the human ASC secretion

Neonatal overfeeding impairs differentiation potential of mice subcutaneous adipose mesenchymal stem cells.

PubMed

Dias, Isabelle; Salviano, Ísis; Mencalha, André; de Carvalho, Simone Nunes; Thole, Alessandra Alves; Carvalho, Laís; Cortez, Erika; Stumbo, Ana Carolina

2018-04-17

Nutritional changes in the development (intrauterine life and postnatal period) may trigger long-term pathophysiological complications such as obesity and cardiovascular disease. Metabolic programming leads to organs and tissues modifications, including adipose tissue, with increased lipogenesis, production of inflammatory cytokines, and decreased glucose uptake. However, stem cells participation in adipose tissue dysfunctions triggered by overfeeding during lactation has not been elucidated. Therefore, this study was the first to evaluate the effect of metabolic programming on adipose mesenchymal stem cells (ASC) from mice submitted to overfeeding during lactation, using the litter reduction model. Cells were evaluated for proliferation capacity, viability, immunophenotyping, and reactive oxygen species (ROS) production. The content of UCP-2 and PGC1-α was determined by Western Blot. ASC differentiation potential in adipogenic and osteogenic environments was also evaluated, as well the markers of adipogenic differentiation (PPAR-γ and FAB4) and osteogenic differentiation (osteocalcin) by RT-qPCR. Results indicated that neonatal overfeeding does not affect ASC proliferation, ROS production, and viability. However, differentiation potential and proteins related to metabolism were altered. ASC from overfed group presented increased adipogenic differentiation, decreased osteogenic differentiation, and also showed increased PGC1-α protein content and reduced UCP-2 expression. Thus, ASC may be involved with the increased adiposity observed in neonatal overfeeding, and its therapeutic potential may be affected.

Infusion of autologous adipose tissue derived neuronal differentiated mesenchymal stem cells and hematopoietic stem cells in post-traumatic paraplegia offers a viable therapeutic approach.

PubMed

Thakkar, Umang G; Vanikar, Aruna V; Trivedi, Hargovind L; Shah, Veena R; Dave, Shruti D; Dixit, Satyajit B; Tiwari, Bharat B; Shah, Harda H

2016-01-01

Spinal cord injury (SCI) is not likely to recover by current therapeutic modalities. Stem cell (SC) therapy (SCT) has promising results in regenerative medicine. We present our experience of co-infusion of autologous adipose tissue derived mesenchymal SC differentiated neuronal cells (N-Ad-MSC) and hematopoietic SCs (HSCs) in a set of patients with posttraumatic paraplegia. Ten patients with posttraumatic paraplegia of mean age 3.42 years were volunteered for SCT. Their mean age was 28 years, and they had variable associated complications. They were subjected to adipose tissue resection for in vitro generation of N-Ad-MSC and bone marrow aspiration for generation of HSC. Generated SCs were infused into the cerebrospinal fluid (CSF) below injury site in all patients. Total mean quantum of SC infused was 4.04 ml with a mean nucleated cell count of 4.5 × 10(4)/μL and mean CD34+ of 0.35%, CD45-/90+ and CD45-/73+ of 41.4%, and 10.04%, respectively. All of them expressed transcription factors beta-3 tubulin and glial fibrillary acid protein. No untoward effect of SCT was noted. Variable and sustained improvement in Hauser's index and American Spinal Injury Association score was noted in all patients over a mean follow-up of 2.95 years. Mean injury duration was 3.42 years against the period of approximately 1-year required for natural recovery, suggesting a positive role of SCs. Co-infusion of N-Ad-MSC and HSC in CSF is safe and viable therapeutic approach for SCIs.

Ligament Tissue Engineering Using a Novel Porous Polycaprolactone Fumarate Scaffold and Adipose Tissue-Derived Mesenchymal Stem Cells Grown in Platelet Lysate.

PubMed

Wagner, Eric R; Bravo, Dalibel; Dadsetan, Mahrokh; Riester, Scott M; Chase, Steven; Westendorf, Jennifer J; Dietz, Allan B; van Wijnen, Andre J; Yaszemski, Michael J; Kakar, Sanjeev

2015-11-01

Surgical reconstruction of intra-articular ligament injuries is hampered by the poor regenerative potential of the tissue. We hypothesized that a novel composite polymer "neoligament" seeded with progenitor cells and growth factors would be effective in regenerating native ligamentous tissue. We synthesized a fumarate-derivative of polycaprolactone fumarate (PCLF) to create macro-porous scaffolds to allow cell-cell communication and nutrient flow. Clinical grade human adipose tissue-derived human mesenchymal stem cells (AMSCs) were cultured in 5% human platelet lysate (PL) and seeded on scaffolds using a dynamic bioreactor. Cell growth, viability, and differentiation were examined using metabolic assays and immunostaining for ligament-related markers (e.g., glycosaminoglycans [GAGs], alkaline phosphatase [ALP], collagens, and tenascin-C). AMSCs seeded on three-dimensional (3D) PCLF scaffolds remain viable for at least 2 weeks with proliferating cells filling the pores. AMSC proliferation rates increased in PL compared to fetal bovine serum (FBS) (p < 0.05). Cells had a low baseline expression of ALP and GAG, but increased expression of total collagen when induced by the ligament and tenogenic growth factor fibroblast growth factor 2 (FGF-2), especially when cultured in the presence of PL (p < 0.01) instead of FBS (p < 0.05). FGF-2 and PL also significantly increased immunostaining of tenascin-C and collagen at 2 and 4 weeks compared with human fibroblasts. Our results demonstrate that AMSCs proliferate and eventually produce a collagen-rich extracellular matrix on porous PCLF scaffolds. This novel scaffold has potential in stem cell engineering and ligament regeneration.

Ligament Tissue Engineering Using a Novel Porous Polycaprolactone Fumarate Scaffold and Adipose Tissue-Derived Mesenchymal Stem Cells Grown in Platelet Lysate

PubMed Central

Wagner, Eric R.; Bravo, Dalibel; Dadsetan, Mahrokh; Riester, Scott M.; Chase, Steven; Westendorf, Jennifer J.; Dietz, Allan B.; van Wijnen, Andre J.; Yaszemski, Michael J.

2015-01-01

Purpose: Surgical reconstruction of intra-articular ligament injuries is hampered by the poor regenerative potential of the tissue. We hypothesized that a novel composite polymer “neoligament” seeded with progenitor cells and growth factors would be effective in regenerating native ligamentous tissue. Methods: We synthesized a fumarate-derivative of polycaprolactone fumarate (PCLF) to create macro-porous scaffolds to allow cell–cell communication and nutrient flow. Clinical grade human adipose tissue-derived human mesenchymal stem cells (AMSCs) were cultured in 5% human platelet lysate (PL) and seeded on scaffolds using a dynamic bioreactor. Cell growth, viability, and differentiation were examined using metabolic assays and immunostaining for ligament-related markers (e.g., glycosaminoglycans [GAGs], alkaline phosphatase [ALP], collagens, and tenascin-C). Results: AMSCs seeded on three-dimensional (3D) PCLF scaffolds remain viable for at least 2 weeks with proliferating cells filling the pores. AMSC proliferation rates increased in PL compared to fetal bovine serum (FBS) (p < 0.05). Cells had a low baseline expression of ALP and GAG, but increased expression of total collagen when induced by the ligament and tenogenic growth factor fibroblast growth factor 2 (FGF-2), especially when cultured in the presence of PL (p < 0.01) instead of FBS (p < 0.05). FGF-2 and PL also significantly increased immunostaining of tenascin-C and collagen at 2 and 4 weeks compared with human fibroblasts. Summary: Our results demonstrate that AMSCs proliferate and eventually produce a collagen-rich extracellular matrix on porous PCLF scaffolds. This novel scaffold has potential in stem cell engineering and ligament regeneration. PMID:26413793

Safety and efficacy of allogeneic adipose tissue-derived mesenchymal stem cells for treatment of dogs with inflammatory bowel disease: Endoscopic and histological outcomes.

PubMed

Pérez-Merino, E M; Usón-Casaús, J M; Duque-Carrasco, J; Zaragoza-Bayle, C; Mariñas-Pardo, L; Hermida-Prieto, M; Vilafranca-Compte, M; Barrera-Chacón, R; Gualtieri, M

2015-12-01

Systemic administration of mesenchymal stem cells (MSCs) has been shown to be safe and efficacious in humans with Crohn's disease. The aim of this study was to evaluate the safety of an intravenous (IV) infusion of adipose tissue-derived mesenchymal stem cells (ASCs) and to assess macroscopic and histological effects in the digestive tract of dogs with inflammatory bowel disease (IBD). Eleven dogs with confirmed IBD received a single ASC infusion (2 × 10(6) cells/kg bodyweight). Full digestive endoscopic evaluation was performed pre-treatment and between 90 and 120 days post-treatment with mucosal changes being assessed using a fit-for-purpose endoscopic scale. Endoscopic biopsies from each digestive section were evaluated histologically according to the World Small Animal Veterinary Association (WSAVA) Gastrointestinal Standardization Group criteria. The pre- and post-treatment canine IBD endoscopic index (CIBDEI) and histological score (HS) were calculated and compared using the Wilcoxon test. Remission was defined as a reduction of >75% of the CIBDEI and HS compared with pre-treatment. No acute reactions to ASC infusion or side effects were reported in any dog. Significant differences between pre- and post-treatment were found in both the CIBDEI (P = 0.004) and HS (P = 0.004). Endoscopic remission occurred in 4/11 dogs with the remaining dogs showing decreased CIBDEI (44.8% to 73.3%). Histological remission was not achieved in any dog, with an average reduction of the pre-treatment HS of 27.2%. In conclusion, a single IV infusion of allogeneic ASCs improved gastrointestinal lesions as assessed macroscopically and slightly reduced gastrointestinal inflammation as evaluated by histopathology in dogs with IBD. Copyright © 2015 Elsevier Ltd. All rights reserved.

Adipose-Derived Mesenchymal Stem Cells in the Regeneration of Vocal Folds: A Study on a Chronic Vocal Fold Scar

PubMed Central

Vassiliki, Kalodimou; Irini, Messini; Nikolaos, Psychalakis; Karampela, Eleftheria; Apostolos, Papalois

2016-01-01

Background. The aim of the study was to assess the histological effects of autologous infusion of adipose-derived stem cells (ADSC) on a chronic vocal fold scar in a rabbit model as compared to an untreated scar as well as in injection of hyaluronic acid. Study Design. Animal experiment. Method. We used 74 New Zealand rabbits. Sixteen of them were used as control/normal group. We created a bilateral vocal fold wound in the remaining 58 rabbits. After 18 months we separated our population into three groups. The first group served as control/scarred group. The second one was injected with hyaluronic acid in the vocal folds, and the third received an autologous adipose-derived stem cell infusion in the scarred vocal folds (ADSC group). We measured the variation of thickness of the lamina propria of the vocal folds and analyzed histopathologic changes in each group after three months. Results. The thickness of the lamina propria was significantly reduced in the group that received the ADSC injection, as compared to the normal/scarred group. The collagen deposition, the hyaluronic acid, the elastin levels, and the organization of elastic fibers tend to return to normal after the injection of ADSC. Conclusions. Autologous injection of adipose-derived stem cells on a vocal fold chronic scar enhanced the healing of the vocal folds and the reduction of the scar tissue, even when compared to other treatments. PMID:26933440

Controlled, blinded force platform analysis of the effect of intraarticular injection of autologous adipose-derived mesenchymal stem cells associated to PRGF-Endoret in osteoarthritic dogs.

PubMed

Vilar, Jose M; Morales, Manuel; Santana, Angelo; Spinella, Giuseppe; Rubio, Mónica; Cuervo, Belen; Cugat, Ramón; Carrillo, Jose M

2013-07-02

Adipose-derived mesenchymal stem cell (ADMSC) therapy in regenerative medicine is a rapidly growing area of research and is currently also being used to treat osteoarthritis (OA). Force platform analysis has been consistently used to verify the efficacy of different therapeutic strategies for the treatment of OA in dogs, but never with AD-MSC.The aim of this study was to use a force platform to measure the efficacy of intraarticular ADMSC administration for limb function improvement in dogs with severe OA. Eight lame dogs with severe hip OA and a control group of 5 sound dogs were used for this study. Results were statistically analyzed to detect a significant increase in peak vertical force (PVF) and vertical impulse (VI) in treated dogs. Mean values of PVF and VI were significantly improved after treatment of the OA groups, reaching 53.02% and 14.84% of body weight, respectively, at day 180, compared with only 43.56% and 12.16% at day 0. This study objectively demonstrated that intraarticular ADMSC therapy resulted in reduced lameness due to OA.

Controlled, blinded force platform analysis of the effect of intraarticular injection of autologous adipose-derived mesenchymal stem cells associated to PRGF-Endoret in osteoarthritic dogs

PubMed Central

2013-01-01

Background Adipose-derived mesenchymal stem cell (ADMSC) therapy in regenerative medicine is a rapidly growing area of research and is currently also being used to treat osteoarthritis (OA). Force platform analysis has been consistently used to verify the efficacy of different therapeutic strategies for the treatment of OA in dogs, but never with AD-MSC. The aim of this study was to use a force platform to measure the efficacy of intraarticular ADMSC administration for limb function improvement in dogs with severe OA. Results Eight lame dogs with severe hip OA and a control group of 5 sound dogs were used for this study. Results were statistically analyzed to detect a significant increase in peak vertical force (PVF) and vertical impulse (VI) in treated dogs. Mean values of PVF and VI were significantly improved after treatment of the OA groups, reaching 53.02% and 14.84% of body weight, respectively, at day 180, compared with only 43.56% and 12.16% at day 0. Conclusion This study objectively demonstrated that intraarticular ADMSC therapy resulted in reduced lameness due to OA. PMID:23819757

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

Closure of a Recurrent Bronchopleural Fistula Using a Matrix Seeded With Patient-Derived Mesenchymal Stem Cells.

PubMed

Aho, Johnathon M; Dietz, Allan B; Radel, Darcie J; Butler, Greg W; Thomas, Mathew; Nelson, Timothy J; Carlsen, Brian T; Cassivi, Stephen D; Resch, Zachary T; Faubion, William A; Wigle, Dennis A

2016-10-01

: Management of recurrent bronchopleural fistula (BPF) after pneumonectomy remains a challenge. Although a variety of devices and techniques have been described, definitive management usually involves closure of the fistula tract through surgical intervention. Standard surgical approaches for BPF incur significant morbidity and mortality and are not reliably or uniformly successful. We describe the first-in-human application of an autologous mesenchymal stem cell (MSC)-seeded matrix graft to repair a multiply recurrent postpneumonectomy BPF. Adipose-derived MSCs were isolated from patient abdominal adipose tissue, expanded, and seeded onto bio-absorbable mesh, which was surgically implanted at the site of BPF. Clinical follow-up and postprocedural radiological and bronchoscopic imaging were performed to ensure BPF closure, and in vitro stemness characterization of patient-specific MSCs was performed. The patient remained clinically asymptomatic without evidence of recurrence on bronchoscopy at 3 months, computed tomographic imaging at 16 months, and clinical follow-up of 1.5 years. There is no evidence of malignant degeneration of MSC populations in situ, and the patient-derived MSCs were capable of differentiating into adipocytes, chondrocytes, and osteocytes using established protocols. Isolation and expansion of autologous MSCs derived from patients in a malnourished, deconditioned state is possible. Successful closure and safety data for this approach suggest the potential for an expanded study of the role of autologous MSCs in regenerative surgical applications for BPF. Bronchopleural fistula is a severe complication of pulmonary resection. Current management is not reliably successful. This work describes the first-in-human application of an autologous mesenchymal stem cell (MSC)-seeded matrix graft to the repair of a large, multiply recurrent postpneumonectomy BPF. Clinical follow-up of 1.5 years without recurrence suggests initial safety and feasibility of

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

Antagonizing Effects of Aspartic Acid against Ultraviolet A-Induced Downregulation of the Stemness of Human Adipose Tissue-Derived Mesenchymal Stem Cells

PubMed Central

Lee, Jienny; Shin, Seoung Woo; Jang, Sunghee; Jung, Eunsun; Kim, Min Hee; Lee, Jongsung

2015-01-01

Ultraviolet A (UVA) irradiation is responsible for a variety of changes in cell biology. The purpose of this study was to investigate effects of aspartic acid on UVA irradiation-induced damages in the stemness properties of human adipose tissue-derived mesenchymal stem cells (hAMSCs). Furthermore, we elucidated the UVA-antagonizing mechanisms of aspartic acid. The results of this study showed that aspartic acid attenuated the UVA-induced reduction of the proliferative potential and stemness of hAMSCs, as evidenced by increased proliferative activity in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and upregulation of stemness-related genes OCT4, NANOG, and SOX2 in response to the aspartic acid treatment. UVA-induced reduction in the mRNA level of hypoxia-inducible factor (HIF)-1α was also significantly recovered by aspartic acid. In addition, the antagonizing effects of aspartic acid against the UVA effects were found to be mediated by reduced production of PGE2 through the inhibition of JNK and p42/44 MAPK. Taken together, these findings show that aspartic acid improves reduced stemness of hAMSCs induced by UVA and its effects are mediated by upregulation of HIF-1α via the inhibition of PGE2-cAMP signaling. In addition, aspartic acid may be used as an antagonizing agent to mitigate the effects of UVA. PMID:25909857

Antagonizing Effects of Aspartic Acid against Ultraviolet A-Induced Downregulation of the Stemness of Human Adipose Tissue-Derived Mesenchymal Stem Cells.

PubMed

Jung, Kwangseon; Cho, Jae Youl; Soh, Young-Jin; Lee, Jienny; Shin, Seoung Woo; Jang, Sunghee; Jung, Eunsun; Kim, Min Hee; Lee, Jongsung

2015-01-01

Ultraviolet A (UVA) irradiation is responsible for a variety of changes in cell biology. The purpose of this study was to investigate effects of aspartic acid on UVA irradiation-induced damages in the stemness properties of human adipose tissue-derived mesenchymal stem cells (hAMSCs). Furthermore, we elucidated the UVA-antagonizing mechanisms of aspartic acid. The results of this study showed that aspartic acid attenuated the UVA-induced reduction of the proliferative potential and stemness of hAMSCs, as evidenced by increased proliferative activity in the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and upregulation of stemness-related genes OCT4, NANOG, and SOX2 in response to the aspartic acid treatment. UVA-induced reduction in the mRNA level of hypoxia-inducible factor (HIF)-1α was also significantly recovered by aspartic acid. In addition, the antagonizing effects of aspartic acid against the UVA effects were found to be mediated by reduced production of PGE2 through the inhibition of JNK and p42/44 MAPK. Taken together, these findings show that aspartic acid improves reduced stemness of hAMSCs induced by UVA and its effects are mediated by upregulation of HIF-1α via the inhibition of PGE2-cAMP signaling. In addition, aspartic acid may be used as an antagonizing agent to mitigate the effects of UVA.

Analysis of migration rate and chemotaxis of human adipose-derived mesenchymal stem cells in response to LPS and LTA in vitro

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

Herzmann, Nicole; Salamon, Achim; Fiedler, Tomas

Mesenchymal stem cells (MSC) are able to stimulate the regeneration of injured tissue. Since bacterial infections are common complications in wound healing, bacterial pathogens and their components come into direct contact with MSC. The interaction with bacterial structures influences the proliferation, differentiation and migratory activity of the MSC, which might be of relevance during regeneration. Studies on MSC migration in response to bacterial components have shown different results depending on the cell type. Here, we analyzed the migration rate and chemotaxis of human adipose-derived MSC (adMSC) in response to the basic cell-wall components lipopolysaccharide (LPS) of Gram-negative bacteria and lipoteichoicmore » acid (LTA) of Gram-positive bacteria in vitro. To this end, we used transwell and scratch assays, as well as a specific chemotaxis assay combined with live-cell imaging. We found no significant influence of LPS or LTA on the migration rate of adMSC in transwell or scratch assays. Furthermore, in the µ-slide chemotaxis assay, the stimulation with LPS did not exert any chemotactic effect on adMSC. - Highlights: • LPS increased the release of IL-6 and IL-8 in adMSC significantly. • The migration rate of adMSC was not influenced by LPS or LTA. • LPS or LTA did not exert a chemotactic effect on adMSC.« less

Cu, Zn-Superoxide Dismutase Increases the Therapeutic Potential of Adipose-derived Mesenchymal Stem Cells by Maintaining Antioxidant Enzyme Levels.

PubMed

Yoo, Dae Young; Kim, Dae Won; Chung, Jin Young; Jung, Hyo Young; Kim, Jong Whi; Yoon, Yeo Sung; Hwang, In Koo; Choi, Jung Hoon; Choi, Goang-Min; Choi, Soo Young; Moon, Seung Myung

2016-12-01

In the present study, we investigated the ability of Cu, Zn-superoxide dismutase (SOD1) to improve the therapeutic potential of adipose tissue-derived mesenchymal stem cells (Ad-MSCs) against ischemic damage in the spinal cord. Animals were divided into four groups: the control group, vehicle (PEP-1 peptide and artificial cerebrospinal fluid)-treated group, Ad-MSC alone group, and Ad-MSC-treated group with PEP-1-SOD1. The abdominal aorta of the rabbit was occluded for 30 min in the subrenal region to induce ischemic damage, and immediately after reperfusion, artificial cerebrospinal fluid or Ad-MSCs (2 × 10 5 ) were administered intrathecally. In addition, PEP-1 or 0.5 mg/kg PEP-1-SOD1 was administered intraperitoneally to the Ad-MSC-treated rabbits. Motor behaviors and NeuN-immunoreactive neurons were significantly decreased in the vehicle-treated group after ischemia/reperfusion. Administration of Ad-MSCs significantly ameliorated the changes in motor behavior and NeuN-immunoreactive neuronal survival. In addition, the combination of PEP-1-SOD1 and Ad-MSCs further increased the ameliorative effects of Ad-MSCs in the spinal cord after ischemia. Furthermore, the administration of Ad-MSCs with PEP-1-SOD1 decreased lipid peroxidation and maintained levels of antioxidants such as SOD1 and glutathione peroxidase compared to the Ad-MSC alone group. These results suggest that combination therapy using Ad-MSCs and PEP-1-SOD1 strongly protects neurons from ischemic damage by modulating the balance of lipid peroxidation and antioxidants.

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.

A Citrus bergamia Extract Decreases Adipogenesis and Increases Lipolysis by Modulating PPAR Levels in Mesenchymal Stem Cells from Human Adipose Tissue

PubMed Central

Lo Furno, Debora; Avola, Rosanna; Bonina, Francesco; Mannino, Giuliana

2016-01-01

The aim of this research was to assess the impact of a well-characterized extract from Citrus bergamia juice on adipogenesis and/or lipolysis using mesenchymal stem cells from human adipose tissue as a cell model. To evaluate the effects on adipogenesis, some cell cultures were treated with adipogenic medium plus 10 or 100 μg/mL of extract. To determine the properties on lipolysis, additional mesenchymal stem cells were cultured with adipogenic medium for 14 days and after this time added with Citrus bergamia for further 14 days. To verify adipogenic differentiation, oil red O staining at 7, 14, 21, and 28 days was performed. Moreover, the expression of peroxisome proliferator-activated receptor gamma (PPAR-γ), adipocytes fatty acid-binding protein (A-FABP), adipose triglyceride lipase (ATGL), hormone-sensitive lipase (HSL), monoglyceride lipase (MGL), 5′-adenosine monophosphate-activated protein kinase (AMPK)α1/2, and pAMPKα1/2 was evaluated by Western blot analysis and the release of glycerol by colorimetric assay. Citrus bergamia extract suppressed the accumulation of intracellular lipids in mesenchymal stem cells during adipogenic differentiation and promoted lipolysis by repressing the expression of adipogenic genes and activating lipolytic genes. Citrus bergamia extract could be a useful natural product for improving adipose mobilization in obesity-related disorders. PMID:27403151

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.

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.

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

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

Current progress in use of adipose derived stem cells in peripheral nerve regeneration

PubMed Central

Zack-Williams, Shomari DL; Butler, Peter E; Kalaskar, Deepak M

2015-01-01

Unlike central nervous system neurons; those in the peripheral nervous system have the potential for full regeneration after injury. Following injury, recovery is controlled by schwann cells which replicate and modulate the subsequent immune response. The level of nerve recovery is strongly linked to the severity of the initial injury despite the significant advancements in imaging and surgical techniques. Multiple experimental models have been used with varying successes to augment the natural regenerative processes which occur following nerve injury. Stem cell therapy in peripheral nerve injury may be an important future intervention to improve the best attainable clinical results. In particular adipose derived stem cells (ADSCs) are multipotent mesenchymal stem cells similar to bone marrow derived stem cells, which are thought to have neurotrophic properties and the ability to differentiate into multiple lineages. They are ubiquitous within adipose tissue; they can form many structures resembling the mature adult peripheral nervous system. Following early in vitro work; multiple small and large animal in vivo models have been used in conjunction with conduits, autografts and allografts to successfully bridge the peripheral nerve gap. Some of the ADSC related neuroprotective and regenerative properties have been elucidated however much work remains before a model can be used successfully in human peripheral nerve injury (PNI). This review aims to provide a detailed overview of progress made in the use of ADSC in PNI, with discussion on the role of a tissue engineered approach for PNI repair. PMID:25621105

"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

Protective effect of melatonin-supported adipose-derived mesenchymal stem cells against small bowel ischemia-reperfusion injury in rat.

PubMed

Chang, Chia-Lo; Sung, Pei-Hsun; Sun, Cheuk-Kwan; Chen, Chih-Hung; Chiang, Hsin-Ju; Huang, Tien-Hung; Chen, Yi-Ling; Zhen, Yen-Yi; Chai, Han-Tan; Chung, Sheng-Ying; Tong, Meng-Shen; Chang, Hsueh-Wen; Chen, Hong-Hwa; Yip, Hon-Kan

2015-09-01

We tested the hypothesis that combined melatonin and autologous adipose-derived mesenchymal stem cells (ADMSC) was superior to either alone against small bowel ischemia-reperfusion (SBIR) injury induced by superior mesenteric artery clamping for 30 min followed by reperfusion for 72 hr. Male adult Sprague Dawley rats (n = 50) were equally categorized into sham-operated controls SC, SBIR, SBIR-ADMSC (1.0 × 10(6) intravenous and 1.0 × 10(6) intrajejunal injection), SBIR-melatonin (intraperitoneal 20 mg/kg at 30 min after SI ischemia and 50 mg/kg at 6 and 18 hr after SI reperfusion), and SBIR-ADMSC-melatonin groups. The results demonstrated that the circulating levels of TNF-α, MPO, LyG6+ cells, CD68+ cells, WBC count, and gut permeability were highest in SBIR and lowest in SC, significantly higher in SBIR-ADMSC group and further increased in SBIR-melatonin group than in the combined therapy group (all P < 0.001). The ischemic mucosal damage score, the protein expressions of inflammation (TNF-α, NF-κB, MMP-9, MPO, and iNOS), oxidative stress (NOX-1, NOX-2, and oxidized protein), apoptosis (APAF-1, mitochondrial Bax, cleaved caspase-3 and PARP), mitochondrial damage (cytosolic cytochrome C) and DNA damage (γ-H2AX) markers, as well as cellular expressions of proliferation (PCNA), apoptosis (caspase-3, TUNEL assay), and DNA damage (γ-H2AX) showed an identical pattern, whereas mitochondrial cytochrome C exhibited an opposite pattern compared to that of inflammation among all groups (all P < 0.001). Besides, antioxidant expressions at protein (NQO-1, GR, and GPx) and cellular (HO-1) levels progressively increased from SC to the combined treatment group (all P < 0.001). In conclusion, combined melatonin-ADMSC treatment offered additive beneficial effect against SBIR injury. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Update on Controls for Isolation and Quantification Methodology of Extracellular Vesicles Derived from Adipose Tissue Mesenchymal Stem Cells

PubMed Central

Franquesa, Marcella; Hoogduijn, Martin J.; Ripoll, Elia; Luk, Franka; Salih, Mahdi; Betjes, Michiel G. H.; Torras, Juan; Baan, Carla C.; Grinyó, Josep M.; Merino, Ana Maria

2014-01-01

The research field on extracellular vesicles (EV) has rapidly expanded in recent years due to the therapeutic potential of EV. Adipose tissue human mesenchymal stem cells (ASC) may be a suitable source for therapeutic EV. A major limitation in the field is the lack of standardization of the challenging techniques to isolate and characterize EV. The aim of our study was to incorporate new controls for the detection and quantification of EV derived from ASC and to analyze the applicability and limitations of the available techniques. ASC were cultured in medium supplemented with 5% of vesicles-free fetal bovine serum. The EV were isolated from conditioned medium by differential centrifugation with size filtration (0.2 μm). As a control, non-conditioned culture medium was used (control medium). To detect EV, electron microscopy, conventional flow cytometry, and western blot were used. The quantification of the EV was by total protein quantification, ExoELISA immunoassay, and Nanosight. Cytokines and growth factors in the EV samples were measured by multiplex bead array kit. The EV were detected by electron microscope. Total protein measurement was not useful to quantify EV as the control medium showed similar protein contents as the EV samples. The ExoELISA kits had technical troubles and it was not possible to quantify the concentration of exosomes in the samples. The use of Nanosight enabled quantification and size determination of the EV. It is, however, not possible to distinguish protein aggregates from EV with this method. The technologies for quantification and characterization of the EV need to be improved. In addition, we detected protein contaminants in the EV samples, which make it difficult to determine the real effect of EV in experimental models. It will be crucial in the future to optimize design novel methods for purification and characterization of EV. PMID:25374572

Direct implantation versus platelet-rich fibrin-embedded adipose-derived mesenchymal stem cells in treating rat acute myocardial infarction.

PubMed

Sun, Cheuk-Kwan; Zhen, Yen-Yi; Leu, Steve; Tsai, Tzu-Hsien; Chang, Li-Teh; Sheu, Jiunn-Jye; Chen, Yung-Lung; Chua, Sarah; Chai, Han-Tan; Lu, Hung-I; Chang, Hsueh-Wen; Lee, Fan-Yen; Yip, Hon-Kan

2014-05-15

This study tested whether adipose-derived mesenchymal stem cells (ADMSC) embedded in platelet-rich fibrin (PRF) scaffold is superior to direct ADMSC implantation in improving left ventricular (LV) performance and reducing LV remodeling in a rat acute myocardial infarction (AMI) model of left anterior descending coronary artery (LAD) ligation. Twenty-eight male adult Sprague Dawley rats equally divided into group 1 [sham control], group 2 (AMI only), group 3 (AMI+direct ADMSC implantation), and group 4 (AMI+PRF-embedded autologous ADMSC) were sacrificed on day 42 after AMI. LV systolic and diastolic dimensions and volumes, and infarct/fibrotic areas were highest in group 2, lowest in group 1 and significantly higher in group 3 than in group 4, whereas LV performance and LV fractional shortening exhibited a reversed pattern (p<0.005). Protein expressions of inflammation (oxidative stress, IL-1β, MMP-9), apoptosis (mitochondrial Bax, cleaved PARP), fibrosis (Smad3, TGF-β), and pressure-overload biomarkers (BNP, MHC-β) displayed a pattern similar to that of LV dimensions, whereas anti-inflammatory (IL-10), anti-apoptotic (Bcl-2), and anti-fibrotic (Smad1/5, BMP-2) indices showed a pattern similar to that of LV performance among the four groups (all p<0.05). Angiogenesis biomarkers at protein (CXCR4, SDF-1α, VEGF), cellular (CD31+, CXCR4+, SDF-1α+), and immunohistochemical (small vessels) levels, and cardiac stem cell markers (C-kit+, Sca-1+) in infarct myocardium were highest in group 4, lowest in group 1, and significantly higher in group 3 than in group 2 (all p<0.005). PRF-embedded ADMSC is superior to direct ADMSC implantation in preserving LV function and attenuating LV remodeling. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Bone Marrow Adipose Tissue and Skeletal Health.

PubMed

Muruganandan, Shanmugam; Govindarajan, Rajgopal; Sinal, Christopher J

2018-05-31

To summarize and discuss recent progress and novel signaling mechanisms relevant to bone marrow adipocyte formation and its physiological/pathophysiological implications for bone remodeling. Skeletal remodeling is a coordinated process entailing removal of old bone and formation of new bone. Several bone loss disorders such as osteoporosis are commonly associated with increased bone marrow adipose tissue. Experimental and clinical evidence supports that a reduction in osteoblastogenesis from mesenchymal stem cells at the expense of adipogenesis, as well as the deleterious effects of adipocyte-derived signaling, contributes to the etiology of osteoporosis as well as bone loss associated with aging, diabetes mellitus, post-menopause, and chronic drug therapy. However, this view is challenged by findings indicating that, in some contexts, bone marrow adipose tissue may have a beneficial impact on skeletal health. Further research is needed to better define the role of marrow adipocytes in bone physiology/pathophysiology and to determine the therapeutic potential of manipulating mesenchymal stem cell differentiation.

[A preliminary study for the effect of nano hydroxyapatite on human adipose-derived mesenchymal stem cells mixture 3D bio-printing].

PubMed

Song, Y; Wang, X F; Wang, Y G; Dong, F; Lv, P J

2016-10-18

To study the effect of nano hydroxyapatite on human adipose-derived mesenchymal stem cells(hASCs) mixture 3D bio-printing for cells' proliferation and osteogenesis. P5 hASCs were used as seed cells, 10 g/L nano hydroxyapatite was added into the cell-sodium alginate-gelatin mixture (concentration: 20 g/L sodium alginate, 80 g/L gelatin; cell density: 1×10 6 /mL), then the mixture was printed by 3D bio-printer as the experimental group. And the cell-sodium alginate-gelatin mixture without nano hydroxyapatite was printed as the control group. Respectively, both the experimental and control groups were detected by microscope, CCK-8, Western blot and PCR at certain time pointsafter being printed, whose cells' proliferation and osteogenic differentiation were analyzed. The microscopic observation and CCK-8 results showed that the cells of the experimental group and the control group both had a good proliferation 24 h and 7 d after being printed. The Western blot results showed that 14 d after printing, the expression of Runt-related transcription factor 2 (RUNX2) had no statistical difference between the experimental group and control group. The PCR results showed that 14 d after printing, the expression of osteogenesis-related genes (RUNX2, osterix, and osteocalcin) was significantly higher in the experimental group than in the control group. Nano hydroxyapatite can increase osteogenic differentiation of the hASCs mixture after bio-printing, in which the cells still have a good proliferation.

