Ulex europaeus I lectin as a marker for vascular endothelium in human tissues.

PubMed

Holthöfer, H; Virtanen, I; Kariniemi, A L; Hormia, M; Linder, E; Miettinen, A

1982-07-01

Ulex europaeus I agglutinin, a lectin specific for some alpha-L-fucose-containing glycocompounds, was used in fluorescence microscopy to stain cryostat sections of human tissues. The endothelium of vessels of all sizes was stained ubiquitously in all tissues studied as judged by double staining with a known endothelial marker, antibodies against human clotting factor VIII. Cultured human umbilical vein endothelial cells, but not fibroblasts, also bound Ulex lectin. The staining was not affected by the blood group type of the tissue donor. In some tissues Ulex lectin presented additional binding to epithelial structures. Also, this was independent on the blood group or the ability of the tissue donor to secrete soluble blood group substances. Lotus tetragonolobus agglutinin, another lectin specific for some alpha-L-fucose-containing moieties failed to react with endothelial cells. Our results suggest that Ulex europaeus I agglutinin is a good histologic marker for endothelium in human tissues.

Connective tissue growth factor and bone morphogenetic protein 2 are induced following myocardial ischemia in mice and humans.

PubMed

Rutkovskiy, Arkady; Sagave, Julia; Czibik, Gabor; Baysa, Anton; Zihlavnikova Enayati, Katarina; Hillestad, Vigdis; Dahl, Christen Peder; Fiane, Arnt; Gullestad, Lars; Gravning, Jørgen; Ahmed, Shakil; Attramadal, Håvard; Valen, Guro; Vaage, Jarle

2017-09-01

We aimed to study the cardiac expression of bone morphogenetic protein 2, its receptor 1 b, and connective tissue growth factor, factors implicated in cardiac embryogenesis, following ischemia/hypoxia, heart failure, and in remodeling hearts from humans and mice. Biopsies from the left ventricle of patients with end-stage heart failure due to dilated cardiomyopathy or coronary artery disease were compared with donor hearts and biopsies from patients with normal heart function undergoing coronary artery bypass grafting. Mouse model of post-infarction remodeling was made by permanent ligation of the left coronary artery. Hearts were analyzed by real-time polymerase chain reaction and Western blotting after 24 hours and after 2 and 4 weeks. Patients with dilated cardiomyopathy and mice post-infarction had increased cardiac expression of connective tissue growth factor. Bone morphogenetic protein 2 was increased in human hearts failing due to coronary artery disease and in mice post-infarction. Gene expression of bone morphogenetic protein receptor 1 beta was reduced in hearts of patients with failure, but increased two weeks following permanent ligation of the left coronary artery in mice. In conclusion, connective tissue growth factor is upregulated in hearts of humans with dilated cardiomyopathy, bone morphogenetic protein 2 is upregulated in remodeling due to myocardial infarction while its receptor 1 b in human failing hearts is downregulated. A potential explanation might be an attempt to engage regenerative processes, which should be addressed by further, mechanistic studies.

Formation of proteoglycan and collagen-rich scaffold-free stiff cartilaginous tissue using two-step culture methods with combinations of growth factors.

PubMed

Miyazaki, Tatsuya; Miyauchi, Satoshi; Matsuzaka, Satoshi; Yamagishi, Chie; Kobayashi, Kohei

2010-05-01

Tissue-engineered cartilage may be expected to serve as an alternative to autologous chondrocyte transplantation treatment. Several methods for producing cartilaginous tissue have been reported. In this study, we describe the production of scaffold-free stiff cartilaginous tissue of pig and human, using allogeneic serum and growth factors. The tissue was formed in a mold using chondrocytes recovered from alginate bead culture and maintained in a medium with transforming growth factor-beta and several other additives. In the case of porcine tissue, the tear strength of the tissue and the contents of proteoglycan (PG) and collagen per unit of DNA increased dose-dependently with transforming growth factor-beta. The length of culture was significantly and positively correlated with thickness, tear strength, and PG and collagen contents. Tear strength showed positive high correlations with both PG and collagen contents. A positive correlation was also seen between PG content and collagen content. Similar results were obtained with human cartilaginous tissue formed from chondrocytes expanded in monolayer culture. Further, an in vivo pilot study using pig articular cartilage defect model demonstrated that the cartilaginous tissue was well integrated with surrounding tissue at 13 weeks after the implantation. In conclusion, we successfully produced implantable scaffold-free stiff cartilaginous tissue, which characterized high PG and collagen contents.

Platelet Glycoprotein lb-1X and Malignancy

DTIC Science & Technology

2011-09-01

Constitutive production and thrombin-induced release of vascular endothelial growth factor by human megakaryocytes and platelets. Proc Natl Acad Sci...JM, Hakim J, de Prost D. Vascular endothelial growth factor production by fibroblasts in response to factor VIIa binding to tissue factor involves...interactions in vitro. (14) The extrinsic pathway of coagulation triggered by factor VII ( FVII ) and tissue factor can be activated in cancer patients. (15

Syndecan-2 Is a Novel Target of Insulin-Like Growth Factor Binding Protein-3 and Is Over-Expressed in Fibrosis

PubMed Central

Ruiz, Ximena D.; Mlakar, Logan R.; Yamaguchi, Yukie; Su, Yunyun; Larregina, Adriana T.; Pilewski, Joseph M.; Feghali-Bostwick, Carol A.

2012-01-01

Extracellular matrix deposition and tissue scarring characterize the process of fibrosis. Transforming growth factor beta (TGFβ) and Insulin-like growth factor binding protein-3 (IGFBP-3) have been implicated in the pathogenesis of fibrosis in various tissues by inducing mesenchymal cell proliferation and extracellular matrix deposition. We identified Syndecan-2 (SDC2) as a gene induced by TGFβ in an IGFBP-3-dependent manner. TGFβ induction of SDC2 mRNA and protein required IGFBP-3. IGFBP-3 independently induced production of SDC2 in primary fibroblasts. Using an ex-vivo model of human skin in organ culture expressing IGFBP-3, we demonstrate that IGFBP-3 induces SDC2 ex vivo in human tissue. We also identified Mitogen-activated protein kinase-interacting kinase (Mknk2) as a gene induced by IGFBP-3. IGFBP-3 triggered Mknk2 phosphorylation resulting in its activation. Mknk2 independently induced SDC2 in human skin. Since IGFBP-3 is over-expressed in fibrotic tissues, we examined SDC2 levels in skin and lung tissues of patients with systemic sclerosis (SSc) and lung tissues of patients with idiopathic pulmonary fibrosis (IPF). SDC2 levels were increased in fibrotic dermal and lung tissues of patients with SSc and in lung tissues of patients with IPF. This is the first report describing elevated levels of SDC2 in fibrosis. Increased SDC2 expression is due, at least in part, to the activity of two pro-fibrotic factors, TGFβ and IGFBP-3. PMID:22900087

Syndecan-2 is a novel target of insulin-like growth factor binding protein-3 and is over-expressed in fibrosis.

PubMed

Ruiz, Ximena D; Mlakar, Logan R; Yamaguchi, Yukie; Su, Yunyun; Larregina, Adriana T; Pilewski, Joseph M; Feghali-Bostwick, Carol A

2012-01-01

Extracellular matrix deposition and tissue scarring characterize the process of fibrosis. Transforming growth factor beta (TGFβ) and Insulin-like growth factor binding protein-3 (IGFBP-3) have been implicated in the pathogenesis of fibrosis in various tissues by inducing mesenchymal cell proliferation and extracellular matrix deposition. We identified Syndecan-2 (SDC2) as a gene induced by TGFβ in an IGFBP-3-dependent manner. TGFβ induction of SDC2 mRNA and protein required IGFBP-3. IGFBP-3 independently induced production of SDC2 in primary fibroblasts. Using an ex-vivo model of human skin in organ culture expressing IGFBP-3, we demonstrate that IGFBP-3 induces SDC2 ex vivo in human tissue. We also identified Mitogen-activated protein kinase-interacting kinase (Mknk2) as a gene induced by IGFBP-3. IGFBP-3 triggered Mknk2 phosphorylation resulting in its activation. Mknk2 independently induced SDC2 in human skin. Since IGFBP-3 is over-expressed in fibrotic tissues, we examined SDC2 levels in skin and lung tissues of patients with systemic sclerosis (SSc) and lung tissues of patients with idiopathic pulmonary fibrosis (IPF). SDC2 levels were increased in fibrotic dermal and lung tissues of patients with SSc and in lung tissues of patients with IPF. This is the first report describing elevated levels of SDC2 in fibrosis. Increased SDC2 expression is due, at least in part, to the activity of two pro-fibrotic factors, TGFβ and IGFBP-3.

Three-dimensional bioprinting of thick vascularized tissues

NASA Astrophysics Data System (ADS)

Kolesky, David B.; Homan, Kimberly A.; Skylar-Scott, Mark A.; Lewis, Jennifer A.