Improved viability and activity of neutrophils differentiated from HL-60 cells by co-culture with adipose tissue-derived mesenchymal stem cells

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

Park, Yoon Shin; Lim, Goh-Woon; Cho, Kyung-Ah

Highlights: Black-Right-Pointing-Pointer Neutropenia is a principal complication of cancer treatment. Black-Right-Pointing-Pointer Co-culture of neutrophils with AD-MSC retained cell survival and proliferation and inhibited neutrophil apoptosis under serum starved conditions. Black-Right-Pointing-Pointer AD-MSC increased functions of neutrophil. Black-Right-Pointing-Pointer AD-MSC promoted the viability of neutrophils by enhancing respiratory burst through the expression of IFN-{alpha}, G-CSF, and TGF-{beta}. Black-Right-Pointing-Pointer AD-MSC can be used to improve immunity for neutropenia treatment. -- Abstract: Neutropenia is a principal complication of cancer treatment. We investigated the supportive effect of adipose tissue-derived mesenchymal stem cells (AD-MSCs) on the viability and function of neutrophils. Neutrophils were derived from HL-60 cellsmore » by dimethylformamide stimulation and cultured with or without AD-MSCs under serum-starved conditions to evaluate neutrophil survival, proliferation, and function. Serum starvation resulted in the apoptosis of neutrophils and decreased cell survival. The co-culture of neutrophils and AD-MSCs resulted in cell survival and inhibited neutrophil apoptosis under serum-starved conditions. The survival rate of neutrophils was prolonged up to 72 h, and the expression levels of interferon (IFN)-{alpha}, granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor, and transforming growth factor (TGF)-{beta} in AD-MSCs were increased after co-culture with neutrophils. AD-MSCs promoted the viability of neutrophils by inhibiting apoptosis as well as enhancing respiratory burst, which could potentially be mediated by the increased expression of IFN-{alpha}, G-CSF, and TGF-{beta}. Thus, we conclude that the use of AD-MSCs may be a promising cell-based therapy for increasing immunity by accelerating neutrophil function.« less

Articular cartilage-derived cells hold a strong osteogenic differentiation potential in comparison to mesenchymal stem cells in vitro

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

Salamon, Achim, E-mail: achim.salamon@med.uni-rostock.de; Jonitz-Heincke, Anika, E-mail: anika.jonitz@med.uni-rostock.de; Adam, Stefanie, E-mail: stefanie.adam@med.uni-rostock.de

Cartilaginous matrix-degenerative diseases like osteoarthritis (OA) are characterized by gradual cartilage erosion, and also by increased presence of cells with mesenchymal stem cell (MSC) character within the affected tissues. Moreover, primary chondrocytes long since are known to de-differentiate in vitro and to be chondrogenically re-differentiable. Since both findings appear to conflict with each other, we quantitatively assessed the mesenchymal differentiation potential of OA patient cartilage-derived cells (CDC) towards the osteogenic and adipogenic lineage in vitro and compared it to that of MSC isolated from adipose tissue (adMSC) of healthy donors. We analyzed expression of MSC markers CD29, CD44, CD105, andmore » CD166, and, following osteogenic and adipogenic induction in vitro, quantified their expression of osteogenic and adipogenic differentiation markers. Furthermore, CDC phenotype and proliferation were monitored. We found that CDC exhibit an MSC CD marker expression pattern similar to adMSC and a similar increase in proliferation rate during osteogenic differentiation. In contrast, the marked reduction of proliferation observed during adipogenic differentiation of adMSC was absent in CDC. Quantification of differentiation markers revealed a strong osteogenic differentiation potential for CDC, however almost no capacity for adipogenic differentiation. Since in the pathogenesis of OA, cartilage degeneration coincides with high bone turnover rates, the high osteogenic differentiation potential of OA patient-derived CDC may affect clinical therapeutic regimens aiming at autologous cartilage regeneration in these patients. - Highlights: • We analyze the mesenchymal differentiation capacity of cartilage-derived cells (CDC). • CDC express mesenchymal stem cell (MSC) markers CD29, CD44, CD105, and CD166. • CDC and MSC proliferation is reduced in adipogenesis and increased in osteogenesis. • Adipogenic differentiation is virtually absent in CDC

Monolayered mesenchymal stem cells repair scarred myocardium after myocardial infarction.

PubMed

Miyahara, Yoshinori; Nagaya, Noritoshi; Kataoka, Masaharu; Yanagawa, Bobby; Tanaka, Koichi; Hao, Hiroyuki; Ishino, Kozo; Ishida, Hideyuki; Shimizu, Tatsuya; Kangawa, Kenji; Sano, Shunji; Okano, Teruo; Kitamura, Soichiro; Mori, Hidezo

2006-04-01

Mesenchymal stem cells are multipotent cells that can differentiate into cardiomyocytes and vascular endothelial cells. Here we show, using cell sheet technology, that monolayered mesenchymal stem cells have multipotent and self-propagating properties after transplantation into infarcted rat hearts. We cultured adipose tissue-derived mesenchymal stem cells characterized by flow cytometry using temperature-responsive culture dishes. Four weeks after coronary ligation, we transplanted the monolayered mesenchymal stem cells onto the scarred myocardium. After transplantation, the engrafted sheet gradually grew to form a thick stratum that included newly formed vessels, undifferentiated cells and few cardiomyocytes. The mesenchymal stem cell sheet also acted through paracrine pathways to trigger angiogenesis. Unlike a fibroblast cell sheet, the monolayered mesenchymal stem cells reversed wall thinning in the scar area and improved cardiac function in rats with myocardial infarction. Thus, transplantation of monolayered mesenchymal stem cells may be a new therapeutic strategy for cardiac tissue regeneration.

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

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.

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

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

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.

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

Morphology and contractile gene expression of adipose-derived mesenchymal stem cells in response to short-term cyclic uniaxial strain and TGF-β1.

PubMed

Rashidi, Neda; Tafazzoli-Shadpour, Mohammad; Haghighipour, Nooshin; Khani, Mohammad-Mehdi

2018-06-27

Previous studies have shown smooth muscle induction in adipose-derived mesenchymal stem cells (ASCs) caused by long-term cyclic stretch. Here we examined the capability of the short-term straining with time steps of 4, 8, 16 and 24 h alone or combined with TGF-β1 on smooth muscle induction of rabbit ASCs. Alterations in cell morphology were quantified through the cell shape index and orientation angle, and expression levels of α-SMA, SM22-α, h-caldesmon and calponin3 markers were examined using the real-time polymerase chain reaction (PCR) method. Moreover, F-actin cytoskeleton organization was observed by fluorescence staining. Mechanical strain either alone or combined with growth factor treatment caused significant up-regulation of both early and intermediate smooth muscle cells (SMCs) specific markers during the initial hours of stimulation peaking in 8 to 16 h. Furthermore, gradual alignment of cells perpendicular to the strain direction during loading time, and cell elongation resembling contractile SMC phenotype, together with alignment and reorganization of F-actin fibers were observed. Considering previously reported protein up-regulation in following days of straining, the effects of short-term cyclic stretch on smooth muscle induction of ASCs were revealed which can be helpful in achieving functional contractile SMCs through synergistic mechano-chemical regulation of ASCs as an appealing cell source for vascular tissue engineering.

Cytotoxic Effects of Nonionic Iodinated Contrast Agent on Human Adipose-derived Mesenchymal Stem Cells.

PubMed

Wu, Tao; Nie, Hai; Dietz, Allan B; Salek, David R; Smith, Jay; van Wijnen, Andre J; Qu, Wenchun

2018-05-31

Transplantation of mesenchymal stem cells (MSCs) is a promising therapy for degenerative spine conditions. However, cell therapy for painful spine degeneration presently requires use of contrast agents during fluoroscopy-guided injections and the effects of these agents on MSCs represents a gap in knowledge. To investigate the biological effects of contrast media that are co-injected with MSCs. Prospective observational study. Academic medical center. Patient-derived clinical-grade culture expanded MSCs. Iohexol(Omnipaque300) was reduced to 12.5%, 25%, 50% and 100% of the stock solution and incubated with MSCs for 30 minutes, 4 hours and 48 hours. We also used complete media and 12.5%, 25%, 50%, 100% of phosphate buffered saline (PBS) as control group. We examined cytotoxicity of Iohexol at different concentrations and exposure duration, as well as the potential for recovery over time. Cell counts, mitochondrial activity, and quantitative real time reverse-transcriptase polymerase chain reaction (qRT-PCR) of related genes were analyzed immediately after exposure (day 0) and after two days of exposure (day 2). Human MSCs exhibit a time- and concentration-dependent cytotoxic response to iodinated CM. A brief 30min exposure did not affect MSCs function and viability. However, extended treatment with iohexol for 4 hours at 50% or higher concentration had a significant impact on both viability and gene expression in MSCs. CM (Omnipaque300) is cytotoxic to MSCs in a time-and concentration-dependent manner. Hence, the concentration of CM that accompanies MSC injections should be carefully considered during mesenchymal stem cell therapy for disc degenerative diseases. Copyright © 2018 American Academy of Physical Medicine and Rehabilitation. Published by Elsevier Inc. All rights reserved.

Intravenous adipose-derived mesenchymal stem cell therapy for the treatment of feline asthma: a pilot study.

PubMed

Trzil, Julie E; Masseau, Isabelle; Webb, Tracy L; Chang, Chee-Hoon; Dodam, John R; Liu, Hong; Quimby, Jessica M; Dow, Steven W; Reinero, Carol R

2016-12-01

The aim of this study was to evaluate the feasibility and efficacy of serially administered adipose-derived mesenchymal stem cells (MSCs) in an experimental feline asthma model. Allergic asthma was acutely induced with Bermuda grass allergen in six purpose-bred cats. Five intravenous infusions of allogeneic MSCs (n = 4; MSC-treated) or saline (n = 2; placebo-treated) were administered over the first 130 days after asthma induction. Infusions contained 2 × 10 6 , 4 × 10 6 , 4.7 × 10 6 , 1 × 10 7 and 1 × 10 7 cryopreserved MSCs/cat. For thoracic imaging additional cats were enrolled as control groups: four untreated, experimentally asthmatic cats (combined with placebo-treated cats), and six healthy, non-asthmatic cats. Outcome measures included airway eosinophilia, pulmonary mechanics, thoracic computed tomography and several immunologic assays. Cats were assessed for 9 months after treatment. At early points, airway eosinophil percentage was not affected by MSC administration (post-treatment average of days 12, 26, 47, 108 and 133 in MSC-treated cats was 41 ± 15% and in placebo-treated cats it was 34 ± 16%). By month 9, eosinophil percentages in all MSC-treated cats decreased to normal reference intervals (MSC-treated 6%; placebo-treated 20%; normal <17%). Diminished airway hyper-responsiveness was noted in all MSC-treated compared with placebo-treated cats at day 133 (dose of methacholine to double baseline airway resistance: MSC-treated median 22.9 mg/ml [range 6.4-64.0]; individual placebo-treated cats 1.1 and 5.0 mg/ml). Lung attenuation (mean ± SEM MSC-treated -865 ± 12 Hounsfield units [HU]; untreated asthmatics -820 ± 11 HU; P = 0.004) and bronchial wall thickening scores (median [interquartile range] MSC-treated 0 [0-1.5]; untreated asthmatic 11.6 [7.3-27.3]; P = 0.010) were significantly reduced in MSC-treated vs untreated asthmatic cats, consistent with decreased airway remodeling at month 9. No clear immunologic mechanisms by which MSCs act

Matrilin-3 codelivery with adipose-derived mesenchymal stem cells promotes articular cartilage regeneration in a rat osteochondral defect model.

PubMed

Muttigi, Manjunatha S; Kim, Byoung Ju; Choi, Bogyu; Yoshie, Arai; Kumar, Hemant; Han, Inbo; Park, Hansoo; Lee, Soo-Hong

2018-03-01

Matrilin-3 is an essential extracellular matrix component present only in cartilaginous tissues. Matrilin-3 exerts chondroprotective effects by regulating an anti-inflammatory function and extracellular matrix components. We hypothesized that the codelivery of matrilin-3 with infrapatellar adipose-tissue-derived mesenchymal stem cells (Ad-MSCs) may enhance articular cartilage regeneration. Matrilin-3 treatment of Ad-MSCs in serum-free media induced collagen II and aggrecan expression, and matrilin-3 in chondrogenic media also enhanced in vitro chondrogenic differentiation. Next, the in vivo effect of matrilin-3 codelivery with Ad-MSCs on cartilage regeneration was assessed in an osteochondral defect model in Sprague Dawley rats: Ad-MSCs and hyaluronic acid were implanted at the defect site with or without matrilin-3 (140, 280, and 700 ng). Safranin O staining revealed that matrilin-3 (140 and 280 ng) treatment significantly improved cartilage regeneration and glycosaminoglycan accumulation. In the animals treated with 140-ng matrilin-3, in particular, the defect site exhibited complete integration with surrounding tissue and a smooth glistening surface. The International Cartilage Repair Society macroscopic and O'Driscoll microscopic scores for regenerated cartilage were furthermore shown to be considerably higher for this group (matrilin-3; 140 ng) compared with the other groups. Furthermore, the defects treated with 140-ng matrilin-3 revealed significant hyaline-like cartilage regeneration in the osteochondral defect model; in contrast, the defects treated with 700-ng matrilin-3 exhibited drastically reduced cartilage regeneration with mixed hyaline-fibrocartilage morphology. Codelivery of matrilin-3 with Ad-MSCs significantly influenced articular cartilage regeneration, supporting the potential use of this tissue-specific protein for a cartilage-targeted stem cell therapy. Copyright © 2017 John Wiley & Sons, Ltd.

Adipose derived mesenchymal stem cells partially rescue mitomycin C treated ARPE19 cells from death in co-culture condition.

PubMed

Singh, Amar K; Srivastava, Girish K; García-Gutiérrez, María T; Pastor, J Carlos

2013-12-01

Age-related macular degeneration is a retinal disease with important damage at the RPE layer. This layer is considered a target for therapeutical approaches. Stem cell transplantation is a promising option for retinal diseases. Adipose derived mesenchymal stem cells secret growth factors which might play a significant role in RPE maintenance. This study aimed to evaluate human AD-MSCs ability to rescue mitomycin C treated dying ARPE19 cells in co-culture condition. ARPE19 cells were treated with MMC (50 μg/ml, 100 μg/ml and 200 μg/ml) for 2 hours to induce cell death. These treated cells were co-cultured with hAD-MSCs in indirect co-culture system for 3 days and 3 weeks. Then the viability, growth and proliferation of these ARPE19 cells were evaluated by a cell viability/cytotoxicity assay kit and Alamar Blue (AB) assay. Untreated ARPE19 cells and human skin fibroblasts (HSF) were used as controls. MMC blocked ARPE19 cell proliferation significantly in 3 days and cells were almost completely dead after 3 weeks. Cell toxicity of MMC increased significantly with concentration. When these cells were co-cultured with hAD-MSCs, a significant growth difference was observed in treated cells compared to untreated cells. hAD-MSCs rescue capacity was also significantly higher than HSF for treated ARPE19 cells. This study showed that hAD-MSCs rescued MMC treated ARPE19 cells from death. It probably occurred due to undefined growth factors secreted by hAD-MSCs in the medium, shared by treated ARPE19 cells in co-culture conditions. This study supports further evaluation of the effect of hAD-MSCs subretinal transplantation over the RPE degeneration process in AMD patients.

Health Span-Extending Activity of Human Amniotic Membrane- and Adipose Tissue-Derived Stem Cells in F344 Rats

PubMed Central

Kim, Dajeong; Kyung, Jangbeen; Park, Dongsun; Choi, Ehn-Kyoung; Kim, Kwang Sei; Shin, Kyungha; Lee, Hangyoung; Shin, Il Seob; Kang, Sung Keun

2015-01-01

Aging brings about the progressive decline in cognitive function and physical activity, along with losses of stem cell population and function. Although transplantation of muscle-derived stem/progenitor cells extended the health span and life span of progeria mice, such effects in normal animals were not confirmed. Human amniotic membrane-derived mesenchymal stem cells (AMMSCs) or adipose tissue-derived mesenchymal stem cells (ADMSCs) (1 × 106 cells per rat) were intravenously transplanted to 10-month-old male F344 rats once a month throughout their lives. Transplantation of AMMSCs and ADMSCs improved cognitive and physical functions of naturally aging rats, extending life span by 23.4% and 31.3%, respectively. The stem cell therapy increased the concentration of acetylcholine and recovered neurotrophic factors in the brain and muscles, leading to restoration of microtubule-associated protein 2, cholinergic and dopaminergic nervous systems, microvessels, muscle mass, and antioxidative capacity. The results indicate that repeated transplantation of AMMSCs and ADMSCs elongate both health span and life span, which could be a starting point for antiaging or rejuvenation effects of allogeneic or autologous stem cells with minimum immune rejection. Significance This study demonstrates that repeated treatment with stem cells in normal animals has antiaging potential, extending health span and life span. Because antiaging and prolonged life span are issues currently of interest, these results are significant for readers and investigators. PMID:26315571

Health Span-Extending Activity of Human Amniotic Membrane- and Adipose Tissue-Derived Stem Cells in F344 Rats.

PubMed

Kim, Dajeong; Kyung, Jangbeen; Park, Dongsun; Choi, Ehn-Kyoung; Kim, Kwang Sei; Shin, Kyungha; Lee, Hangyoung; Shin, Il Seob; Kang, Sung Keun; Ra, Jeong Chan; Kim, Yun-Bae

2015-10-01

Aging brings about the progressive decline in cognitive function and physical activity, along with losses of stem cell population and function. Although transplantation of muscle-derived stem/progenitor cells extended the health span and life span of progeria mice, such effects in normal animals were not confirmed. Human amniotic membrane-derived mesenchymal stem cells (AMMSCs) or adipose tissue-derived mesenchymal stem cells (ADMSCs) (1×10(6) cells per rat) were intravenously transplanted to 10-month-old male F344 rats once a month throughout their lives. Transplantation of AMMSCs and ADMSCs improved cognitive and physical functions of naturally aging rats, extending life span by 23.4% and 31.3%, respectively. The stem cell therapy increased the concentration of acetylcholine and recovered neurotrophic factors in the brain and muscles, leading to restoration of microtubule-associated protein 2, cholinergic and dopaminergic nervous systems, microvessels, muscle mass, and antioxidative capacity. The results indicate that repeated transplantation of AMMSCs and ADMSCs elongate both health span and life span, which could be a starting point for antiaging or rejuvenation effects of allogeneic or autologous stem cells with minimum immune rejection. This study demonstrates that repeated treatment with stem cells in normal animals has antiaging potential, extending health span and life span. Because antiaging and prolonged life span are issues currently of interest, these results are significant for readers and investigators. ©AlphaMed Press.

[Multifunctional nano-vector for gene delivery into human adipose derived mesenchymal stem cells and in vitro cellular magnetic resonance imaging].

PubMed

Pang, Pengfei; Li, Bing; Hu, Xiaojun; Kang, Zhuang; Guan, Shouhai; Gong, Faming; Meng, Xiaochun; Li, Dan; Huang, Mingsheng; Shan, Hong

2014-04-08

To examine the feasibility and efficacy of using superparamagnetic iron oxide nanoparticles coated with polyethylene glycol-grafted polyethylenimine (PEG-g-PEI-SPION) as a carrier for gene delivery into human adipose derived mesenchymal stem cells (hADMSCs) and in vitro cellular magnetic resonance imaging (MRI). PEG-g-PEI-SPION was synthesized as previously reported. Gel electrophoresis was performed to assess the pDNA condensation capacity of PEG-g-PEI-SPION. The particle size and zeta potential of PEG-g-PEI-SPION/pDNA complexes were determined by dynamic light scattering. Cytotoxicity of PEG-g-PEI-SPION was evaluated by CCK-8 assay with hADMSCs. Gene transfection efficiency of PEG-g-PEI-SPION in hADMSCs was quantified by flow cytometry. The cellular internalization of PEG-g-PEI-SPION/pDNA nanocomplexes was studied by confocal laser scanning microscopy and Prussian blue staining. MRI function of PEG-g-PEI-SPION was studied by in vitro cellular MRI scanning. PEG-g-PEI-SPION condensed pDNA to form stable complexes of 80-100 nm in diameter and showed low cytotoxicity in hADMSCs. At the optimal N/P ratio of 20, PEG-g-PEI-SPION/pDNA obtained the highest transfection efficiency of 22.8% ± 3.6% in hADMSCs. And it was higher than that obtained with lipofectamine 11.2% ± 2.6% (P < 0.05). Furthermore, hADMSCs labeled with PEG-g-PEI-SPION showed sensitive low signal intensity on MRI T2-weighted images in vitro. PEG-g-PEI-SPION is an efficient and MRI-visible nano-vector for gene delivery into hADMSCs.

Tracing the destiny of mesenchymal stem cells from embryo to adult bone marrow and white adipose tissue via Pdgfrα expression.

PubMed

Miwa, Hiroyuki; Era, Takumi

2018-01-29

Mesenchymal stem cells (MSCs) are somatic stem cells that can be derived from adult bone marrow (BM) and white adipose tissue (WAT), and that display multipotency and self-renewal capacity. Although MSCs are essential for tissue formation and have already been used in clinical therapy, the origins and markers of these cells remain unknown. In this study, we first investigated the developmental process of MSCs in mouse embryos using the gene encoding platelet-derived growth factor receptor α ( Pdgfra ) as a marker. We then traced cells expressing Pdgfra and other genes (brachyury, Sox1 and Pmx1 ) in various mutant mouse embryos until the adult stage. This tracing of MSC origins and destinies indicates that embryonic MSCs emerge in waves and that almost all adult BM MSCs and WAT MSCs originate from mesoderm and embryonic Pdgfrα-positive cells. Furthermore, we demonstrate that adult Pdgfrα-positive cells are involved in some pathological conditions. © 2018. Published by The Company of Biologists Ltd.

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.

Uremic Toxins Affect the Imbalance of Redox State and Overexpression of Prolyl Hydroxylase 2 in Human Adipose Tissue-Derived Mesenchymal Stem Cells Involved in Wound Healing.

PubMed

Khanh, Vuong Cat; Ohneda, Kinuko; Kato, Toshiki; Yamashita, Toshiharu; Sato, Fujio; Tachi, Kana; Ohneda, Osamu

2017-07-01

Chronic kidney disease (CKD) results in a delay in wound healing because of its complications such as uremia, anemia, and fluid overload. Mesenchymal stem cells (MSCs) are considered to be a candidate for wound healing because of the ability to recruit many types of cells. However, it is still unclear whether the CKD-adipose tissue-derived MSCs (CKD-AT-MSCs) have the same function in wound healing as healthy donor-derived normal AT-MSCs (nAT-MSCs). In this study, we found that uremic toxins induced elevated reactive oxygen species (ROS) expression in nAT-MSCs, resulting in the reduced expression of hypoxia-inducible factor-1α (HIF-1α) under hypoxic conditions. Consistent with the uremic-treated AT-MSCs, there was a definite imbalance of redox state and high expression of ROS in CKD-AT-MSCs isolated from early-stage CKD patients. In addition, a transplantation study clearly revealed that nAT-MSCs promoted the recruitment of inflammatory cells and recovery from ischemia in the mouse flap model, whereas CKD-AT-MSCs had defective functions and the wound healing process was delayed. Of note, the expression of prolyl hydroxylase domain 2 (PHD2) is selectively increased in CKD-AT-MSCs and its inhibition can restore the expression of HIF-1α and the wound healing function of CKD-AT-MSCs. These results indicate that more studies about the functions of MSCs from CKD patients are required before they can be applied in the clinical setting.

Vitamin D: Correlation with biochemical and body composition changes in a southern Brazilian population and induction of cytotoxicity in mesenchymal stem cells derived from human adipose tissue.

PubMed

Pesarini, João Renato; Oliveira, Rodrigo Juliano; Pessatto, Lucas Roberto; Antoniolli-Silva, Andréia Conceição Milan Brochado; Felicidade, Ingrid; Nardi, Nance Beyer; Camassola, Melissa; Mantovani, Mário Sérgio; Ribeiro, Lúcia Regina

2017-07-01

Studies have shown that metabolic disorders, serum inflammatory markers and weight gain (obesity) are correlated with vitamin D deficiency. Therefore, the present study correlated the serum calcidiol (s25(OH)D 3 ) levels in a sample of individuals from southern Brazil with variables related to metabolic disorders, obesity and lifestyle habits and assessed the cytotoxic effect of calcitriol on adipose tissue-derived mesenchymal stem cells (ADSCs). The results showed a 79.23% prevalence of hypovitaminosis D in the study population and a correlation (p<0.05) between a low serum vitamin D concentration and an elevated low-density lipoprotein cholesterol (LDL-c) level. Univariate linear regression analysis using 25(OH)D 3 as a regressor showed a negative association (p<0.05) with an indoor work environment (β=-2.305), increased body fat (β=-0.095), age (β=-0.065) and high-density lipoprotein cholesterol (HDL-c; β=-0.109). An in vitro 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay performed with ADSCs using five calcitriol concentrations (15.625, 31.25, 62.5, 125 and 250nM) indicated cytotoxic potential (p<0.05) at the 62.5nM concentration at 48 and 72h and at the 125 and 250nM concentrations at 24, 48 and 72h. The results reported herein corroborate one another and suggest a key association between vitamin D deficiency and the development of obesity because ADSCs are involved in adipose tissue hyperplasia and differentiate into adipocytes that can sequester the bioavailable vitamin D necessary for homeostasis. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Behaviour of adipose-derived canine mesenchymal stem cells after superparamagnetic iron oxide nanoparticles labelling for magnetic resonance imaging.

PubMed

Kolecka, Malgorzata Anna; Arnhold, Stefan; Schmidt, Martin; Reich, Christine; Kramer, Martin; Failing, Klaus; von Pückler, Kerstin

2017-02-24

Therapy with mesenchymal stem cells (MSCs) has been reported to provide beneficial effects in the treatment of neurological and orthopaedic disorders in dogs. The exact mechanism of action is poorly understood. Magnetic resonance imaging (MRI) gives the opportunity to observe MSCs after clinical administration. To visualise MSCs with the help of MRI, labelling with an MRI contrast agent is necessary. However, it must be clarified whether there is any negative influence on cell function and viability after labelling prior to clinical administration. For the purpose of the study, seven samples with canine adipose-derived stem cells were incubated with superparamagnetic iron oxide nanoparticles (SPIO: 319.2 μg/mL Fe) for 24 h. The internalisation of the iron particles occurred via endocytosis. SPIO particles were localized as free clusters in the cytoplasm or within lysosomes depending on the time of investigation. The efficiency of the labelling was investigated using Prussian blue staining and MACS assay. After 3 weeks the percentage of SPIO labelled canine stem cells decreased. Phalloidin staining showed no negative effect on the cytoskeleton. Labelled cells underwent osteogenic and adipogenic differentiation. Chondrogenic differentiation occurred to a lesser extent compared with a control sample. MTT-Test and wound healing assay showed no influence of labelling on the proliferation. The duration of SPIO labelling was assessed using a 1 Tesla clinical MRI scanner and T2 weighted turbo spin echo and T2 weighted gradient echo MRI sequences 1, 2 and 3 weeks after labelling. The hypointensity caused by SPIO lasted for 3 weeks in both sequences. An Endorem labelling concentration of 319.2 μg/mL Fe (448 μg/mL SPIO) had no adverse effects on the viability of canine ASCs. Therefore, this contrast agent could be used as a model for iron oxide labelling agents. However, the tracking ability in vivo has to be evaluated in further studies.

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

Isolation of adipose derived stem cells and their induction to a chondrogenic phenotype

PubMed Central

Estes, Bradley T.; Diekman, Brian O.; Gimble, Jeffrey M.; Guilak, Farshid

2011-01-01

Summary The ability to isolate, expand, and differentiate adult stem cells into a chondrogenic lineage is an important step in the development of tissue engineering approaches for cartilage repair or regeneration for the treatment of joint injury or osteoarthritis, or for application in plastic or reconstructive surgery. Adipose-derived stem cells (ASCs) provide an abundant and easily accessible source of adult stem cells for use in such regenerative approaches. This protocol describes the isolation of ASCs from liposuction aspirate, as well as cell culture conditions for growth factor based induction of ASCs into chondrocyte-like cells. These methods are similar to those used for bone marrow mesenchymal stem cells but distinct due to the unique properties of ASCs. Investigators can expect consistent ASC differentiation, allowing for slight variation due to donor and serum lot effects. Approximately 10–12 weeks are needed for ASC isolation and the characterization of chondrocyte-like cells, which is also described. PMID:20595958

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.

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.

Effect of Uniaxial Tensile Cyclic Loading Regimes on Matrix Organization and Tenogenic Differentiation of Adipose-Derived Stem Cells Encapsulated within 3D Collagen Scaffolds

PubMed Central

Stasuk, Alexander

2017-01-01

Adipose-derived mesenchymal stem cells have become a popular cell choice for tendon repair strategies due to their relative abundance, ease of isolation, and ability to differentiate into tenocytes. In this study, we investigated the solo effect of different uniaxial tensile strains and loading frequencies on the matrix directionality and tenogenic differentiation of adipose-derived stem cells encapsulated within three-dimensional collagen scaffolds. Samples loaded at 0%, 2%, 4%, and 6% strains and 0.1 Hz and 1 Hz frequencies for 2 hours/day over a 7-day period using a custom-built uniaxial tensile strain bioreactor were characterized in terms of matrix organization, cell viability, and musculoskeletal gene expression profiles. The results displayed that the collagen fibers of the loaded samples exhibited increased matrix directionality with an increase in strain values. Gene expression analyses demonstrated that ASC-encapsulated collagen scaffolds loaded at 2% strain and 0.1 Hz frequency showed significant increases in extracellular matrix genes and tenogenic differentiation markers. Importantly, no cross-differentiation potential to osteogenic, chondrogenic, and myogenic lineages was observed at 2% strain and 0.1 Hz frequency loading condition. Thus, 2% strain and 0.1 Hz frequency were identified as the appropriate mechanical loading regime to induce tenogenic differentiation of adipose-derived stem cells cultured in a three-dimensional environment. PMID:29375625

Skeletal tissue engineering using mesenchymal or embryonic stem cells: clinical and experimental data.

PubMed

Gamie, Zakareya; MacFarlane, Robert J; Tomkinson, Alicia; Moniakis, Alexandros; Tran, Gui Tong; Gamie, Yehya; Mantalaris, Athanasios; Tsiridis, Eleftherios

2014-11-01

Mesenchymal stem cells (MSCs) can be obtained from a wide variety of tissues for bone tissue engineering such as bone marrow, adipose, birth-associated, peripheral blood, periosteum, dental and muscle. MSCs from human fetal bone marrow and embryonic stem cells (ESCs) are also promising cell sources. In vitro, in vivo and clinical evidence was collected using MEDLINE® (1950 to January 2014), EMBASE (1980 to January 2014) and Google Scholar (1980 to January 2014) databases. Enhanced results have been found when combining bone marrow-derived mesenchymal stem cells (BMMSCs) with recently developed scaffolds such as glass ceramics and starch-based polymeric scaffolds. Preclinical studies investigating adipose tissue-derived stem cells and umbilical cord tissue-derived stem cells suggest that they are likely to become promising alternatives. Stem cells derived from periosteum and dental tissues such as the periodontal ligament have an osteogenic potential similar to BMMSCs. Stem cells from human fetal bone marrow have demonstrated superior proliferation and osteogenic differentiation than perinatal and postnatal tissues. Despite ethical concerns and potential for teratoma formation, developments have also been made for the use of ESCs in terms of culture and ideal scaffold.

Validation of a novel animal model for sciatic nerve repair with an adipose-derived stem cell loaded fibrin conduit.

PubMed

Saller, Maximilian M; Huettl, Rosa-Eva; Mayer, Julius M; Feuchtinger, Annette; Krug, Christian; Holzbach, Thomas; Volkmer, Elias

2018-05-01

Despite the regenerative capabilities of peripheral nerves, severe injuries or neuronal trauma of critical size impose immense hurdles for proper restoration of neuro-muscular circuitry. Autologous nerve grafts improve re-establishment of connectivity, but also comprise substantial donor site morbidity. We developed a rat model which allows the testing of different cell applications, i.e., mesenchymal stem cells, to improve nerve regeneration in vivo. To mimic inaccurate alignment of autologous nerve grafts with the injured nerve, a 20 mm portion of the sciatic nerve was excised, and sutured back in place in reversed direction. To validate the feasibility of our novel model, a fibrin gel conduit containing autologous undifferentiated adipose-derived stem cells was applied around the coaptation sites and compared to autologous nerve grafts. After evaluating sciatic nerve function for 16 weeks postoperatively, animals were sacrificed, and gastrocnemius muscle weight was determined along with morphological parameters (g-ratio, axon density & diameter) of regenerating axons. Interestingly, the addition of undifferentiated adipose-derived stem cells resulted in a significantly improved re-myelination, axon ingrowth and functional outcome, when compared to animals without a cell seeded conduit. The presented model thus displays several intriguing features: it imitates a certain mismatch in size, distribution and orientation of axons within the nerve coaptation site. The fibrin conduit itself allows for an easy application of cells and, as a true critical-size defect model, any observed improvement relates directly to the performed intervention. Since fibrin and adipose-derived stem cells have been approved for human applications, the technique can theoretically be performed on humans. Thus, we suggest that the model is a powerful tool to investigate cell mediated assistance of peripheral nerve regeneration.

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

Increasing of blastocyst rate and gene expression in co-culture of bovine embryos with adult adipose tissue-derived mesenchymal stem cells.

PubMed

Miranda, Moysés S; Nascimento, Hamilton S; Costa, Mayra P R; Costa, Nathália N; Brito, Karynne N L; Lopes, Cinthia T A; Santos, Simone S D; Cordeiro, Marcela S; Ohashi, Otávio M

2016-10-01

Despite advances in the composition of defined embryo culture media, co-culture with somatic cells is still used for bovine in vitro embryo production (IVEP) in many laboratories worldwide. Granulosa cells are most often used for this purpose, although recent work suggests that co-culture with stem cells of adult or embryonic origin or their derived biomaterials may improve mouse, cattle, and pig embryo development. In experiment 1, in vitro produced bovine embryos were co-cultured in the presence of two concentrations of bovine adipose tissue-derived mesenchymal cells (b-ATMSCs; 10 3 and 10 4 cells/mL), in b-ATMSC preconditioned medium (SOF-Cond), or SOF alone (control). In experiment 2, co-culture with 10 4 b-ATMSCs/mL was compared to the traditional granulosa cell co-culture system (Gran). In experiment 1, co-culture with 10 4 b-ATMSCs/mL improved blastocyst rates in comparison to conditioned and control media (p < 0.05). Despite that it did not show difference with 10 3 b-ATMSCs/mL (p = 0.051), group 10 4 b-ATMSCs/mL yielded higher results of blastocyst production. In experiment 2, when compared to group Gran, co-culture with 10 4 b-ATMSCs/mL improved not only blastocyst rates but also quality as assessed by increased total cell numbers and mRNA expression levels for POU5F1 and G6PDH (p < 0.05). Co-culture of bovine embryos with b-ATMSCs was more beneficial than the traditional co-culture system with granulosa cells. We speculate that the microenvironmental modulatory potential of MSCs, by means of soluble substances and exosome secretions, could be responsible for the positive effects observed. Further experiments must be done to evaluate if this beneficial effect in vitro also translates to an increase in offspring following embryo transfer. Moreover, this study provides an interesting platform to study the basic requirements during preimplantation embryo development, which, in turn, may aid the improvement of embryo culture protocols in bovine

[Osteogenesis of human adipose-derived mesenchymal stem cells-biomaterial mixture in vivo after 3D bio-printing].