2016-03-01

The advancement of tissue and, ultimately, organ engineering requires the ability to pattern human tissues composed of cells, extracellular matrix, and vasculature with controlled microenvironments that can be sustained over prolonged time periods. To date, bioprinting methods have yielded thin tissues that only survive for short durations. To improve their physiological relevance, we report a method for bioprinting 3D cell-laden, vascularized tissues that exceed 1 cm in thickness and can be perfused on chip for long time periods (>6 wk). Specifically, we integrate parenchyma, stroma, and endothelium into a single thick tissue by coprinting multiple inks composed of human mesenchymal stem cells (hMSCs) and human neonatal dermal fibroblasts (hNDFs) within a customized extracellular matrix alongside embedded vasculature, which is subsequently lined with human umbilical vein endothelial cells (HUVECs). These thick vascularized tissues are actively perfused with growth factors to differentiate hMSCs toward an osteogenic lineage in situ. This longitudinal study of emergent biological phenomena in complex microenvironments represents a foundational step in human tissue generation.

Biology of soft tissue wound healing and regeneration--consensus report of Group 1 of the 10th European Workshop on Periodontology.

PubMed

Hämmerle, Christoph H F; Giannobile, William V

2014-04-01

The scope of this consensus was to review the biological processes of soft tissue wound healing in the oral cavity and to histologically evaluate soft tissue healing in clinical and pre-clinical models. To review the current knowledge regarding the biological processes of soft tissue wound healing at teeth, implants and on the edentulous ridge. Furthermore, to review soft tissue wound healing at these sites, when using barrier membranes, growth and differentiation factors and soft tissue substitutes. Searches of the literature with respect to recessions at teeth and soft tissue deficiencies at implants, augmentation of the area of keratinized tissue and soft tissue volume were conducted. The available evidence was collected, categorized and summarized. Oral mucosal and skin wound healing follow a similar pattern of the four phases of haemostasis, inflammation, proliferation and maturation/matrix remodelling. The soft connective tissue determines the characteristics of the overlaying oral epithelium. Within 7-14 days, epithelial healing of surgical wounds at teeth is completed. Soft tissue healing following surgery at implants requires 6-8 weeks for maturation. The resulting tissue resembles scar tissue. Well-designed pre-clinical studies providing histological data have been reported describing soft tissue wound healing, when using barrier membranes, growth and differentiation factors and soft tissue substitutes. Few controlled clinical studies with low numbers of patients are available for some of the treatments reviewed at teeth. Whereas, histological new attachment has been demonstrated in pre-clinical studies resulting from some of the treatments reviewed, human histological data commonly report a lack of new attachment but rather long junctional epithelial attachment and connective tissue adhesion. Regarding soft tissue healing at implants human data are very scarce. Oral soft tissue healing at teeth, implants and the edentulous ridge follows the same phases as skin wound healing. Histological studies in humans have not reported new attachment formation at teeth for the indications studied. Human histological data of soft tissue wound healing at implants are limited. The use of barriers membranes, growth and differentiation factors and soft tissue substitutes for the treatment of localized gingival/mucosal recessions, insufficient amount of keratinized tissue and insufficient soft tissue volume is at a developing stage. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Differences in prostate and adipose tissue basic fibroblast growth factor: analysis of preliminary results.

PubMed

Mydlo, J H; Kral, J G; Macchia, R J

1997-09-01

Basic fibroblast growth factor (bFGF or FGF-2) is mitogenic to human prostate epithelial and stromal cells, and it is reported to be elevated in the serum and urine of patients with various cancers, including prostate cancer. Obesity, with increased body fat, is a risk factor for prostate cancer through unknown mechanisms. Because adipose tissue is a source of FGF-2, we determined the quantity and quality of activity of FGF-2 in omental adipose tissue and compared it with normal and cancerous prostate tissues. Using heparin-Sepharose chromatography, we extracted proteins from human omental adipose tissue, adenocarcinoma of the prostate, and benign prostatic hypertrophic (BPH) tissues. Each of the mitogenic proteins eluted with NaCl concentrations between 1.4 M and 1.8 M, similar to control FGF-2. Using FGF-2 antisera (which inhibited the mitogenic activity of the proteins), we performed Western blot analysis to confirm their homology to FGF-2. We also assessed recovery, mitogenicity, and angiogenicity of each of the proteins using thymidine incorporation into human umbilical vein endothelial cells and the chorioallantoic membrane assay. There was greater recovery of FGF-2 from omental adipose tissue compared with cancerous or BPH homogenates (40 micrograms [2.0 micrograms/g] versus 25 micrograms [1.25 micrograms/g] and 20 micrograms [1.0 microgram/g], respectively). Moreover. FGF-2 from adipose tissue had greater mitogenic activity (96.2% versus 74.8% and 54%; P < 0.05) and a greater angiogenic activity (5.1 vessels versus 2.9 and 1.8 vessels; P < 0.05) on the chorioallantoic assay. We suggest that human omental adipose tissue FGF-2 may demonstrate greater mitogenic and angiogenic activity than either BPH or prostate cancer tissue FGF-2. It is not known whether FGF-2 from adipose tissue qualitatively or quantitatively may underlie the relationship between obesity and prostate cancer.

The use of bone marrow stromal cells (bone marrow-derived multipotent mesenchymal stromal cells) for alveolar bone tissue engineering: basic science to clinical translation.

PubMed

Kagami, Hideaki; Agata, Hideki; Inoue, Minoru; Asahina, Izumi; Tojo, Arinobu; Yamashita, Naohide; Imai, Kohzoh

2014-06-01

Bone tissue engineering is a promising field of regenerative medicine in which cultured cells, scaffolds, and osteogenic inductive signals are used to regenerate bone. Human bone marrow stromal cells (BMSCs) are the most commonly used cell source for bone tissue engineering. Although it is known that cell culture and induction protocols significantly affect the in vivo bone forming ability of BMSCs, the responsible factors of clinical outcome are poorly understood. The results from recent studies using human BMSCs have shown that factors such as passage number and length of osteogenic induction significantly affect ectopic bone formation, although such differences hardly affected the alkaline phosphatase activity or gene expression of osteogenic markers. Application of basic fibroblast growth factor helped to maintain the in vivo osteogenic ability of BMSCs. Importantly, responsiveness of those factors should be tested under clinical circumstances to improve the bone tissue engineering further. In this review, clinical application of bone tissue engineering was reviewed with putative underlying mechanisms.

Curcumin inhibits TGF-β1-induced connective tissue growth factor expression through the interruption of Smad2 signaling in human gingival fibroblasts.

PubMed

Chen, Jung-Tsu; Wang, Chen-Ying; Chen, Min-Huey

2018-01-13

Many fibrotic processes are associated with an increased level of transforming growth factor-β1 (TGF-β1). TGF-β1 can increase synthesis of matrix proteins and enhance secretion of protease inhibitors, resulting in matrix accumulation. Connective tissue growth factor (CTGF) is a downstream profibrotic effector of TGF-β1 and is associated with the fibrosis in several human organs. Curcumin has been applied to reduce matrix accumulation in fibrotic diseases. This study was aimed to evaluate whether curcumin could suppress TGF-β1-induced CTGF expression and its related signaling pathway involving in this inhibitory action in primary human gingival fibroblasts. The differences in CTGF expression among three types of gingival overgrowth and normal gingival tissues were assessed by immunohistochemistry. Gingival fibroblast viability in cultured media with different concentrations of curcumin was studied by MTT assay. The effect of curcumin on TGF-β1-induced CTGF expression in primary human gingival fibroblasts was examined by immunoblotting. Moreover, the proteins involved in TGF-β1 signaling pathways including TGF-β1 receptors and Smad2 were also analyzed by immunoblotting. CTGF was highly expressed in fibroblasts, epithelial cells and some of endothelial cells, smooth muscle cells, and inflammatory cells in phenytoin-induced gingival overgrowth tissues rather than in those of hereditary and inflammatory gingival overgrowth tissues. Moreover, CTGF expression in the epithelial and connective tissue layers was higher in phenytoin-induced gingival overgrowth tissues than in normal gingival tissues. Curcumin was nontoxic and could reduce TGF-β1-induced CTGF expression by attenuating the phosphorylation and nuclear translocation of Smad2. Curcumin can suppress TGF-β1-induced CTGF expression through the interruption of Smad2 signaling. Copyright © 2018. Published by Elsevier B.V.

Isolation and characterization of canine perivascular stem/stromal cells for bone tissue engineering.

PubMed

James, Aaron W; Zhang, Xinli; Crisan, Mihaela; Hardy, Winters R; Liang, Pei; Meyers, Carolyn A; Lobo, Sonja; Lagishetty, Venu; Childers, Martin K; Asatrian, Greg; Ding, Catherine; Yen, Yu-Hsin; Zou, Erin; Ting, Kang; Peault, Bruno; Soo, Chia

2017-01-01

For over 15 years, human subcutaneous adipose tissue has been recognized as a rich source of tissue resident mesenchymal stem/stromal cells (MSC). The isolation of perivascular progenitor cells from human adipose tissue by a cell sorting strategy was first published in 2008. Since this time, the interest in using pericytes and related perivascular stem/stromal cell (PSC) populations for tissue engineering has significantly increased. Here, we describe a set of experiments identifying, isolating and characterizing PSC from canine tissue (N = 12 canine adipose tissue samples). Results showed that the same antibodies used for human PSC identification and isolation are cross-reactive with canine tissue (CD45, CD146, CD34). Like their human correlate, canine PSC demonstrate characteristics of MSC including cell surface marker expression, colony forming unit-fibroblast (CFU-F) inclusion, and osteogenic differentiation potential. As well, canine PSC respond to osteoinductive signals in a similar fashion as do human PSC, such as the secreted differentiation factor NEL-Like Molecule-1 (NELL-1). Nevertheless, important differences exist between human and canine PSC, including differences in baseline osteogenic potential. In summary, canine PSC represent a multipotent mesenchymogenic cell source for future translational efforts in tissue engineering.

The effects of different preservation processes on the total protein and growth factor content in a new biological product developed from human amniotic membrane.

PubMed

Russo, Alessandra; Bonci, Paola; Bonci, Paolo

2012-06-01

The aim of this work is to quantify the total protein and growth factors content in a tissue-suspension obtained from processed human amniotic membrane (hAM). hAM was collected, frozen, freeze dried, powdered and sterilized by γ-irradiation. At each step of the process, samples were characterized for the total protein amounts by a Bradford protein assay and for the growth factor concentrations by ELISA test of the tissue suspensions. Frozen-hAM samples show higher release of total proteins and specific growth factors in the tissue suspension in comparison with freeze-dried hAM. We observed that even if the protein extraction is hindered once the tissue is dried, the powdering process allows a greater release in the tissue suspension of total proteins and growth factors after tissue re-solubilization in comparison with only the freeze-drying process (+91 ± 13% for EGF, +16 ± 4% for HGF, +11 ± 5% for FGF, +16 ± 9% for TGF-β1), and a greater release of EGF (85 ± 10%) in comparison with only the freezing process, because proteins become much readily solubilized in the solution. According with these results, we describe a protocol to obtain a new sterile biological product from hAM tissue, with well-known effects of thermal, mechanical and physical processes on the total protein and grow factors contents.

Approaches to improve angiogenesis in tissue-engineered skin.

PubMed

Sahota, Parbinder S; Burn, J Lance; Brown, Nicola J; MacNeil, Sheila

2004-01-01

A problem with tissue-engineered skin is clinical failure due to delays in vascularization. The aim of this study was to explore a number of simple strategies to improve angiogenesis/vascularization using a tissue-engineered model of skin to which small vessel human dermal microvascular endothelial cells were added. For the majority of these studies, a modified Guirguis chamber was used, which allowed the investigation of several variables within the same experiment using the same human dermis; cell type, angiogenic growth factors, the influence of keratinocytes and fibroblasts, mechanical penetration of the human dermis, the site of endothelial cell addition, and the influence of hypoxia were all examined. A qualitative scoring system was used to assess the impact of these factors on the penetration of endothelial cells throughout the dermis. Similar results were achieved using freshly isolated small vessel human dermal microvascular endothelial cells or an endothelial cell line and a minimum cell seeding density was identified. Cell penetration was not influenced by the addition of angiogenic growth factors (vascular endothelial growth factor and basic fibroblast growth factor); similarly, including epidermal keratinocytes or dermal fibroblasts did not encourage endothelial cell entry, and neither did mechanical introduction of holes throughout the dermis. Two factors were identified that significantly enhanced endothelial cell penetration into the dermis: hypoxia and the site of endothelial cell addition. Endothelial cells added from the papillary surface entered into the dermis much more effectively than when cells were added to the reticular surface of the dermis. We conclude that this model is valuable in improving our understanding of how to enhance vascularization of tissue-engineered grafts.

Investigation of scattering coefficients and anisotropy factors of human cancerous and normal prostate tissues using Mie theory

NASA Astrophysics Data System (ADS)

Pu, Yang; Chen, Jun; Wang, Wubao

2014-02-01

The scattering coefficient, μs, the anisotropy factor, g, the scattering phase function, p(θ), and the angular dependence of scattering intensity distributions of human cancerous and normal prostate tissues were systematically investigated as a function of wavelength, scattering angle and scattering particle size using Mie theory and experimental parameters. The Matlab-based codes using Mie theory for both spherical and cylindrical models were developed and applied for studying the light propagation and the key scattering properties of the prostate tissues. The optical and structural parameters of tissue such as the index of refraction of cytoplasm, size of nuclei, and the diameter of the nucleoli for cancerous and normal human prostate tissues obtained from the previous biological, biomedical and bio-optic studies were used for Mie theory simulation and calculation. The wavelength dependence of scattering coefficient and anisotropy factor were investigated in the wide spectral range from 300 nm to 1200 nm. The scattering particle size dependence of μs, g, and scattering angular distributions were studied for cancerous and normal prostate tissues. The results show that cancerous prostate tissue containing larger size scattering particles has more contribution to the forward scattering in comparison with the normal prostate tissue. In addition to the conventional simulation model that approximately considers the scattering particle as sphere, the cylinder model which is more suitable for fiber-like tissue frame components such as collagen and elastin was used for developing a computation code to study angular dependence of scattering in prostate tissues. To the best of our knowledge, this is the first study to deal with both spherical and cylindrical scattering particles in prostate tissues.

Growth factors in urologic tissues: detection, characterization, and clinical applications.

PubMed

Mydlo, J H; Macchia, R J

1992-12-01

During the last two decades, enormous strides have been made in understanding cellular and molecular biology. The direction of treatment of many neoplasms and other diseases are starting at the microscopic level. Growth factors are polypeptides that play a part in the development and maintenance of living tissues. We, as well as others, have investigated the role that growth factors play particularly in urologic tissues, both benign and malignant. We review several well-known growth factors and their function in prostate, kidney, and bladder tissues, as well as their functions in other regulating processes of the human body, and also the use of growth factors as tumor markers, and antibodies to growth factors as possible treatment of disease.

Human corpus luteum: presence of epidermal growth factor receptors and binding characteristics

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

Ayyagari, R.R.; Khan-Dawood, F.S.

Epidermal growth factor receptors are present in many reproductive tissues but have not been demonstrated in the human corpus luteum. To determine the presence of epidermal growth factor receptors and its binding characteristics, we carried out studies on the plasma cell membrane fraction of seven human corpora lutea (days 16 to 25) of the menstrual cycle. Specific epidermal growth factor receptors were present in human corpus luteum. Insulin, nerve growth factor, and human chorionic gonadotropin did not competitively displace epidermal growth factor binding. The optimal conditions for corpus luteum-epidermal growth factor receptor binding were found to be incubation for 2more » hours at 4 degrees C with 500 micrograms plasma membrane protein and 140 femtomol /sup 125/I-epidermal growth factor per incubate. The number (mean +/- SEM) of epidermal growth factor binding sites was 12.34 +/- 2.99 X 10(-19) mol/micrograms protein; the dissociation constant was 2.26 +/- 0.56 X 10(-9) mol/L; the association constant was 0.59 +/- 0.12 X 10(9) L/mol. In two regressing corpora lutea obtained on days 2 and 3 of the menstrual cycle, there was no detectable specific epidermal growth factor receptor binding activity. Similarly no epidermal growth factor receptor binding activity could be detected in ovarian stromal tissue. Our findings demonstrate that specific receptors for epidermal growth factor are present in the human corpus luteum. The physiologic significance of epidermal growth factor receptors in human corpus luteum is unknown, but epidermal growth factor may be involved in intragonadal regulation of luteal function.« less

Preliminary results comparing the recovery of basic fibroblast growth factor (FGF-2) in adipose tissue and benign and malignant renal tissue.

PubMed

Mydlo, J H; Kral, J G; Macchia, R J

1998-06-01

Basic fibroblast growth factor (bFGF or FGF-2) is mitogenic to numerous epithelial, mesodermal and endothelial cells, and thus may play a role in the neovascularity and progression of several tumors. Furthermore, FGF-2 is reported to be elevated in the serum and urine of patients with various cancers, including renal cancer. Obesity, with increased body fat, is a risk factor for renal cancer through unknown mechanisms. Since adipose tissue is a source of FGF-2, we determined the quantity and quality of activity of FGF-2 in omental adipose tissue and compared it to normal and cancerous renal tissue. Using heparin-Sepharose chromatography we extracted proteins from human omental adipose tissue, renal cell carcinoma (RCC) and benign renal tissue (BRT). Using FGF-2 antisera we performed western blot analysis to confirm their homology to FGF-2. We also assessed recovery, mitogenicity and angiogenicity of each of the proteins using thymidine incorporation into human umbilical vein endothelial cells (HUVEC) and the chorioallantoic membrane (CAM) assay. Each of the three purified mitogenic proteins eluted with NaCl concentrations between 1.4 M. and 1.8 M., similar to control FGF-2. There was greater recovery of FGF-2 from omental adipose tissue compared with renal cell carcinoma or benign renal tissue (42 microg. vs. 24 microg. and 18 microg., respectively; ANOVA p <0.05). Moreover, FGF-2 from adipose tissue had greater mitogenic activity (96.% versus 68% and 38%; p <0.05) and greater angiogenic activity (5.5 vessels versus 2.7 and 1.6 vessels; p <0.05) on the CAM assay. We suggest that human omental adipose tissue FGF-2 may demonstrate greater mitogenic and angiogenic activity than either benign or cancerous renal tissue FGF-2. It is not known if FGF-2 from adipose tissue may play a role in the relationship between obesity and renal cancer.

Dosimetry of {sup 210}Po in humans, caribou, and wolves in northern Canada

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

Thomas, P.A.