PubMed

Song, Yang; Wang, Xiao-fei; Wang, Yu-guang; Sun, Yu-chun; Lv, Pei-jun

2016-02-18

To construct human adipose-derived mesenchymal stem cells (hASCs)-biomaterial mixture 3D bio-printing body and detect its osteogenesis in vivo, and to establish a guideline of osteogenesis in vivo by use of 3D bio-printing technology preliminarily. P4 hASCs were used as seed cells, whose osteogenic potential in vitro was tested by alkaline phosphatase (ALP) staining and alizarin red staining after 14 d of osteogenic induction. The cells were added into 20 g/L sodium alginate and 80 g/L gelatin mixture (cell density was 1 × 10(6)/mL), and the cell-sodium alginate-gelatin mixture was printed by Bioplotter 3D bio-printer (Envision company, Germany), in which the cells'survival rate was detected by live- dead cell double fluorescence staining. Next, the printing body was osteogenically induced for 1 week to gain the experimental group; and the sodium alginate-gelatin mixture without cells was also printed to gain the control group. Both the experimental group and the control group were implanted into the back of the nude mice. After 6 weeks of implantation, the samples were collected, HE staining, Masson staining, immunohistochemical staining and Inveon Micro CT test were preformed to analyze their osteogenic capability. The cells'survival rate was 89%± 2% after printing. Six weeks after implantation, the samples of the control group were mostly degraded, whose shape was irregular and gel-like; the samples of the experimental group kept their original size and their texture was tough. HE staining and Masson staining showed that the bone-like tissue and vessel in-growth could be observed in the experimental group 6 weeks after implantation, immunohistochemical staining showed that the result of osteocalcin was positive, and Micro CT results showed that samples of the experimental group had a higher density and the new bone volume was 18% ± 1%. hASCs -biomaterial mixture 3D bio-printing body has capability of ectopic bone formation in nude mice, and it is feasible to

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

Therapeutic Effect of Human Adipose Tissue-Derived Mesenchymal Stem Cells in Experimental Corneal Failure Due to Limbal Stem Cell Niche Damage.

PubMed

Galindo, Sara; Herreras, José M; López-Paniagua, Marina; Rey, Esther; de la Mata, Ana; Plata-Cordero, María; Calonge, Margarita; Nieto-Miguel, Teresa

2017-10-01

Limbal stem cells are responsible for the continuous renewal of the corneal epithelium. The destruction or dysfunction of these stem cells or their niche induces limbal stem cell deficiency (LSCD) leading to visual loss, chronic pain, and inflammation of the ocular surface. To restore the ocular surface in cases of bilateral LSCD, an extraocular source of stem cells is needed to avoid dependence on allogeneic limbal stem cells that are difficult to obtain, isolate, and culture. The aim of this work was to test the tolerance and the efficacy of human adipose tissue-derived mesenchymal stem cells (hAT-MSCs) to regenerate the ocular surface in two experimental models of LSCD that closely resemble different severity grades of the human pathology. hAT-MSCs transplanted to the ocular surface of the partial and total LSCD models developed in rabbits were well tolerated, migrated to inflamed tissues, reduced inflammation, and restrained the evolution of corneal neovascularization and corneal opacity. The expression profile of the corneal epithelial cell markers CK3 and E-cadherin, and the limbal epithelial cell markers CK15 and p63 was lost in the LSCD models, but was partially recovered after hAT-MSC transplantation. For the first time, we demonstrated that hAT-MSCs improve corneal and limbal epithelial phenotypes in animal LSCD models. These results support the potential use of hAT-MSCs as a novel treatment of ocular surface failure due to LSCD. hAT-MSCs represent an available, non-immunogenic source of stem cells that may provide therapeutic benefits in addition to reduce health care expenses. Stem Cells 2017;35:2160-2174. © 2017 AlphaMed Press.

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.

Differentiation of Mesenchymal Stem Cells Towards Nephrogenic Lineage and Their Enhanced Resistance to Oxygen Peroxide-induced Oxidative Stress.

PubMed

Tayyeb, Asima; Shahzad, Naveed; Ali, Gibran

2017-07-01

Mesenchymal stem cells (MSCs) have been publicized to ameliorate kidney injury both in vitro and in vivo. However, very less is known if MSCs can be differentiated towards renal lineages and their further application potential in kidney injuries. The present study developed a conditioning system of growth factors fibroblast growth factor 2, transforming growth factor-β2, and leukemia inhibitory factor for in vitro differentiation of MSCs isolated from different sources towards nephrogenic lineage. Less invasively isolated adipose-derived MSCs were also compared to bone marrow-derived MSCs for their differentiation potential to induce renal cell. Differentiated MSCs were further evaluated for their resistance to oxidative stress induced by oxygen peroxide. A combination of growth factors successfully induced differentiation of MSCs. Both types of differentiated cells showed significant expression of pronephrogenic markers (Wnt4, Wt1, and Pax2) and renal epithelial markers (Ecad and ZO1). In contrast, expression of mesenchymal stem cells marker Oct4 and Vim were downregulated. Furthermore, differentiated adipose-derived MSCs and bone marrow-derived MSCs showed enhanced and comparable resistance to oxygen peroxide-induced oxidative stress. Adipose-derived MSC provides a promising alternative to bone marrow-derived MSC as a source of autologous stem cells in human kidney injuries. In addition, differentiated MSCs with further in vivo investigations may serve as a cell source for tissue engineering or cell therapy in different renal ailments.

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

Epigenetic Plasticity Drives Adipogenic and Osteogenic Differentiation of Marrow-derived Mesenchymal Stem Cells*

PubMed Central

Meyer, Mark B.; Benkusky, Nancy A.; Sen, Buer; Rubin, Janet; Pike, J. Wesley

2016-01-01

Terminal differentiation of multipotent stem cells is achieved through a coordinated cascade of activated transcription factors and epigenetic modifications that drive gene transcription responsible for unique cell fate. Within the mesenchymal lineage, factors such as RUNX2 and PPARγ are indispensable for osteogenesis and adipogenesis, respectively. We therefore investigated genomic binding of transcription factors and accompanying epigenetic modifications that occur during osteogenic and adipogenic differentiation of mouse bone marrow-derived mesenchymal stem cells (MSCs). As assessed by ChIP-sequencing and RNA-sequencing analyses, we found that genes vital for osteogenic identity were linked to RUNX2, C/EBPβ, retinoid X receptor, and vitamin D receptor binding sites, whereas adipocyte differentiation favored PPARγ, retinoid X receptor, C/EBPα, and C/EBPβ binding sites. Epigenetic marks were clear predictors of active differentiation loci as well as enhancer activities and selective gene expression. These marrow-derived MSCs displayed an epigenetic pattern that suggested a default preference for the osteogenic pathway; however, these patterns were rapidly altered near the Adipoq, Cidec, Fabp4, Lipe, Plin1, Pparg, and Cebpa genes during adipogenic differentiation. Surprisingly, we found that these cells also exhibited an epigenetic plasticity that enabled them to trans-differentiate from adipocytes to osteoblasts (and vice versa) after commitment, as assessed by staining, gene expression, and ChIP-quantitative PCR analysis. The osteogenic default pathway may be subverted during pathological conditions, leading to skeletal fragility and increased marrow adiposity during aging, estrogen deficiency, and skeletal unloading. Taken together, our data provide an increased mechanistic understanding of the epigenetic programs necessary for multipotent differentiation of MSCs that may prove beneficial in the development of therapeutic strategies. PMID:27402842

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.

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.

Anti-Tumor Effect of Adipose Tissue Derived-Mesenchymal Stem Cells Expressing Interferon-β and Treatment with Cisplatin in a Xenograft Mouse Model for Canine Melanoma

PubMed Central

Ahn, Jin ok; Lee, Hee woo; Seo, Kyoung won; Kang, Sung keun; Ra, Jeong chan; Youn, Hwa young

2013-01-01

Adipose tissue-derived mesenchymal stem cells (AT-MSCs) are attractive cell-therapy vehicles for the delivery of anti-tumor molecules into the tumor microenvironment. The innate tropism of AT-MSCs for tumors has important implications for effective cellular delivery of anti-tumor molecules, including cytokines, interferon, and pro-drugs. The present study was designed to determine the possibility that the combination of stem cell-based gene therapy with low-dose cisplatin would improve therapeutic efficacy against canine melanoma. The IFN-β transduced canine AT-MSCs (cAT-MSC-IFN-β) inhibited the growth of LMeC canine melanoma cells in direct and indirect in vitro co-culture systems. In animal experiments using BALB/c nude mouse xenografts, which developed by injecting LMeC cells, the combination treatment of cAT-MSC-IFN-β and low-dose cisplatin significantly reduced tumor volume compared with the other treatment groups. Fluorescent microscopic analysis with a TUNEL (terminal deoxynucleotidyl transferase-mediated nick-end labeling) assay of tumor section provided evidence for homing of cAT-MSC-IFN-β to the tumor site and revealed that the combination treatment of cAT-MSC-IFN-β with low-dose cisplatin induced high levels of cell apoptosis. These findings may prove useful in further explorations of the application of these combined approaches to the treatment of malignant melanoma and other tumors. PMID:24040358

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.

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.

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...

Paracrine-Mediated Neuroprotection and Neuritogenesis of Axotomised Retinal Ganglion Cells by Human Dental Pulp Stem Cells: Comparison with Human Bone Marrow and Adipose-Derived Mesenchymal Stem Cells

PubMed Central

Mead, Ben; Logan, Ann; Berry, Martin

2014-01-01

We have investigated and compared the neurotrophic activity of human dental pulp stem cells (hDPSC), human bone marrow-derived mesenchymal stem cells (hBMSC) and human adipose-derived stem cells (hAMSC) on axotomised adult rat retinal ganglion cells (RGC) in vitro in order to evaluate their therapeutic potential for neurodegenerative conditions of RGC. Using the transwell system, RGC survival and length/number of neurites were quantified in coculture with stem cells in the presence or absence of specific Fc-receptor inhibitors to determine the role of NGF, BDNF, NT-3, VEGF, GDNF, PDGF-AA and PDGF-AB/BB in stem cell-mediated RGC neuroprotection and neuritogenesis. Conditioned media, collected from cultured hDPSC/hBMSC/hAMSC, were assayed for the secreted growth factors detailed above using ELISA. PCR array determined the hDPSC, hBMSC and hAMSC expression of genes encoding 84 growth factors and receptors. The results demonstrated that hDPSC promoted significantly more neuroprotection and neuritogenesis of axotomised RGC than either hBMSC or hAMSC, an effect that was neutralized after the addition of specific Fc-receptor inhibitors. hDPSC secreted greater levels of various growth factors including NGF, BDNF and VEGF compared with hBMSC/hAMSC. The PCR array confirmed these findings and identified VGF as a novel potentially therapeutic hDPSC-derived neurotrophic factor (NTF) with significant RGC neuroprotective properties after coculture with axotomised RGC. In conclusion, hDPSC promoted significant multi-factorial paracrine-mediated RGC survival and neurite outgrowth and may be considered a potent and advantageous cell therapy for retinal nerve repair. PMID:25290916

The effect of autologous adipose derived mesenchymal stem cell therapy in the treatment of a large osteochondral defect of the knee following unsuccessful surgical intervention of osteochondritis dissecans - a case study.

PubMed

Freitag, Julien; Shah, Kiran; Wickham, James; Boyd, Richard; Tenen, Abi

2017-07-14

A prospective analysis of the effect of autologous adipose derived mesenchymal stem cell (MSC) therapy in the treatment of an osteochondral defect of the knee with early progressive osteoarthritis following unsuccessful surgical intervention of osteochondritis dissecans (OCD). After failed conventional management of OCD a patient undergoes intra-articular MSC therapy. Patient outcome measures included the Numeric Pain Rating Scale (NPRS), the Western Ontario and McMaster Universities Arthritis Index (WOMAC) and the Knee Injury and Osteoarthritis Outcome Score (KOOS). Structural outcome was assessed using MRI with the novel technique of T2 mapping used to indicate cartilage quality. Following MSC therapy the patient reported improvement in pain and function as measured by NPRS, WOMAC and KOOS. Repeat MRI analysis showed regeneration of cartilage. MRI T2 mapping indicated hyaline like cartilage regrowth. In this report, the use of MSCs, after unsuccessful conventional OCD management, resulted in structural, functional and pain improvement. These results highlight the need to further study the regenerative potential of MSC therapy. Australian and New Zealand Clinical Trial Registry Number - ACTRN12615000258550 (Date registered 19/03/2015 - retrospectively registered).

Preclinical Biosafety Evaluation of Genetically Modified Human Adipose Tissue-Derived Mesenchymal Stem Cells for Clinical Applications to Brainstem Glioma.

PubMed

Choi, Seung Ah; Yun, Jun-Won; Joo, Kyeung Min; Lee, Ji Yeoun; Kwak, Pil Ae; Lee, Young Eun; You, Ji-Ran; Kwon, Euna; Kim, Woo Ho; Wang, Kyu-Chang; Phi, Ji Hoon; Kang, Byeong-Cheol; Kim, Seung-Ki

2016-06-15

Stem-cell based gene therapy is a promising novel therapeutic approach for inoperable invasive tumors, including brainstem glioma. Previously, we demonstrated the therapeutic potential of human adipose tissue-derived mesenchymal stem cells (hAT-MSC) genetically engineered to express a secreted form of tumor necrosis factor-related apoptosis-inducing ligand (sTRAIL) against brainstem glioma. However, safety concerns should be comprehensively investigated before clinical applications of hAT-MSC.sTRAIL. At first, we injected stereotactically low (1.2 × 10(5) cells/18 μL), medium (2.4 × 10(5)/18 μL), or high dose (3.6 × 10(5)/18 μL) of hAT-MSC.sTRAIL into the brainstems of immunodeficient mice reflecting the plan of the future clinical trial. Local toxicity, systemic toxicity, secondary tumor formation, and biodistribution of hAT-MSC.sTRAIL were investigated. Next, presence of hAT-MSC.sTRAIL was confirmed in the brain and major organs at 4, 9, and 14 weeks in brainstem glioma-bearing mice. In the 15-week subchronic toxicity test, no serious adverse events in terms of body weight, food consumption, clinical symptom, urinalysis, hematology, clinical chemistry, organ weight, and histopathology were observed. In the 26-week tumorigenicity test, hAT-MSC.sTRAIL made no detectable tumors, whereas positive control U-87 MG cells made huge tumors in the brainstem. No remaining hAT-MSC.sTRAIL was observed in any organs examined, including the brainstem at 15 or 26 weeks. In brainstem glioma-bearing mice, injected hAT-MSC.sTRAIL was observed, but gradually decreased over time in the brain. The mRNA of human specific GAPDH and TRAIL was not detected in all major organs. These results indicate that the hAT-MSC.sTRAIL could be applicable to the future clinical trials in terms of biosafety.

Effect of berberine on the viability of adipose tissue-derived mesenchymal stem cells in nutrients deficient condition.

PubMed

Ghorbani, Ahmad; Baradaran Rahimi, Vafa; Sadeghnia, Hamid Reza; Hosseini, Azar

2018-03-01

This study was designed to examine whether berberine protects rat adipose tissue-derived stem cells (ASCs) against glucose and serum deprivation (GSD)-induced cell death. ASCs were cultured for 24 h in GSD condition in the presence of berberine and then cell viability, apoptosis and generation of reactive oxygen species (ROS) were evaluated. The GSD condition significantly decreased ASCs viability and increased ROS generation and apoptosis. Incubation with 0.75-3 μM berberine partially increased cell viability and decreased ROS generation and apoptosis in GSD condition. In conclusion, berberine partially protects ASCs in nutrients deficient condition and may help ASCs to preserve their survival during cell therapy of ischemia.

Inhibiting glycogen synthase kinase-3 and transforming growth factor-β signaling to promote epithelial transition of human adipose mesenchymal stem cells.

PubMed

Setiawan, Melina; Tan, Xiao-Wei; Goh, Tze-Wei; Hin-Fai Yam, Gary; Mehta, Jodhbir S

2017-09-02

This study was aimed to investigate the epithelial differentiation of human adipose-derived mesenchymal stem cells (ADSCs) by inhibiting glycogen synthase kinase-3 (GSK3) and transforming growth factor β (TGFβ) signaling. STEMPRO human ADSCs at passage 2 were treated with CHIR99021 (GSK3 inhibitor), E-616452 (TGFβ1 receptor kinase inhibitor), A-83-01 (TGFβ type 1 receptor inhibitor), valproic acid (histone deacetylase inhibitor), tranylcypromine (monoamine oxidase inhibitor) and all-trans retinoic acid for 72 h. The mesenchymal-epithelial transition was shown by down-regulation of mesenchymal genes (Slug, Zinc Finger E-box Binding Homeobox 1 ZEB1, integrin α5 ITGA5 and vimentin VIM) and up-regulation of epithelial genes (E-cadherin, Epithelial Cell Adhesion Molecule EpCAM, Zonula Occludens-1 ZO-1, occludin, deltaN p63 δNp63, Transcription Factor 4 TCF4 and Twist Family bHLH Transcription Factor TWIST), compared to untreated ADSCs. Cell morphology and stress fiber pattern were examined and the treated cells became less migratory in scratch wound closure assay. The formation of cell junction complexes was observed under transmission electron microscopy. Global gene expression using GeneChip ® Human Genome U133 Array (Affymetrix) showed that the treatment up-regulated 540 genes (containing genes for cell cycle, cytoskeleton reorganization, chemotaxis, epithelium development and regulation of cell migration) and down-regulated 483 genes. Human ADSCs were transited to epithelial lineage by inhibiting GSK3 and TGFβ signaling. It can be an adult stem cell source for epithelial cell-based therapy. Copyright © 2017 Elsevier Inc. All rights reserved.

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.

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

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.

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.

Canine Adipose-Derived Stem Cells: Purinergic Characterization and Neurogenic Potential for Therapeutic Applications.

PubMed

Roszek, Katarzyna; Makowska, Noemi; Czarnecka, Joanna; Porowińska, Dorota; Dąbrowski, Marcin; Danielewska, Justyna; Nowak, Wiesław

2017-01-01

The presented results evidence that canine adipose-derived stem cells (ADSCs) represent the premature population of stem cells with great biological potential and properties. ADCS are easy to obtain and culture, able to differentiate into the neurogenic lineage as well as it is easy to control their proliferation rate with nucleotides and nucleosides or analogues. We report that in vitro cultured canine ADSCs response to adenosine- and ATP-mediated stimulation. Differences in canine ADSCs and human mesenchymal stem cells in ecto-nucleotidase activity have been observed. The ecto-nucleotidase activity changes during ADSCs in vitro transdifferentiation into neurogenic lineage are fast and simple to analyze. Therefore, the simple analysis of ecto-enzymes activity allows for verification of the stem cells quality: their stemness or initiation of the differentiation process. The biological potential of the cells isolated from canine fat, as well as the good quality control of this cell culture, make them a promising tool for both experimental and therapeutic usage. J. Cell. Biochem. 118: 58-65, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

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.

Exosomes derived from mesenchymal non-small cell lung cancer cells promote chemoresistance.

PubMed

Lobb, Richard J; van Amerongen, Rosa; Wiegmans, Adrian; Ham, Sunyoung; Larsen, Jill E; Möller, Andreas

2017-08-01

Non-small cell lung cancer (NSCLC) is the most common lung cancer type and the most common cause of mortality in lung cancer patients. NSCLC is often associated with resistance to chemotherapeutics and together with rapid metastatic spread, results in limited treatment options and poor patient survival. NSCLCs are heterogeneous, and consist of epithelial and mesenchymal NSCLC cells. Mesenchymal NSCLC cells are thought to be responsible for the chemoresistance phenotype, but if and how this phenotype can be transferred to other NSCLC cells is currently not known. We hypothesised that small extracellular vesicles, exosomes, secreted by mesenchymal NSCLC cells could potentially transfer the chemoresistance phenotype to surrounding epithelial NSCLC cells. To explore this possibility, we used a unique human bronchial epithelial cell (HBEC) model in which the parental cells were transformed from an epithelial to mesenchymal phenotype by introducing oncogenic alterations common in NSCLC. We found that exosomes derived from the oncogenically transformed, mesenchymal HBECs could transfer chemoresistance to the parental, epithelial HBECs and increase ZEB1 mRNA, a master EMT transcription factor, in the recipient cells. Additionally, we demonstrate that exosomes from mesenchymal, but not epithelial HBECs contain the ZEB1 mRNA, thereby providing a potential mechanism for the induction of a mesenchymal phenotype in recipient cells. Together, this work demonstrates for the first time that exosomes derived from mesenchymal, oncogenically transformed lung cells can transfer chemoresistance and mesenchymal phenotypes to recipient cells, likely via the transfer of ZEB1 mRNA in exosomes. © 2017 UICC.

Perivascular Stem Cells: A Prospectively Purified Mesenchymal Stem Cell Population for Bone Tissue Engineering

PubMed Central

James, Aaron W.; Zara, Janette N.; Zhang, Xinli; Askarinam, Asal; Goyal, Raghav; Chiang, Michael; Yuan, Wei; Chang, Le; Corselli, Mirko; Shen, Jia; Pang, Shen; Stoker, David; Wu, Ben

2012-01-01

Adipose tissue is an ideal source of mesenchymal stem cells for bone tissue engineering: it is largely dispensable and readily accessible with minimal morbidity. However, the stromal vascular fraction (SVF) of adipose tissue is a heterogeneous cell population, which leads to unreliable bone formation. In the present study, we prospectively purified human perivascular stem cells (PSCs) from adipose tissue and compared their bone-forming capacity with that of traditionally derived SVF. PSCs are a population (sorted by fluorescence-activated cell sorting) of pericytes (CD146+CD34−CD45−) and adventitial cells (CD146−CD34+CD45−), each of which we have previously reported to have properties of mesenchymal stem cells. Here, we found that PSCs underwent osteogenic differentiation in vitro and formed bone after intramuscular implantation without the need for predifferentiation. We next sought to optimize PSCs for in vivo bone formation, adopting a demineralized bone matrix for osteoinduction and tricalcium phosphate particle formulation for protein release. Patient-matched, purified PSCs formed significantly more bone in comparison with traditionally derived SVF by all parameters. Recombinant bone morphogenetic protein 2 increased in vivo bone formation but with a massive adipogenic response. In contrast, recombinant Nel-like molecule 1 (NELL-1; a novel osteoinductive growth factor) selectively enhanced bone formation. These studies suggest that adipose-derived human PSCs are a new cell source for future efforts in skeletal regenerative medicine. Moreover, PSCs are a stem cell-based therapeutic that is readily approvable by the U.S. Food and Drug Administration, with potentially increased safety, purity, identity, potency, and efficacy. Finally, NELL-1 is a candidate growth factor able to induce human PSC osteogenesis. PMID:23197855

Mesenchymal stem cells from adipose and bone marrow promote angiogenesis via distinct cytokine and protease expression mechanisms

PubMed Central

Kachgal, Suraj; Putnam, Andrew J.

2012-01-01

Using a fibrin-based angiogenesis model, we have established that there is no canonical mechanism used by ECs to degrade the surrounding extracellular matrix (ECM), but rather the set of proteases used is dependent on the mural cells providing the angiogenic cues. Mesenchymal stem cells (MSCs) originating from different tissues, which are thought to be phenotypically similar, promote angiogenesis through distinct mechanisms. Specifically, adipose-derived stem cells (ASCs) promote utilization of the plasminogen activator-plasmin axis by ECs as the primary means of vessel invasion and elongation in fibrin. Matrix metalloproteinases (MMPs) serve a purpose in regulating capillary diameter and possibly in stabilizing the nascent vessels. These proteolytic mechanisms are more akin to those involved in fibroblast-mediated angiogenesis than to those in bone marrow-derived stem cell (BMSC)-mediated angiogenesis. In addition, expression patterns of angiogenic factors such as urokinase plasminogen activator (uPA), hepatocyte growth factor (HGF), and tumor necrosis factor alpha (TNFα) were similar for ASC and fibroblast-mediated angiogenesis, and in direct contrast to BMSC-mediated angiogenesis. The present study illustrates that the nature of the heterotypic interactions between mural cells and endothelial cells depend on the identity of the mural cell used. Even MSCs which are shown to behave phenotypically similar do not stimulate angiogenesis via the same mechanisms. PMID:21104120

Protective/restorative Role of the Adipose Tissue-derived Mesenchymal Stem Cells on the Radioiodine-induced Salivary Gland Damage in Rats

PubMed Central

Saylam, Güleser; Bayır, Ömer; Pınarlı, Ferda Alparslan; Han, Ünsal; Korkmaz, Mehmet Hakan; Sancaktar, Mehmet Eser; Tatar, İlkan; Sargon, Mustafa Fevzi; Tatar, Emel Çadallı

2017-01-01

Abstract Background To analyze protective/regenerative effects of adipose tissue-derived mesenchymal stem cells (ADMSC) on 131I-Radioiodine (RAI)-induced salivary gland damage in rats. Materials and Methods Study population consisted of controls (n:6) and study groups (n:54): RAI (Group 1), ADMSC (Group 2), amifostine (Group 3), RAI+amifostine (Group 4), concomitant RAI+ADMSC (Group 5) and RAI+ADMSC after 48 h (Group 6). We used light microscopy (LM), transmission electron microscopy (TEM), and salivary gland scintigraphy (SGS), and analyzed data statistically. Results We observed the homing of ADMSC in salivary glands at 1st month on LM. RAI exposure affected necrosis, periductal fibrosis, periductal sclerosis, vascular sclerosis and the total sum score were in a statistically significant manner (P < 0.05). Intragroup comparisons with LM at 1st and 6th months revealed statistically significant improvements in Group 6 (P < 0.05) but not in Groups 4 and 5. Intergroup comparisons of the total score showed that Groups 4 and 5 in 1st month and Group 6 in 6th month had the lowest values. TEM showed vacuolization, edema, and fibrosis at 1st month, and an improvement in damage in 6th month in Groups 5 and 6. SGSs revealed significant differences for the maximum secretion ratio (Smax) (P = 0.01) and the gland-to-background ratio at a maximum count (G/BGmax) (P = 0. 01) at 1st month, for G/BGmax (P = 0.01), Smax (P = 0.01) and the time to reach the maximum count ratio over the time to reach the minimum count (Tmax/Tmin) (P = 0.03) at 6th month. 1st and 6th month scans showed differences for Smax and G/BGmax (P = 0.04), but not for Tmax/Tmin (p > 0.05). We observed a significant deterioration in gland function in group 1, whereas, mild to moderate deteriorations were seen in protective treatment groups. Conclusions Our results indicated that ADMSC might play a promising role as a protective/regenerative agent against RAI-induced salivary gland dysfunction. PMID:28959167

Mesenchymal stem cell-derived extracellular vesicles attenuate kidney inflammation.

PubMed

Eirin, Alfonso; Zhu, Xiang-Yang; Puranik, Amrutesh S; Tang, Hui; McGurren, Kelly A; van Wijnen, Andre J; Lerman, Amir; Lerman, Lilach O

2017-07-01

Mesenchymal stem/stromal cells (MSCs) have distinct capability for renal repair, but may have safety concerns. MSC-derived extracellular vesicles emerged as a novel noncellular alternative. Using a porcine model of metabolic syndrome and renal artery stenosis we tested whether extracellular vesicles attenuate renal inflammation, and if this capacity is mediated by their cargo of the anti-inflammatory cytokine interleukin (IL) 10. Pigs with metabolic syndrome were studied after 16 weeks of renal artery stenosis untreated or treated four weeks earlier with a single intrarenal delivery of extracellular vesicles harvested from adipose tissue-derived autologous MSCs. Lean and sham metabolic syndrome animals served as controls (seven each). Five additional pigs with metabolic syndrome and renal artery stenosis received extracellular vesicles with pre-silenced IL10 (IL10 knock-down). Single-kidney renal blood flow, glomerular filtration rate, and oxygenation were studied in vivo and renal injury pathways ex vivo. Retention of extracellular vesicles in the stenotic kidney peaked two days after delivery and decreased thereafter. Four weeks after injection, extracellular vesicle fragments colocalized with stenotic-kidney tubular cells and macrophages, indicating internalization or fusion. Extracellular vesicle delivery attenuated renal inflammation, and improved medullary oxygenation and fibrosis. Renal blood flow and glomerular filtration rate fell in metabolic syndrome and renal artery stenosis compared to metabolic syndrome, but was restored in pigs treated with extracellular vesicles. These renoprotective effects were blunted in pigs treated with IL10-depleted extracellular vesicles. Thus, extracellular vesicle-based regenerative strategies might be useful for patients with metabolic syndrome and renal artery stenosis. Copyright © 2017 International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Ex vivo generation of glucose sensitive insulin secreting mesenchymal stem cells derived from human adipose tissue.

PubMed

Dave, Shruti D; Vanikar, Aruna V; Trivedi, Hargovind L

2012-03-01

Diabetics are incapable of producing insulin/have autoimmune mechanisms making it ineffective to control glucose secretion. We present a prospective study of glucose-sensitive insulin-secreting mesenchymal stem cells (IS-MSC) generated from human adipose tissue (h-AD) sans xenogenic material. Ten grams h-AD from donor anterior abdominal wall was collected in proliferation medium composed of α-Minimum Essential Media (α-MEM), albumin, fibroblast-growth factor and antibiotics, minced, incubated in collagenase-I at 37°C with shaker and centrifuged. Supernatant and pellets were separately cultured in proliferation medium on cell+ plates at 37°C with 5% CO(2) for 10 days. Cells were harvested by trypsinization, checked for viability, sterility, counts, flow-cytometry (CD45(-)/90(+)/73(+)), and differentiated into insulin-expressing cells using medium composed of DMEM, gene expressing up-regulators and antibiotics for 3 days. They were studied for transcriptional factors Pax-6, Isl-1, pdx-1 (immunofluorescence). C-peptide and insulin were measured by chemiluminescence. In vitro glucose sensitivity assay was carried out by measuring levels of insulin and C-peptide secretion in absence of glucose followed by 2 hours incubation after glucose addition. Mean IS-AD-MSC quantum was 3.21 ml, cell count, 1.5 ×10(3) cells/μl), CD45(-)/90(+)/73(+) cells were 44.37% /25.52%. All of them showed presence of pax-6, pdx-1, and Isl-1. Mean C-Peptide and insulin levels were 0.36 ng/ml and 234 μU/ml, respectively, pre-glucose and 0.87 ng/ml and 618.3 μU/ml post-glucose additions. The mean rise in secretion levels was 2.42 and 2.65 fold, respectively. Insulin-secreting h-AD-MSC can be generated safely and effectively showing in vitro glucose responsive alteration in insulin and C-peptide secretion levels.

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.

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.

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.

Proportion of collagen type II in the extracellular matrix promotes the differentiation of human adipose-derived mesenchymal stem cells into nucleus pulposus cells.

PubMed

Tao, Yiqing; Zhou, Xiaopeng; Liu, Dongyu; Li, Hao; Liang, Chengzhen; Li, Fangcai; Chen, Qixin

2016-01-01

During degeneration process, the catabolism of collagen type II and anabolism of collagen type I in nucleus pulposus (NP) may influence the bioactivity of transplanted cells. Human adipose-derived mesenchymal stem cells (hADMSCs) were cultured as a micromass or in a series of gradual proportion hydrogels of a mix of collagen types I and II. Cell proliferation and cytotoxicity were detected using CCK-8 and LDH assays respectively. The expression of differentiation-related genes and proteins, including SOX9, aggrecan, collagen type I, and collagen type II, was examined using RT-qPCR and Western blotting. Novel phenotypic genes were also detected by RT-qPCR and western blotting. Alcian blue and dimethylmethylene blue assays were used to investigate sulfate proteoglycan expression, and PI3K/AKT, MAPK/ERK, and Smad signaling pathways were examined by Western blotting. The results showed collagen hydrogels have good biocompatibility, and cell proliferation increased after collagen type II treatment. Expressions of SOX9, aggrecan, and collagen type II were increased in a collagen type II dependent manner. Sulfate proteoglycan synthesis increased in proportion to collagen type II concentration. Only hADMSCs highly expressed NP cell marker KRT19 in collagen type II culture. Additionally, phosphorylated Smad3, which is associated with phosphorylated ERK, was increased after collagen type II-stimulation. The concentration and type of collagen affect hADMSC differentiation into NP cells. Collagen type II significantly ameliorates hADMSC differentiation into NP cells and promotes extracellular matrix synthesis. Therefore, anabolism of collagen type I and catabolism of type II may attenuate the differentiation and biosynthesis of transplanted stem cells. © 2016 International Union of Biochemistry and Molecular Biology.

Development of a rapid culture method to induce adipocyte differentiation of human bone marrow-derived mesenchymal stem cells

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

Ninomiya, Yuichi; Sugahara-Yamashita, Yzumi; Nakachi, Yutaka

2010-04-02

Human mesenchymal stem cells (hMSCs) derived from bone marrow are multipotent stem cells that can regenerate mesenchymal tissues such as adipose, bone or muscle. It is thought that hMSCs can be utilized as a cell resource for tissue engineering and as human models to study cell differentiation mechanisms, such as adipogenesis, osteoblastogenesis and so on. Since it takes 2-3 weeks for hMSCs to differentiate into adipocytes using conventional culture methods, the development of methods to induce faster differentiation into adipocytes is required. In this study we optimized the culture conditions for adipocyte induction to achieve a shorter cultivation time formore » the induction of adipocyte differentiation in bone marrow-derived hMSCs. Briefly, we used a cocktail of dexamethasone, insulin, methylisobutylxanthine (DIM) plus a peroxisome proliferator-activated receptor {gamma} agonist, rosiglitazone (DIMRo) as a new adipogenic differentiation medium. We successfully shortened the period of cultivation to 7-8 days from 2-3 weeks. We also found that rosiglitazone alone was unable to induce adipocyte differentiation from hMSCs in vitro. However, rosiglitazone appears to enhance hMSC adipogenesis in the presence of other hormones and/or compounds, such as DIM. Furthermore, the inhibitory activity of TGF-{beta}1 on adipogenesis could be investigated using DIMRo-treated hMSCs. We conclude that our rapid new culture method is very useful in measuring the effect of molecules that affect adipogenesis in hMSCs.« less

Additional benefit of combined therapy with melatonin and apoptotic adipose-derived mesenchymal stem cell against sepsis-induced kidney injury.

PubMed

Chen, Hong-Hwa; Lin, Kun-Chen; Wallace, Christopher G; Chen, Yen-Ta; Yang, Chih-Chao; Leu, Steve; Chen, Yi-Ching; Sun, Cheuk-Kwan; Tsai, Tzu-Hsien; Chen, Yung-Lung; Chung, Sheng-Ying; Chang, Chia-Lo; Yip, Hon-Kan

2014-08-01

This study tested whether combined therapy with melatonin and apoptotic adipose-derived mesenchymal stem cells (A-ADMSCs) offered additional benefit in ameliorating sepsis-induced acute kidney injury. Adult male Sprague-Dawley rats (n = 65) were randomized equally into five groups: Sham controls (SC), sepsis induced by cecal-ligation and puncture (CLP), CLP-melatonin, CLP-A-ADMSC, and CLP-melatonin-A-ADMSC. Circulating TNF-α level at post-CLP 6 hr was highest in CLP and lowest in SC groups, higher in CLP-melatonin than in CLP-A-ADMSC and CLP-melatonin-A-ADMSC groups (all P < 0.001). Immune reactivity as reflected in the number of splenic helper-, cytoxic-, and regulatory-T cells at post-CLP 72 hr exhibited the same pattern as that of circulating TNF-α among all groups (P < 0.001). The histological scoring of kidney injury and the number of F4/80+ and CD14+ cells in kidney were highest in CLP and lowest in SC groups, higher in CLP-melatonin than in CLP-A-ADMSC and CLP-melatonin-A-ADMSC groups, and higher in CLP-A-ADMSC than in CLP-melatonin-A-ADMSC groups (all P < 0.001). Changes in protein expressions of inflammatory (RANTES, TNF-1α, NF-κB, MMP-9, MIP-1, IL-1β), apoptotic (cleaved caspase 3 and PARP, mitochondrial Bax), fibrotic (Smad3, TGF-β) markers, reactive-oxygen-species (NOX-1, NOX-2), and oxidative stress displayed a pattern identical to that of kidney injury score among the five groups (all P < 0.001). Expressions of antioxidants (GR+, GPx+, HO-1, NQO-1+) were lowest in SC group and highest in CLP-melatonin-A-ADMSC group, lower in CLP than in CLP-melatonin and CLP-A-ADMSC groups, and lower in CLP-melatonin- than in CLP-A-ADMSC-tretaed animals (all P < 0.001). In conclusion, combined treatment with melatonin and A-ADMSC was superior to A-ADMSC alone in protecting the kidneys from sepsis-induced injury. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

In Vitro Expression of Cytokeratin 19 in Adipose-Derived Stem Cells Is Induced by Epidermal Growth Factor.