1994-06-01

Effective doses from {sup 210}Po intake with caribou meat were determined for human residents in Baker Lake and Snowdrift in the Northwest Territories of Canada and compared to doses calculated from reported {sup 210}Po tissue activities in Alaskan and British residents. Effective doses were calculated to separate body tissues, using ICRP 60 human weighting factors and the ICRP 30 metabolic model for {sup 210}Po. Baker Lake and Alaskan effective doses were similar at 0.4 mSv y{sup {minus}1} and slightly higher than Snowdrift doses (0.3 mSv y{sup {minus}1}). Alaskan tissue activities indicated higher effective doses to liver, bone surfaces and redmore » marrow and lower doses to spleen than the {sup 210}Po metabolic model (ICRP 1979a) predicts. Effective doses to Baker Lake and Snowdrift caribou and wolves, calculated from tissue activities, ranged from 7-20 mSv y{sup {minus}1} using human weighting factors for comparison to human doses only. Effective doses to northern Canadians and wildlife were, respectively, 7-11% and 1.8-5 times an estimated human background of 4 mSv y{sup {minus}} from all sources. 51 refs., 2 figs., 9 tabs.« less

Tissue engineering a human phalanx.

PubMed

Landis, W J; Chubinskaya, S; Tokui, T; Wada, Y; Isogai, N; Jacquet, R

2017-08-01

A principal purpose of tissue engineering is the augmentation, repair or replacement of diseased or injured human tissue. This study was undertaken to determine whether human biopsies as a cell source could be utilized for successful engineering of human phalanges consisting of both bone and cartilage. This paper reports the use of cadaveric human chondrocytes and periosteum as a model for the development of phalanx constructs. Two factors, osteogenic protein-1 [OP-1/bone morphogenetic protein-7 (BMP7)], alone or combined with insulin-like growth factor (IGF-1), were examined for their potential enhancement of chondrocytes and their secreted extracellular matrices. Design of the study included culture of chondrocytes and periosteum on biodegradable polyglycolic acid (PGA) and poly-l-lactic acid (PLLA)-poly-ε-caprolactone (PCL) scaffolds and subsequent implantation in athymic nu/nu (nude) mice for 5, 20, 40 and 60 weeks. Engineered constructs retrieved from mice were characterized with regard to genotype and phenotype as a function of developmental (implantation) time. Assessments included gross observation, X-ray radiography or microcomputed tomography, histology and gene expression. The resulting data showed that human cell-scaffold constructs could be successfully developed over 60 weeks, despite variability in donor age. Cartilage formation of the distal phalanx models enhanced with both OP-1 and IGF-1 yielded more cells and extracellular matrix (collagen and proteoglycans) than control chondrocytes without added factors. Summary data demonstrated that human distal phalanx models utilizing cadaveric chondrocytes and periosteum were successfully fabricated and OP-1 and OP-1/IGF-1 accelerated construct development and mineralization. The results suggest that similar engineering and transplantation of human autologous tissues in patients are clinically feasible. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Human adipose tissue-derived mesenchymal stem cells differentiate into insulin, somatostatin, and glucagon expressing cells

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

Timper, Katharina; Seboek, Dalma; Eberhardt, Michael

2006-03-24

Mesenchymal stem cells (MSC) from mouse bone marrow were shown to adopt a pancreatic endocrine phenotype in vitro and to reverse diabetes in an animal model. MSC from human bone marrow and adipose tissue represent very similar cell populations with comparable phenotypes. Adipose tissue is abundant and easily accessible and could thus also harbor cells with the potential to differentiate in insulin producing cells. We isolated human adipose tissue-derived MSC from four healthy donors. During the proliferation period, the cells expressed the stem cell markers nestin, ABCG2, SCF, Thy-1 as well as the pancreatic endocrine transcription factor Isl-1. The cellsmore » were induced to differentiate into a pancreatic endocrine phenotype by defined culture conditions within 3 days. Using quantitative PCR a down-regulation of ABCG2 and up-regulation of pancreatic developmental transcription factors Isl-1, Ipf-1, and Ngn3 were observed together with induction of the islet hormones insulin, glucagon, and somatostatin.« less

Mesenchymal stem cells derived from human exocrine pancreas express transcription factors implicated in beta-cell development.

PubMed

Baertschiger, Reto M; Bosco, Domenico; Morel, Philippe; Serre-Beinier, Veronique; Berney, Thierry; Buhler, Leo H; Gonelle-Gispert, Carmen

2008-07-01

Transplantation of in vitro generated islets or insulin-producing cells represents an attractive option to overcome organ shortage. The aim of this study was to isolate, expand, and characterize cells from human exocrine pancreas and analyze their potential to differentiate into beta cells. Fibroblast-like cells growing out of human exocrine tissue were characterized by flow cytometry and by their capacity to differentiate into mesenchymal cell lineages. During cell expansion and after differentiation toward beta cells, expression of transcription factors of endocrine pancreatic progenitors was analyzed by reverse transcription polymerase chain reaction. Cells emerged from 14/18 human pancreatic exocrine fractions and were expanded up to 40 population doublings. These cells displayed surface antigens similar to mesenchymal stem cells from bone marrow. A culture of these cells in adipogenic and chondrogenic differentiation media allowed differentiation into adipocyte- and chondrocyte-like cells. During expansion, cells expressed transcription factors implicated in islet development such as Isl1, Nkx2.2, Nkx6.1, nestin, Ngn3, Pdx1, and NeuroD. Activin A and hepatocyte growth factor induced an expression of insulin, glucagon, and glucokinase. Proliferating cells with characteristics of mesenchymal stem cells and endocrine progenitors were isolated from exocrine tissue. Under specific conditions, these cells expressed little insulin. Human pancreatic exocrine tissue might thus be a source of endocrine cell progenitors.

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

PubMed

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

2016-04-15

Using a perfusion decellularization protocol, we developed a decellularized skin/adipose tissue flap (DSAF) comprising extracellular matrix (ECM) and intact vasculature. Our DSAF had a dominant vascular pedicle, microcirculatory vascularity, and a sensory nerve network and retained three-dimensional (3D) nanofibrous structures well. DSAF, which was composed of collagen and laminin with well-preserved growth factors (e.g., vascular endothelial growth factor, basic fibroblast growth factor), was successfully repopulated with human adipose-derived stem cells (hASCs) and human umbilical vein endothelial cells (HUVECs), which integrated with DSAF and formed 3D aggregates and vessel-like structures in vitro. We used microsurgery techniques to re-anastomose the recellularized DSAF into nude rats. In vivo, the engineered flap construct underwent neovascularization and constructive remodeling, which was characterized by the predominant infiltration of M2 macrophages and significant adipose tissue formation at 3months postoperatively. Our results indicate that DSAF co-cultured with hASCs and HUVECs is a promising platform for vascularized soft tissue flap engineering. This platform is not limited by the flap size, as the entire construct can be immediately perfused by the recellularized vascular network following simple re-integration into the host using conventional microsurgical techniques. Significant soft tissue loss resulting from traumatic injury or tumor resection often requires surgical reconstruction using autologous soft tissue flaps. However, the limited availability of qualitative autologous flaps as well as the donor site morbidity significantly limits this approach. Engineered soft tissue flap grafts may offer a clinically relevant alternative to the autologous flap tissue. In this study, we engineered vascularized soft tissue free flap by using skin/adipose flap extracellular matrix scaffold (DSAF) in combination with multiple types of human cells. Following vascular reanastomosis in the recipient site, the engineered products successful regenerated large-scale fat tissue in vivo. This approach may provide a translatable platform for composite soft tissue free flap engineering for microsurgical reconstruction. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Phase transitions in oleic acid and in human breast tissue as studied by Raman spectroscopy and Raman imaging.

PubMed

Brozek-Pluska, Beata; Jablonska-Gajewicz, Joanna; Kordek, Radzislaw; Abramczyk, Halina

2011-05-12

We present the results of differential scanning calorimetry (DSC) and Raman studies in the temperature range of 293-77 K on vibrational properties of the oleic acid and the human breast tissue as a function of temperature. We have found that vibrational properties are very sensitive indicators to specify phases and phase transitions at the molecular level. We have found that water content confined in the cancerous tissue is markedly different from that in the noncancerous tissue. The OH stretching vibrations of water are useful as potential Raman biomarkers to distinguish between the cancerous and the noncancerous human breast tissues. Our results provide experimental evidence on the role of lipid profile and cell hydration as factors of particular significance in differentiation of the noncancerous and cancerous breast tissues.

General equation for the differential pathlength factor of the frontal human head depending on wavelength and age.

PubMed

Scholkmann, Felix; Wolf, Martin

2013-10-01

Continuous-wave near-infrared spectroscopy and near-infrared imaging enable the measurement of relative concentration changes in oxy- and deoxyhemoglobin and thus hemodynamics and oxygenation. The accuracy of determined changes depends mainly on the modeling of the light transport through the probed tissue. Due to the highly scattering nature of tissue, the light path is longer than the source-detector separation (d). This is incorporated in modeling by multiplying d by a differential pathlength factor (DPF) which depends on several factors such as wavelength, age of the subject, and type of tissue. In the present work, we derive a general DPF equation for the frontal human head, incorporating dependency on wavelength and age, based on published data. We validated the equation using different data sets of experimentally determined DPFs from six independent studies.

HIF isoforms in the skin differentially regulate systemic arterial pressure

PubMed Central

Cowburn, Andrew S.; Takeda, Norihiko; Boutin, Adam T.; Kim, Jung-Whan; Sterling, Jane C.; Nakasaki, Manando; Southwood, Mark; Goldrath, Ananda W.; Jamora, Colin; Nizet, Victor; Chilvers, Edwin R.; Johnson, Randall S.