PubMed

Chen, Shangliang; Wang, Mingzhu; Chen, Xinglu; Chen, Shaolian; Liu, Li; Zhu, Jianbin; Wang, Jinhui; Yang, Xiaorong; Cai, Xiangsheng

2018-06-21

BACKGROUND Cytokeratin 19 (CK19) is a typical epithelial marker. In this study, we determined whether epidermal growth factor (EGF) or basic fibroblast growth factor (bFGF) could enhance CK19 expression in adipose-derived stem cells (ADSCs), thereby inducing the differentiation of ADSCs into epithelial-like cells. MATERIAL AND METHODS ADSCs were isolated from perinephric fat, and the expression of CD29, CD90, and CD105 was confirmed. Following isolation, ADSCs were cultured in static medium or medium containing EGF or bFGF. RESULTS Flow cytometry revealed that EGF and bFGF could alter mesenchymal stem cell markers as well as the cell cycle of ADSCs. Western blotting and immunofluorescence revealed that after 14 days, EGF treatment enhanced the expression of CK19 in ADSCs. CONCLUSIONS Our findings offer important insight for the clinical use of ADSCs in the generation of epithelial-like cells in the future.

Purification and differentiation of human adipose-derived stem cells by membrane filtration and membrane migration methods

PubMed Central

Lin, Hong Reng; Heish, Chao-Wen; Liu, Cheng-Hui; Muduli, Saradaprasan; Li, Hsing-Fen; Higuchi, Akon; Kumar, S. Suresh; Alarfaj, Abdullah A.; Munusamy, Murugan A.; Hsu, Shih-Tien; Chen, Da-Chung; Benelli, Giovanni; Murugan, Kadarkarai; Cheng, Nai-Chen; Wang, Han-Chow; Wu, Gwo-Jang

2017-01-01

Human adipose-derived stem cells (hADSCs) are easily isolated from fat tissue without ethical concerns, but differ in purity, pluripotency, differentiation ability, and stem cell marker expression, depending on the isolation method. We isolated hADSCs from a primary fat tissue solution using: (1) conventional culture, (2) a membrane filtration method, (3) a membrane migration method where the primary cell solution was permeated through membranes, adhered hADSCs were cultured, and hADSCs migrated out from the membranes. Expression of mesenchymal stem cell markers and pluripotency genes, and osteogenic differentiation were compared for hADSCs isolated by different methods using nylon mesh filter membranes with pore sizes ranging from 11 to 80 μm. hADSCs isolated by the membrane migration method had the highest MSC surface marker expression and efficient differentiation into osteoblasts. Osteogenic differentiation ability of hADSCs and MSC surface marker expression were correlated, but osteogenic differentiation ability and pluripotent gene expression were not. PMID:28071738

Hanging drop culture enhances differentiation of human adipose-derived stem cells into anterior neuroectodermal cells using small molecules.

PubMed

Amirpour, Noushin; Razavi, Shahnaz; Esfandiari, Ebrahim; Hashemibeni, Batoul; Kazemi, Mohammad; Salehi, Hossein

2017-06-01

Inspired by in vivo developmental process, several studies were conducted to design a protocol for differentiating of mesenchymal stem cells into neural cells in vitro. Human adipose-derived stem cells (hADSCs) as mesenchymal stem cells are a promising source for this purpose. At current study, we applied a defined neural induction medium by using small molecules for direct differentiation of hADSCs into anterior neuroectodermal cells. Anterior neuroectodermal differentiation of hADSCs was performed by hanging drop and monolayer protocols. At these methods, three small molecules were used to suppress the BMP, Nodal, and Wnt signaling pathways in order to obtain anterior neuroectodermal (eye field) cells from hADSCs. After two and three weeks of induction, the differentiated cells with neural morphology expressed anterior neuroectodermal markers such as OTX2, SIX3, β-TUB III and PAX6. The protein expression of such markers was confirmed by real time, RT-PCR and immunocytochemistry methods According to our data, it seems that the hanging drop method is a proper approach for neuroectodermal induction of hADSCs. Considering wide availability and immunosuppressive properties of hADSCs, these cells may open a way for autologous cell therapy of neurodegenerative disorders. Copyright © 2017 ISDN. Published by Elsevier Ltd. All rights reserved.

Characterisation of insulin-producing cells differentiated from tonsil derived mesenchymal stem cells.

PubMed

Kim, So-Yeon; Kim, Ye-Ryung; Park, Woo-Jae; Kim, Han Su; Jung, Sung-Chul; Woo, So-Youn; Jo, Inho; Ryu, Kyung-Ha; Park, Joo-Won

2015-01-01

Tonsil-derived (T-) mesenchymal stem cells (MSCs) display mutilineage differentiation potential and self-renewal capacity and have potential as a banking source. Diabetes mellitus is a prevalent disease in modern society, and the transplantation of pancreatic progenitor cells or various stem cell-derived insulin-secreting cells has been suggested as a novel therapy for diabetes. The potential of T-MSCs to trans-differentiate into pancreatic progenitor cells or insulin-secreting cells has not yet been investigated. We examined the potential of human T-MSCs to trans-differentiate into pancreatic islet cells using two different methods based on β-mercaptoethanol and insulin-transferin-selenium, respectively. First, we compared the efficacy of the two methods for inducing differentiation into insulin-producing cells. We demonstrated that the insulin-transferin-selenium method is more efficient for inducing differentiation into insulin-secreting cells regardless of the source of the MSCs. Second, we compared the differentiation potential of two different MSC types: T-MSCs and adipose-derived MSCs (A-MSCs). T-MSCs had a differentiation capacity similar to that of A-MSCs and were capable of secreting insulin in response to glucose concentration. Islet-like clusters differentiated from T-MSCs had lower synaptotagmin-3, -5, -7, and -8 levels, and consequently lower secreted insulin levels than cells differentiated from A-MSCs. These results imply that T-MSCs can differentiate into functional pancreatic islet-like cells and could provide a novel, alternative cell therapy for diabetes mellitus. Copyright © 2015 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

Plasma Rich in Growth Factors Induces Cell Proliferation, Migration, Differentiation, and Cell Survival of Adipose-Derived Stem Cells

PubMed Central

Mellado-López, Maravillas; Griffeth, Richard J.; Meseguer-Ripolles, Jose; García, Montserrat

2017-01-01

Adipose-derived stem cells (ASCs) are a promising therapeutic alternative for tissue repair in various clinical applications. However, restrictive cell survival, differential tissue integration, and undirected cell differentiation after transplantation in a hostile microenvironment are complications that require refinement. Plasma rich in growth factors (PRGF) from platelet-rich plasma favors human and canine ASC survival, proliferation, and delaying human ASC senescence and autophagocytosis in comparison with serum-containing cultures. In addition, canine and human-derived ASCs efficiently differentiate into osteocytes, adipocytes, or chondrocytes in the presence of PRGF. PRGF treatment induces phosphorylation of AKT preventing ASC death induced by lethal concentrations of hydrogen peroxide. Indeed, AKT inhibition abolished the PRGF apoptosis prevention in ASC exposed to 100 μM of hydrogen peroxide. Here, we show that canine ASCs respond to PRGF stimulus similarly to the human cells regarding cell survival and differentiation postulating the use of dogs as a suitable translational model. Overall, PRGF would be employed as a serum substitute for mesenchymal stem cell amplification to improve cell differentiation and as a preconditioning agent to prevent oxidative cell death. PMID:29270200

Plasma Rich in Growth Factors Induces Cell Proliferation, Migration, Differentiation, and Cell Survival of Adipose-Derived Stem Cells.

PubMed

Mellado-López, Maravillas; Griffeth, Richard J; Meseguer-Ripolles, Jose; Cugat, Ramón; García, Montserrat; Moreno-Manzano, Victoria

2017-01-01

Adipose-derived stem cells (ASCs) are a promising therapeutic alternative for tissue repair in various clinical applications. However, restrictive cell survival, differential tissue integration, and undirected cell differentiation after transplantation in a hostile microenvironment are complications that require refinement. Plasma rich in growth factors (PRGF) from platelet-rich plasma favors human and canine ASC survival, proliferation, and delaying human ASC senescence and autophagocytosis in comparison with serum-containing cultures. In addition, canine and human-derived ASCs efficiently differentiate into osteocytes, adipocytes, or chondrocytes in the presence of PRGF. PRGF treatment induces phosphorylation of AKT preventing ASC death induced by lethal concentrations of hydrogen peroxide. Indeed, AKT inhibition abolished the PRGF apoptosis prevention in ASC exposed to 100  μ M of hydrogen peroxide. Here, we show that canine ASCs respond to PRGF stimulus similarly to the human cells regarding cell survival and differentiation postulating the use of dogs as a suitable translational model. Overall, PRGF would be employed as a serum substitute for mesenchymal stem cell amplification to improve cell differentiation and as a preconditioning agent to prevent oxidative cell death.

Adipose-derived mesenchymal stem cells (AdMSC) for the treatment of secondary-progressive multiple sclerosis: A triple blinded, placebo controlled, randomized phase I/II safety and feasibility study.

PubMed

Fernández, Oscar; Izquierdo, Guillermo; Fernández, Victoria; Leyva, Laura; Reyes, Virginia; Guerrero, Miguel; León, Antonio; Arnaiz, Carlos; Navarro, Guillermo; Páramo, Maria Dolores; Cuesta, Antonio De la; Soria, Bernat; Hmadcha, Abdelkrim; Pozo, David; Fernandez-Montesinos, Rafael; Leal, Maria; Ochotorena, Itziar; Gálvez, Patricia; Geniz, Maria Angeles; Barón, Francisco Javier; Mata, Rosario; Medina, Cristina; Caparrós-Escudero, Carlos; Cardesa, Ana; Cuende, Natividad

2018-01-01

Currently available treatments for secondary progressive multiple sclerosis(SPMS) have limited efficacy and/or safety concerns. Adipose-mesenchymal derived stem cells(AdMSCs) represent a promising option and can be readily obtained using minimally invasive procedures. In this triple-blind, placebo-controlled study, cell samples were obtained from consenting patients by lipectomy and subsequently expanded. Patients were randomized to a single infusion of placebo, low-dose(1x106cells/kg) or high-dose(4x106cells/kg) autologous AdMSC product and followed for 12 months. Safety was monitored recording adverse events, laboratory parameters, vital signs and spirometry. Expanded disability status score (EDSS), magnetic-resonance-imaging, and other measures of possible treatment effects were also recorded. Thirty-four patients underwent lipectomy for AdMSCs collection, were randomized and thirty were infused (11 placebo, 10 low-dose and 9 high-dose); 4 randomized patients were not infused because of karyotype abnormalities in the cell product. Only one serious adverse event was observed in the treatment arms (urinary infection, considered not related to study treatment). No other safety parameters showed changes. Measures of treatment effect showed an inconclusive trend of efficacy. Infusion of autologous AdMSCs is safe and feasible in patients with SPMS. Larger studies and probably treatment at earlier phases would be needed to investigate the potential therapeutic benefit of this technique.

Overexpressed human heme Oxygenase-1 decreases adipogenesis in pigs and porcine adipose-derived stem cells.

PubMed

Park, Eun Jung; Koo, Ok Jae; Lee, Byeong Chun

2015-11-27

Adipose-derived mesenchymal stem cells (ADSC) are multipotent, which means they are able to differentiate into several lineages in vivo and in vitro under proper conditions. This indicates it is possible to determine the direction of differentiation of ADSC by controlling the microenvironment. Heme oxygenase 1 (HO-1), a type of antioxidant enzyme, attenuates adipogenicity and obesity. We produced transgenic pigs overexpressing human HO-1 (hHO-1-Tg), and found that these animals have little fatty tissue when autopsied. To determine whether overexpressed human HO-1 suppresses adipogenesis in pigs, we analyzed body weight increases of hHO-1-Tg pigs and wild type (WT) pigs of the same strain, and induced adipogenic differentiation of ADSC derived from WT and hHO-1-Tg pigs. The hHO-1-Tg pigs had lower body weights than WT pigs from 16 weeks of age until they died. In addition, hHO-1-Tg ADSC showed reduced adipogenic differentiation and expression of adipogenic molecular markers such as PPARγ and C/EBPα compared to WT ADSC. These results suggest that HO-1 overexpression reduces adipogenesis both in vivo and in vitro, which could support identification of therapeutic targets of obesity and related metabolic diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

Apoptotic adipose-derived mesenchymal stem cell therapy protects against lung and kidney injury in sepsis syndrome caused by cecal ligation puncture in rats

PubMed Central

2013-01-01

Introduction We tested the hypothesis that apoptotic adipose-derived mesenchymal stem cells (A-ADMSC) are superior to healthy (H)-ADMSC in attenuating cecal ligation puncture (CLP)-induced sepsis-mediated lung and kidney injuries. Methods Adult male rats divided into group 1 (sham controls), group 2 (CLP), group 3 [CLP + H-ADMSC administered at 0.5, 6, and 18 hours after CLP], and group 4 [CLP + A-ADMSC administered as in group 3] were sacrificed 72 hours after CLP with blood, lung, and kidney collected for studies. Results White blood cell (WBC) count, circulating TNF-α and creatinine levels were higher in groups 2 and 3 than in groups 1 and 4 (all P < 0.001). Kidney and lung damage scores were highest in group 2, lowest in group 1, significantly higher in group 3 than in group 4 (all P < 0.0001). Protein expressions of inflammatory (ICAM-1, MMP-9, TNF-α, NF-κB), oxidative, and apoptotic (Bax, caspase-3, PARP) biomarkers were higher in groups 2 and 3 than groups 1 and 4, whereas anti-apoptotic (Bcl-2) and mitochondrial integrity (cytochrome-C) biomarkers were lower in groups 2 and 3 than in groups 1 and 4 (all P < 0.001). Expressions of anti-oxidant biomarkers at protein (GR, GPx, NQO-1, HO-1) and cellular (GR, GPx) levels were highest in group 4 (all P < 0.001). The number of inflammatory cells (CD3+) in lungs and levels of DNA damage marker (γ-H2AX) in kidneys were higher in groups 2 and 3 than in groups 1 and 4 (all P < 0.001). Conclusions A-ADMSC therapy was superior to H-ADMSC therapy in protecting major organs from damage in rats with CLP-induced sepsis syndrome. PMID:24451364

Upregulation of CC Chemokine Receptor 7 (CCR7) Enables Migration of Xenogeneic Human Adipose-Derived Mesenchymal Stem Cells to Rat Secondary Lymphoid Organs.

PubMed

Ma, Tian; Luan, Shao-Liang; Huang, Hong; Sun, Xing-Kun; Yang, Yan-Mei; Zhang, Hui; Han, Wei-Dong; Li, Hong; Han, Yan

2016-12-30

BACKGROUND CC chemokine receptor 7 (CCR7) expression is vital for cell migration to secondary lymphoid organs (SLOs). Our previous work showed that inducing CCR7 expression enabled syngeneic mesenchymal stem cells (MSCs) to migrate into SLOs, resulting in enhanced immunosuppressive performance in mice. Given that human adipose-derived stem cells (hASCs) are widely used in clinical therapy, we further investigated whether upregulation of CCR7 enables xenogeneic hASCs to migrate to rat SLOs. MATERIAL AND METHODS hASCs rarely express CCR7; therefore, hASCs were transfected with lentivirus encoding rat CCR7 (rCCR7) plus green fluorescence protein (GFP) or GFP alone. CCR7 mRNA and cell surface expression of rCCR7-hASCs and GFP-hASCs were examined by reverse transcription-polymerase chain reaction (RT-PCR) and flow cytometry (FCM), respectively. The phenotype, differentiation, and proliferation capacity of each cell type was also determined. To examine migration, rCCR7-hASCs and GFP-hASCs were injected intravenously into Lewis rats, and the proportion of GFP-positive cells in the spleen and lymph nodes was determined with FCM. RESULTS mRNA and cell surface protein expression of CCR7 was essentially undetectable in hASCs and GFP-ASCs; however, CCR7 was highly expressed in rCCR7-ASCs. rCCR7-hASCs, GFP-hASCs, and hASCs shared a similar immunophenotype, and maintained the ability of multilineage differentiation and proliferation. In addition, the average proportion of GFP-positive cells was significantly higher following transplantation of rCCR7-hASCs compared with GFP-hASCs (p<0.01). CONCLUSIONS These results suggest that upregulation of rat CCR7 expression does not change the phenotype, differentiation, or proliferation capacity of hASCs, but does enable efficient migration of hASCs to rat SLOs.

Hypoxia enhances the protective effects of placenta-derived mesenchymal stem cells against scar formation through hypoxia-inducible factor-1α.

PubMed

Du, Lili; Lv, Runxiao; Yang, Xiaoyi; Cheng, Shaohang; Xu, Jing; Ma, Tingxian

2016-06-01

To explore the effect of placenta-derived mesenchymal stem cells on scar formation as well as the underlying mechanism. The isolated placenta-derived mesenchymal stem cells from mice were distributed in the wounded areas of scalded mouse models, attenuated inflammatory responses and decreased the deposition of collagens, thus performing a beneficial effect against scar formation. Hypoxia enhanced the protective effect of placenta-derived mesenchymal stem cells and hypoxia-inducible factor-1α was involved in the protective effect of placenta-derived mesenchymal stem cells in hypoxic condition. Hypoxia enhanced the protective effect of placenta-derived mesenchymal stem cells through hypoxia-inducible factor-1α and PMSCs may have a potential application in the treatment of wound.

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

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

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

The efficacy of polycaprolactone/hydroxyapatite scaffold in combination with mesenchymal stem cells for bone tissue engineering.

PubMed

Chuenjitkuntaworn, Boontharika; Osathanon, Thanaphum; Nowwarote, Nunthawan; Supaphol, Pitt; Pavasant, Prasit

2016-01-01

Major drawbacks of using an autograft are the possibilities of insufficient bony source and patient's morbidity after operation. Bone tissue engineering technology, therefore, has been applied for repairing bony defects. Previous study showed that a novel fabricated 3D-Polycaprolactone/Hydroxyapatite (PCL/HAp) scaffold possessed a good biocompatibility for bone cells. This study aimed to determine the ability of PCL/HAp for supporting cell growth, gene expression, and osteogenic differentiation in three types of mesenchymal stem cells, including bone marrow-derived mesenchymal stem cells (BMSCs), dental pulp stem cells (DPSCs), and adiposed-derived mesenchymal stem cells (ADSCs). These were assessed by cell viability assay (MTT), reverse-transcription polymerase chain reaction (RT-PCR) analysis, alkaline phosphatase activity, and osteogenic differentiation by alizarin red-S staining. The results showed that PCL/HAp scaffold could support growth of all three types of mesenchymal stem cells. In addition, DPSCs with PCL/HAp showed the highest level of calcium deposition compared to other groups. In conclusion, DPSCs exhibited a better compatibility with these scaffolds compared to BMSCs and ADSCs. However, the PCL/HAp could be a good candidate scaffold for all tested mesenchymal stem cells in bone tissue engineering. © 2015 Wiley Periodicals, Inc.

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.

Protective effect of adipose-derived mesenchymal stem cells against acute kidney injury induced by ischemia-reperfusion in Sprague-Dawley rats.

PubMed

Sheashaa, Hussein; Lotfy, Ahmed; Elhusseini, Fatma; Aziz, Azza Abdel; Baiomy, Azza; Awad, Samah; Alsayed, Aziza; El-Gilany, Abdel-Hady; Saad, Mohamed-Ahdy A A; Mahmoud, Khaled; Zahran, Faten; Salem, Dalia A; Sarhan, Ahmed; Ghaffar, Hassan Abdel; Sobh, Mohamed

2016-05-01

Acute kidney injury (AKI) is a complex clinical condition associated with significant morbidity and mortality and lacking effective management. Ischemia-reperfusion injury (IRI) remains one of the leading causes of AKI in native and transplanted kidneys. The aim of this study was to evaluate the efficacy of adipose-derived mesenchymal stem cells (ADSCs) in the prevention of renal IRI in rats. The study was conducted on male Sprague-Dawley rats (n=72) weighing 250-300 g. Rats were randomly assigned to three main groups: i) Sham-operated control group (n=24); ii) positive control group, in which rats were subjected to IRI and were administered culture media following 4 h of IRI (n=24); and iii) ADSC group (n=24), in which rats were administered 1×10 6 ADSCs via the tail vein following 4 h of IRI. Each main group was further divided according to the timing after IRI into four equal-sized subgroups. Renal function was tested via the measurement of serum creatinine levels and creatinine clearance. In addition, malondialdehyde (MDA) levels were determined in serum and renal tissue homogenate as an indicator of oxidative stress. Histopathological changes were analyzed in different regions of the kidney, namely the cortex, outer stripe of the outer medulla (OSOM), inner stripe of the outer medulla (ISOM) and inner medulla. In each region, the scoring system considered active injury changes, regenerative changes and chronic changes. The ADSCs were assessed and their differentiation capability was verified. IRI resulted in a significant increase in serum creatinine, serum and tissue MDA levels and a significant reduction in creatinine clearance compared with those in sham-operated rats,. These changes were attenuated by the use of ADSCs. The prominent histopathological changes in the cortex, ISOM and OSOM were reflected in the injury score, which was significantly evident in the positive control group. The use of ADSCs was associated with significantly lowered injury scores

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

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

Transplantation of autologous bone marrow-derived mesenchymal stem cells for traumatic brain injury☆

PubMed Central

Jiang, Jindou; Bu, Xingyao; Liu, Meng; Cheng, Peixun

2012-01-01

Results from the present study demonstrated that transplantation of autologous bone marrow-derived mesenchymal stem cells into the lesion site in rat brain significantly ameliorated brain tissue pathological changes and brain edema, attenuated glial cell proliferation, and increased brain-derived neurotrophic factor expression. In addition, the number of cells double-labeled for 5-bromodeoxyuridine/glial fibrillary acidic protein and cells expressing nestin increased. Finally, blood vessels were newly generated, and the rats exhibited improved motor and cognitive functions. These results suggested that transplantation of autologous bone marrow-derived mesenchymal stem cells promoted brain remodeling and improved neurological functions following traumatic brain injury. PMID:25806058

The Use Of Laser Irradiation To Stimulate Adipose Derived Stem Cell Proliferation And Differentiation For Use In Autologous Grafts

NASA Astrophysics Data System (ADS)

Abrahamse, Heidi

2009-09-01

Stem cells are characterized by the qualities of self-renewal, long term viability, and the ability to differentiate into various cell types. Historically, stem cells have been isolated from the inner cell mass of blastocysts and harvesting these cells resulted in the death of the embryo leading to religious, political and ethical issues. The identification and subsequent isolation of adult stem cells from bone marrow stroma have been welcomed as an alternate source for stem cells. The clinical use of Mesenchymal Stem Cells (MSCs) presented problems such as limited cell number, pain and morbidity upon isolation. Adipose tissue is derived from the mesenchyme, is easily isolated, a reliable source of stem cells and able to differentiate into different cell types including smooth muscle. Over the past few years, the identification and characterization of stem cells has led the potential use of these cells as a promising alternative to cell replacement therapy. Smooth muscle is a major component of human tissues and is essential for the normal functioning of many different organs. Low intensity laser irradiation has been shown to increase viability, protein expression and migration of stem cells in vitro, and to stimulate proliferation of various types of stem cells. In addition, the use of laser irradiation to stimulate differentiation in the absence of growth factors has also been demonstrated in normal human neural progenitor cells (NHNPCs) in vitro where NHNPCs are not only capable of being sustained by light in the absence of growth factors, but that they are also able to differentiate normally as assessed by neurite formation. Our work has focused on the ability of laser irradiation to proliferate adipose derived stem cells (ADSCs), maintain ADSC character and increase the rate and maintenance of differentiation of ADSCs into smooth muscle and skin fibroblast cells. Current studies are also investigating the effect of different irradiation wavelengths and

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.

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.

Influence of Different ECM-Like Hydrogels on Neurite Outgrowth Induced by Adipose Tissue-Derived Stem Cells

PubMed Central

Oliveira, E.; Assunção-Silva, R. C.; Teixeira, F. G.

2017-01-01

Mesenchymal stem cells (MSCs) have been proposed for spinal cord injury (SCI) applications due to their capacity to secrete growth factors and vesicles—secretome—that impacts important phenomena in SCI regeneration. To improve MSC survival into SCI sites, hydrogels have been used as transplantation vehicles. Herein, we hypothesized if different hydrogels could interact differently with adipose tissue-derived MSCs (ASCs). The efficacy of three natural hydrogels, gellan gum (functionalized with a fibronectin peptide), collagen, and a hydrogel rich in laminin epitopes (NVR-gel) in promoting neuritogenesis (alone and cocultured with ASCs), was evaluated in the present study. Their impact on ASC survival, metabolic activity, and gene expression was also evaluated. Our results indicated that all hydrogels supported ASC survival and viability, being this more evident for the functionalized GG hydrogels. Moreover, the presence of different ECM-derived biological cues within the hydrogels appears to differently affect the mRNA levels of growth factors involved in neuronal survival, differentiation, and axonal outgrowth. All the hydrogel-based systems supported axonal growth mediated by ASCs, but this effect was more robust in functionalized GG. The data herein presented highlights the importance of biological cues within hydrogel-based biomaterials as possible modulators of ASC secretome and its effects for SCI applications. PMID:29333166

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

Restoring the IL-1β/NF-κB-induced impaired chondrogenesis by diallyl disulfide in human adipose-derived mesenchymal stem cells via attenuation of reactive oxygen species and elevation of antioxidant enzymes.

PubMed

Bahrampour Juybari, Kobra; Kamarul, Tunku; Najafi, Mohammad; Jafari, Davood; Sharifi, Ali Mohammad

2018-03-26

Strategies based on mesenchymal stem cell (MSC) therapy for restoring injured articular cartilage are not effective enough in osteoarthritis (OA). Due to the enhanced inflammation and oxidative stress in OA microenvironment, differentiation of MSCs into chondrocytes would be impaired. This study aims to explore the effects of diallyl disulfide (DADS) on IL-1β-mediated inflammation and oxidative stress in human adipose derived mesenchymal stem cells (hADSCs) during chondrogenesis. MTT assay was employed to examine the effects of various concentrations of DADS on the viability of hADSCs at different time scales to obtain non-cytotoxic concentration range of DADS. The effects of DADS on IL-1β-induced intracellular ROS generation and lipid peroxidation were evaluated in hADSCs. Western blotting was used to analyze the protein expression levels of IκBα (np), IκBα (p), NF-κB (np) and NF-κB (p). Furthermore, the gene expression levels of antioxidant enzymes in hADSCs and chondrogenic markers at days 7, 14 and 21 of differentiation were measured using qRT-PCR. The results showed that addition of DADS significantly enhanced the mRNA expression levels of antioxidant enzymes as well as reduced ROS elevation, lipid peroxidation, IκBα activation and NF-κB nuclear translocation in hADSCs treated with IL-1β. In addition, DADS could significantly increase the expression levels of IL-1β-induced impaired chondrogenic marker genes in differentiated hADSCs. Treatment with DADS may provide an effective approach to prevent the pro-inflammatory cytokines and oxidative stress as catabolic causes of chondrocyte cell death and enhance the protective anabolic effects by promoting chondrogenesis associated gene expressions in hADSCs exposed to OA condition.

Adipogenesis and epicardial adipose tissue: A novel fate of the epicardium induced by mesenchymal transformation and PPARγ activation

PubMed Central

Yamaguchi, Yukiko; Cavallero, Susana; Patterson, Michaela; Shen, Hua; Xu, Jian; Kumar, S. Ram; Sucov, Henry M.

2015-01-01

The hearts of many mammalian species are surrounded by an extensive layer of fat called epicardial adipose tissue (EAT). The lineage origins and determinative mechanisms of EAT development are unclear, in part because mice and other experimentally tractable model organisms are thought to not have this tissue. In this study, we show that mouse hearts have EAT, localized to a specific region in the atrial–ventricular groove. Lineage analysis indicates that this adipose tissue originates from the epicardium, a multipotent epithelium that until now is only established to normally generate cardiac fibroblasts and coronary smooth muscle cells. We show that adoption of the adipocyte fate in vivo requires activation of the peroxisome proliferator activated receptor gamma (PPARγ) pathway, and that this fate can be ectopically induced in mouse ventricular epicardium, either in embryonic or adult stages, by expression and activation of PPARγ at times of epicardium–mesenchymal transformation. Human embryonic ventricular epicardial cells natively express PPARγ, which explains the abundant presence of fat seen in human hearts at birth and throughout life. PMID:25646471

In-vitro generation of interleukin-10 secreting B-regulatory cells from donor adipose tissue derived mesenchymal stem cells and recipient peripheral blood mononuclear cells for potential cell therapy.

PubMed

Gupte, Kunal S; Vanikar, Aruna V; Trivedi, Hargovind L; Patel, Chetan N; Patel, Jignesh V

2017-02-01

Interleukin-10 secreting B-cells are a major subset of B-regulatory cells (B-regs), commonly recognized as CD19 + /38 hi /24 hi /IL10 + . They carry out immunomodulation by release of specific cytokines and/or cell-to-cell contact. We have generated B-regs in-vitro from donor adipose tissue derived mesenchymal stem cells (AD-MSC) and renal allograft recipient (RAR) peripheral blood mononuclear cells (PBMC) for potential cell therapy. Mononuclear cells separated by density gradient centrifugation from 50 ml anti-coagulated blood of 15-RAR and respective donors were analysed for baseline B-regs using appropriate antibodies. Equal amount (20 × 10 6  cells/ml) of stimulator (irradiated at 7.45 Gy/min for 10 min) and responder (non-irradiated) cells were co-cultured with in-vitro generated AD-MSC (1 × 10 6  cells/ml) in proliferation medium containing lipopolysaccharide from E. coli K12 strain at 37 °C with 5% CO 2 . Cells were harvested on day-7 and analyzed for viability, sterility, quantity, morphology and phenotyping. In-vitro generated B-reg levels were compared with baseline B-regs. In-vitro generated B-reg count increased to 16.75% from baseline count of 3.35%. B-regs can be successfully generated in-vitro from donor AD-MSC and RAR PBMC for potential cell therapy. Copyright © 2017 Chang Gung University. Published by Elsevier B.V. All rights reserved.

Differentiation of Human Adipose Derived Stem Cells into Smooth Muscle Cells Is Modulated by CaMKIIγ

PubMed Central

Aji, Kaisaier; Maimaijiang, Munila; Aimaiti, Abudusaimi; Rexiati, Mulati; Azhati, Baihetiya; Tusong, Hamulati

2016-01-01

The multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII) is known to participate in maintenance and switches of smooth muscle cell (SMC) phenotypes. However, which isoform of CaMKII is involved in differentiation of adult mesenchymal stem cells into contractile SMCs remains unclear. In the present study, we detected γ isoform of CaMKII in differentiation of human adipose derived stem cells (hASCs) into SMCs that resulted from treatment with TGF-β1 and BMP4 in combination for 7 days. The results showed that CaMKIIγ increased gradually during differentiation of hASCs as determined by real-time PCR and western blot analysis. The siRNA-mediated knockdown of CaMKIIγ decreased the protein levels and transcriptional levels of smooth muscle contractile markers (a-SMA, SM22a, calponin, and SM-MHC), while CaMKIIγ overexpression increases the transcriptional and protein levels of smooth muscle contractile markers. These results suggested that γ isoform of CaMKII plays a significant role in smooth muscle differentiation of hASCs. PMID:27493668

Mesenchymal stem cell therapy for cutaneous radiation syndrome.

PubMed

Akita, Sadanori; Akino, Kozo; Hirano, Akiyoshi; Ohtsuru, Akira; Yamashita, Shunichi

2010-06-01

Systemic and local radiation injuries caused by nuclear power reactor accidents, therapeutic irradiation, or nuclear terrorism should be prevented or properly treated in order to improve wound management and save lives. Currently, regenerative surgical modalities should be attempted with temporal artificial dermis impregnated and sprayed with a local angiogenic factor such as basic fibroblast growth factor, and secondary reconstruction can be a candidate for demarcation and saving the donor morbidity. Human mesenchymal stem cells and 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 were tested for differentiation and local stimulation effects in the radiation-exposed wounds. The perforator flap and artificial dermal template with growth factor were successful for reconstruction in patients who were suffering from complex underlying disease. Patients were uneventfully treated with minimal morbidities. In the experiments, the hMSCs are strongly proliferative even after 20 Gy irradiation in vitro. In vivo, 4 Gy rat whole body irradiation demonstrated that sustained marrow stromal (mesenchymal stem) cells survived in the bone marrow. Immediate artificial dermis application impregnated with cells and the cytokine over the 20 Gy irradiated skin and soft tissues demonstrated the significantly improved fat angiogenesis, architected dermal reconstitution, and less inflammatory epidermal recovery. Detailed understanding of underlying diseases and rational reconstructive procedures brings about good outcomes for difficult irradiated wound healing. Adipose-derived stem cells are also implicated in the limited local injuries for short cell harvesting and processing time in the same subject.

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

Generation of a transplantable erythropoietin-producer derived from human mesenchymal stem cells.

PubMed

Yokoo, Takashi; Fukui, Akira; Matsumoto, Kei; Ohashi, Toya; Sado, Yoshikazu; Suzuki, Hideaki; Kawamura, Tetsuya; Okabe, Masataka; Hosoya, Tatsuo; Kobayashi, Eiji

2008-06-15

Differentiation of autologous stem cells into functional transplantable tissue for organ regeneration is a promising regenerative therapeutic approach for cancer, diabetes, and many human diseases. Yet to be established, however, is differentiation into tissue capable of producing erythropoietin (EPO), which has a critical function in anemia. We report a novel EPO-producing organ-like structure (organoid) derived from human mesenchymal stem cells. Using our previously established relay culture system, a human mesenchymal stem cell-derived, human EPO-competent organoid was established in rat omentum. The organoid-derived levels of human EPO increased in response to anemia induced by rapid blood withdrawal. In addition, the presence of an organoid in rats suppressed for native (rat) EPO production enhanced recovery from anemia when compared with control animals lacking the organoid. Together these results confirmed the generation of a stem cell-derived organoid that is capable of producing EPO and sensitive to physiological regulation.

Distinct requirements for cranial ectoderm and mesenchyme-derived wnts in specification and differentiation of osteoblast and dermal progenitors.

PubMed

Goodnough, L Henry; Dinuoscio, Gregg J; Ferguson, James W; Williams, Trevor; Lang, Richard A; Atit, Radhika P

2014-02-01

The cranial bones and dermis differentiate from mesenchyme beneath the surface ectoderm. Fate selection in cranial mesenchyme requires the canonical Wnt effector molecule β-catenin, but the relative contribution of Wnt ligand sources in this process remains unknown. Here we show Wnt ligands are expressed in cranial surface ectoderm and underlying supraorbital mesenchyme during dermal and osteoblast fate selection. Using conditional genetics, we eliminate secretion of all Wnt ligands from cranial surface ectoderm or undifferentiated mesenchyme, to uncover distinct roles for ectoderm- and mesenchyme-derived Wnts. Ectoderm Wnt ligands induce osteoblast and dermal fibroblast progenitor specification while initiating expression of a subset of mesenchymal Wnts. Mesenchyme Wnt ligands are subsequently essential during differentiation of dermal and osteoblast progenitors. Finally, ectoderm-derived Wnt ligands provide an inductive cue to the cranial mesenchyme for the fate selection of dermal fibroblast and osteoblast lineages. Thus two sources of Wnt ligands perform distinct functions during osteoblast and dermal fibroblast formation.

Paracrine Activity from Adipose-Derived Stem Cells on In Vitro Wound Healing in Human Tympanic Membrane Keratinocytes.

PubMed

Ong, Huan Ting; Redmond, Sharon L; Marano, Robert J; Atlas, Marcus D; von Unge, Magnus; Aabel, Peder; Dilley, Rodney J

2017-03-15

Stem cell therapies for tympanic membrane repair have shown initial experimental success using mesenchymal stem cells in rat models to promote healing; however, the mechanisms providing this benefit are not known. We investigated in vitro the paracrine effects of human adipose-derived stem cells (ADSCs) on wound healing mechanisms for human tympanic membrane-derived keratinocytes (hTM) and immortalized human keratinocytes (HaCaT). ADSC conditioned media (CM ADSC ) were assessed for paracrine activity on keratinocyte proliferation and migration, with hypoxic conditions for ADSC culture used to generate contrasting effects on cytokine gene expression. Keratinocytes cultured in CM ADSC showed a significant increase in cell number compared to serum-free cultures and further significant increases in hypoxic CM ADSC . Assessment of ADSC gene expression on a cytokine array showed a range of wound healing cytokines expressed and under stringent hypoxic and serum-free conditions was upregulated (VEGF A, MMP9, Tissue Factor, PAI-1) or downregulated (CXCL5, CCL7, TNF-α). Several of these may contribute to the activity of conditioned media on the keratinocytes with potential applications in TM perforation repair. VEGFA protein was confirmed by immunoassay to be increased in conditioned media. Together with gene regulation associated with hypoxia in ADSCs, this study has provided several strong leads for a stem cell-derived approach to TM wound healing.

Role of mesenchymal stem cells versus angiotensin converting enzyme inhibitor in kidney repair.

PubMed

Ahmed, Hanaa H; Toson, Elshahat A; El-Mezayen, Hatem A; Rashed, Laila A; Elsherbiny, Eslam S

2017-07-01

The current study sought to clarify the role of bone marrow derived mesenchymal stem cells (BM-MSCs) and adipose tissue derived mesenchymal stem cells (AD-MSCs) in repressing nephropathy in the experimental model. Moreover, the aim of this work was extended to compare between stem cells role and angiotensin converting enzyme inhibitor in kidney repair. Isolation and preparation of MSCs culture, flow cytometry using CD34, CD44 and CD105 cell surface markers, biochemical analyses for determination of serum creatinine, urea, transforming growth factor β (TGF-β), cystatin C (CYS-C) and urinary N-Acetyl-ß-D-Glucosaminidase (UNAG), and histopathological investigation of kidney tissue sections were performed. The results of the present study revealed that single intravenous infusion of MSCs either derived from bone marrow or adipose tissue was able to enhance renal reparative processes through significantly decreased serum creatinine, urea, TGF-β and CYS-C levels as well as UNAG level and significantly increase glomerular filtration rate. Additionally, the histopathological investigations of kidney tissues showed that MSCs have significant regenerative effects as evidenced by the decrease in focal inflammatory cells infiltration, focal interstitial nephritis and congested glomeruli as well as degenerated tubules. The current data provided distinct evidence about the favourable impact of AD-MSCs and BM-MSCs in attenuation of cyclosporine-induced nephropathy in rats through their ability to promote functional and structural kidney repair via transdifferentiation. © 2016 Asian Pacific Society of Nephrology.

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.

[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.

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

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.

I.V. infusion of brain-derived neurotrophic factor gene-modified human mesenchymal stem cells protects against injury in a cerebral ischemia model in adult rat.

PubMed

Nomura, T; Honmou, O; Harada, K; Houkin, K; Hamada, H; Kocsis, J D

2005-01-01

I.V. delivery of mesenchymal stem cells prepared from adult bone marrow reduces infarction size and ameliorates functional deficits in rat cerebral ischemia models. Administration of the brain-derived neurotrophic factor to the infarction site has also been demonstrated to be neuroprotective. To test the hypothesis that brain-derived neurotrophic factor contributes to the therapeutic benefits of mesenchymal stem cell delivery, we compared the efficacy of systemic delivery of human mesenchymal stem cells and human mesenchymal stem cells transfected with a fiber-mutant F/RGD adenovirus vector with a brain-derived neurotrophic factor gene (brain-derived neurotrophic factor-human mesenchymal stem cells). A permanent middle cerebral artery occlusion was induced by intraluminal vascular occlusion with a microfilament. Human mesenchymal stem cells and brain-derived neurotrophic factor-human mesenchymal stem cells were i.v. injected into the rats 6 h after middle cerebral artery occlusion. Lesion size was assessed at 6 h, 1, 3 and 7 days using MR imaging, and histological methods. Functional outcome was assessed using the treadmill stress test. Both human mesenchymal stem cells and brain-derived neurotrophic factor-human mesenchymal stem cells reduced lesion volume and elicited functional improvement compared with the control sham group, but the effect was greater in the brain-derived neurotrophic factor-human mesenchymal stem cell group. ELISA analysis of the infarcted hemisphere revealed an increase in brain-derived neurotrophic factor in the human mesenchymal stem cell groups, but a greater increase in the brain-derived neurotrophic factor-human mesenchymal stem cell group. These data support the hypothesis that brain-derived neurotrophic factor contributes to neuroprotection in cerebral ischemia and cellular delivery of brain-derived neurotrophic factor can be achieved by i.v. delivery of human mesenchymal stem cells.

Generation of functional islets from human umbilical cord and placenta derived mesenchymal stem cells.

PubMed

Kadam, Sachin; Govindasamy, Vijayendran; Bhonde, Ramesh

2012-01-01

Bone marrow-derived mesenchymal stem cells (BM-MSCs) have been used for allogeneic application in tissue engineering but have certain drawbacks. Therefore, mesenchymal stem cells (MSCs) derived from other adult tissue sources have been considered as an alternative. The human umbilical cord and placenta are easily available noncontroversial sources of human tissue, which are often discarded as biological waste, and their collection is noninvasive. These sources of MSCs are not subjected to ethical constraints, as in the case of embryonic stem cells. MSCs derived from umbilical cord and placenta are multipotent and have the ability to differentiate into various cell types crossing the lineage boundary towards endodermal lineage. The aim of this chapter is to provide a detailed reproducible cookbook protocol for the isolation, propagation, characterization, and differentiation of MSCs derived from human umbilical cord and placenta with special reference to harnessing their potential towards pancreatic/islet lineage for utilization as a cell therapy product. We show here that mesenchymal stromal cells can be extensively expanded from umbilical cord and placenta of human origin retaining their multilineage differentiation potential in vitro. Our report indicates that postnatal tissues obtained as delivery waste represent a rich source of mesenchymal stromal cells, which can be differentiated into functional islets employing three-stage protocol developed by our group. These islets could be used as novel in vitro model for screening hypoglycemics/insulin secretagogues, thus reducing animal experimentation for this purpose and for the future human islet transplantation programs to treat diabetes.

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

Impact of apoptotic adipose-derived mesenchymal stem cells on attenuating organ damage and reducing mortality in rat sepsis syndrome induced by cecal puncture and ligation.

PubMed

Chang, Chia-Lo; Leu, Steve; Sung, Hsin-Ching; Zhen, Yen-Yi; Cho, Chung-Lung; Chen, Angela; Tsai, Tzu-Hsien; Chung, Sheng-Ying; Chai, Han-Tan; Sun, Cheuk-Kwan; Yen, Chia-Hung; Yip, Hon-Kan

2012-12-07

We tested whether apoptotic adipose-derived mesenchymal stem cells (A-ADMSCs) were superior to healthy (H)-ADMSCs at attenuating organ damage and mortality in sepsis syndrome following cecal ligation and puncture (CLP). Adult male rats were categorized into group 1 (sham control), group 2 (CLP), group 3 [CLP + H-ADMSC administered 0.5, 6, and 18 h after CLP], group 4 [CLP + A-ADMSC administered as per group 3]. Circulating peak TNF-α level, at 6 h, was highest in groups 2 and 3, and higher in group 4 than group 1 (p < 0.0001). Immune reactivity (indicated by circulating and splenic helper-, cytoxic-, and regulatory-T cells) at 24 and 72 h exhibited the same pattern as TNF-α amongst the groups (all p < 0.0001). The mononuclear-cell early and late apoptosis level and organ damage parameters of liver (AST, ALT), kidney (creatinine) and lung (arterial oxygen saturation) also displayed a similar pattern to TNF-α levels (all p < 0.001). Protein levels of inflammatory (TNF-α, MMP-9, NF-κB, ICAM-1), oxidative (oxidized protein) and apoptotic (Bax, caspase-3, PARP) biomarkers were higher in groups 2 and 3 than group 1, whereas anti-apoptotic (Bcl-2) biomarker was lower in groups 2 and 3 than in group 1 but anti-oxidant (GR, GPx, HO-1, NQO-1) showed an opposite way of Bcl-2; these patterns were reversed for group 4 (all p < 0.001). Mortality was highest in group 3 and higher in group 2 than group 4 than group 1 (all p < 0.001). A-ADMSC therapy protected major organs from damage and improved prognosis in rats with sepsis syndrome.

Preservation media, durations and cell concentrations of short-term storage affect key features of human adipose-derived mesenchymal stem cells for therapeutic application.

PubMed

Zhang, Fengli; Ren, Huaijuan; Shao, Xiaohu; Zhuang, Chao; Chen, Yantian; Qi, Nianmin

2017-01-01

Adipose-derived mesenchymal stem cells (ADSCs) have shown great potential in the treatment of various diseases. However, the optimum short-term storage condition of ADSCs in 2∼8 °C is rarely reported. This study aimed at optimizing a short-term storage condition to ensure the viability and function of ADSCs before transplantation. Preservation media and durations of storage were evaluated by cell viability, apoptosis, adhesion ability and colony-forming unit (CFU) capacity of ADSCs. The abilities of cell proliferation and differentiation were used to optimize cell concentrations. Optimized preservation condition was evaluated by cell surface markers, cell cycle and immunosuppressive capacity. A total of 5% human serum albumin in multiple electrolytes (ME + HSA) was the optimized medium with high cell viability, low cluster rate, good adhesion ability and high CFU capacity of ADSCs. Duration of storage should be limited to 24 h to ensure the quality of ADSCs before transplantation. A concentration of 5 × 10 6 cells/ml was the most suitable cell concentration with low late stage apoptosis, rapid proliferation and good osteogenic and adipogenic differentiation ability. This selected condition did not change surface markers, cell cycle, indoleamine 2, 3-dioxygenase 1 (IDO1) gene expression and kynurenine (Kyn) concentration significantly. In this study, ME + HSA was found to be the best medium, most likely due to the supplement of HSA which could protect cells, the physiological pH (7.4) of ME and sodium gluconate ingredient in ME which could provide energy for cells. Duration should be limited to 24 h because of reduced nutrient supply and increased waste and lactic acid accumulation during prolonged storage. To keep cell proliferation and limit lactic acid accumulation, the proper cell concentration is 5× 10 6 cells/ml. Surface markers, cell cycle and immunosuppressive capacity did not change significantly after storage using the optimized condition, which

Gene expression profile in mesenchymal stem cells derived from dental tissues and bone marrow

PubMed Central

Kim, Su-Hwan; Kim, Young-Sung; Lee, Su-Yeon; Kim, Kyoung-Hwa; Lee, Yong-Moo; Kim, Won-Kyung

2011-01-01

Purpose The aim of this study is to compare the gene expression profile in mesenchymal stem cells derived from dental tissues and bone marrow for characterization of dental stem cells. Methods We employed GeneChip analysis to the expression levels of approximately 32,321 kinds of transcripts in 5 samples of bone-marrow-derived mesenchymal stem cells (BMSCs) (n=1), periodontal ligament stem cells (PDLSCs) (n=2), and dental pulp stem cells (DPSCs) (n=2). Each cell was sorted by a FACS Vantage Sorter using immunocytochemical staining of the early mesenchymal stem cell surface marker STRO-1 before the microarray analysis. Results We identified 379 up-regulated and 133 down-regulated transcripts in BMSCs, 68 up-regulated and 64 down-regulated transcripts in PDLSCs, and 218 up-regulated and 231 down-regulated transcripts in DPSCs. In addition, anatomical structure development and anatomical structure morphogenesis gene ontology (GO) terms were over-represented in all three different mesenchymal stem cells and GO terms related to blood vessels, and neurons were over-represented only in DPSCs. Conclusions This study demonstrated the genome-wide gene expression patterns of STRO-1+ mesenchymal stem cells derived from dental tissues and bone marrow. The differences among the expression profiles of BMSCs, PDLSCs, and DPSCs were shown, and 999 candidate genes were found to be definitely up- or down-regulated. In addition, GOstat analyses of regulated gene products provided over-represented GO classes. These data provide a first step for discovering molecules key to the characteristics of dental stem cells. PMID:21954424

Comparative Analysis of Human Mesenchymal Stem Cells from Bone Marrow, Adipose Tissue, and Umbilical Cord Blood as Sources of Cell Therapy

PubMed Central

Jin, Hye Jin; Bae, Yun Kyung; Kim, Miyeon; Kwon, Soon-Jae; Jeon, Hong Bae; Choi, Soo Jin; Kim, Seong Who; Yang, Yoon Sun; Oh, Wonil; Chang, Jong Wook

2013-01-01

Various source-derived mesenchymal stem cells (MSCs) have been considered for cell therapeutics in incurable diseases. To characterize MSCs from different sources, we compared human bone marrow (BM), adipose tissue (AT), and umbilical cord blood-derived MSCs (UCB-MSCs) for surface antigen expression, differentiation ability, proliferation capacity, clonality, tolerance for aging, and paracrine activity. Although MSCs from different tissues have similar levels of surface antigen expression, immunosuppressive activity, and differentiation ability, UCB-MSCs had the highest rate of cell proliferation and clonality, and significantly lower expression of p53, p21, and p16, well known markers of senescence. Since paracrine action is the main action of MSCs, we examined the anti-inflammatory activity of each MSC under lipopolysaccharide (LPS)-induced inflammation. Co-culture of UCB-MSCs with LPS-treated rat alveolar macrophage, reduced expression of inflammatory cytokines including interleukin-1α (IL-1α), IL-6, and IL-8 via angiopoietin-1 (Ang-1). Using recombinant Ang-1 as potential soluble paracrine factor or its small interference RNA (siRNA), we found that Ang-1 secretion was responsible for this beneficial effect in part by preventing inflammation. Our results demonstrate that primitive UCB-MSCs have biological advantages in comparison to adult sources, making UCB-MSCs a useful model for clinical applications of cell therapy. PMID:24005862

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.

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.

Does the liposuction method influence the phenotypic characteristic of human adipose-derived stem cells?

PubMed

Bajek, Anna; Gurtowska, Natalia; Gackowska, Lidia; Kubiszewska, Izabela; Bodnar, Magdalena; Marszałek, Andrzej; Januszewski, Rafał; Michalkiewicz, Jacek; Drewa, Tomasz

2015-05-14

Adipose-derived stem cells (ASCs) possess a high differentiation and proliferation potential. However, the phenotypic characterization of ASCs is still difficult. Until now, there is no extensive analysis of ASCs markers depending on different liposuction methods. Therefore, the aim of the present study was to analyse 242 surface markers and determine the differences in the phenotypic pattern between ASCs obtained during mechanical and ultrasound-assisted liposuction. ASCs were isolated from healthy donors, due to mechanical and ultrasound-assisted liposuction and cultured in standard medium to the second passage. Differentiation potential and markers expression was evaluated to confirm the mesenchymal nature of cells. Then, the BD LyoplateTM Human Cell Surface Marker Screening Panel was used. Results shown that both population of ASCs are characterized by high expression of markers specific for ASCs: cluster of differentiation (CD)9, CD10, CD34, CD44, CD49d, CD54, CD55, CD59, CD71 and low expression of CD11a, CD11c and CD144. Moreover, we have noticed significant differences in antigen expression in 58 markers from the 242 studied. Presented study shows for the first time that different liposuction methods are not a significant factor which can influence the expression of human ASCs surface markers. © 2015 The Authors.

Adipose tissue-organotypic culture system as a promising model for studying adipose tissue biology and regeneration

PubMed Central

Uchihashi, Kazuyoshi; Aoki, Shigehisa; Sonoda, Emiko; Yamasaki, Fumio; Piao, Meihua; Ootani, Akifumi; Yonemitsu, Nobuhisa; Sugihara, Hajime

2009-01-01

Adipose tissue consists of mature adipocytes, preadipocytes and mesenchymal stem cells (MSCs), but a culture system for analyzing their cell types within the tissue has not been established. We have recently developed “adipose tissue-organotypic culture system” that maintains unilocular structure, proliferative ability and functions of mature adipocytes for a long term, using three-dimensional collagen gel culture of the tissue fragments. In this system, both preadipocytes and MSCs regenerate actively at the peripheral zone of the fragments. Our method will open up a new way for studying both multiple cell types within adipose tissue and the cell-based mechanisms of obesity and metabolic syndrome. Thus, it seems to be a promising model for investigating adipose tissue biology and regeneration. In this article, we introduce adipose tissue-organotypic culture, and propose two theories regarding the mechanism of tissue regeneration that occurs specifically at peripheral zone of tissue fragments in vitro. PMID:19794899

Derivation of Multipotent Mesenchymal Precursors from Human Embryonic Stem Cells

PubMed Central

Barberi, Tiziano; Willis, Lucy M; Socci, Nicholas D; Studer, Lorenz

2005-01-01

Background Human embryonic stem cells provide access to the earliest stages of human development and may serve as a source of specialized cells for regenerative medicine. Thus, it becomes crucial to develop protocols for the directed differentiation of embryonic stem cells into tissue-restricted precursors. Methods and Findings Here, we present culture conditions for the derivation of unlimited numbers of pure mesenchymal precursors from human embryonic stem cells and demonstrate multilineage differentiation into fat, cartilage, bone, and skeletal muscle cells. Conclusion Our findings will help to elucidate the mechanism of mesoderm specification during embryonic stem cell differentiation and provide a platform to efficiently generate specialized human mesenchymal cell types for future clinical applications. PMID:15971941

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

[Analysis of factors related to the number of mesenchymal stem cells derived from synovial fluid of the temporomandibular joint].

PubMed

Sun, Y P; Zheng, Y H; Zhang, Z G

2017-06-09

Objective: To analyze related factors on the number of mesenchymal stem cells in the synovial fluid of the temporomandibular joint (TMJ) and provide an research basis for understanding of the source and biological role of mesenchymal stem cells derived from synovial fluid in TMJ. Methods: One hundred and twenty-two synovial fluid samples from 91 temporomandibular disorders (TMD) patients who visited in Department of TMJ Center, Guanghua School of Stomatology, Hospital of Stomatology, Sun Yat-sen University from March 2013 to December 2013 were collected in this study, and 6 TMJ synovial fluid samples from 6 normal volunteers who were studying in the North Campus of Sun Yat-sen University were also collected, so did their clinical information. Then the relation between the number of mesenchymal stem cells derived from synovial fluid and the health status of the joints, age of donor, disc perforation, condylar bony destruction, blood containing and visual analogue scale score of pain were investigated using Mann-Whitney U test and Spearman rank correlation test. Results: The number of mesenchymal stem cells derived from synovial fluid had no significant relation with visual analogue scale score of pain ( r= 0.041, P= 0.672), blood containing ( P= 0.063), condylar bony destruction ( P= 0.371). Linear correlation between the number of mesenchymal stem cells derived from synovial fluid and age of donor was very week ( r= 0.186, P= 0.043). The number of mesenchymal stem cells up-regulated when the joint was in a disease state ( P= 0.001). The disc perforation group had more mesenchymal stem cells in synovial fluid than without disc perforation group ( P= 0.042). Conclusions: The number of mesenchymal stem cells derived from synovial fluid in TMJ has no correlation with peripheral blood circulation and condylar bony destruction, while has close relation with soft tissue structure damage of the joint.

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.

Tracking and Therapeutic Value of Human Adipose Tissue–derived Mesenchymal Stem Cell Transplantation in Reducing Venous Neointimal Hyperplasia Associated with Arteriovenous Fistula

PubMed Central

Yang, Binxia; Brahmbhatt, Akshaar; Nieves Torres, Evelyn; Thielen, Brian; McCall, Deborah L.; Engel, Sean; Bansal, Aditya; Pandey, Mukesh K.; Dietz, Allan B.; Leof, Edward B.; DeGrado, Timothy R.; Mukhopadhyay, Debabrata

2016-01-01

Purpose To determine if adventitial transplantation of human adipose tissue–derived mesenchymal stem cells (MSCs) to the outflow vein of B6.Cg-Foxn1nu/J mice with arteriovenous fistula (AVF) at the time of creation would reduce monocyte chemoattractant protein-1 (Mcp-1) gene expression and venous neointimal hyperplasia. The second aim was to track transplanted zirconium 89 (89Zr)–labeled MSCs serially with positron emission tomography (PET) for 21 days. Materials and Methods All animal experiments were performed according to protocols approved by the institutional animal care and use committee. Fifty B6.Cg-Foxn1nu/J mice were used to accomplish the study aims. Green fluorescent protein was used to stably label 2.5 × 105 MSCs, which were injected into the adventitia of the outflow vein at the time of AVF creation in the MSC group. Eleven mice died after AVF placement. Animals were sacrificed on day 7 after AVF placement for real-time polymerase chain reaction (n = 6 for MSC and control groups) and histomorphometric (n = 6 for MSC and control groups) analyses and on day 21 for histomorphometric analysis only (n = 6 for MSC and control groups). In a separate group of experiments (n = 3), animals with transplanted 89Zr-labeled MSCs were serially imaged with PET for 3 weeks. Multiple comparisons were performed with two-way analysis of variance, followed by the Student t test with post hoc Bonferroni correction. Results In vessels with transplanted MSCs compared with control vessels, there was a significant decrease in Mcp-1 gene expression (day 7: mean reduction, 62%; P = .029), with a significant increase in the mean lumen vessel area (day 7: mean increase, 176% [P = .013]; day 21: mean increase, 415% [P = .011]). Moreover, this was accompanied by a significant decrease in Ki-67 index (proliferation on day 7: mean reduction, 81% [P = .0003]; proliferation on day 21: mean reduction, 60%, [P = .016]). Prolonged retention of MSCs at the adventitia was evidenced by

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

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

Selection of Reference Genes for Quantitative Gene Expression in Porcine Mesenchymal Stem Cells Derived from Various Sources along with Differentiation into Multilineages

PubMed Central

Lee, Won-Jae; Jeon, Ryoung-Hoon; Jang, Si-Jung; Park, Ji-Sung; Lee, Seung-Chan; Baregundi Subbarao, Raghavendra; Lee, Sung-Lim; Park, Bong-Wook; King, William Allan; Rho, Gyu-Jin

2015-01-01

The identification of stable reference genes is a prerequisite for ensuring accurate validation of gene expression, yet too little is known about stable reference genes of porcine MSCs. The present study was, therefore, conducted to assess the stability of reference genes in porcine MSCs derived from bone marrow (BMSCs), adipose (AMSCs), and skin (SMSCs) with their in vitro differentiated cells into mesenchymal lineages such as adipocytes, osteocytes, and chondrocytes. Twelve commonly used reference genes were investigated for their threshold cycle (Ct) values by qRT-PCR. The Ct values of candidate reference genes were analyzed by geNorm software to clarify stable expression regardless of experimental conditions. Thus, Pearson's correlation was applied to determine correlation between the three most stable reference genes (NF3) and optimal number of reference genes (NFopt). In assessment of stability of reference gene across experimental conditions by geNorm analysis, undifferentiated MSCs and each differentiated status into mesenchymal lineages showed slightly different results but similar patterns about more or less stable rankings. Furthermore, Pearson's correlation revealed high correlation (r > 0.9) between NF3 and NFopt. Overall, the present study showed that HMBS, YWHAZ, SDHA, and TBP are suitable reference genes for qRT-PCR in porcine MSCs. PMID:25972899

Human Adipose-Derived Mesenchymal Stem Cells Respond to Short-Term Hypoxia by Secreting Factors Beneficial for Human Islets In Vitro and Potentiate Antidiabetic Effect In Vivo

PubMed Central

Schive, Simen W.; Mirlashari, Mohammad Reza; Hasvold, Grete; Wang, Mengyu; Josefsen, Dag; Gullestad, Hans Petter; Korsgren, Olle; Foss, Aksel; Kvalheim, Gunnar; Scholz, Hanne

2017-01-01

Adipose-derived mesenchymal stem cells (ASCs) release factors beneficial for islets in vitro and protect against hyperglycemia in rodent models of diabetes. Oxygen tension has been shown to induce metabolic changes and alter ASCs’ release of soluble factors. The effects of hypoxia on the antidiabetic properties of ASCs have not been explored. To investigate this, we incubated human ASCs for 48 h in 21% (normoxia) or 1% O2 (hypoxia) and compared viability, cell growth, surface markers, differentiation capability, and soluble factors in the conditioned media (CM). Human islets were exposed to CM from ASCs incubated in either normoxia or hypoxia, and islet function and apoptosis after culture with or without proinflammatory cytokines were measured. To test hypoxic preconditioned ASCs’ islet protective effects in vivo, ASCs were incubated for 48 h in normoxia or hypoxia before being injected into Balb/c Rag 1–/– immunodeficient mice with streptozotocin-induced insulitis. Progression of diabetes and insulin content of pancreas were measured. We found that incubation in hypoxia was well tolerated by ASCs and that levels of VEGF-A, FGF-2, and bNGF were elevated in CM from ASCs incubated in hypoxia compared to normoxia, while levels of HGF, IL-8, and CXCL1 were reduced. CM from ASCs incubated in hypoxia significantly improved human islet function and reduced apoptosis after culture, and reduced cytokine-induced apoptosis. In our mouse model, pancreas insulin content was higher in both groups receiving ASCs compared to control, but the mice receiving preconditioned ASCs had lower random and fasting blood glucose, as well as improved oral glucose tolerance compared to untreated mice. In conclusion, our in vitro results indicate that the islet protective potential of ASCs improves in hypoxia, and we give insight into factors involved in this. Finally we show that hypoxic preconditioning potentiates ASCs’ antidiabetic effect in vivo. PMID:28713640

Hyaluronic acid effect on adipose-derived stem cells. Biological in vitro evaluation.

PubMed

Moreno, A; Martínez, A; Olmedillas, S; Bello, S; de Miguel, F

2015-01-01

To evaluate the in vitro effects of hyaluronic acid (HA) on adipose-derived stem cells (ASC) in order to consider the possibility of their combined used in the treatment of knee arthrosis. The ASC cells were grown both in the presence and absence of AH, and several studies were carried out: proliferation (WST8) and cell viability studies (Alamar Blue® and Trypan Blue), possible chondrogenic differentiation (collagen type 2 expression) by RT-PCR, AH receptor expression (CD44) by flow cytometry and RT-QPCR, and expression of inflammatory and anti-inflammatory factors (IL-6, TGFß, IL-10) by RT-QPCR. The number of ASC significantly increased after 7 days with HA (158±39%, p <0.05). Additionally, the cell viability of the ASC treated with HA after 1, 3, 5 and 7 days was similar to that of the control cells, being considered non-toxic. There were no changes observed in the expression of CD44 and chondrogenic differentiation. TGFß expression was not modified after AH treatment, but there was a 4-fold decrease in IL-6 expression and IL-10 expression increased up to 2-fold compared to control cells. Hyaluronic acid favours ASC proliferation without causing cellular toxicity, and inducing an anti-inflammatory profile in these cells. Hyaluronic acid appears to be a suitable vehicle for the intra-articular administration of mesenchymal stem cells. Copyright © 2014 SECOT. Published by Elsevier Espana. All rights reserved.

Age of the Donor Reduces the Ability of Human Adipose-Derived Stem Cells to Alleviate Symptoms in the Experimental Autoimmune Encephalomyelitis Mouse Model

PubMed Central

Scruggs, Brittni A.; Semon, Julie A.; Zhang, Xiujuan; Zhang, Shijia; Bowles, Annie C.; Pandey, Amitabh C.; Imhof, Kathleen M.P.; Kalueff, Allan V.; Gimble, Jeffrey M.

2013-01-01

There is a significant clinical need for effective therapies for primary progressive multiple sclerosis, which presents later in life (i.e., older than 50 years) and has symptoms that increase in severity without remission. With autologous mesenchymal stem cell therapy now in the early phases of clinical trials for all forms of multiple sclerosis (MS), it is necessary to determine whether autologous stem cells from older donors have therapeutic effectiveness. In this study, the therapeutic efficacy of human adipose-derived mesenchymal stem cells (ASCs) from older donors was directly compared with that of cells from younger donors for disease prevention. Mice were induced with chronic experimental autoimmune encephalomyelitis (EAE) using the myelin oligodendrocyte glycoprotein35–55 peptide and treated before disease onset with ASCs derived from younger (<35 years) or older (>60 years) donors. ASCs from older donors failed to ameliorate the neurodegeneration associated with EAE, and mice treated with older donor cells had increased central nervous system inflammation, demyelination, and splenocyte proliferation in vitro compared with the mice receiving cells from younger donors. Therefore, the results of this study demonstrated that donor age significantly affects the ability of human ASCs to provide neuroprotection, immunomodulation, and/or remyelination in EAE mice. The age-related therapeutic differences corroborate recent findings that biologic aging occurs in stem cells, and the differences are supported by evidence in this study that older ASCs, compared with younger donor cells, secrete less hepatocyte growth factor and other bioactive molecules when stimulated in vitro. These results highlight the need for evaluation of autologous ASCs derived from older patients when used as therapy for MS. PMID:24018793

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

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.

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

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.

Adult hippocampus derived soluble factors induce a neuronal-like phenotype in mesenchymal stem cells.

PubMed

Rivera, Francisco J; Sierralta, Walter D; Minguell, Jose J; Aigner, Ludwig

2006-10-02

Bone marrow-derived mesenchymal stem cells (MSCs) are not restricted in their differentiation fate to cells of the mesenchymal lineage. They acquire a neural phenotype in vitro and in vivo after transplantation in the central nervous system. Here we investigated whether soluble factors derived from different brain regions are sufficient to induce a neuronal phenotype in MSCs. We incubated bone marrow-derived MSCs in conditioned medium (CM) derived from adult hippocampus (HCM), cortex (CoCM) or cerebellum (CeCM) and analyzed the cellular morphology and the expression of neuronal and glial markers. In contrast to muscle derived conditioned medium, which served as control, conditioned medium derived from the different brain regions induced a neuronal morphology and the expression of the neuronal markers GAP-43 and neurofilaments in MSCs. Hippocampus derived conditioned medium had the strongest activity. It was independent of NGF or BDNF; and it was restricted to the neuronal differentiation fate, since no induction of the astroglial marker GFAP was observed. The work indicates that soluble factors present in the brain are sufficient to induce a neuronal phenotype in MSCs.

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.

Neural Crest-Derived Mesenchymal Cells Require Wnt Signaling for Their Development and Drive Invagination of the Telencephalic Midline

PubMed Central

Choe, Youngshik; Zarbalis, Konstantinos S.; Pleasure, Samuel J.

2014-01-01

Embryonic neural crest cells contribute to the development of the craniofacial mesenchyme, forebrain meninges and perivascular cells. In this study, we investigated the function of ß-catenin signaling in neural crest cells abutting the dorsal forebrain during development. In the absence of ß-catenin signaling, neural crest cells failed to expand in the interhemispheric region and produced ectopic smooth muscle cells instead of generating dermal and calvarial mesenchyme. In contrast, constitutive expression of stabilized ß-catenin in neural crest cells increased the number of mesenchymal lineage precursors suggesting that ß-catenin signaling is necessary for the expansion of neural crest-derived mesenchymal cells. Interestingly, the loss of neural crest-derived mesenchymal stem cells (MSCs) leads to failure of telencephalic midline invagination and causes ventricular system defects. This study shows that ß-catenin signaling is required for the switch of neural crest cells to MSCs and mediates the expansion of MSCs to drive the formation of mesenchymal structures of the head. Furthermore, loss of these structures causes striking defects in forebrain morphogenesis. PMID:24516524

Research Advancements in Porcine Derived Mesenchymal Stem Cells

PubMed Central

Bharti, Dinesh; Shivakumar, Sharath Belame; Subbarao, Raghavendra Baregundi; Rho, Gyu-Jin

2016-01-01

In the present era of stem cell biology, various animals such as Mouse, Bovine, Rabbit and Porcine have been tested for the efficiency of their mesenchymal stem cells (MSCs) before their actual use for stem cell based application in humans. Among them pigs have many similarities to humans in the form of organ size, physiology and their functioning, therefore they have been considered as a valuable model system for in vitro studies and preclinical assessments. Easy assessability, few ethical issues, successful MSC isolation from different origins like bone marrow, skin, umbilical cord blood, Wharton’s jelly, endometrium, amniotic fluid and peripheral blood make porcine a good model for stem cell therapy. Porcine derived MSCs (pMSCs) have shown greater in vitro differentiation and transdifferention potential towards mesenchymal lineages and specialized lineages such as cardiomyocytes, neurons, hepatocytes and pancreatic beta cells. Immunomodulatory and low immunogenic profiles as shown by autologous and heterologous MSCs proves them safe and appropriate models for xenotransplantation purposes. Furthermore, tissue engineered stem cell constructs can be of immense importance in relation to various osteochondral defects which are difficult to treat otherwise. Using pMSCs successful treatment of various disorders like Parkinson’s disease, cardiac ischemia, hepatic failure, has been reported by many studies. Here, in this review we highlight current research findings in the area of porcine mesenchymal stem cells dealing with their isolation methods, differentiation ability, transplantation applications and their therapeutic potential towards various diseases. PMID:26201864

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.

Pretreatment of Adipose Derived Stem Cells with Curcumin Facilitates Myocardial Recovery via Antiapoptosis and Angiogenesis

PubMed Central

Liu, Jianfeng; Zhu, Ping; Song, Peng; Xiong, Weiping; Chen, Haixu; Peng, Wenhui; Wang, Shuxia; Li, Shan; Fu, Zhiqing; Wang, Yutang; Wang, Haibin

2015-01-01

The poor survival rate of transplanted stem cells in ischemic myocardium has limited their therapeutic efficacy. Curcumin has potent antioxidant property. This study investigates whether prior curcumin treatment protects stem cells from oxidative stress injury and improves myocardial recovery following cells transplantation. Autologous Sprague-Dawley rat adipose derived mesenchymal stem cells (ADSCs) were pretreated with or without curcumin. The hydrogen peroxide/serum deprivation (H2O2/SD) medium was used to mimic the ischemic condition in vitro. Cytoprotective effects of curcumin on ADSCs were evaluated. Curcumin pretreatment significantly increased cell viability and VEGF secretion, and decreased cell injury and apoptosis via regulation of PTEN/Akt/p53 and HO-1 signal proteins expression. The therapeutic potential of ADSCs implantation was investigated in myocardial ischemia-reperfusion injury (IRI) model. Transplantation of curcumin pretreated ADSCs not only resulted in better heart function, higher cells retention, and smaller infarct size, but also decreased myocardial apoptosis, promoted neovascularization, and increased VEGF level in ischemic myocardium. Together, priming of ADSCs with curcumin improved tolerance to oxidative stress injury and resulted in enhancement of their therapeutic potential of ADSCs for myocardial repair. Curcumin pretreatment is a promising adjuvant strategy for stem cells transplantation in myocardial restoration. PMID:26074974

Bilaminar Device of Poly(Lactic-co-Glycolic Acid)/Collagen Cultured With Adipose-Derived Stem Cells for Dermal Regeneration.