2013-01-01

Vascular flow through tissues is regulated via a number of homeostatic mechanisms. Localized control of tissue blood flow, or autoregulation, is a key factor in regulating tissue perfusion and oxygenation. We show here that the net balance between two hypoxia-inducible factor (HIF) transcription factor isoforms, HIF-1α and HIF-2α, is an essential mechanism regulating both local and systemic blood flow in the skin of mice. We also show that balance of HIF isoforms in keratinocyte-specific mutant mice affects thermal adaptation, exercise capacity, and systemic arterial pressure. The two primary HIF isoforms achieve these effects in opposing ways that are associated with HIF isoform regulation of nitric oxide production. We also show that a correlation exists between altered levels of HIF isoforms in the skin and the degree of idiopathic hypertension in human subjects. Thus, the balance between HIF-1α and HIF-2α expression in keratinocytes is a control element of both tissue perfusion and systemic arterial pressure, with potential implications in human hypertension. PMID:24101470

Biological properties of dehydrated human amnion/chorion composite graft: implications for chronic wound healing

PubMed Central

Koob, Thomas J; Rennert, Robert; Zabek, Nicole; Massee, Michelle; Lim, Jeremy J; Temenoff, Johnna S; Li, William W; Gurtner, Geoffrey

2013-01-01

Human amnion/chorion tissue derived from the placenta is rich in cytokines and growth factors known to promote wound healing; however, preservation of the biological activities of therapeutic allografts during processing remains a challenge. In this study, PURION® (MiMedx, Marietta, GA) processed dehydrated human amnion/chorion tissue allografts (dHACM, EpiFix®, MiMedx) were evaluated for the presence of growth factors, interleukins (ILs) and tissue inhibitors of metalloproteinases (TIMPs). Enzyme-linked immunosorbent assays (ELISA) were performed on samples of dHACM and showed quantifiable levels of the following growth factors: platelet-derived growth factor-AA (PDGF-AA), PDGF-BB, transforming growth factor α (TGFα), TGFβ1, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), placental growth factor (PLGF) and granulocyte colony-stimulating factor (GCSF). The ELISA assays also confirmed the presence of IL-4, 6, 8 and 10, and TIMP 1, 2 and 4. Moreover, the relative elution of growth factors into saline from the allograft ranged from 4% to 62%, indicating that there are bound and unbound fractions of these compounds within the allograft. dHACM retained biological activities that cause human dermal fibroblast proliferation and migration of human mesenchymal stem cells (MSCs) in vitro. An in vivo mouse model showed that dHACM when tested in a skin flap model caused mesenchymal progenitor cell recruitment to the site of implantation. The results from both the in vitro and in vivo experiments clearly established that dHACM contains one or more soluble factors capable of stimulating MSC migration and recruitment. In summary, PURION® processed dHACM retains its biological activities related to wound healing, including the potential to positively affect four distinct and pivotal physiological processes intimately involved in wound healing: cell proliferation, inflammation, metalloproteinase activity and recruitment of progenitor cells. This suggests a paracrine mechanism of action for dHACM when used for wound healing applications. PMID:23902526

Manifestation of cryptic fibroblast tissue factor occurs at detergent concentrations which dissolve the plasma membrane.

PubMed

Carson, S D

1996-04-01

Cultured fibroblasts treated with increasing concentrations of detergents expressed only encrypted levels of tissue factor activity (measured by fX activation in the presence of fVIIa), characteristic of undamaged cells, until each detergent reached a critical concentration at which the cryptic tissue factor activity was manifested. Beyond the narrow ranges of concentrations over which the detergents stimulated tissue factor activity, the detergents were inhibitory. Studies with Triton X-100 and octyl glucoside revealed that manifestation of tissue factor activity coincided with breakdown of the plasma membrane. The magnitude of the increased tissue factor activity differed among detergents, with octyl glucoside giving the largest response. The tissue factor that was active after Triton X-100 treatment remained mostly associated with the insoluble cell residue, whereas the concentration of octyl glucoside which stimulated activity released tissue factor activity into the supernatant. Radiolabeled antibody against human tissue factor was used to show that a small percentage of the total accessible tissue factor remained in the insoluble fraction after treatment with either non-ionic detergent. Chromatographic analysis of lipids extracted from cells treated with detergents and dansyl chloride showed dansyl-reactivity of phosphatidylserine on intact cells, and solubilization of membrane lipids at sublytic concentrations of detergents. These findings reveal that there is a critical level of detergent-induced membrane damage at which tissue factor activity is maximally expressed, in essentially an all-or-none manner. The results are consistent with a major role for phospholipid asymmetry in regulation of tissue factor specific activity, but require either maintenance of asymmetry during sublytic detergent perturbation of the plasma membrane or additional control mechanisms.

21 CFR 1270.33 - Records, general requirements.

Code of Federal Regulations, 2010 CFR

2010-04-01

... assure freedom from risk factors for and clinical evidence of HIV infection, hepatitis B, and hepatitis C... performed and to relate the records to the particular tissue involved. (b) All human tissue shall be...

Evaluation of human recession defects treated with coronally advanced flaps and either purified recombinant human platelet-derived growth factor-BB with beta tricalcium phosphate or connective tissue: a histologic and microcomputed tomographic examination.

PubMed

McGuire, Michael K; Scheyer, Todd; Nevins, Myron; Schupbach, Peter

2009-02-01

The current study examined the histologic and microcomputed tomographic (micro CT) outcomes of the treatment of gingival recession defects with either a subepithelial connective tissue graft (CTG) or 0.3 mg/mL recombinant human platelet-derived growth factor (rhPDGF-BB) on a beta tricalcium phosphate (beta-TCP) matrix. Gingival recession defects were surgically created in six premolar teeth with no more than 3 mm of keratinized marginal tissue, an osseous crest 2 to 3 mm apical to the newly created gingival margin, and recession depth of at least 3 mm. The defects were left untouched for 2 months; then, four defects were grafted with rhPDGF-BB + beta-TCP + a wound healing dressing, and two defects received CTGs. A coronally advanced flap covered each grafted site. Nine months later, sections were obtained for examination. All four sites treated with rhPDGF-BB + beta-TCP showed connective tissue fibers (Sharpey fibers) perpendicularly inserting into newly formed cementum and alveolar bone. In the two sites treated with CTGs, a long junctional epithelium was seen coronal to the osseous crest and connective tissue fibers ran parallel to the adjacent root surfaces, with no evidence of insertion into cementum or bone. There was no evidence of regeneration of cementum, inserting connective tissue fibers, or supporting alveolar bone. Regeneration of the periodontium in gingival recession defects is possible through growth factor-mediated therapy.

AAV vector encoding human VEGF165-transduced pectineus muscular flaps increase the formation of new tissue through induction of angiogenesis in an in vivo chamber for tissue engineering: A technique to enhance tissue and vessels in microsurgically engineered tissue.

PubMed

Moimas, Silvia; Manasseri, Benedetto; Cuccia, Giuseppe; Stagno d'Alcontres, Francesco; Geuna, Stefano; Pattarini, Lucia; Zentilin, Lorena; Giacca, Mauro; Colonna, Michele R

2015-01-01

In regenerative medicine, new approaches are required for the creation of tissue substitutes, and the interplay between different research areas, such as tissue engineering, microsurgery and gene therapy, is mandatory. In this article, we report a modification of a published model of tissue engineering, based on an arterio-venous loop enveloped in a cross-linked collagen-glycosaminoglycan template, which acts as an isolated chamber for angiogenesis and new tissue formation. In order to foster tissue formation within the chamber, which entails on the development of new vessels, we wondered whether we might combine tissue engineering with a gene therapy approach. Based on the well-described tropism of adeno-associated viral vectors for post-mitotic tissues, a muscular flap was harvested from the pectineus muscle, inserted into the chamber and transduced by either AAV vector encoding human VEGF165 or AAV vector expressing the reporter gene β-galactosidase, as a control. Histological analysis of the specimens showed that muscle transduction by AAV vector encoding human VEGF165 resulted in enhanced tissue formation, with a significant increase in the number of arterioles within the chamber in comparison with the previously published model. Pectineus muscular flap, transduced by adeno-associated viral vectors, acted as a source of the proangiogenic factor vascular endothelial growth factor, thus inducing a consistent enhancement of vessel growth into the newly formed tissue within the chamber. In conclusion, our present findings combine three different research fields such as microsurgery, tissue engineering and gene therapy, suggesting and showing the feasibility of a mixed approach for regenerative medicine.

Evaluation of human epidermal growth factor receptor 2 (HER2) single nucleotide polymorphisms (SNPs) in normal and breast tumor tissues and their link with breast cancer prognostic factors.

PubMed

Furrer, Daniela; Lemieux, Julie; Côté, Marc-André; Provencher, Louise; Laflamme, Christian; Barabé, Frédéric; Jacob, Simon; Michaud, Annick; Diorio, Caroline

2016-12-01

Amplification of the human epidermal growth factor receptor 2 (HER2) gene is associated with worse prognosis and decreased overall survival in breast cancer patients. The HER2 gene contains several polymorphisms; two of the best-characterized HER2 polymorphisms are Ile655Val and Ala1170Pro. The aim of this study was to evaluate the association between these two HER2 polymorphisms in normal breast and breast cancer tissues and known breast cancer prognostic factors in a retrospective cohort study of 73 women with non-metastatic HER2-positive breast cancer. HER2 polymorphisms were assessed in breast cancer tissue and normal breast tissue using TaqMan assay. Ala1170Pro polymorphism in normal breast tissue was associated with age at diagnosis (p = 0.007), tumor size (p = 0.004) and lymphovascular invasion (p = 0.06). Similar significant associations in cancer tissues were observed. No association between the Ile655Val polymorphism and prognostic factors were observed. However, we found significant differences in the distribution of Ile655Val (p = 0.03) and Ala1170Pro (p = 0.01) genotypes between normal breast and breast tumor tissues. This study demonstrates that only the Ala1170Pro polymorphism is associated with prognostic factors in HER2-positive breast cancer patients. Moreover, our results suggest that both HER2 polymorphisms could play a significant role in carcinogenesis in non-metastatic HER2-positive breast cancer women. Copyright © 2016 Elsevier Ltd. All rights reserved.

Human breast tissue disposition and bioactivity of limonene in women with early-stage breast cancer.

PubMed

Miller, Jessica A; Lang, Julie E; Ley, Michele; Nagle, Ray; Hsu, Chiu-Hsieh; Thompson, Patricia A; Cordova, Catherine; Waer, Amy; Chow, H-H Sherry

2013-06-01

Limonene is a bioactive food component found in citrus peel oil that has shown chemopreventive and chemotherapeutic activities in preclinical studies. We conducted an open-label pilot clinical study to determine the human breast tissue disposition of limonene and its associated bioactivity. We recruited 43 women with newly diagnosed operable breast cancer electing to undergo surgical excision to take 2 grams of limonene daily for two to six weeks before surgery. Blood and breast tissue were collected to determine drug/metabolite concentrations and limonene-induced changes in systemic and tissue biomarkers of breast cancer risk or carcinogenesis. Limonene was found to preferentially concentrate in the breast tissue, reaching high tissue concentration (mean = 41.3 μg/g tissue), whereas the major active circulating metabolite, perillic acid, did not concentrate in the breast tissue. Limonene intervention resulted in a 22% reduction in cyclin D1 expression (P = 0.002) in tumor tissue but minimal changes in tissue Ki67 and cleaved caspase-3 expression. No significant changes in serum leptin, adiponectin, TGF-β1, insulin-like growth factor binding protein-3 (IGFBP-3), and interleukin-6 (IL-6) levels were observed following limonene intervention. There was a small but statistically significant postintervention increase in insulin-like growth factor I (IGF-I) levels. We conclude that limonene distributed extensively to human breast tissue and reduced breast tumor cyclin D1 expression that may lead to cell-cycle arrest and reduced cell proliferation. Furthermore, placebo-controlled clinical trials and translational research are warranted to establish limonene's role for breast cancer prevention or treatment.

Comparative Expression Analysis of Cytochrome P450 1A1, Cytochrome P450 1B1 and Nuclear Receptors in the Female Genital and Colorectal Tissues of Human and Pigtailed Macaque

PubMed Central

Hu, Minlu; Zhou, Tian; Pearlman, Andrew P; Paton, Dorothy L; Rohan, Lisa C

2017-01-01

Summary This manuscript summarizes our recent progress in examine the CYP1A1 and CYP1B1 as well as a number of nuclear receptors in the female genital and colorectal tissues of human and pigtailed macaque. Understanding the nuclear receptor mediated regulation of CYP1A1 and 1B1 expression in these tissues is necessary for identifying cancer risk factors and developing CYP1A1/1B1-targeted anti-cancer therapeutics. However, there is a lack of systematic and comparative analysis of the expression profile of CYP1A1, 1B1 and NRs in the female genital and colorectal tissues of human and clinically relevant animal models. The current study aims to fill this gap. We found CYP1A1, CYP1B1 and a number of nuclear receptors were expressed in the female genital and colorectal tissues of human and macaque. However, the mRNA level and protein localization of these CYP enzymes and NRs depended on the type of tissue examined. Cytochrome P450 (CYP) 1A1 and CYP1B1 activate hormonal and environmental procarcinogens, and are associated with carcinogenesis in female genital and colorectal tissues. Understanding the nuclear receptor (NR) mediated regulation of CYP expression in these tissues is necessary for identifying cancer risk factors and developing CYP1A1/1B1-targeted anti-cancer therapeutics. The study aims to analyze the expression profile of CYP1A1, 1B1 and NRs in the female genital and colorectal tissues of human and pigtailed macaques. We found that compared to the liver, human CYP1A1 mRNA level in the genital and colorectal tissues was significantly lower, while the CYP1B1 level was significantly higher. CYP1A1 protein was mainly localized in the plasma membrane of the uterine and endocervical epithelial cells. The CYP1B1 protein was concentrated in the nucleus of genital and colorectal tissues. Fourteen NRs in the genital tract and 12 NRs in colorectal tissue were expressed at levels similar to or higher than the liver. The expression and localization of CYP1A1, CYP1B1, and NRs in macaque tissues were usually comparable to those of human tissues. In addition, menopause did not significantly alter the ectocervical mRNA levels of CYP1A1, CYP1B1, or NRs. PMID:29276805

Comparative Expression Analysis of Cytochrome P450 1A1, Cytochrome P450 1B1 and Nuclear Receptors in the Female Genital and Colorectal Tissues of Human and Pigtailed Macaque.

PubMed

Hu, Minlu; Zhou, Tian; Pearlman, Andrew P; Paton, Dorothy L; Rohan, Lisa C

2016-01-01

This manuscript summarizes our recent progress in examine the CYP1A1 and CYP1B1 as well as a number of nuclear receptors in the female genital and colorectal tissues of human and pigtailed macaque. Understanding the nuclear receptor mediated regulation of CYP1A1 and 1B1 expression in these tissues is necessary for identifying cancer risk factors and developing CYP1A1/1B1-targeted anti-cancer therapeutics. However, there is a lack of systematic and comparative analysis of the expression profile of CYP1A1, 1B1 and NRs in the female genital and colorectal tissues of human and clinically relevant animal models. The current study aims to fill this gap. We found CYP1A1, CYP1B1 and a number of nuclear receptors were expressed in the female genital and colorectal tissues of human and macaque. However, the mRNA level and protein localization of these CYP enzymes and NRs depended on the type of tissue examined. Cytochrome P450 (CYP) 1A1 and CYP1B1 activate hormonal and environmental procarcinogens, and are associated with carcinogenesis in female genital and colorectal tissues. Understanding the nuclear receptor (NR) mediated regulation of CYP expression in these tissues is necessary for identifying cancer risk factors and developing CYP1A1/1B1-targeted anti-cancer therapeutics. The study aims to analyze the expression profile of CYP1A1, 1B1 and NRs in the female genital and colorectal tissues of human and pigtailed macaques. We found that compared to the liver, human CYP1A1 mRNA level in the genital and colorectal tissues was significantly lower, while the CYP1B1 level was significantly higher. CYP1A1 protein was mainly localized in the plasma membrane of the uterine and endocervical epithelial cells. The CYP1B1 protein was concentrated in the nucleus of genital and colorectal tissues. Fourteen NRs in the genital tract and 12 NRs in colorectal tissue were expressed at levels similar to or higher than the liver. The expression and localization of CYP1A1, CYP1B1, and NRs in macaque tissues were usually comparable to those of human tissues. In addition, menopause did not significantly alter the ectocervical mRNA levels of CYP1A1, CYP1B1, or NRs.

Comparative transcriptional analysis of three human ligaments with distinct biomechanical properties

PubMed Central

Lorda-Diez, Carlos I; Canga-Villegas, Ana; Cerezal, Luis; Plaza, Santiago; Hurlé, Juan M; García-Porrero, Juan A; Montero, Juan A

2013-01-01

One major aim of regenerative medicine targeting the musculoskeletal system is to provide complementary and/or alternative therapeutic approaches to current surgical therapies, often involving the removal and prosthetic substitution of damaged tissues such as ligaments. For these approaches to be successful, detailed information regarding the cellular and molecular composition of different musculoskeletal tissues is required. Ligaments have often been considered homogeneous tissues with common biomechanical properties. However, advances in tissue engineering research have highlighted the functional relevance of the organisational and compositional differences between ligament types, especially in those with higher risks of injury. The aim of this study was to provide information concerning the relative expression levels of a subset of key genes (including extracellular matrix components, transcription factors and growth factors) that confer functional identity to ligaments. We compared the transcriptomes of three representative human ligaments subjected to different biomechanical demands: the anterior cruciate ligament (ACL); the ligamentum teres of the hip (LT); and the iliofemoral ligament (IL). We revealed significant differences in the expression of type I collagen, elastin, fibromodulin, biglycan, transforming growth factor β1, transforming growth interacting factor 1, hypoxia-inducible factor 1-alpha and transforming growth factor β-induced gene between the IL and the other two ligaments. Thus, considerable molecular heterogeneity can exist between anatomically distinct ligaments with differing biomechanical demands. However, the LT and ACL were found to show remarkable molecular homology, suggesting common functional properties. This finding provides experimental support for the proposed role of the LT as a hip joint stabiliser in humans. PMID:24128114

Marker of cemento-periodontal ligament junction associated with periodontal regeneration.