PubMed

Domingues, Juliana A; Cherutti, Giselle; Motta, Adriana C; Hausen, Moema A; Oliveira, Rômulo T D; Silva-Zacarin, Elaine C M; Barbo, Maria Lourdes P; Duek, Eliana A R

2016-10-01

Several materials are commercially available as substitutes for skin. However, new strategies are needed to improve the treatment of skin wounds. In this study, we developed and characterized a new device consisting of poly(lactic-co-glycolic acid) (PLGA) and collagen associated with mesenchymal stem cells derived from human adipose tissue. To develop the bilaminar device, we initially obtained a membrane of PLGA by dissolving the copolymer in chloroform and then produced a collagen type I scaffold by freeze-drying. The materials were characterized physically by gel permeation chromatography, scanning electron microscopy, and mass loss. Biological activity was assessed by cell proliferation assay. A preliminary study in vivo was performed with a pig model in which tissue regeneration was assessed macroscopically and histologically, the commercial device Integra being used as a control. The PLGA/collagen bilaminar material was porous, hydrolytically degradable, and compatible with skin growth. The polymer complex allowed cell adhesion and proliferation, making it a potentially useful cell carrier. In addition, the transparency of the material allowed monitoring of the lesion when the dressings were changed. Xenogeneic mesenchymal cells cultured on the device (PLGA/collagen/ASC) showed a reduced granulomatous reaction to bovine collagen, down-regulation of α-SMA, enhancement in the number of neoformed blood vessels, and collagen organization as compared with normal skin; the device was superior to other materials tested (PLGA/collagen and Integra) in its ability to stimulate the formation of new cutaneous tissue. Copyright © 2016 International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.

Suppression of in vitro murine T cell proliferation by human adipose tissue-derived mesenchymal stem cells is dependent mainly on cyclooxygenase-2 expression

PubMed Central

Kim, Jin-Hee; Lee, Yong-Taek; Hong, Jun Man

2013-01-01

Mesenchymal stem cells (MSCs) of human origin have been frequently applied to experimental animal models to evaluate their immunomodulatory functions. MSCs are known to be activated by cytokines from T cells, predominantly by interferon-γ (IFN-γ), in conjunction with other cytokines such as tumor necrosis factor-α (TNF-α) and interlukin-1β. Because IFN-γ is not cross-reactive between human and mouse species, the manner in which human MSCs administered in experimental animals are activated and stimulated to function has been questioned. In the present study, we established MSCs from human adipose tissue. They successfully suppressed the proliferation of not only human peripheral blood mononuclear cells but also mouse splenic T cells. When these human MSCs were stimulated with a culture supernatant of mouse T cells or recombinant murine TNF-α, they expressed cyclooxygenase-2 (COX-2), but not indoleamine 2,3-dioxygenase. The dominant role of COX-2 in suppressing mouse T cell proliferation was validated by the addition of COX-2 inhibitor in the co-culture, wherein the suppressed proliferation was almost completely recovered. In conclusion, human MSCs in a murine environment were activated, at least in part, by TNF-α and mainly used COX-2 as a tool for the suppression of in vitro T cell proliferation. These results should be considered when interpreting results for human MSCs in experimental animals. PMID:24386599

Combination therapy of exendin-4 and allogenic adipose-derived mesenchymal stem cell preserved renal function in a chronic kidney disease and sepsis syndrome setting in rats

PubMed Central

Chen, Chih-Hung; Cheng, Ben-Chung; Chen, Kuan-Hung; Shao, Pei-Lin; Sung, Pei-Hsun; Chiang, Hsin-Ju; Yang, Chih-Chao; Lin, Kun-Chen; Sun, Cheuk-Kwan; Sheu, Jiunn-Jye; Chang, Hsueh-Wen; Lee, Mel S.; Yip, Hon-Kan

2017-01-01

Combined therapy with exendin-4 (Ex4) and allogenic adipose-derived mesenchymal stem cells (ADMSC) was tested against either therapy alone for protecting kidney function against chronic kidney disease (CKD) complicated by sepsis syndrome (SS) [i.e., by intraperitoneal injection of cecal-derived bacteria (1.0 × 104) cells/milliliter/total 5.0 cc].Adult-male-Sprague Dawley rats (n=36) were equally divided into group 1 (sham-control), group 2 (CKD), group 3 (CKD-SS), group 4 (CKD-SS-Ex4), group 5 (CKD-SS-ADMSC) and group 6 (CKD-SS-Ex4-ADMSC). At day 42 after CKD induction SS was induced. Thirty-minutes after SS induction, ADMSCs (2.0 ×106 cells) were intravenously administered to groups 5 and 6. Ex4 (10 μg/kg) was intraperitoneally administered groups 4 and 6 at 30 min and days 1 to 5 after SS induction. Animals were euthanized at day 47 after CKD induction. Kidney-injury score, collagen-deposition area, and creatinine/BUN levels were lowest in group 1, highest in group 3 and significantly higher in group 2 than in groups 4 to 6 in a progressively increasing manner (all P<0.0001). Protein expressions of inflammatory (MMP-9/TNF-α/NF-κB/IL-1ß/ICAM-1), oxidative-stress (NOX-1/NOX-2/oxidized protein), apoptotic (mitochondrial-Bax/cleaved-caspase-3/cleaved-PARP) and fibrotic/DNA-damaged (Smad3/TGF-ß/γ-H2AX) biomarkers showed an identical pattern, whereas anti-fibrotic (BMP-2/Smad1/5), anti-apoptotic/endothelial-integrity (Bcl-2/eNOS) and podocyte-integrity (ZO-1/p-cadherin) biomarkers exhibited an opposite pattern of kidney-injury score among the six groups (all P>0.0001). Cellular expressions of inflammatory (CD14/CD68) and glomerulus/tubular-injury (WT-1/KIM-1) biomarkers displayed an identical pattern, whereas glomerulus/podocyte-component (dystroglycan/nephrin/ZO-1/fibronectin/p-cadherin) biomarkers showed an opposite kidney-injury score among the six groups (all P<0.0001). In conclusion, Ex4-ADMSC therapy effectively preserved renal function in the CKD

The synergistic effects of shear stress and cyclic hydrostatic pressure modulate chondrogenic induction of human mesenchymal stem cells.

PubMed

Hosseini, Motahare-Sadat; Tafazzoli-Shadpour, Mohammad; Haghighipour, Nooshin; Aghdami, Naser; Goodarzi, Alireza

2015-10-01

In this study, we examined chondrogenic regulation of 2 types of mesenchymal stem cells seeded on the bioengineered substrate in monolayer cultures under mechanically defined conditions to mimic the in vivo microenvironment of chondrocytes within articular cartilage tissues. Human adipose-derived mesenchymal stem cells (ASCs) and bone marrow mesenchymal stem cells (BSCs) were exposed to 0.2 Pa shear stress, 3 MPa cyclic hydrostatic pressure, and combined loading with different sequences on chemically designed medical-grade silicone rubber, while no soluble growth factors were added to the culture medium. The expression levels of chondrogenic-specific genes of SOX9, aggrecan, and type II collagen (Col II) were measured. Results were compared to those of cells treated by biological growth factor. Gene expression patterns were dependent on the loading regime. Moreover, the source of mesenchymal stem cells (adipose or bone marrow) was influential in gene expression. Overall, enhanced expression of chondrogenic markers was found through application of mechanical stimuli. The response was generally found to be significantly promoted when the 2 loading regimes were superimposed. Differentiation of ASCs was shown by a modest increase in gene expression profiles. In general, BSCs expressed higher levels of chondrogenic gene expression than ASCs after 3 weeks. A greater effect on Col II and SOX9 mRNA expression was observed when combined loadings were applied. Results may be applied in determining the proper loading sequence for obtaining functional target cells in cartilage engineering applications.

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.

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

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

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.

The Role of Paracrine and Autocrine Signaling in the Early Phase of Adipogenic Differentiation of Adipose-derived Stem Cells

PubMed Central

Hemmingsen, Mette; Vedel, Søren; Skafte-Pedersen, Peder; Sabourin, David; Collas, Philippe; Bruus, Henrik; Dufva, Martin

2013-01-01

Introduction High cell density is known to enhance adipogenic differentiation of mesenchymal stem cells, suggesting secretion of signaling factors or cell-contact-mediated signaling. By employing microfluidic biochip technology, we have been able to separate these two processes and study the secretion pathways. Methods and results Adipogenic differentiation of human adipose-derived stem cells (ASCs) cultured in a microfluidic system was investigated under perfusion conditions with an adipogenic medium or an adipogenic medium supplemented with supernatant from differentiating ASCs (conditioned medium). Conditioned medium increased adipogenic differentiation compared to adipogenic medium with respect to accumulation of lipid-filled vacuoles and gene expression of key adipogenic markers (C/EBPα, C/EBPβ, C/EBPδ, PPARγ, LPL and adiponectin). The positive effects of conditioned medium were observed early in the differentiation process. Conclusions Using different cell densities and microfluidic perfusion cell cultures to suppress the effects of cell-released factors, we have demonstrated the significant role played by auto- or paracrine signaling in adipocyte differentiation. The cell-released factor(s) were shown to act in the recruitment phase of the differentiation process. PMID:23723991

Curcumin-induced heme oxygenase-1 expression prevents H2O2-induced cell death in wild type and heme oxygenase-2 knockout adipose-derived mesenchymal stem cells.

PubMed

Cremers, Niels A J; Lundvig, Ditte M S; van Dalen, Stephanie C M; Schelbergen, Rik F; van Lent, Peter L E M; Szarek, Walter A; Regan, Raymond F; Carels, Carine E; Wagener, Frank A D T G

2014-10-08

Mesenchymal stem cell (MSC) administration is a promising adjuvant therapy to treat tissue injury. However, MSC survival after administration is often hampered by oxidative stress at the site of injury. Heme oxygenase (HO) generates the cytoprotective effector molecules biliverdin/bilirubin, carbon monoxide (CO) and iron/ferritin by breaking down heme. Since HO-activity mediates anti-apoptotic, anti-inflammatory, and anti-oxidative effects, we hypothesized that modulation of the HO-system affects MSC survival. Adipose-derived MSCs (ASCs) from wild type (WT) and HO-2 knockout (KO) mice were isolated and characterized with respect to ASC marker expression. In order to analyze potential modulatory effects of the HO-system on ASC survival, WT and HO-2 KO ASCs were pre-treated with HO-activity modulators, or downstream effector molecules biliverdin, bilirubin, and CO before co-exposure of ASCs to a toxic dose of H2O2. Surprisingly, sensitivity to H2O2-mediated cell death was similar in WT and HO-2 KO ASCs. However, pre-induction of HO-1 expression using curcumin increased ASC survival after H2O2 exposure in both WT and HO-2 KO ASCs. Simultaneous inhibition of HO-activity resulted in loss of curcumin-mediated protection. Co-treatment with glutathione precursor N-Acetylcysteine promoted ASC survival. However, co-incubation with HO-effector molecules bilirubin and biliverdin did not rescue from H2O2-mediated cell death, whereas co-exposure to CO-releasing molecules-2 (CORM-2) significantly increased cell survival, independently from HO-2 expression. Summarizing, our results show that curcumin protects via an HO-1 dependent mechanism against H2O2-mediated apoptosis, and likely through the generation of CO. HO-1 pre-induction or administration of CORMs may thus form an attractive strategy to improve MSC therapy.

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.

Enhanced adipogenic differentiation of bovine bone marrow-derived mesenchymal stem cells

USDA-ARS?s Scientific Manuscript database

Until now, the isolation and characterization of bovine bone marrow-derived mesenchymal stem cells (bBM-MSCs) have not been established, which prompted us to optimize the differentiation protocol for bBM-MSCs. In this study, bBM-MSCs were freshly isolated from three 6-month-old cattle and used for p...

Adipose tissue-derived mesenchymal stem cells in long-term dialysis patients display downregulation of PCAF expression and poor angiogenesis activation.

PubMed

Yamanaka, Shuichiro; Yokote, Shinya; Yamada, Akifumi; Katsuoka, Yuichi; Izuhara, Luna; Shimada, Yohta; Omura, Nobuo; Okano, Hirotaka James; Ohki, Takao; Yokoo, Takashi

2014-01-01

We previously demonstrated that mesenchymal stem cells (MSCs) differentiate into functional kidney cells capable of urine and erythropoietin production, indicating that they may be used for kidney regeneration. However, the viability of MSCs from dialysis patients may be affected under uremic conditions. In this study, we isolated MSCs from the adipose tissues of end-stage kidney disease (ESKD) patients undergoing long-term dialysis (KD-MSCs; mean: 72.3 months) and from healthy controls (HC-MSCs) to compare their viability. KD-MSCs and HC-MSCs were assessed for their proliferation potential, senescence, and differentiation capacities into adipocytes, osteoblasts, and chondrocytes. Gene expression of stem cell-specific transcription factors was analyzed by PCR array and confirmed by western blot analysis at the protein level. No significant differences of proliferation potential, senescence, or differentiation capacity were observed between KD-MSCs and HC-MSCs. However, gene and protein expression of p300/CBP-associated factor (PCAF) was significantly suppressed in KD-MSCs. Because PCAF is a histone acetyltransferase that mediates regulation of hypoxia-inducible factor-1α (HIF-1α), we examined the hypoxic response in MSCs. HC-MSCs but not KD-MSCs showed upregulation of PCAF protein expression under hypoxia. Similarly, HIF-1α and vascular endothelial growth factor (VEGF) expression did not increase under hypoxia in KD-MSCs but did so in HC-MSCs. Additionally, a directed in vivo angiogenesis assay revealed a decrease in angiogenesis activation of KD-MSCs. In conclusion, long-term uremia leads to persistent and systematic downregulation of PCAF gene and protein expression and poor angiogenesis activation of MSCs from patients with ESKD. Furthermore, PCAF, HIF-1α, and VEGF expression were not upregulated by hypoxic stimulation of KD-MSCs. These results suggest that the hypoxic response may be blunted in MSCs from ESKD patients.

Adipose Tissue-Derived Mesenchymal Stem Cells in Long-Term Dialysis Patients Display Downregulation of PCAF Expression and Poor Angiogenesis Activation

PubMed Central

Yamanaka, Shuichiro; Yokote, Shinya; Yamada, Akifumi; Katsuoka, Yuichi; Izuhara, Luna; Shimada, Yohta; Omura, Nobuo; Okano, Hirotaka James; Ohki, Takao; Yokoo, Takashi

2014-01-01

We previously demonstrated that mesenchymal stem cells (MSCs) differentiate into functional kidney cells capable of urine and erythropoietin production, indicating that they may be used for kidney regeneration. However, the viability of MSCs from dialysis patients may be affected under uremic conditions. In this study, we isolated MSCs from the adipose tissues of end-stage kidney disease (ESKD) patients undergoing long-term dialysis (KD-MSCs; mean: 72.3 months) and from healthy controls (HC-MSCs) to compare their viability. KD-MSCs and HC-MSCs were assessed for their proliferation potential, senescence, and differentiation capacities into adipocytes, osteoblasts, and chondrocytes. Gene expression of stem cell-specific transcription factors was analyzed by PCR array and confirmed by western blot analysis at the protein level. No significant differences of proliferation potential, senescence, or differentiation capacity were observed between KD-MSCs and HC-MSCs. However, gene and protein expression of p300/CBP-associated factor (PCAF) was significantly suppressed in KD-MSCs. Because PCAF is a histone acetyltransferase that mediates regulation of hypoxia-inducible factor-1α (HIF-1α), we examined the hypoxic response in MSCs. HC-MSCs but not KD-MSCs showed upregulation of PCAF protein expression under hypoxia. Similarly, HIF-1α and vascular endothelial growth factor (VEGF) expression did not increase under hypoxia in KD-MSCs but did so in HC-MSCs. Additionally, a directed in vivo angiogenesis assay revealed a decrease in angiogenesis activation of KD-MSCs. In conclusion, long-term uremia leads to persistent and systematic downregulation of PCAF gene and protein expression and poor angiogenesis activation of MSCs from patients with ESKD. Furthermore, PCAF, HIF-1α, and VEGF expression were not upregulated by hypoxic stimulation of KD-MSCs. These results suggest that the hypoxic response may be blunted in MSCs from ESKD patients. PMID:25025381

Adipose tissue in myocardial infarction.

PubMed

Su, Leon; Siegel, John E; Fishbein, Michael C

2004-01-01

The histologic evolution of myocardial infarction (MI) has been studied in some detail. However, there is little mention of the presence of adipose tissue in healed MI(HMI). Ninety-one hearts explanted during 1997-2001 were examined to determine the extent of adipose tissue within HMI. The medical records, surgical pathology reports, and all histologic sections of the explanted heart, from patients undergoing heart transplantation for ischemic heart disease, were reviewed. Adipose tissue within the areas of HMI was quantified. The location of the HMI, the age and gender of the patient, age of HMI, and whether the patient was treated with coronary artery bypass surgery (CABG) were noted. Of the 91 hearts examined, 168 HMIs were identified; 141 (84%) contained some mature fat within the HMI. Adipose tissue increased with increasing age, in males, and in those patients who had CABG surgery. The amount of adipose tissue was not related to the location or age of the HMI. Adipose tissue is a prevalent histological finding in HMIs. The pathogenesis of adipose tissue is unknown, but may be influenced by current medical therapy for ischemic heart disease, thus explaining why adipose tissue in HMIs was not reported until 1997. The presence of fat supports the speculation that a regenerative cell, or multipotent stem cell, exists within the heart, and under the influence of microenvironmental or therapeutic factors can differentiate into fat, other mesenchymal tissues, and potentially even myocardium.

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

Preconditioning of adipose tissue-derived mesenchymal stem cells with deferoxamine increases the production of pro-angiogenic, neuroprotective and anti-inflammatory factors: Potential application in the treatment of diabetic neuropathy.

PubMed

Oses, Carolina; Olivares, Belén; Ezquer, Marcelo; Acosta, Cristian; Bosch, Paul; Donoso, Macarena; Léniz, Patricio; Ezquer, Fernando

2017-01-01

Diabetic neuropathy (DN) is one of the most frequent and troublesome complications of diabetes mellitus. Evidence from diabetic animal models and diabetic patients suggests that reduced availability of neuroprotective and pro-angiogenic factors in the nerves in combination with a chronic pro-inflammatory microenvironment and high level of oxidative stress, contribute to the pathogenesis of DN. Mesenchymal stem cells (MSCs) are of great interest as therapeutic agents for regenerative purposes, since they can secrete a broad range of cytoprotective and anti-inflammatory factors. Therefore, the use of the MSC secretome may represent a promising approach for DN treatment. Recent data indicate that the paracrine potential of MSCs could be boosted by preconditioning these cells with an environmental or pharmacological stimulus, enhancing their therapeutic efficacy. In the present study, we observed that the preconditioning of human adipose tissue-derived MSCs (AD-MSCs) with 150μM or 400μM of the iron chelator deferoxamine (DFX) for 48 hours, increased the abundance of the hypoxia inducible factor 1 alpha (HIF-1α) in a concentration dependent manner, without affecting MSC morphology and survival. Activation of HIF-1α led to the up-regulation of the mRNA levels of pro-angiogenic factors like vascular endothelial growth factor alpha and angiopoietin 1. Furthermore this preconditioning increased the expression of potent neuroprotective factors, including nerve growth factor, glial cell-derived neurotrophic factor and neurotrophin-3, and cytokines with anti-inflammatory activity like IL4 and IL5. Additionally, we observed that these molecules, which could also be used as therapeutics, were also increased in the secretome of MSCs preconditioned with DFX compared to the secretome obtained from non-preconditioned cells. Moreover, DFX preconditioning significantly increased the total antioxidant capacity of the MSC secretome and they showed neuroprotective effects when

Preconditioning of adipose tissue-derived mesenchymal stem cells with deferoxamine increases the production of pro-angiogenic, neuroprotective and anti-inflammatory factors: Potential application in the treatment of diabetic neuropathy

PubMed Central

Oses, Carolina; Olivares, Belén; Ezquer, Marcelo; Acosta, Cristian; Bosch, Paul; Donoso, Macarena; Léniz, Patricio

2017-01-01

Diabetic neuropathy (DN) is one of the most frequent and troublesome complications of diabetes mellitus. Evidence from diabetic animal models and diabetic patients suggests that reduced availability of neuroprotective and pro-angiogenic factors in the nerves in combination with a chronic pro-inflammatory microenvironment and high level of oxidative stress, contribute to the pathogenesis of DN. Mesenchymal stem cells (MSCs) are of great interest as therapeutic agents for regenerative purposes, since they can secrete a broad range of cytoprotective and anti-inflammatory factors. Therefore, the use of the MSC secretome may represent a promising approach for DN treatment. Recent data indicate that the paracrine potential of MSCs could be boosted by preconditioning these cells with an environmental or pharmacological stimulus, enhancing their therapeutic efficacy. In the present study, we observed that the preconditioning of human adipose tissue-derived MSCs (AD-MSCs) with 150μM or 400μM of the iron chelator deferoxamine (DFX) for 48 hours, increased the abundance of the hypoxia inducible factor 1 alpha (HIF-1α) in a concentration dependent manner, without affecting MSC morphology and survival. Activation of HIF-1α led to the up-regulation of the mRNA levels of pro-angiogenic factors like vascular endothelial growth factor alpha and angiopoietin 1. Furthermore this preconditioning increased the expression of potent neuroprotective factors, including nerve growth factor, glial cell-derived neurotrophic factor and neurotrophin-3, and cytokines with anti-inflammatory activity like IL4 and IL5. Additionally, we observed that these molecules, which could also be used as therapeutics, were also increased in the secretome of MSCs preconditioned with DFX compared to the secretome obtained from non-preconditioned cells. Moreover, DFX preconditioning significantly increased the total antioxidant capacity of the MSC secretome and they showed neuroprotective effects when

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.

A simple modification of the separation method reduces heterogeneity of adipose-derived stem cells.

PubMed

Griesche, Nadine; Luttmann, Werner; Luttmann, Arlette; Stammermann, Thekla; Geiger, Helmut; Baer, Patrick C

2010-01-01

High hopes are put into the use of mesenchymal stem cells (MSCs) in various approaches for tissue engineering and regenerative medicine. MSCs are derived from different tissues with only small differences in their phenotype or their differentiation potential, but higher differences in the cell yield. Since fat is easily accessible and contains a high amount of MSCs to be isolated, adipose-derived stem cells (ASCs) are very promising for clinical approaches. ASCs are not a completely homogeneous cell population. Our study was initiated to explore an easy and convenient method to reduce heterogeneity. We tested different isolation methods: (1) the standard isolation method for ASCs based on plastic attachment, (2) the standard method with an initial washing step after 60 min of adherence and (3) immunomagnetic isolation by 4 typical markers (CD49a, CD90, CD105 and CD271). Cells isolated by these methods were evaluated using quantitative PCR and flow cytometry as well as by their differentiation potential. Washing led to a significantly lower expression of desmin, smA and six2, and a higher expression of the stem cell markers nestin, oct-4 and sall-1, compared to standard isolated cells, while the immunomagnetically isolated cells showed no significant changes. All cells independent of the isolation method could be induced to differentiate into adipocytes and osteoblasts. Our study demonstrates that a simple washing step reduces heterogeneity of cultured ASCs according to PCR analysis, whereas the immunomagnetic isolation only showed minor advantages compared to the standard method, but the disadvantage of significantly lower cell yields in the primary isolates. Copyright 2010 S. Karger AG, Basel.

The suitability of human adipose-derived stem cells for the engineering of ligament tissue.

PubMed

Eagan, Michael J; Zuk, Patricia A; Zhao, Ke-Wei; Bluth, Benjamin E; Brinkmann, Elyse J; Wu, Benjamin M; McAllister, David R

2012-10-01

Rupture of the anterior cruciate ligament (ACL) is the one of the most common sports-related injuries. With its poor healing capacity, surgical reconstruction using either autografts or allografts is currently required to restore function. However, serious complications are associated with graft reconstructions and the number of such reconstructions has steadily risen over the years, necessitating the search for an alternative approach to ACL repair. Such an approach may likely be tissue engineering. Recent engineering approaches using ligament-derived fibroblasts have been promising, but the slow growth rate of such fibroblasts in vitro may limit their practical application. More promising results are being achieved using bone marrow mesenchymal stem cells (MSCs). The adipose-derived stem cell (ASC) is often proposed as an alternative choice to the MSC and, as such, may be a suitable stem cell for ligament engineering. However, the use of ASCs in ligament engineering still remains relatively unexplored. Therefore, in this study, the potential use of human ASCs in ligament tissue engineering was initially explored by examining their ability to express several ligament markers under growth factor treatment. ASC populations treated for up to 4 weeks with TGFβ1 or IGF1 did not show any significant and consistent upregulation in the expression of collagen types 1 and 3, tenascin C and scleraxis. While treatment with EGF or bFGF resulted in increased tenascin C expression, increased expression of collagens 1 and 3 were never observed. Therefore, simple in vitro treatment of human ASC populations with growth factors may not stimulate their ligament differentiative potential. Copyright © 2011 John Wiley & Sons, Ltd.

High density lipoprotein promotes proliferation of adipose-derived stem cells via S1P1 receptor and Akt, ERK1/2 signal pathways.

PubMed

Shen, Haitao; Zhou, Enchen; Wei, Xiujing; Fu, Zhiwei; Niu, Chenguang; Li, Yang; Pan, Bing; Mathew, Anna V; Wang, Xu; Pennathur, Subramaniam; Zheng, Lemin; Wang, Yongyu

2015-05-15

Adipose-derived stem cells (ADSC) are non-hematopoietic mesenchymal stem cells that have shown great promise in their ability to differentiate into multiple cell lineages. Their ubiquitous nature and the ease of harvesting have attracted the attention of many researchers, and they pose as an ideal candidate for applications in regenerative medicine. Several reports have demonstrated that transplanting ADSC can promote repair of injured tissue and angiogenesis in animal models. Survival of these cells after transplant remains a key limiting factor for the success of ADSC transplantation. Circulating factors like High Density Lipoprotein (HDL) has been known to promote survival of other stems cells like bone marrow derived stem cells and endothelial progenitor cells, both by proliferation and by inhibiting cell apoptosis. The effect of HDL on transplanted adipose-derived stem cells in vivo is largely unknown. This study focused on exploring the effects of plasma HDL on ADSC and delineating the mechanisms involved in their proliferation after entering the bloodstream. Using the MTT and BrdU assays, we tested the effects of HDL on ADSC proliferation. We probed the downstream intracellular Akt and ERK1/2 signaling pathways and expression of cyclin proteins in ADSC using western blot. Our study found that HDL promotes proliferation of ADSC, by binding to sphingosine-1- phosphate receptor-1(S1P1) on the cell membrane. This interaction led to activation of intracellular Akt and ERK1/2 signaling pathways, resulting in increased expression of cyclin D1 and cyclin E, and simultaneous reduction in expression of cyclin-dependent kinase inhibitors p21 and p27, therefore promoting cell cycle progression and cell proliferation. These studies raise the possibility that HDL may be a physiologic regulator of stem cells and increasing HDL concentrations may be valuable strategy to promote ADSC transplantation.

Meeting report of the 2016 bone marrow adiposity meeting.

PubMed

van der Eerden, Bram; van Wijnen, André

2017-10-02

There is considerable interest in the physiology and pathology, as well as the cellular and molecular biology, of bone marrow adipose tissue (BMAT). Because bone marrow adiposity is linked not only to systemic energy metabolism, but also to both bone marrow and musculoskeletal disorders, this biologic compartment has become of major interest to investigators from diverse disciplines. Bone marrow adiposity represents a virtual multi-tissue endocrine organ, which encompasses cells from multiple developmental lineages (e.g., mesenchymal, myeloid, lymphoid) and occupies all the non-osseous and non-cartilaginous space within long bones. A number of research groups are now focusing on bone marrow adiposity to understand a range of clinical afflictions associated with bone marrow disorders and to consider mechanisms-based strategies for future therapies.

Can photobiomodulation associated with implantation of mesenchymal adipose-derived stem cells attenuate the expression of MMPs and decrease degradation of type II collagen in an experimental model of osteoarthritis?

PubMed

Stancker, Tatiane Garcia; Vieira, Stella Souza; Serra, Andrey Jorge; do Nascimento Lima, Rafael; Dos Santos Feliciano, Regiane; Silva, José Antônio; Dos Santos, Solange Almeida; Dos Santos Vieira, Marcia Ataize; Simões, Maíra Cecília Brandão; Leal-Junior, Ernesto Cesar; de Tarso Camillo de Carvalho, Paulo

2018-03-08

This study aimed to determine whether photobiomodulation therapy (PBMT) could improve the bioavailability and chondroprotective benefits of mesenchymal stem cells injected into the knees of rats used as an experimental model of osteoarthritis (OA) as well as reduce the expression of matrix metalloproteinases (MMPs) and degradation of type II collagen (COL2-1) in the cartilage. Adipose-derived stem/stromal cells (ADSCs) were collected from three male Fischer 344 rats and characterized by flow cytometry. Fifty female Fischer 344 rats were distributed into five groups of 10 animals each. These groups were as follows: control, OA, OA PBMT, OA ADSC, and OA ADSC PBMT. OA was induced in the animals using a 4% papain solution. Animals from the OA ADSC and OA ADSC PBMT groups received an intra-articular injection of 10 × 10 6 ADSCs and were treated with PBMT by irradiation (wavelength: 808 nm, power: 50 mW, energy: 42 J, energy density: 71.2 J/cm 2 , spot size: 0.028). Euthanasia was performed 7 days after the first treatment. The use of PBMT alone and the injection of ADSCs resulted in downregulation of pro-inflammatory cytokines and MPs in cartilage compared to the OA group. PBMT and ADSCs caused upregulation of tissue inhibitors of MPs 1 and 2 and mRNA and protein expression of COL2-1 in cartilage compared to the OA group. The intra-articular injection of ADSCs and PBMT prevented joint degeneration resulting from COL2-1 degradation and modulated inflammation by downregulating cytokines and MMPs in the OA group.

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

Adipose tissue-derived stem cell secreted IGF-1 protects myoblasts from the negative effect of myostatin.

PubMed

Gehmert, Sebastian; Wenzel, Carina; Loibl, Markus; Brockhoff, Gero; Huber, Michaela; Krutsch, Werner; Nerlich, Michael; Gosau, Martin; Klein, Silvan; Schreml, Stephan; Prantl, Lukas; Gehmert, Sanga

2014-01-01

Myostatin, a TGF-β family member, is associated with inhibition of muscle growth and differentiation and might interact with the IGF-1 signaling pathway. Since IGF-1 is secreted at a bioactive level by adipose tissue-derived mesenchymal stem cells (ASCs), these cells (ASCs) provide a therapeutic option for Duchenne Muscular Dystrophy (DMD). But the protective effect of stem cell secreted IGF-1 on myoblast under high level of myostatin remains unclear. In the present study murine myoblasts were exposed to myostatin under presence of ASCs conditioned medium and investigated for proliferation and apoptosis. The protective effect of IGF-1 was further examined by using IGF-1 neutralizing and receptor antibodies as well as gene silencing RNAi technology. MyoD expression was detected to identify impact of IGF-1 on myoblasts differentiation when exposed to myostatin. IGF-1 was accountable for 43.6% of the antiapoptotic impact and 48.8% for the proliferative effect of ASCs conditioned medium. Furthermore, IGF-1 restored mRNA and protein MyoD expression of myoblasts under risk. Beside fusion and transdifferentiation the beneficial effect of ASCs is mediated by paracrine secreted cytokines, particularly IGF-1. The present study underlines the potential of ASCs as a therapeutic option for Duchenne muscular dystrophy and other dystrophic muscle diseases.

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.

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.

Human adipose mesenchymal stem cells as potent anti-fibrosis therapy for systemic sclerosis.

PubMed

Maria, Alexandre T J; Toupet, Karine; Maumus, Marie; Fonteneau, Guillaume; Le Quellec, Alain; Jorgensen, Christian; Guilpain, Philippe; Noël, Danièle

2016-06-01

Displaying immunosuppressive and trophic properties, mesenchymal stem/stromal cells (MSC) are being evaluated as promising therapeutic options in a variety of autoimmune and degenerative diseases. Although benefits may be expected in systemic sclerosis (SSc), a rare autoimmune disease with fibrosis-related mortality, MSC have yet to be evaluated in this specific condition. While autologous approaches could be inappropriate because of functional alterations in MSC from patients, the objective of the present study was to evaluate allogeneic and xenogeneic MSC in the HOCl-induced model of diffuse SSc. We also questioned the source of human MSC and compared bone marrow- (hBM-MSC) and adipose-derived MSC (hASC). HOCl-challenged BALB/c mice received intravenous injection of BM-MSC from syngeneic BALB/c or allogeneic C57BL/6 mice, and xenogeneic hBM-MSC or hASC (3 donors each). Skin thickness was measured during the experiment. At euthanasia, histology, immunostaining, collagen determination and RT-qPCR were performed in skin and lungs. Xenogeneic hBM-MSC were as effective as allogeneic or syngeneic BM-MSC in decreasing skin thickness, expression of Col1, Col3, α-Sma transcripts, and collagen content in skin and lungs. This anti-fibrotic effect was not associated with MSC migration to injured skin or with long-term MSC survival. Interestingly, compared with hBM-MSC, hASC were significantly more efficient in reducing skin fibrosis, which was related to a stronger reduction of TNFα, IL1β, and enhanced ratio of Mmp1/Timp1 in skin and lung tissues. Using primary cells isolated from 3 murine and 6 human individuals, this preclinical study demonstrated similar therapeutic effects using allogeneic or xenogeneic BM-MSC while ASC exerted potent anti-inflammatory and remodeling properties. This sets the proof-of-concept prompting to evaluate the therapeutic efficacy of allogeneic ASC in SSc patients. Copyright © 2016 Elsevier Ltd. All rights reserved.

Long Term Study of Protective Mechanisms of Human Adipose Derived Mesenchymal Stem Cells on Cisplatin Induced Kidney injury in Sprague-Dawley Rats.

PubMed

Elhusseini, Fatma M; Saad, Mohamed-Ahdy A A; Anber, Nahla; Elghannam, Doaa; Sobh, Mohamed-Ahmed; Alsayed, Aziza; El-Dusoky, Sara; Sheashaa, Hussein; Abdel-Ghaffar, Hassan; Sobh, Mohamed

2016-01-01

Long-term evaluation of cisplatin induced nephrotoxicity and the probable renal protective activities of stem cells are lacking up until now. We evaluated the early and long-term role of human adipose derived mesenchymal stem cells (ADMSCs) in prevention or amelioration of cisplatin induced acute kidney injury (AKI) in Sprague-Dawley rats. For this, we determined the kidney tissue level of oxidative stress markers in conjugation with a renal histopathological scoring system of both acute and chronic renal changes. This study used eighty Sprague-Dawley (SD) rats weighing 250-300g. They were assigned into four equal groups (each group n=20): (I) Negative control group, rats injected with single dose of 1 ml normal saline. (II) Positive control cisplatin, rats injected with a single dose of 5 mg/kg I.P in 1 ml saline. (III) Cisplatin and culture media group, rats injected with 0.5 ml of culture media single dose into the tail vein and (IV) Cisplatin and ADMSCs group, rats injected with a single dose of 0.5 ml of culture media containing 5 x10(6)ADMSCs into the tail vein one day after cisplatin administration. Each main group was further divided according to the timing of sacrifice into four subgroups (each subgroup n=5). Rats in the subgroup A were sacrificed after 4 days; subgroup B were sacrificed after 7 days; subgroup C were sacrificed after 11 days; and subgroup D were sacrificed after 30 days. Before sacrifice, 24 hrs.-urine was collected using a metabolic cage. Renal function was evaluated through blood urea nitrogen (BUN), serum creatinine and creatinine clearance. Kidney tissue homogenate oxidative stress parameters, Malondialdehyde (MDA), Superoxide dismutase (SOD) and Glutathione (GSH) were determined. In addition, histopathological analysis for active injury, regenerative and chronic changes was performed. ADMSCs were characterized and their capability of differentiation was proved. Cisplatin induced a significant increase in plasma creatinine and tissue

Regeneration of skin tissue promoted by mesenchymal stem cells seeded in nanostructured membrane.

PubMed

Souza, C M C O; Mesquita, L A F; Souza, D; Irioda, A C; Francisco, J C; Souza, C F; Guarita-Souza, L C; Sierakowski, M-R; Carvalho, K A T

2014-01-01

The mesenchymal stem cell therapy has proven to be an effective option in the treatment of skin injuries. The combination of these cells with nanostructured membranes seems to be the future for tissues recovery. The aim of this project was to use biomolecules of polysaccharides to be incorporated on regenerated cellulose membranes and to prospect the improvement as bioactive wound dressings with mesenchymal stem cells. The biocomposites were obtained after defibrillation with the use of never-dried bacterial cellulose to form a pulp, and, after the films were regenerated, in the presence of gellan gum with or without fluconazole. Membrane atomic force microscopy was performed for comparison of their structures. Adipose-derived mesenchymal stem cells were obtained from human adipose tissue liposuction in accordance with Zuk et al. The flow cytometric analysis and induction tests for adipocytes and osteocytes were performed. In vitro assays were performed on different membranes to evaluate the ability of these cells to adhere at 2 hours and proliferate at 7 days; the results were obtained by use of the MTT cell counting technique. In vivo testing allowed us to observe cell migration and participation in wound-healing by fluorescence labeling of the cells with BrdU. The bioactive curative, seeded with cells, was tested in skin burned in a murine model. The bacterial cellulose with gelan gum membrane incorporated with fluconazole presented the best performance in adhesion and proliferation tests. The cells can be identified in burned host tissue after occurrence of the wound. Copyright © 2014 Elsevier Inc. All rights reserved.

Identification of Regulatory Factors for Mesenchymal Stem Cell-Derived Salivary Epithelial Cells in a Co-Culture System

PubMed Central

Park, Yun-Jong; Koh, Jin; Gauna, Adrienne E.; Chen, Sixue; Cha, Seunghee

2014-01-01

Patients with Sjögren’s syndrome or head and neck cancer patients who have undergone radiation therapy suffer from severe dry mouth (xerostomia) due to salivary exocrine cell death. Regeneration of the salivary glands requires a better understanding of regulatory mechanisms by which stem cells differentiate into exocrine cells. In our study, bone marrow-derived mesenchymal stem cells were co-cultured with primary salivary epithelial cells from C57BL/6 mice. Co-cultured bone marrow-derived mesenchymal stem cells clearly resembled salivary epithelial cells, as confirmed by strong expression of salivary gland epithelial cell-specific markers, such as alpha-amylase, muscarinic type 3 receptor, aquaporin-5, and cytokeratin 19. To identify regulatory factors involved in this differentiation, transdifferentiated mesenchymal stem cells were analyzed temporarily by two-dimensional-gel-electrophoresis, which detected 58 protein spots (>1.5 fold change, p<0.05) that were further categorized into 12 temporal expression patterns. Of those proteins only induced in differentiated mesenchymal stem cells, ankryin-repeat-domain-containing-protein 56, high-mobility-group-protein 20B, and transcription factor E2a were selected as putative regulatory factors for mesenchymal stem cell transdifferentiation based on putative roles in salivary gland development. Induction of these molecules was confirmed by RT-PCR and western blotting on separate sets of co-cultured mesenchymal stem cells. In conclusion, our study is the first to identify differentially expressed proteins that are implicated in mesenchymal stem cell differentiation into salivary gland epithelial cells. Further investigation to elucidate regulatory roles of these three transcription factors in mesenchymal stem cell reprogramming will provide a critical foundation for a novel cell-based regenerative therapy for patients with xerostomia. PMID:25402494

Human adipose-derived mesenchymal stem cells in vitro: evaluation of an optimal expansion medium preserving stemness.

PubMed

Baer, Patrick C; Griesche, Nadine; Luttmann, Werner; Schubert, Ralf; Luttmann, Arlette; Geiger, Helmut

2010-01-01

The potential of cultured adipose-derived stem cells (ASC) in regenerative medicine and new cell therapeutic concepts has been shown recently by many investigations. However, while the method of isolation of ASC from liposuction aspirates depending on plastic adhesion is well established, a standard expansion medium optimally maintaining the undifferentiated state has not been described. We cultured ASC in five commonly used culture media (two laboratory-made media and three commercially available media) and compared them with a standard medium. We analyzed the effects on cell morphology, proliferation, hepatocyte growth factor (HGF) expression, stem cell marker profile and differentiation potential. Proliferation was measured with a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and a fluorescent assay. Release of HGF was assessed by an immunoassay. Expression of characteristic stem cell-related transcription factors and markers was evaluated by quantitative polymerase chain reaction (qPCR) (Nanog, Sox-2, Rex-1, nestin and Oct-4) and flow cytometry (CD44, CD73, CD90, CD105 and CD166), and differentiation was shown by adipogenic medium. The morphology and expansion of ASC were significantly affected by the media used, whereas none of the media influenced the ASC potential to differentiate into adipocytes. Furthermore, two of the media induced an increase in expression of transcription factors, an increased secretion of HGF and a decrease in CD105 expression. Culture of ASC in one of these two media before using the cells in cell therapeutic approaches may have a benefit on their regenerative potential.

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.

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.

Gene expression analysis of human adipose tissue-derived stem cells during the initial steps of in vitro osteogenesis.

PubMed

Robert, Anny Waloski; Angulski, Addeli Bez Batti; Spangenberg, Lucia; Shigunov, Patrícia; Pereira, Isabela Tiemy; Bettes, Paulo Sergio Loiacono; Naya, Hugo; Correa, Alejandro; Dallagiovanna, Bruno; Stimamiglio, Marco Augusto

2018-03-16

Mesenchymal stem cells (MSCs) have been widely studied with regard to their potential use in cell therapy protocols and regenerative medicine. However, a better comprehension about the factors and molecular mechanisms driving cell differentiation is now mandatory to improve our chance to manipulate MSC behavior and to benefit future applications. In this work, we aimed to study gene regulatory networks at an early step of osteogenic differentiation. Therefore, we analyzed both the total mRNA and the mRNA fraction associated with polysomes on human adipose tissue-derived stem cells (hASCs) at 24 h of osteogenesis induction. The RNA-seq results evidenced that hASC fate is not compromised with osteogenesis at this time and that 21 days of continuous cell culture stimuli are necessary for full osteogenic differentiation of hASCs. Furthermore, early stages of osteogenesis induction involved gene regulation that was linked to the management of cell behavior in culture, such as the control of cell adhesion and proliferation. In conclusion, although discrete initial gene regulation related to osteogenesis occur, the first 24 h of induction is not sufficient to trigger and drive in vitro osteogenic differentiation of hASCs.

Genetic modification of human mesenchymal stem cells helps to reduce adiposity and improve glucose tolerance in an obese diabetic mouse model.

PubMed

Sen, Sabyasachi; Domingues, Cleyton C; Rouphael, Carol; Chou, Cyril; Kim, Chul; Yadava, Nagendra

2015-12-09

Human mesenchymal stem cells (MSCs) are multipotent cells that can differentiate into fat, muscle, bone and cartilage cells. Exposure of subcutaneous abdominal adipose tissue derived AD-MSCs to high glucose (HG) leads to superoxide accumulation and up-regulation of inflammatory molecules. Our aim was to inquire how HG exposure affects MSCs differentiation and whether the mechanism is reversible. We exposed human adipose tissue derived MSCs to HG (25 mM) and compared it to normal glucose (NG, 5.5 mM) exposed cells at 7, 10 and 14 days. We examined mitochondrial superoxide accumulation (Mitosox-Red), cellular oxygen consumption rate (OCR, Seahorse) and gene expression. HG increased reactive superoxide (ROS) accumulation noted by day 7 both in cytosol and mitochondria. The OCR between the NG and HG exposed groups however did not change until 10 days at which point OCR of HG exposed cells were reduced significantly. We noted that HG exposure upregulated mRNA expression of adipogenic (PPARG, FABP-4, CREBP alpha and beta), inflammatory (IL-6 and TNF alpha) and antioxidant (SOD2 and Catalase) genes. Next, we used AdSOD2 to upregulate SOD2 prior to HG exposure and thereby noted reduction in superoxide generation. SOD2 upregulation helped reduce mRNA over-expression of PPARG, FABP-4, IL-6 and TNFα. In a series of separate experiments, we delivered the eGFP and SOD2 upregulated MSCs (5 days post ex-vivo transduction) and saline intra-peritoneally (IP) to obese diabetic (db/db) mice. We confirmed homing-in of eGFP labeled MSCs, delivered IP, to different inflamed fat pockets, particularly omental fat. Mice receiving SOD2-MSCs showed progressive reduction in body weight and improved glucose tolerance (GTT) at 4 weeks, post MSCs transplantation compared to the GFP-MSC group (control). High glucose evokes superoxide generation, OCR reduction and adipogenic differentiation. Mitochondrial superoxide dismutase upregulation quenches excess superoxide and reduces adipocyte

Isolation, Characterization and Growth Kinetic Comparison of Bone Marrow and Adipose Tissue Mesenchymal Stem Cells of Guinea Pig.

PubMed

Aliborzi, Ghaem; Vahdati, Akbar; Mehrabani, Davood; Hosseini, Seyed Ebrahim; Tamadon, Amin

2016-05-30

Mesenchymal stem cells (MSCs) from different sources have different characteristics. Moreover, MSCs are not isolated and characterized in Guinea pig for animal model of cell therapy. was the isolating of bone marrow MSCs (BM-MSCs) and adipose tissue MSCs (AT-MSCs) from Guinea pig and assessing their characteristics. In this study, bone marrow and adipose tissue were collected from three Guinea pigs and cultured and expanded through eight passages. BM-MSCs and AT-MSCs at passages 2, 5 and 8 were seeded in 24-well plates in triplicate. Cells were counted from each well 1~7 days after seeding to determine population doubling time (PDT) and cell growth curves. Cells of passage 3 were cultured in osteogenic and adipogenic differentiation media. BM-MSCs and AT-MSCs attached to the culture flask and displayed spindle-shaped morphology. Proliferation rate of AT-MSCs in the analyzed passages was more than BM-MSCs. The increase in the PDT of MSCs occurs with the increase in the number of passages. Moreover, after culture of BM-MSCs and AT-MSCs in differentiation media, the cells differentiated toward osteoblasts and adipocytes as verified by Alizarin Red staining and Oil Red O staining, respectively. BM-MSCs and AT-MSCs of Guinea pig could be valuable source of multipotent stem cells for use in experimental and preclinical studies in animal models.

Adipogenic Differentiation of Mesenchymal Stem Cells Alters Their Immunomodulatory Properties in a Tissue-Specific Manner.

PubMed

Munir, Hafsa; Ward, Lewis S C; Sheriff, Lozan; Kemble, Samuel; Nayar, Saba; Barone, Francesca; Nash, Gerard B; McGettrick, Helen M

2017-06-01

Chronic inflammation is associated with formation of ectopic fat deposits that might represent damage-induced aberrant mesenchymal stem cell (MSC) differentiation. Such deposits are associated with increased levels of inflammatory infiltrate and poor prognosis. Here we tested the hypothesis that differentiation from MSC to adipocytes in inflamed tissue might contribute to chronicity through loss of immunomodulatory function. We assessed the effects of adipogenic differentiation of MSC isolated from bone marrow or adipose tissue on their capacity to regulate neutrophil recruitment by endothelial cells and compared the differentiated cells to primary adipocytes from adipose tissue. Bone marrow derived MSC were immunosuppressive, inhibiting neutrophil recruitment to TNFα-treated endothelial cells (EC), but MSC-derived adipocytes were no longer able to suppress neutrophil adhesion. Changes in IL-6 and TGFβ1 signalling appeared critical for the loss of the immunosuppressive phenotype. In contrast, native stromal cells, adipocytes derived from them, and mature adipocytes from adipose tissue were all immunoprotective. Thus disruption of normal tissue stroma homeostasis, as occurs in chronic inflammatory diseases, might drive "abnormal" adipogenesis which adversely influences the behavior of MSC and contributes to pathogenic recruitment of leukocytes. Interestingly, stromal cells programmed in native fat tissue retain an immunoprotective phenotype. Stem Cells 2017;35:1636-1646. © 2017 The Authors STEM CELLS published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

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

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.

Protection against acetaminophen-induced acute liver failure by omentum adipose tissue derived stem cells through the mediation of Nrf2 and cytochrome P450 expression.

PubMed

Huang, Yu-Jen; Chen, Poda; Lee, Chih-Yuan; Yang, Sin-Yu; Lin, Ming-Tsan; Lee, Hsuan-Shu; Wu, Yao-Ming

2016-01-19

Acetaminophen (APAP) overdose causes acute liver failure (ALF) in animals and humans via the rapid depletion of intracellular glutathione (GSH) and the generation of excess reactive oxygen species (ROS) that damage hepatocytes. Stem cell therapy is a potential treatment strategy for ALF. We isolated mesenchymal stem cells (MSCs) from mice omentum adipose tissue-derived stem cells (ASCs) and transplanted them into a mouse model of APAP-induced ALF to explore their therapeutic potential. In addition, we performed in vitro co-culture studies with omentum-derived ASCs and primary isolated hepatocytes to demonstrate the hepatoprotective effect of omentum-derived ASCs on hepatocytes that were subjected to APAP-induced damage. ASC transplantation significantly improved the survival rate of mice with ALF and attenuated the severity of APAP-induced liver damage by suppressing cytochrome P450 activity to reduce the accumulation of toxic nitrotyrosine and the upregulation of NF-E2-related factor 2 (Nrf2) expression, resulting in an increase in the subsequent antioxidant activity. These effects protected the hepatocytes from APAP-induced damage through the suppression of downstream MAPK signal activation and inflammatory cytokine production. our results demonstrate that omentum-derived ASCs are an alternative source of ASCs that regulate the antioxidant response and may represent a beneficial therapeutic strategy for ALF.

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

Genetic expression of adipose derived stem cell and smooth muscle cell markers to monitor differentiation potential following low intensity laser irradiation

NASA Astrophysics Data System (ADS)

Abrahamse, Heidi

2014-02-01

Mesenchymal stem cells (MSCs) have the capacity to differentiate into a variety of cell types that could potentially be used in tissue engineering and regenerative medicine. Low intensity laser irradiation (LILI) has been shown to induce a significant increase in cell viability and proliferation. Growth factors such as retinoic acid (RA) and transforming growth factor β1 (TGF-β1) play important roles in the differentiation of cells. The aim of this study was to investigate whether LILI in combination with growth factors could induce the differentiation of adipose derived stem cells (ADSCs) cocultured with smooth muscle cells (SMCs). The study used primary and continuous ADSC cell lines and a SMC line (SKUT-1) as control. Cells were co-cultured directly at a ratio of 1:1 using established methods, with and without growth factors and then exposed to LILI at 5 J/cm2 using a 636 nm diode laser. The cellular morphology, viability and proliferation of the co-cultures were assessed over a period of one week. The study also monitored the expression of cell specific markers over the same period of time. Genetic expression of the markers for both adipose derived stem cells (β1 Integrin and Thymidine 1) and smooth muscle cells (Heavy Myosin Chain) was monitored using flow cytometry. Cell viability and proliferation increased significantly in the co-cultured groups that were exposed to laser alone, as well as in combination with growth factors. Furthermore, there was a significant decrease in the expression of stem cell markers in the ADSCs over time. The results indicate that LILI in combination with growth factors not only increases the viability and proliferation of co-cultured cells but also decreases the expression of ADSC stem cell markers. This could indicate the possible differentiation of ADSCs into SMCs.

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

Differential Response of Human Adipose Tissue-Derived Mesenchymal Stem Cells, Dermal Fibroblasts, and Keratinocytes to Burn Wound Exudates: Potential Role of Skin-Specific Chemokine CCL27

PubMed Central

van den Broek, Lenie J.; Kroeze, Kim L.; Waaijman, Taco; Breetveld, Melanie; Sampat-Sardjoepersad, Shakun C.; Niessen, Frank B.; Middelkoop, Esther; Scheper, Rik J.

2014-01-01

Many cell-based regenerative medicine strategies toward tissue-engineered constructs are currently being explored. Cell–cell interactions and interactions with different biomaterials are extensively investigated, whereas very few studies address how cultured cells will interact with soluble wound-healing mediators that are present within the wound bed after transplantation. The aim of this study was to determine how adipose tissue-derived mesenchymal stem cells (ASC), dermal fibroblasts, and keratinocytes will react when they come in contact with the deep cutaneous burn wound bed. Burn wound exudates isolated from deep burn wounds were found to contain many cytokines, including chemokines and growth factors related to inflammation and wound healing. Seventeen mediators were identified by ELISA (concentration range 0.0006–9 ng/mg total protein), including the skin-specific chemokine CCL27. Burn wound exudates activated both ASC and dermal fibroblasts, but not keratinocytes, to increase secretion of CXCL1, CXCL8, CCL2, and CCL20. Notably, ASC but not fibroblasts or keratinocytes showed significant increased secretion of vascular endothelial growth factor (5-fold) and interleukin-6 (253-fold), although when the cells were incorporated in bi-layered skin substitute (SS) these differences were less pronounced. A similar discrepancy between ASC and dermal fibroblast mono-cultures was observed when recombinant human-CCL27 was used instead of burn wound exudates. Although CCL27 did not stimulate the secretion of any of the wound-healing mediators by keratinocytes, these cells, in contrast to ASC or dermal fibroblasts, showed increased proliferation and migration. Taken together, these results indicate that on transplantation, keratinocytes are primarily activated to promote wound closure. In contrast, dermal fibroblasts and, in particular, ASC respond vigorously to factors present in the wound bed, leading to increased secretion of angiogenesis/granulation tissue

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

Meeting report of the 2016 bone marrow adiposity meeting

PubMed Central

van der Eerden, Bram; van Wijnen, André

2017-01-01

Abstract There is considerable interest in the physiology and pathology, as well as the cellular and molecular biology, of bone marrow adipose tissue (BMAT). Because bone marrow adiposity is linked not only to systemic energy metabolism, but also to both bone marrow and musculoskeletal disorders, this biologic compartment has become of major interest to investigators from diverse disciplines. Bone marrow adiposity represents a virtual multi-tissue endocrine organ, which encompasses cells from multiple developmental lineages (e.g., mesenchymal, myeloid, lymphoid) and occupies all the non-osseous and non-cartilaginous space within long bones. A number of research groups are now focusing on bone marrow adiposity to understand a range of clinical afflictions associated with bone marrow disorders and to consider mechanisms-based strategies for future therapies. PMID:28410005

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.

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.

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

Combined influence of basal media and fibroblast growth factor on the expansion and differentiation capabilities of adipose-derived stem cells.

PubMed

Ahearne, Mark; Lysaght, Joanne; Lynch, Amy P

2014-01-01

Interest in adipose-derived stem cells (ASCs) has increased in recent years due to their multi-linage differentiation capabilities. While much work has been done to optimize the differentiation media, few studies have focused on examining the influence of different expansion media on cell behavior. In this study, three basal media (low glucose Dulbecco's modified Eagle's medium (DMEM), high glucose DMEM and DMEM-F12) supplemented with or without fibroblast growth factor 2 (FGF) were examined to assess their suitability for expanding ASCs. Flow cytometry, colony-forming unit assays (CFU-Fs) and differentiation assays were utilized to study cell behavior. High glucose media CFU-Fs produced fewest colonies while the addition of FGF increased colony size. By passage 2, the majority of cells were positive for CD44, 45, 73, 90 and 105 and negative for CD14, 31 and 45, indicating a mesenchymal phenotype. A sub-population of CD34 positive cells was present among passage 2 cells; however, by passage 4 the cells were negative for CD34. FGF has a negative effective on passage 4 ASC adipogenesis and high glucose media plus FGF-enhanced osteogenic capacity of passage 4 ASCs. FGF supplemented basal media were most suitable for chondrogenesis. High glucose media plus FGF appeared to be the most beneficial for priming ASCs to induce a keratocyte phenotype. These findings demonstrate the reciprocal effect FGF and basal media have on ASCs. This research has implications for those interested regenerating bone, cartilage, cornea or adipose tissues.

FGF8 signaling sustains progenitor status and multipotency of cranial neural crest-derived mesenchymal cells in vivo and in vitro

PubMed Central

Shao, Meiying; Liu, Chao; Song, Yingnan; Ye, Wenduo; He, Wei; Yuan, Guohua; Gu, Shuping; Lin, Congxin; Ma, Liang; Zhang, Yanding; Tian, Weidong; Hu, Tao; Chen, YiPing

2015-01-01

The cranial neural crest (CNC) cells play a vital role in craniofacial development and regeneration. They are multi-potent progenitors, being able to differentiate into various types of tissues. Both pre-migratory and post-migratory CNC cells are plastic, taking on diverse fates by responding to different inductive signals. However, what sustains the multipotency of CNC cells and derivatives remains largely unknown. In this study, we present evidence that FGF8 signaling is able to sustain progenitor status and multipotency of CNC-derived mesenchymal cells both in vivo and in vitro. We show that augmented FGF8 signaling in pre-migratory CNC cells prevents cell differentiation and organogenesis in the craniofacial region by maintaining their progenitor status. CNC-derived mesenchymal cells with Fgf8 overexpression or control cells in the presence of exogenous FGF8 exhibit prolonged survival, proliferation, and multi-potent differentiation capability in cell cultures. Remarkably, exogenous FGF8 also sustains the capability of CNC-derived mesenchymal cells to participate in organogenesis such as odontogenesis. Furthermore, FGF8-mediated signaling strongly promotes adipogenesis but inhibits osteogenesis of CNC-derived mesenchymal cells in vitro. Our results reveal a specific role for FGF8 in the maintenance of progenitor status and in fate determination of CNC cells, implicating a potential application in expansion and fate manipulation of CNC-derived cells in stem cell-based craniofacial regeneration. PMID:26243590

Mesenchymal Stem Cell-Derived Factors Restore Function to Human Frataxin-Deficient Cells.

PubMed

Kemp, Kevin; Dey, Rimi; Cook, Amelia; Scolding, Neil; Wilkins, Alastair

2017-08-01

Friedreich's ataxia is an inherited neurological disorder characterised by mitochondrial dysfunction and increased susceptibility to oxidative stress. At present, no therapy has been shown to reduce disease progression. Strategies being trialled to treat Friedreich's ataxia include drugs that improve mitochondrial function and reduce oxidative injury. In addition, stem cells have been investigated as a potential therapeutic approach. We have used siRNA-induced knockdown of frataxin in SH-SY5Y cells as an in vitro cellular model for Friedreich's ataxia. Knockdown of frataxin protein expression to levels detected in patients with the disorder was achieved, leading to decreased cellular viability, increased susceptibility to hydrogen peroxide-induced oxidative stress, dysregulation of key anti-oxidant molecules and deficiencies in both cell proliferation and differentiation. Bone marrow stem cells are being investigated extensively as potential treatments for a wide range of neurological disorders, including Friedreich's ataxia. The potential neuroprotective effects of bone marrow-derived mesenchymal stem cells were therefore studied using our frataxin-deficient cell model. Soluble factors secreted by mesenchymal stem cells protected against cellular changes induced by frataxin deficiency, leading to restoration in frataxin levels and anti-oxidant defences, improved survival against oxidative stress and stimulated both cell proliferation and differentiation down the Schwann cell lineage. The demonstration that mesenchymal stem cell-derived factors can restore cellular homeostasis and function to frataxin-deficient cells further suggests that they may have potential therapeutic benefits for patients with Friedreich's ataxia.

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.

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

Percutaneous intraportal application of adipose tissue-derived mesenchymal stem cells using a balloon occlusion catheter in a porcine model of liver fibrosis.

PubMed

Avritscher, Rony; Abdelsalam, Mohamed E; Javadi, Sanaz; Ensor, Joe; Wallace, Michael J; Alt, Eckhard; Madoff, David C; Vykoukal, Jody V

2013-12-01

To investigate the safety and effectiveness of a novel endovascular approach for therapeutic cell delivery using a balloon occlusion catheter in a large animal model of liver fibrosis. Transcatheter arterial embolization with ethiodized oil (Ethiodol) and ethanol was used to induce liver damage in 11 pigs. Mesenchymal stem cells (MSCs) were harvested from adipose tissue and engineered to express green fluorescent protein (GFP). A balloon occlusion catheter was positioned in the bilateral first-order portal vein branches 2 weeks after embolization to allow intraportal application of MSCs in six experimental animals. MSCs were allowed to dwell for 10 minutes using prolonged balloon inflation. Five control animals received a sham injection of normal saline in a similar fashion. Hepatic venous pressure gradient (HVPG) was measured immediately before necropsy. Specimens from all accessible lobes were obtained with ultrasound-guided percutaneous 18-gauge biopsy 2 hours after cell application. All animals were euthanized within 4 weeks. Fluorescent microscopy was used to assess the presence and distribution of cells. Liver injury and fibrosis were successfully induced in all animals. MSCs (6-10 × 10(7)) were successfully delivered into the portal vein in the six experimental animals. Cell application was not associated with vascular complications. HVPG showed no instances of portal hypertension. GFP-expressing MSCs were visualized in biopsy specimens and were distributed primarily within the sinusoidal spaces; however, 4 weeks after implantation, MSCs could not be identified in histologic specimens. A percutaneous endovascular approach for cell delivery using a balloon occlusion catheter proved safe for intraportal MSC application in a large animal model of liver fibrosis. © 2013 SIR Published by SIR All rights reserved.

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.

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.

The anti-fibrotic effects of mesenchymal stem cells on irradiated lungs via stimulating endogenous secretion of HGF and PGE2

PubMed Central

Dong, Li-Hua; Jiang, Yi-Yao; Liu, Yong-Jun; Cui, Shuang; Xia, Cheng-Cheng; Qu, Chao; Jiang, Xin; Qu, Ya-Qin; Chang, Peng-Yu; Liu, Feng

2015-01-01

Radiation-induced pulmonary fibrosis is a common disease and has a poor prognosis owing to the progressive breakdown of gas exchange regions in the lung. Recently, a novel strategy of administering mesenchymal stem cells for pulmonary fibrosis has achieved high therapeutic efficacy. In the present study, we attempted to use human adipose tissue-derived mesenchymal stem cells to prevent disease in Sprague-Dawley rats that received semi-thoracic irradiation (15 Gy). To investigate the specific roles of mesenchymal stem cells in ameliorating radiation-induced pulmonary fibrosis, we treated control groups of irradiated rats with human skin fibroblasts or phosphate-buffered saline. After mesenchymal stem cells were infused, host secretions of hepatocyte growth factor (HGF) and prostaglandin E2 (PGE2) were elevated compared with those of the controls. In contrast, tumour necrosis factor-alpha (TNF-α) and transforming growth factor-beta1 (TGF-β1) levels were decreased after infusion of mesenchymal stem cells. Consequently, the architecture of the irradiated lungs was preserved without marked activation of fibroblasts or collagen deposition within the injured sites. Moreover, mesenchymal stem cells were able to prevent the irradiated type II alveolar epithelial cells from undergoing epithelial-mesenchymal transition. Collectively, these data confirmed that mesenchymal stem cells have the potential to limit pulmonary fibrosis after exposure to ionising irradiation. PMID:25736907

GMP-grade human fetal liver-derived mesenchymal stem cells for clinical transplantation.

PubMed

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

2015-01-01

Stem cell therapy seems a promising avenue in regenerative medicine. Within various stem cells, mesenchymal stem cells have progressively used for cellular therapy. Because of the age-related decreasing in the frequency and differentiating capacity of adult MSCs, fetal tissues such as fetal liver, lung, pancreas, spleen, etc. have been introduced as an alternative source of MSCs for cellular therapy. On the other hand, using stem cells as advanced therapy medicinal products, must be performed in compliance with cGMP as a quality assurance system to ensure the safety, quality, and identity of cell products during translation from the basic stem cell sciences into clinical cell transplantation. In this chapter the authors have demonstrated the manufacturing of GMP-grade human fetal liver-derived mesenchymal stem cells.

The Effects of Cyclic Hydrostatic Pressure on Chondrogenesis and Viability of Human Adipose- and Bone Marrow-Derived Mesenchymal Stem Cells in Three-Dimensional Agarose Constructs

PubMed Central

Puetzer, Jennifer; Williams, John; Gillies, Allison; Bernacki, Susan

2013-01-01

This study investigates the effects of cyclic hydrostatic pressure (CHP) on chondrogenic differentiation of human adipose-derived stem cells (hASCs) in three-dimensional (3-D) agarose constructs maintained in a complete growth medium without soluble chondrogenic inducing factors. hASCs were seeded in 2% agarose hydrogels and exposed to 7.5 MPa CHP for 4 h per day at a frequency of 1 Hz for up to 21 days. On days 0, 7, 14, and 21, the expression levels of collagen II, Sox9, aggrecan, and cartilage oligomeric matrix protein (COMP) were examined by real-time reverse transcriptase–polymerase chain reaction analysis. Gene expression analysis found collagen II mRNA expression in only the CHP-loaded construct at day 14 and at no other time during the study. CHP-loaded hASCs exhibited upregulated mRNA expression of Sox9, aggrecan, and COMP at day 7 relative to unloaded controls, suggesting that CHP initiated chondrogenic differentiation of hASCs in a manner similar to human bone marrow-derived mesenchymal stem cells (hMSC). By day 14, however, loaded hASC constructs exhibited significantly lower mRNA expression of the chondrogenic markers than unloaded controls. Additionally, by day 21, the samples exhibited little measurable mRNA expression at all, suggesting a decreased viability. Histological analysis validated the lack of mRNA expression at day 21 for both the loaded and unloaded control samples with a visible decrease in the cell number and change in morphology. A comparative study with hASCs and hMSCs further examined long-term cell viability in 3-D agarose constructs of both cell types. Decreased cell metabolic activity was observed throughout the 21-day experimental period in both the CHP-loaded and control constructs of both hMSCs and hASCs, suggesting a decrease in cell metabolic activity, alluding to a decrease in cell viability. This suggests that a 2% agarose hydrogel may not optimally support hASC or hMSC viability in a complete growth medium in the

Potential use of mesenchymal stem cells in human meniscal repair: current insights

PubMed Central

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

2017-01-01

The menisci of the human knee play an important role in maintaining normal functions to provide stability and nutrition to the articular cartilage, and to absorb shock. Once injured, these important structures have very limited natural healing potential. Unfortunately, the traditional arthroscopic meniscectomy performed on these damaged menisci may predispose the joint toward early development of osteoarthritis. Although a very limited number of studies are available, mesenchymal stem cells (MSCs) have been investigated as an alternative therapeutic modality to repair human knee meniscal tears. This review summarizes the results of published applications of MSCs in human patients, which showed that the patients who received MSCs (autologous adipose tissue-derived stem cells or culture-expanded bone marrow-derived stem cells) presented symptomatic improvements, along with magnetic resonance imaging evidences of the meniscal repair. PMID:28356779

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.

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.

Isolation and cellular properties of mesenchymal cells derived from the decidua of human term placenta.

PubMed

Kanematsu, Daisuke; Shofuda, Tomoko; Yamamoto, Atsuyo; Ban, Chiaki; Ueda, Takafumi; Yamasaki, Mami; Kanemura, Yonehiro

2011-09-01

The clinical promise of cell-based therapies is generally recognized, and has driven an intense search for good cell sources. In this study, we isolated plastic-adherent cells from human term decidua vera, called decidua-derived-mesenchymal cells (DMCs), and compared their properties with those of bone marrow-derived-mesenchymal stem cells (BM-MSCs). The DMCs strongly expressed the mesenchymal cell marker vimentin, but not cytokeratin 19 or HLA-G, and had a high proliferative potential. That is, they exhibited a typical fibroblast-like morphology for over 30 population doublings. Cells phenotypically identical to the DMCs were identified in the decidua vera, and genotyping confirmed that the DMCs were derived from the maternal components of the fetal adnexa. Flow cytometry analysis showed that the expression pattern of CD antigens on the DMCs was almost identical to that on BM-MSCs, but some DMCs expressed the CD45 antigen, and over 50% of them also expressed anti-fibroblast antigen. In vitro, the DMCs showed good differentiation into chondrocytes and moderate differentiation into adipocytes, but scant evidence of osteogenesis, compared with the BM-MSCs. Gene expression analysis showed that, compared with BM-MSCs, the DMCs expressed higher levels of TWIST2 and RUNX2 (which are associated with early mesenchymal development and/or proliferative capacity), several matrix metalloproteinases (MMP1, 3, 10, and 12), and cytokines (BMP2 and TGFB2), and lower levels of MSX2, interleukin 26, and HGF. Although DMCs did not show the full multipotency of BM-MSCs, their higher proliferative ability indicates that their cultivation would require less maintenance. Furthermore, the use of DMCs avoids the ethical concerns associated with the use of embryonic tissues, because they are derived from the maternal portion of the placenta, which is otherwise discarded. Thus, the unique properties of DMCs give them several advantages for clinical use, making them an interesting and

Human gingiva-derived mesenchymal stem cells are superior to bone marrow-derived mesenchymal stem cells for cell therapy in regenerative medicine

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

Tomar, Geetanjali B.; Srivastava, Rupesh K.; Gupta, Navita

2010-03-12

Mesenchymal stem cells (MSCs) are capable of self-renewal and differentiation into multiple cell lineages. Presently, bone marrow is considered as a prime source of MSCs; however, there are some drawbacks and limitations in use of these MSCs for cell therapy. In this study, we demonstrate that human gingival tissue-derived MSCs have several advantages over bone marrow-derived MSCs. Gingival MSCs are easy to isolate, homogenous and proliferate faster than bone marrow MSCs without any growth factor. Importantly, gingival MSCs display stable morphology and do not loose MSC characteristic at higher passages. In addition, gingival MSCs maintain normal karyotype and telomerase activitymore » in long-term cultures, and are not tumorigenic. Thus, we reveal that human gingiva is a better source of MSCs than bone marrow, and large number of functionally competent clinical grade MSCs can be generated in short duration for cell therapy in regenerative medicine and tissue engineering.« less

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

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.

Enhancement of the repair of dog alveolar cleft by an autologous iliac bone, bone marrow-derived mesenchymal stem cell, and platelet-rich fibrin mixture.