PubMed

Hara, Ryohko; Wato, Masahiro; Tanaka, Akio

2005-06-01

The purpose of this study was to identify factors promoting formation of the cemento-periodontal ligament junction. Regeneration of the cemento-periodontal ligament junction is an important factor in recovery of the connective tissue attachment to the cementum and it is important to identify all specific substances that promote its formation. To clarify the substances involved in cemento-periodontal ligament junction formation, we produced a monoclonal antibody (mAb) to human cemento-periodontal ligament junction (designated as the anti-TAP mAb) and examined its immunostaining properties and reactive antigen. Hybridomas producing monoclonal antibody against human cemento-periodontal ligament junction antigens were established by fusing P3U1 mouse myeloma cells with spleen cells from BALB/c mice immunized with homogenized human cemento-periodontal ligament junction. The mAb, the anti-TAP mAb for cemento-periodontal ligament junction, was then isolated. The immunoglobulin class and light chain of the mAb were examined using an isotyping kit. Before immunostaining, antigen determination using an enzymatic method or heating was conducted. Human teeth, hard tissue-forming lesions, and animal tissues were immunostained by the anti-TAP mAb. The anti-TAP mAb was positive in human cemento-periodontal ligament junction and predentin but negative in all other human and animal tissues examined. In the cemento-osseous lesions, the anti-TAP mAb was positive in the peripheral area of the cementum and cementum-like hard tissues and not in the bone and bone-like tissues. The anti-TAP mAb showed IgM (kappa) and recognized phosphoprotein. The anti-TAP mAb is potentially useful for developing new agents promoting cementogenesis and periodontal regeneration.

Transforming growth factor beta-1 expression in macrophages of human chronic periapical diseases.

PubMed

Liang, Z-Z; Li, J; Huang, S-G

2017-03-30

The objective of this study was to observe the distribution of macrophages (MPs) expressing transforming growth factor beta-1 (TGF-β1) in tissue samples from patients with different human chronic periapical diseases. In this study, samples were collected from 75 volunteers, who were divided into three groups according to classified standards, namely, healthy control (N = 25), periapical granuloma (N = 25), and periapical cyst (N = 25). The samples were fixed in 10% buffered formalin for more than 48 h, dehydrated, embedded, and stained with hematoxylin and eosin for histopathology. Double immunofluorescence was conducted to analyze the expression of TGF-β-CD14 double-positive MPs in periapical tissues. The number of double-positive cells (cells/mm 2 ) were significantly higher in the chronic periapical disease tissues (P < 0.01) compared to that in the control tissue; in addition, the density of TGF-β1-CD14 double positive cells was significantly higher in the periapical cyst group than in the periapical granuloma group (P < 0.01). The number of TGF-β1 expressing macrophages varied with human chronic periapical diseases. The TGF-β1-CD14 double-positive cells might play an important role in the pathology of human chronic periapical diseases.

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 of SC-CO 2 method for delipidation and decellularization of adipose tissue whilst retaining its ECM and its subsequent utilization as a bioactive surface coating material for soft tissue engineering, angiogenesis and wound healing applications. Copyright © 2017 Elsevier B.V. All rights reserved.

Vascular tissue engineering by computer-aided laser micromachining.

PubMed

Doraiswamy, Anand; Narayan, Roger J

2010-04-28

Many conventional technologies for fabricating tissue engineering scaffolds are not suitable for fabricating scaffolds with patient-specific attributes. For example, many conventional technologies for fabricating tissue engineering scaffolds do not provide control over overall scaffold geometry or over cell position within the scaffold. In this study, the use of computer-aided laser micromachining to create scaffolds for vascular tissue networks was investigated. Computer-aided laser micromachining was used to construct patterned surfaces in agarose or in silicon, which were used for differential adherence and growth of cells into vascular tissue networks. Concentric three-ring structures were fabricated on agarose hydrogel substrates, in which the inner ring contained human aortic endothelial cells, the middle ring contained HA587 human elastin and the outer ring contained human aortic vascular smooth muscle cells. Basement membrane matrix containing vascular endothelial growth factor and heparin was to promote proliferation of human aortic endothelial cells within the vascular tissue networks. Computer-aided laser micromachining provides a unique approach to fabricate small-diameter blood vessels for bypass surgery as well as other artificial tissues with complex geometries.

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.

Endometrial stem cell differentiation into smooth muscle cell: a novel approach for bladder tissue engineering in women.

PubMed

Shoae-Hassani, Alireza; Sharif, Shiva; Seifalian, Alexander M; Mortazavi-Tabatabaei, Seyed Abdolreza; Rezaie, Sassan; Verdi, Javad

2013-10-01

To investigate manufacturing smooth muscle cells (SMCs) for regenerative bladder reconstruction from differentiation of endometrial stem cells (EnSCs), as the recent discovery of EnSCs from the lining of women's uteri, opens up the possibility of using these cells for tissue engineering applications, such as building up natural tissue to repair prolapsed pelvic floors as well as building urinary bladder wall. Human EnSCs that were positive for cluster of differentiation 146 (CD146), CD105 and CD90 were isolated and cultured in Dulbecco's modified Eagle/F12 medium supplemented with myogenic growth factors. The myogenic factors included: transforming growth factor β, platelet-derived growth factor, hepatocyte growth factor and vascular endothelial growth factor. Differentiated SMCs on bioabsorbable polyethylene-glycol and collagen hydrogels were checked for SMC markers by real-time reverse-transcriptase polymerase chain reaction (RT-PCR), western blot (WB) and immunocytochemistry (ICC) analyses. Histology confirmed the growth of SMCs in the hydrogel matrices. The myogenic growth factors decreased the proliferation rate of EnSCs, but they differentiated the human EnSCs into SMCs more efficiently on hydrogel matrices and expressed specific SMC markers including α-smooth muscle actin, desmin, vinculin and calponin in RT-PCR, WB and ICC experiments. The survival rate of cultures on the hydrogel-coated matrices was significantly higher than uncoated cultures. Human EnSCs were successfully differentiated into SMCs, using hydrogels as scaffold. EnSCs may be used for autologous bladder wall regeneration without any immunological complications in women. Currently work is in progress using bioabsorbable nanocomposite materials as EnSC scaffolds for developing urinary bladder wall tissue. © 2013 The Authors. BJU International © 2013 BJU International.

Tissue mechanics and fibrosis.

PubMed

Wells, Rebecca G

2013-07-01

Mechanical forces are essential to the development and progression of fibrosis, and are likely to be as important as soluble factors. These forces regulate the phenotype and proliferation of myofibroblasts and other cells in damaged tissues, the activation of growth factors, the structure and mechanics of the matrix, and, potentially, tissue patterning. Better understanding of the variety and magnitude of forces, the characteristics of those forces in biological tissues, and their impact on fibrosis in multiple tissues is needed and may lead to identification of important new therapeutic targets. This article is part of a Special Issue entitled: Fibrosis: Translation of basic research to human disease. Copyright © 2013 Elsevier B.V. All rights reserved.

Vasohibin 2 promotes human luminal breast cancer angiogenesis in a non-paracrine manner via transcriptional activation of fibroblast growth factor 2.

PubMed

Tu, Min; Lu, Cheng; Lv, Nan; Wei, Jishu; Lu, Zipeng; Xi, Chunhua; Chen, Jianmin; Guo, Feng; Jiang, Kuirong; Li, Qiang; Wu, Junli; Song, Guoxin; Wang, Shui; Gao, Wentao; Miao, Yi

2016-12-28

Vasohibin 2 (VASH2) is an angiogenic factor and cancer-related protein that acts via paracrine mechanisms. Here, we investigated the angiogenic function and mechanism of action of VASH2 in 200 human breast cancer tissues by performing immunohistochemical staining, western blot, indirect sandwich enzyme-linked immunosorbent assay (ELISA), and a semi-quantitative sandwich-based antibody array. Breast cancer cells stably overexpressing VASH2 or with knocked-down VASH2 were established and used for in vivo and in vitro models. In human luminal tissue, but not in HER2-positive or basal-like breast cancer tissues, VASH2 was positively correlated with CD31-positive microvascular density, induced angiogenesis in xenograft tumors, and promoted human umbilical vein endothelial cell tube formation in vitro. VASH2 expression was absent in the concentrated conditioned medium collected from knocked-down VASH2 and VASH2-overexpressing luminal breast cancer cells. Further, VASH2 regulated the expression of fibroblast growth factor 2 (FGF2) in human luminal breast cancer cells, and the pro-angiogenic effect induced by VASH2 overexpression was blocked by FGF2 neutralization in vitro. Additionally, dual luciferase reporter assay and Chromatin immunoprecipitation analysis results showed that FGF2 promoter was transcriptionally activated by VASH2 via histone modifications. In conclusion, VASH2 expression is positively correlated with FGF2 expression and promotes angiogenesis in human luminal breast cancer by transcriptional activation of fibroblast growth factor 2 through non-paracrine mechanisms. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

A role of low dose chemical mixtures in adipose tissue in carcinogenesis.