PubMed

Yuanzheng, Chen; Yan, Gao; Ting, Li; Yanjie, Fu; Peng, Wu; Nan, Bai

2015-05-01

Autologous bone graft has been regarded as the criterion standard for the repair of alveolar cleft. However, the most prominent issue in alveolar cleft treatment is the high absorption rate of the bone graft. The authors' objective was to investigate the effects of an autologous iliac bone, bone marrow-derived mesenchymal stem cell, and platelet-rich fibrin mixture on the repair of dog alveolar cleft. Twenty beagle dogs with unilateral alveolar clefts created by surgery were divided randomly into four groups: group A underwent repair with an autologous iliac bone, bone marrow-derived mesenchymal stem cell, and platelet-rich fibrin mixture; group B underwent repair with autologous iliac bone and bone marrow-derived mesenchymal stem cells; group C underwent repair with autologous iliac bone and platelet-rich fibrin; and group D underwent repair with autologous iliac bone as the control. One day and 6 months after transplantation, the transplant volumes and bone mineral density were assessed by quantitative computed tomography. All of the transplants were harvested for hematoxylin and eosin staining 6 months later. Bone marrow-derived mesenchymal stem cells and platelet-rich fibrin transplants formed the greatest amounts of new bone among the four groups. The new bone formed an extensive union with the underlying maxilla in groups A, B, and C. Transplants with the bone marrow-derived mesenchymal stem cells, platelet-rich fibrin, and their mixture retained the majority of their initial volume, whereas the transplants in the control group showed the highest absorption rate. Bone mineral density of transplants with the bone marrow-derived mesenchymal stem cells, platelet-rich fibrin, and their mixture 6 months later was significantly higher than in the control group (p < 0.05), and was the highest in bone marrow-derived mesenchymal stem cells and platelet-rich fibrin mixed transplants. Hematoxylin and eosin staining showed that the structure of new bones formed the best

Exosomes derived from human adipose mensenchymal stem cells accelerates cutaneous wound healing via optimizing the characteristics of fibroblasts.

PubMed

Hu, Li; Wang, Juan; Zhou, Xin; Xiong, Zehuan; Zhao, Jiajia; Yu, Ran; Huang, Fang; Zhang, Handong; Chen, Lili

2016-09-12

Prolonged healing and scar formation are two major challenges in the treatment of soft tissue trauma. Adipose mesenchymal stem cells (ASCs) play an important role in tissue regeneration, and recent studies have suggested that exosomes secreted by stem cells may contribute to paracrine signaling. In this study, we investigated the roles of ASCs-derived exosomes (ASCs-Exos) in cutaneous wound healing. We found that ASCs-Exos could be taken up and internalized by fibroblasts to stimulate cell migration, proliferation and collagen synthesis in a dose-dependent manner, with increased genes expression of N-cadherin, cyclin-1, PCNA and collagen I, III. In vivo tracing experiments demonstrated that ASCs-Exos can be recruited to soft tissue wound area in a mouse skin incision model and significantly accelerated cutaneous wound healing. Histological analysis showed increased collagen I and III production by systemic administration of exosomes in the early stage of wound healing, while in the late stage, exosomes might inhibit collagen expression to reduce scar formation. Collectively, our findings indicate that ASCs-Exos can facilitate cutaneous wound healing via optimizing the characteristics of fibroblasts. Our results provide a new perspective and therapeutic strategy for the use of ASCs-Exos in soft tissue repair.

Iota-carrageenan/chitosan/gelatin scaffold for the osteogenic differentiation of adipose-derived MSCs in vitro.

PubMed

Li, Junjie; Yang, Boguang; Qian, Yufeng; Wang, Qiyu; Han, Ruijin; Hao, Tong; Shu, Yao; Zhang, Yabin; Yao, Fanglian; Wang, Changyong

2015-10-01

In this study, we have developed ι-carrageenan/chitosan/gelatin (CCG) scaffold containing multiple functional groups (-NH2 , -OH, -COOH, and -SO3 H) to resemble the native extracellular matrix (ECM), using the ion-shielding technology and ultrasonic dispersion method. Fourier transform infrared spectroscopy (FTIR) of the CCG scaffolds suggests that the formation of CCG network involves electrostatic interactions between ι-carrageenan (ι-CA) and chitosan/gelatin, and the covalent cross-linking among amino groups of chitosan and/or gelatin. Scanning electron microscopic (SEM) observation reveals that the porous structure of scaffolds can be modulated by the ratio of ι-CA to chitosan/gelatin. The swelling ratio of the hydrogels increases as the ι-CA contents increase. Using differential scanning calorimetry, we found that the double helix structure of ι-CA is only stabilized at low contents of ι-CA in the CCG scaffolds (e.g., 5 wt %). The scaffolds containing 5% ι-CA showed the best protein adsorption capacity (4.46 ± 0.63 μg protein/mg scaffold) and elastic modulus (5.37 ± 1.03 MPa). In addition, the CCG scaffolds exhibit excellent support for adipose-derived mesenchymal stem cells (ADMSCs) attachment and proliferation, and they can improve the osteogenic differentiation and neovascularization capacities of ADMSCs. Overall, we conclude that the CCG may represent an ideal scaffold material for bone tissue engineering. © 2014 Wiley Periodicals, Inc.

Evaluation of gold nanotracers to track adipose-derived stem cells in a PEGylated fibrin gel for dermal tissue engineering applications

PubMed Central

Chung, Eunna; Nam, Seung Yun; Ricles, Laura M; Emelianov, Stanislav Y; Suggs, Laura J

2013-01-01

Evaluating the regenerative capacity of a tissue-engineered device in a noninvasive and synchronous manner is critical to determining the mechanisms for success in clinical applications. In particular, directly tracking implanted cells in a three-dimensional (3D) scaffold is desirable in that it enables the monitoring of cellular activity in a specific and localized manner. The authors’ group has previously demonstrated that the PEGylation of fibrin results in a 3D scaffold that supports morphologic and phenotypic changes in mesenchymal stem cells that may be advantageous in wound healing applications. Recently, the authors have evaluated adipose-derived stem cells (ASCs) as a mesenchymal cell source to regenerate skin and blood vessels due to their potential for proliferation, differentiation, and production of growth factors. However, tracking and monitoring ASCs in a 3D scaffold, such as a PEGylated fibrin gel, have not yet been fully investigated. In the current paper, nanoscale gold spheres (20 nm) as cell tracers for ASCs cultured in a PEGylated fibrin gel were evaluated. An advanced dual-imaging modality combining ultrasound and photoacoustic imaging was utilized to monitor rat ASCs over time. The ASCs took up gold nanotracers and could be detected up to day 16 with high sensitivity using photoacoustic imaging. There were no detrimental effects on ASC morphology, network formation, proliferation, and protein expression/secretion (ie, smooth muscle α-actin, vascular endothelial growth factor, matrix metalloproteinase-2, and matrix metalloproteinase-9) associated with gold nanotracers. Therefore, utilization of gold nanotracers can be an effective strategy to monitor the regenerative process of a stem cell source in a 3D gel for vascular and dermal tissue engineering applications. PMID:23345978

Enhancement of osteogenic differentiation of rat adipose tissue-derived mesenchymal stem cells by zinc sulphate under electromagnetic field via the PKA, ERK1/2 and Wnt/β-catenin signaling pathways

PubMed Central

Fathi, Ezzatollah; Farahzadi, Raheleh

2017-01-01

Zinc ion as an essential trace element and electromagnetic fields (EMFs) has been reported to be involved in the regulation of bone metabolism. The aim of this study was to elucidate the effects of zinc sulphate (ZnSO4) on the osteogenic differentiation of adipose tissue-derived mesenchymal stem cells (ADSCs) in the presence of EMF as a strategy in osteoporosis therapy. Alkaline phophatase (ALP) activity measurement, calcium assay and expression of several osteoblastic marker genes were examined to assess the effect of ZnSO4 on the osteogenic differentiation of ADSCs under EMF. The expression of cAMP and PKA was evaluated by ELISA. The expression of β-catenin, Wnt1, Wnt3a, low-density lipoprotein receptor-related protein 5 (LRP5) and reduced dickkopf1 (DKK1) genes were used to detect the Wnt/β-catenin pathway. It was found that ZnSO4, in the presence of EMF, resulted in an increase in the expression of osteogenic genes, ALP activity and calcium levels. EMF, in the presence of ZnSO4, increased the cAMP level and protein kinase A (PKA) activity. Treatment of ADSCs with (MAPK)/ERK kinase 1/2 inhibitor, or PKA inhibitor, significantly inhibited the promotion of osteogenic markers, indicating that the induction of osteogenesis was dependent on the ERK and PKA signaling pathways. Real-time PCR analysis showed that ZnSO4, in the presence of EMF, increased the mRNA expressions of β-catenin, Wnt1, Wnt3a, LRP5 and DKK1. In this study, it was shown that 0.432 μg/ml ZnSO4, in the presence of 50 Hz, 20 mT EMF, induced the osteogenic differentiation of ADSCs via PKA, ERK1/2 and Wnt/β-catenin signaling pathways. PMID:28339498

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

Evaluation of the effects of different culture media on the myogenic differentiation potential of adipose tissue- or bone marrow-derived human mesenchymal stem cells.

PubMed

Stern-Straeter, Jens; Bonaterra, Gabriel Alejandro; Juritz, Stephanie; Birk, Richard; Goessler, Ulrich Reinhart; Bieback, Karen; Bugert, Peter; Schultz, Johannes; Hörmann, Karl; Kinscherf, Ralf; Faber, Anne

2014-01-01

The creation of functional muscles/muscle tissue from human stem cells is a major goal of skeletal muscle tissue engineering. Mesenchymal stem cells (MSCs) from fat/adipose tissue (AT-MSCs), as well as bone marrow (BM-MSCs) have been shown to bear myogenic potential, which makes them candidate stem cells for skeletal muscle tissue engineering applications. The aim of this study was to analyse the myogenic differentiation potential of human AT-MSCs and BM-MSCs cultured in six different cell culture media containing different mixtures of growth factors. The following cell culture media were used in our experiments: mesenchymal stem cell growth medium (MSCGM)™ as growth medium, MSCGM + 5-azacytidine (5-Aza), skeletal muscle myoblast cell growth medium (SkGM)-2 BulletKit™, and 5, 30 and 50% conditioned cell culture media, i.e., supernatant of human satellite cell cultures after three days in cell culture mixed with MSCGM. Following the incubation of human AT-MSCs or BM-MSCs for 0, 4, 8, 11, 16 or 21 days with each of the cell culture media, cell proliferation was measured using the alamarBlue® assay. Myogenic differentiation was evaluated by quantitative gene expression analyses, using quantitative RT-PCR (qRT-PCR) and immunocytochemical staining (ICC), using well-defined skeletal markers, such as desmin (DES), myogenic factor 5 (MYF5), myosin, heavy chain 8, skeletal muscle, perinatal (MYH8), myosin, heavy chain 1, skeletal muscle, adult (MYH1) and skeletal muscle actin-α1 (ACTA1). The highest proliferation rates were observed in the AT-MSCs and BM-MSCs cultured with SkGM-2 BulletKit medium. The average proliferation rate was higher in the AT-MSCs than in the BM-MSCs, taking all six culture media into account. qRT-PCR revealed the expression levels of the myogenic markers, ACTA1, MYH1 and MYH8, in the AT-MSC cell cultures, but not in the BM-MSC cultures. The muscle-specific intermediate filament, DES, was only detected (by ICC) in the AT-MSCs, but not in the BM

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

Comparison of differentiation potential of male mouse adipose tissue and bone marrow derived-mesenchymal stem cells into germ cells

PubMed Central

Hosseinzadeh Shirzeily, Maryam; Pasbakhsh, Parichehr; Amidi, Fardin; Mehrannia, Kobra; Sobhani, Aligholi

2013-01-01

Background: Recent publications about differentiation of stem cells to germ cells have motivated researchers to make new approaches to infertility. In vitro production of germ cells improves understanding differentiation process of male and female germ cells. Due to the problem of using embryonic stem cells (ESC), it’s necessary the mentioned cells be replaced with some adult multi-potent stem cells in laboratories. Objective: The aim of this study was to obtain germ cells from appropriate source beyond ESC and compare differential potentials of adipocytes derived stem cells (ADMSCs) with bone marrow derived stem cells (BMMSCs). Materials and Methods: To find multi-potential entity, after providing purified ADMSCs and BMMSCs, differentiation to osteoblast and adipocyte was confirmed by using appropriate culture medium. To confirm mesenchymal lineage production superficial markers (expression of CD90 and CD44 and non-expression of CD45 and CD31) were investigated by flowcytometry. Then the cells were differentiated to germ cells in inductive medium containing retinoic acid for 7days. To evaluate germ cells characteristic markers [Dazl (Deleted in azoospermia-like), Mvh (Mouse vasa homolog gene), Stra8 (Stimulated by retinoic acid) and Scp3 (Synaptonemal complex protein 3)] flowcytometry, imunoflorescence and real time PCR were used. Results: Both types of cells were able to differentiate into osteoblast and adipocyte cells and presentation of stem cell superficial markers (CD90, CD44) and absence of endothelial and blood cell markers (CD31, CD45) were confirmative The flowcytometry, imunoflorescence and real time PCR results showed remarkable expression of germ cells characteristic markers (Mvh, Dazl, Stra8, and Scp3). Conclusion: It was found that although ADMSCs were attained easier and also cultured and differentiated rapidly, germ cell markers were expressed in BMMSCs significantly more than ADMSCs. This article extracted from M.Sc. thesis. (Maryam

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

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.

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.

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

Clonal analysis of synovial fluid stem cells to characterize and identify stable mesenchymal stromal cell/mesenchymal progenitor cell phenotypes in a porcine model: a cell source with enhanced commitment to the chondrogenic lineage.

PubMed

Ando, Wataru; Kutcher, Josh J; Krawetz, Roman; Sen, Arindom; Nakamura, Norimasa; Frank, Cyril B; Hart, David A

2014-06-01

Previous studies have demonstrated that porcine synovial membrane stem cells can adhere to a cartilage defect in vivo through the use of a tissue-engineered construct approach. To optimize this model, we wanted to compare effectiveness of tissue sources to determine whether porcine synovial fluid, synovial membrane, bone marrow and skin sources replicate our understanding of synovial fluid mesenchymal stromal cells or mesenchymal progenitor cells from humans both at the population level and the single-cell level. Synovial fluid clones were subsequently isolated and characterized to identify cells with a highly characterized optimal phenotype. The chondrogenic, osteogenic and adipogenic potentials were assessed in vitro for skin, bone marrow, adipose, synovial fluid and synovial membrane-derived stem cells. Synovial fluid cells then underwent limiting dilution analysis to isolate single clonal populations. These clonal populations were assessed for proliferative and differentiation potential by use of standardized protocols. Porcine-derived cells demonstrated the same relationship between cell sources as that demonstrated previously for humans, suggesting that the pig may be an ideal preclinical animal model. Synovial fluid cells demonstrated the highest chondrogenic potential that was further characterized, demonstrating the existence of a unique clonal phenotype with enhanced chondrogenic potential. Porcine stem cells demonstrate characteristics similar to those in human-derived mesenchymal stromal cells from the same sources. Synovial fluid-derived stem cells contain an inherent phenotype that may be optimal for cartilage repair. This must be more fully investigated for future use in the in vivo tissue-engineered construct approach in this physiologically relevant preclinical porcine model. Copyright © 2014 International Society for Cellular Therapy. Published by Elsevier Inc. All rights reserved.

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.

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

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.

Human adipose-derived stem cells (ADSC) and human periodontal ligament stem cells (PDLSC) as cellular substrates of a toxicity prediction assay.

PubMed

Corrêa, Natássia Caroline Resende; Kuligovski, Crisciele; Paschoal, Ariane Caroline Campos; Abud, Ana Paula Ressetti; Rebelatto, Carmen Lucia Kuniyoshi; Leite, Lidiane Maria Boldrini; Senegaglia, Alexandra Cristina; Dallagiovanna, Bruno; Aguiar, Alessandra Melo de

2018-02-01

With the increasing need to develop in vitro assays to replace animal use, human stem cell-derived methods are emerging and showing outstanding contributions to the toxicological screening of substances. Adult human stem cells such as adipose-derived stem cells (ADSC) and periodontal ligament stem cells (PDLSC) were used as cell substrates for a cytotoxicity assay and toxicity prediction using the neutral red uptake (NRU) assay. First, primary cell cultures from three independent donors, from each tissue source, were characterized as mesenchymal stem cells (MSC) by plastic adherence and appropriate immunophenotype for MSC markers (positive for CD90, CD73, and CD105 and negative for CD11b, CD34, CD45, HLADR, and CD19). Furthermore, ADSC and PDLSC were able to differentiate into adipocytes and osteoblasts when maintained under the same culture conditions previously established for the NRU assay. NRU assays for three reference test substances were performed. R 2 was higher than 0.85 for all conditions, showing the feasibility to calculate IC 50 values. The IC 50 values were then used to predict the LD 50 of the test substances, which were comparable to previous results and the ICCVAM standard test report. Primary ADSC and PDLSC showed the potential to be considered as additional models for use in cytotoxicity assays. Copyright © 2017 Elsevier Inc. All rights reserved.

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.

Treatment With Human Wharton’s Jelly-Derived Mesenchymal Stem Cells Attenuates Sepsis-Induced Kidney Injury, Liver Injury, and Endothelial Dysfunction

PubMed Central

Cóndor, José M.; Rodrigues, Camila E.; de Sousa Moreira, Roberto; Canale, Daniele; Volpini, Rildo A.; Shimizu, Maria H.M.; Camara, Niels O.S.; Noronha, Irene de L.

2016-01-01

The pathophysiology of sepsis involves complex cytokine and inflammatory mediator networks. Downregulation of endothelial nitric oxide synthase contributes to sepsis-induced endothelial dysfunction. Human Wharton’s jelly-derived mesenchymal stem cells (WJ-MSCs) are known to reduce expression of proinflammatory cytokines and markers of apoptosis. We hypothesized that treatment with WJ-MSCs would protect renal, hepatic, and endothelial function in a cecal ligation and puncture (CLP) model of sepsis in rats. Rats were randomly divided into three groups: sham-operated rats; rats submitted to CLP and left untreated; and rats submitted to CLP and intraperitoneally injected, 6 hours later, with 1 × 106 WJ-MSCs. The glomerular filtration rate (GFR) was measured at 6 and 24 hours after CLP or sham surgery. All other studies were conducted at 24 hours after CLP or sham surgery. By 6 hours, GFR had decreased in the CLP rats. At 24 hours, Klotho renal expression significantly decreased. Treatment with WJ-MSCs improved the GFR; improved tubular function; decreased the CD68-positive cell count; decreased the fractional interstitial area; decreased expression of nuclear factor κB and of cytokines; increased expression of eNOS, vascular endothelial growth factor, and Klotho; attenuated renal apoptosis; ameliorated hepatic function; increased glycogen deposition in the liver; and improved survival. Sepsis-induced acute kidney injury is a state of Klotho deficiency, which WJ-MSCs can attenuate. Klotho protein expression was higher in WJ-MSCs than in human adipose-derived MSCs. Because WJ-MSCs preserve renal and hepatic function, they might play a protective role in sepsis. Significance Sepsis is the leading cause of death in intensive care units. Although many different treatments for sepsis have been tested, sepsis-related mortality rates remain high. It was hypothesized in this study that treatment with human Wharton’s jelly-derived mesenchymal stem cells (WJ-MSCs) would

The use of small interfering RNAs to inhibit adipocyte differentiation in human preadipocytes and fetal-femur-derived mesenchymal cells

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

Xu, Y.; Mirmalek-Sani, S.-H.; Yang, X.

2006-06-10

RNA interference (RNAi) has been used in functional genomics and offers innovative approaches in the development of novel therapeutics. Human mesenchymal stem cells offer a unique cell source for tissue engineering/regeneration strategies. The current study examined the potential of small interfering RNAs (siRNA) against human peroxisome proliferator activated receptor gamma (PPAR{gamma}) to suppress adipocyte differentiation (adipogenesis) in human preadipocytes and fetal-femur-derived mesenchymal cells. Adipogenesis was investigated using cellular and biochemical analysis. Transient transfection with PPAR{gamma}-siRNA using a liposomal-based strategy resulted in a significant inhibition of adipogenesis in human preadipocytes and fetal-femur-derived mesenchymal cells, compared to controls (cell, liposomal and negativemore » siRNA). The inhibitory effect of PPAR{gamma}-siRNA was supported by testing human PPAR{gamma} mRNA and adipogenic associated genes using reverse transcription polymerase chain reaction (RT-PCR) to adiponectin receptor 1 and 2 as well as examination of fatty acid binding protein 3 (FABP{sub 3}) expression, an adipocyte-specific marker. The current studies indicate that PPAR{gamma}-siRNA is a useful tool to study adipogenesis in human cells, with potential applications both therapeutic and in the elucidation of mesenchymal cell differentiation in the modulation of cell differentiation in human mesenchymal cells.« less

Differential marker expression by cultures rich in mesenchymal stem cells

PubMed Central

2013-01-01

Background Mesenchymal stem cells have properties that make them amenable to therapeutic use. However, the acceptance of mesenchymal stem cells in clinical practice requires standardized techniques for their specific isolation. To date, there are no conclusive marker (s) for the exclusive isolation of mesenchymal stem cells. Our aim was to identify markers differentially expressed between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. We compared and contrasted the phenotype of tissue cultures in which mesenchymal stem cells are rich and rare. By initially assessing mesenchymal stem cell differentiation, we established that bone marrow and breast adipose cultures are rich in mesenchymal stem cells while, in our hands, foreskin fibroblast and olfactory tissue cultures contain rare mesenchymal stem cells. In particular, olfactory tissue cells represent non-stem cell mesenchymal cells. Subsequently, the phenotype of the tissue cultures were thoroughly assessed using immuno-fluorescence, flow-cytometry, proteomics, antibody arrays and qPCR. Results Our analysis revealed that all tissue cultures, regardless of differentiation potential, demonstrated remarkably similar phenotypes. Importantly, it was also observed that common mesenchymal stem cell markers, and fibroblast-associated markers, do not discriminate between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. Examination and comparison of the phenotypes of mesenchymal stem cell and non-stem cell mesenchymal cell cultures revealed three differentially expressed markers – CD24, CD108 and CD40. Conclusion We indicate the importance of establishing differential marker expression between mesenchymal stem cells and non-stem cell mesenchymal cells in order to determine stem cell specific markers. PMID:24304471

Mesenchymal stem cell-derived microparticles ameliorate peritubular capillary rarefaction via inhibition of endothelial-mesenchymal transition and decrease tubulointerstitial fibrosis in unilateral ureteral obstruction.

PubMed

Choi, Hoon Young; Lee, Hyun Gyu; Kim, Beom Seok; Ahn, Sun Hee; Jung, Ara; Lee, Mirae; Lee, Jung Eun; Kim, Hyung Jong; Ha, Sung Kyu; Park, Hyeong Cheon

2015-03-11

Microparticles (MPs) derived from kidney-derived mesenchymal stem cells (KMSCs) have recently been reported to ameliorate rarefaction of peritubular capillaries (PTC) in ischemic kidneys via delivery of proangiogenic effectors. This study aimed to investigate whether KMSC-derived MPs show anti-fibrotic effects by ameliorating endothelial-to-mesenchymal transition (EndoMT) in human umbilical vein endothelial cells (HUVEC) in vitro and by preserving PTC in kidneys with unilateral ureteral obstruction (UUO) in vivo. MPs isolated from the supernatants of KMSC were co-cultured with HUVEC to assess their in vitro biologic effects on endothelial cells. Mice were treated with MPs via the tail vein after UUO injury to assess their anti-fibrotic and PTC sparing effects. Renal tubulointerstitial damage and inflammatory cell infiltration were examined with Masson's trichrome, F4/80 and α-smooth muscle actin (α-SMA) staining and PTC rarefaction index was determined by CD31 staining. KMSC-derived MPs significantly ameliorated EndoMT and improved in vitro proliferation of TGF-β1 treated HUVEC. In vivo administration of KMSC-derived MPs significantly inhibited EndoMT of PTC endothelial cells and improved PTC rarefaction in UUO kidneys. Furthermore, administration of KMSC-derived MPs inhibited inflammatory cell infiltration as well as tubulointerstitial fibrosis in UUO mice as demonstrated by decreased F4/80 and α-SMA-positive cells and Masson's trichrome staining, respectively. Our results suggest that KMSC-derived MPs ameliorate PTC rarefaction via inhibition of EndoMT and protect against progression of renal damage by inhibiting tubulointerstitial fibrosis.

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.

Parathyroid Hormone Directs Bone Marrow Mesenchymal Cell Fate.

PubMed

Fan, Yi; Hanai, Jun-Ichi; Le, Phuong T; Bi, Ruiye; Maridas, David; DeMambro, Victoria; Figueroa, Carolina A; Kir, Serkan; Zhou, Xuedong; Mannstadt, Michael; Baron, Roland; Bronson, Roderick T; Horowitz, Mark C; Wu, Joy Y; Bilezikian, John P; Dempster, David W; Rosen, Clifford J; Lanske, Beate

2017-03-07

Intermittent PTH administration builds bone mass and prevents fractures, but its mechanism of action is unclear. We genetically deleted the PTH/PTHrP receptor (PTH1R) in mesenchymal stem cells using Prx1Cre and found low bone formation, increased bone resorption, and high bone marrow adipose tissue (BMAT). Bone marrow adipocytes traced to Prx1 and expressed classic adipogenic markers and high receptor activator of nuclear factor kappa B ligand (Rankl) expression. RANKL levels were also elevated in bone marrow supernatant and serum, but undetectable in other adipose depots. By cell sorting, Pref1 + RANKL + marrow progenitors were twice as great in mutant versus control marrow. Intermittent PTH administration to control mice reduced BMAT significantly. A similar finding was noted in male osteoporotic patients. Thus, marrow adipocytes exhibit osteogenic and adipogenic characteristics, are uniquely responsive to PTH, and secrete RANKL. These studies reveal an important mechanism for PTH's therapeutic action through its ability to direct mesenchymal cell fate. Copyright © 2017 Elsevier Inc. All rights reserved.

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.

A Prospective, Nonrandomized, no Placebo-Controlled, Phase I/II Clinical Trial Assessing the Safety and Efficacy of Intramuscular Injection of Autologous Adipose Tissue-Derived Mesenchymal Stem Cells in Patients With Severe Buerger’s Disease

PubMed Central

Ra, Jeong Chan; Jeong, Euicheol C.; Kang, Sung Keun; Lee, Seog Ju; Choi, Kyoung Ho

2017-01-01

Buerger’s disease is a rare and severe disease affecting the blood vessels of the limbs. Adipose tissue-derived mesenchymal stem cells (ADSCs) have the potential to cure Buerger’s disease when developed as a stem cell drug. In the present study, we conducted a prospective, nonrandomized, no placebo-controlled, phase I/II clinical trial with a 2-year follow-up questionnaire survey. A total of 17 patients were intramuscularly administered autologous ADSCs at a dose of 5 million cells/kg. The incidence of adverse events (AEs), adverse drug reaction (ADR), and serious adverse events (SAEs) was monitored. No ADRs and SAEs related to stem cell treatment occurred during the 6-month follow-up. In terms of efficacy, the primary endpoint was increase in total walking distance (TWD). The secondary endpoint was improvement in rest pain, increase in pain-free walking distance (PFWD), toe–brachial pressure index (TBPI), transcutaneous oxygen pressure (TcPO2), and arterial brachial pressure index (ABPI). ADSCs demonstrated significant functional improvement results including increased TWD, PFWD, and rest pain reduction. No amputations were reported during the 6-month clinical trial period and in the follow-up questionnaire survey more than 2 years after the ADSC injection. In conclusion, intramuscular injection of ADSCs is very safe and is shown to prompt functional improvement in patients with severe Buerger’s disease at a dosage of 300 million cells per 60 kg of body weight. However, the confirmatory therapeutic efficacy and angiogenesis need further study. PMID:28713639

Generation and characterisation of human umbilical cord derived mesenchymal stem cells by explant method.

PubMed

Yusoff, Z; Maqbool, M; George, E; Hassan, R; Ramasamy, R

2016-06-01

Mesenchymal stem cells (MSCs) derived from human umbilical cord (UC) have been considered as an important tool for treating various malignancies, tissue repair and organ regeneration. Umbilical cord-derived mesenchymal stem cells (UC-MSCs) are better alternative to MSCs that derived from bone marrow (BM-MSCs) as they are regarded as medical waste with little ethical concern for research and easily culture-expanded. In this present study, the foetal distal end of human UC was utilised to generate MSC by explant method. Upon in vitro culture, adherent cells with fibroblastic morphology were generated with rapid growth kinetics. Under the respective inductive conditions, these cells were capable of differentiating into adipocytes and osteocytes; express an array of standard MSC's surface markers CD29, CD73, CD90, CD106 and MHC-class I. Further assessment of immunosuppression activity revealed that MSCs generated from UC had profoundly inhibited the proliferation of mitogen-activated T lymphocytes in a dosedependent manner. The current laboratory findings have reinforced the application of explant method to generate UCMSCs thus, exploring an ideal platform to fulfil the increasing demand of MSCs for research and potential clinical use.

Induced Pluripotent Stem Cell Derived Mesenchymal Stem Cells for Attenuating Age-Related Bone Loss

DTIC Science & Technology

2012-07-01

Mesenchymal stem cell (MSC) differentiation towards the bone forming osteoblastic lineage decreases as a function of age and may contribute to age-related...problem of age-related reduced availability of MSC we propose to examine the bone anabolic potential of induced pluripotent stem cell (iPS) derived MSC

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

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.

Defined xenogeneic-free and hypoxic environment provides superior conditions for long-term expansion of human adipose-derived stem cells.

PubMed

Yang, Sufang; Pilgaard, Linda; Chase, Lucas G; Boucher, Shayne; Vemuri, Mohan C; Fink, Trine; Zachar, Vladimir

2012-08-01

Development and implementation of therapeutic protocols based on stem cells or tissue-engineered products relies on methods that enable the production of substantial numbers of cells while complying with stringent quality and safety demands. In the current study, we aimed to assess the benefits of maintaining cultures of adipose-derived stem cells (ASCs) in a defined culture system devoid of xenogeneic components (xeno-free) and hypoxia over a 49-day growth period. Our data provide evidence that conditions involving StemPro mesenchymal stem cells serum-free medium (SFM) Xeno-Free and hypoxia (5% oxygen concentration) in the culture atmosphere provide a superior proliferation rate compared to a standard growth environment comprised of alpha-modified Eagle medium (A-MEM) supplemented with fetal calf serum (FCS) and ambient air (20% oxygen concentration) or that of A-MEM supplemented with FCS and hypoxia. Furthermore, a flow cytometric analysis and in vitro differentiation assays confirmed the immunophenotype stability and maintained multipotency of ASCs when expanded under xeno-free conditions and hypoxia. In conclusion, our data demonstrate that growth conditions utilizing a xeno-free and hypoxic environment not only provide an improved environment for the expansion of ASCs, but also set the stage as a culture system with the potential broad spectrum utility for regenerative medicine and tissue engineering applications.

Transgene Reactivation in Induced Pluripotent Stem Cell Derivatives and Reversion to Pluripotency of Induced Pluripotent Stem Cell-Derived Mesenchymal Stem Cells

PubMed Central

Galat, Yekaterina; Perepitchka, Mariana; Jennings, Lawrence J.; Iannaccone, Philip M.; Hendrix, Mary J.C.

2016-01-01

Induced pluripotent stem cells (iPSCs) have enormous potential in regenerative medicine and disease modeling. It is now felt that clinical trials should be performed with iPSCs derived with nonintegrative constructs. Numerous studies, however, including those describing disease models, are still being published using cells derived from iPSCs generated with integrative constructs. Our experimental work presents the first evidence of spontaneous transgene reactivation in vitro in several cellular types. Our results show that the transgenes were predominantly silent in parent iPSCs, but in mesenchymal and endothelial iPSC derivatives, the transgenes experienced random upregulation of Nanog and c-Myc. Additionally, we provide evidence of spontaneous secondary reprogramming and reversion to pluripotency in mesenchymal stem cells derived from iPSCs. These findings strongly suggest that the studies, which use cellular products derived from iPSCs generated with retro- or lentiviruses, should be evaluated with consideration of the possibility of transgene reactivation. The in vitro model described here provides insight into the earliest events of culture transformation and suggests the hypothesis that reversion to pluripotency may be responsible for the development of tumors in cell replacement experiments. The main goal of this work, however, is to communicate the possibility of transgene reactivation in retro- or lenti-iPSC derivatives and the associated loss of cellular fidelity in vitro, which may impact the outcomes of disease modeling and related experimentation. PMID:27193052

Self-assembled adult adipose-derived stem cell spheroids combined with biomaterials promote wound healing in a rat skin repair model.

PubMed

Hsu, Shan-Hui; Hsieh, Pai-Shan

2015-01-01

Adult adipose-derived stem cells (ASCs) are a type of multipotent mesenchymal stem cells (MSCs) with easy availability and serve as a potential cell source for cell-based therapy. Three-dimensional MSC spheroids may be derived from the self-assembly of individual MSCs grown on certain polymer membranes. In this study, we demonstrated that the self-assembled ASC spheroids on chitosan-hyaluronan membranes expressed more cytokine genes (fibroblast growth factor 1, vascular endothelial growth factor, and chemokine [C-C motif] ligand 2) as well as migration-associated genes (chemokine [C-X-C motif] receptor type 4 and matrix metalloprotease 1) compared with ASC dispersed single cells grown on culture dish. To evaluate the in vivo effects of these spheroids, we applied ASC single cells and ASC spheroids in a designed rat skin repair model. Wounds of 15 × 15 mm were created on rat dorsal skin, where ASCs were administered and covered with hyaluronan gel/chitosan sponge to maintain a moist environment. Results showed that skin wounds treated with ASC spheroids had faster wound closure and a significantly higher ratio of angiogenesis. Tracking of fluorescently labeled ASCs showed close localization of ASC spheroids to microvessels, suggesting enhanced angiogenesis through paracrine effects. Based on the in vitro and in vivo results, the self-assembled ASC spheroids may be a promising cellular source for skin tissue engineering and wound regeneration. © 2014 by the Wound Healing Society.

Polyurethane/polylactide-based biomaterials combined with rat olfactory bulb-derived glial cells and adipose-derived mesenchymal stromal cells for neural regenerative medicine applications.

PubMed

Grzesiak, Jakub; Marycz, Krzysztof; Szarek, Dariusz; Bednarz, Paulina; Laska, Jadwiga

2015-01-01

Research concerning the elaboration and application of biomaterial which may support the nerve tissue regeneration is currently one of the most promising directions. Biocompatible polymer devices are noteworthy group among the numerous types of potentially attractive biomaterials for regenerative medicine application. Polylactides and polyurethanes may be utilized for developing devices for supporting the nerve regeneration, like nerve guide conduits or bridges connecting the endings of broken nerve tracts. Moreover, the combination of these biomaterial devices with regenerative cell populations, like stem or precursor cells should significantly improve the final therapeutic effect. Therefore, the composition and structure of final device should support the proper adhesion and growth of cells destined for clinical application. In current research, the three polymer mats elaborated for connecting the broken nerve tracts, made from polylactide, polyurethane and their blend were evaluated both for physical properties and in vitro, using the olfactory-bulb glial cells and mesenchymal stem cells. The evaluation of Young's modulus, wettability and roughness of obtained materials showed the differences between analyzed samples. The analysis of cell adhesion, proliferation and morphology showed that the polyurethane-polylactide blend was the most neutral for cells in culture, while in the pure polymer samples there were significant alterations observed. Our results indicated that polyurethane-polylactide blend is an optimal composition for culturing and delivery of glial and mesenchymal stem cells. Copyright © 2015. Published by Elsevier B.V.

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

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.

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