PubMed

Lee, Duk-Hee; Jacobs, David R; Park, Ho Yong; Carpenter, David O

2017-11-01

The Halifax project recently hypothesized a composite carcinogenic potential of the mixture of low dose chemicals which are commonly encountered environmentally, yet which are not classified as human carcinogens. A long neglected but important fact is that adipose tissue is an important exposure source for chemical mixtures. In fact, findings from human studies based on several persistent organic pollutants in general populations with only background exposure should be interpreted from the viewpoint of chemical mixtures because serum concentrations of these chemicals can be seen as surrogates for chemical mixtures in adipose tissue. Furthermore, in conditions such as obesity with dysfunctional adipocytes or weight loss in which lipolysis is increased, the amount of the chemical mixture released from adipose tissue to circulation is increased. Thus, both obesity and weight loss can enhance the chance of chemical mixtures reaching critical organs, however paradoxical this idea may be when fat mass is the only factor considered. The complicated, interrelated dynamics of adipocytes and chemical mixtures can explain puzzling findings related to body weight among cancer patients, including the obesity paradox. The contamination of fat in human diet with chemical mixtures, occurring for reasons similar to contamination of human adipose tissue, may be a missing factor which affects the association between dietary fat intake and cancer. The presence of chemical mixtures in adipose tissue should be considered in future cancer research, including clinical trials on weight management among cancer survivors. Copyright © 2017 Elsevier Ltd. All rights reserved.

The Human Cutaneous Chemokine System

PubMed Central

McCully, Michelle L.; Moser, Bernhard

2011-01-01

Irrespective of the immune status, the vast majority of all lymphocytes reside in peripheral tissues whereas those present in blood only amount to a small fraction of the total. It has been estimated that T cells in healthy human skin outnumber those present in blood by at least a factor of two. How lymphocytes within these two compartments relate to each other is not well understood. However, mounting evidence suggest that the study of T cell subsets present in peripheral blood does not reflect the function of their counterparts at peripheral sites. This is especially true under steady-state conditions whereby long-lived memory T cells in healthy tissues, notably those in epithelial tissues at body surfaces, are thought to fulfill a critical immune surveillance function by contributing to the first line of defense against a series of local threats, including microbes, tumors, and toxins, and by participating in wound healing. The relative scarcity of information regarding peripheral T cells and the factors regulating their localization is primarily due to inherent difficulties in obtaining healthy tissue for the extraction and study of immune cells on a routine basis. This is most certainly true for humans. Here, we review our current understanding of T cell homing to human skin and compare it when possible with gut-selective homing. We also discuss candidate chemokines that may account for the tissue selectivity in this process and present a model whereby CCR8, and its ligand CCL1, selectively regulate the homeostatic migration of memory lymphocytes to skin tissue. PMID:22566823

[Obesity and bone metabolism].

PubMed

Holecki, Michał; Zahorska-Markiewicz, Barbara; Wiecek, Andrzej; Nieszporek, Teresa; Zak-Gołab, Agnieszka

2008-01-01

Both bone and adipose tissue change their size, shape and distribution during the whole human being's life. Many factors, including genetic factors, hormones and activity of nervous system are responsible for these changes. It is generally accepted that obesity has a protective effect on bone tissue. On the other hand some authors present an opposite results--the lack of beneficial effect of obesity on development of osteoporosis fractures. The aim of this article was to present and discuss the relations between adipose tissue and bone metabolism.

TH-AB-209-12: Tissue Equivalent Phantom with Excised Human Tissue for Assessing Clinical Capabilities of Coherent Scatter Imaging Applications

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

Albanese, K; Morris, R; Spencer, J

Purpose: Previously we reported the development of anthropomorphic tissue-equivalent scatter phantoms of the human breast. Here we present the first results from the scatter imaging of the tissue equivalent breast phantoms for breast cancer diagnosis. Methods: A breast phantom was designed to assess the capability of coded aperture coherent x-ray scatter imaging to classify different types of breast tissue (adipose, fibroglandular, tumor). The phantom geometry was obtained from a prone breast geometry scanned on a dedicated breast CT system. The phantom was 3D printed using the segmented DICOM breast CT data. The 3D breast phantom was filled with lard (asmore » a surrogate for adipose tissue) and scanned in different geometries alongside excised human breast tissues (obtained from lumpectomy and mastectomy procedures). The raw data were reconstructed using a model-based reconstruction algorithm and yielded the location and form factor (i.e., momentum transfer (q) spectrum) of the materials that were imaged. The measured material form factors were then compared to the ground truth measurements acquired by x-ray diffraction (XRD) imaging. Results: Our scatter imaging system was able to define the location and composition of the various materials and tissues within the phantom. Cancerous breast tissue was detected and classified through automated spectral matching and an 86% correlation threshold. The total scan time for the sample was approximately 10 minutes and approaches workflow times for clinical use in intra-operative or other diagnostic tasks. Conclusion: This work demonstrates the first results from an anthropomorphic tissue equivalent scatter phantom to characterize a coherent scatter imaging system. The functionality of the system shows promise in applications such as intra-operative margin detection or virtual biopsy in the diagnosis of breast cancer. Future work includes using additional patient-derived tissues (e.g., human fat), and modeling additional organs (e.g., lung).« less

Physiologically Based Pharmacokinetic Model for Terbinafine in Rats and Humans

PubMed Central

Hosseini-Yeganeh, Mahboubeh; McLachlan, Andrew J.

2002-01-01

The aim of this study was to develop a physiologically based pharmacokinetic (PB-PK) model capable of describing and predicting terbinafine concentrations in plasma and tissues in rats and humans. A PB-PK model consisting of 12 tissue and 2 blood compartments was developed using concentration-time data for tissues from rats (n = 33) after intravenous bolus administration of terbinafine (6 mg/kg of body weight). It was assumed that all tissues except skin and testis tissues were well-stirred compartments with perfusion rate limitations. The uptake of terbinafine into skin and testis tissues was described by a PB-PK model which incorporates a membrane permeability rate limitation. The concentration-time data for terbinafine in human plasma and tissues were predicted by use of a scaled-up PB-PK model, which took oral absorption into consideration. The predictions obtained from the global PB-PK model for the concentration-time profile of terbinafine in human plasma and tissues were in close agreement with the observed concentration data for rats. The scaled-up PB-PK model provided an excellent prediction of published terbinafine concentration-time data obtained after the administration of single and multiple oral doses in humans. The estimated volume of distribution at steady state (Vss) obtained from the PB-PK model agreed with the reported value of 11 liters/kg. The apparent volume of distribution of terbinafine in skin and adipose tissues accounted for 41 and 52%, respectively, of the Vss for humans, indicating that uptake into and redistribution from these tissues dominate the pharmacokinetic profile of terbinafine. The PB-PK model developed in this study was capable of accurately predicting the plasma and tissue terbinafine concentrations in both rats and humans and provides insight into the physiological factors that determine terbinafine disposition. PMID:12069977

Expression of transforming growth factor alpha and epidermal growth factor receptor messenger RNA in neoplastic and nonneoplastic human kidney tissue.

PubMed

Mydlo, J H; Michaeli, J; Cordon-Cardo, C; Goldenberg, A S; Heston, W D; Fair, W R

1989-06-15

Using Northern blot analysis, we have demonstrated that mRNA for transforming growth factor alpha (TGF-alpha) was expressed in five malignant kidney tissue specimens but was not detected in their autologous nonneoplastic homologues. In addition, the expression of epidermal growth factor (EGF) receptor mRNA in these malignant tissues was 2- to 3-fold greater than in nontransformed tissues. In two cases examined using immunohistochemistry, we were able to correlate the increased expression of the mRNA with an increase in protein expression. Since TGF-alpha is known to bind to the EGF receptor, the finding of an increased expression of both TGF-alpha and EGF receptor mRNA in kidney tumor tissue suggests that interaction between TGF-alpha and the EGF receptor may play a role in promoting transformation and/or proliferation of kidney neoplasms, perhaps by an autocrine mechanism.

Granulocyte-Colony Stimulating Factor Receptor, Tissue Factor, and VEGF-R Bound VEGF in Human Breast Cancer In Loco.

PubMed

Wojtukiewicz, Marek Z; Sierko, Ewa; Skalij, Piotr; Kamińska, Magda; Zimnoch, Lech; Brekken, Ralf A; Thorpe, Philip E

2016-01-01

Doxorubicin and docetaxel-based chemotherapy regimens used in breast cancer patients are associated with high risk of febrile neutropenia (FN). Granulocyte colony-stimulating factors (G-CSF) are recommended for both treating and preventing chemotherapy-induced neutropenia. Increased thrombosis incidence in G-CSF treated patients was reported; however, the underlying mechanisms remain unclear. The principal activator of blood coagulation in cancer is tissue factor (TF). It additionally contributes to cancer progression and stimulates angiogenesis. The main proangiogenic factor is vascular endothelial growth factor (VEGF). The aim of the study was to evaluate granulocyte-colony stimulating factor receptor (G-CSFR), tissue factor (TF) expression and vascular endothelial growth factor receptor (VEGF-R) bound VEGF in human breast cancer in loco. G-CSFR, TF and VEGFR bound VEGF (VEGF: VEGFR) were assessed in 28 breast cancer tissue samples. Immunohistochemical (IHC) methodologies according to ABC technique and double staining IHC procedure were employed utilizing antibodies against G-CSFR, TF and VEGF associated with VEGFR (VEGF: VEGFR). Expression of G-CSFR was demonstrated in 20 breast cancer tissue specimens (71%). In 6 cases (21%) the expression was strong (IRS 9-12). Strong expression of TF was observed in all investigated cases (100%). Moreover, expression of VEGF: VEGFR was visualized in cancer cells (IRS 5-8). No presence of G-CSFR, TF or VEGF: VEGFR was detected on healthy breast cells. Double staining IHC studies revealed co-localization of G-CSFR and TF, G-CSFR and VEGF: VEGFR, as well as TF and VEGF: VEGFR on breast cancer cells and ECs. The results of the study indicate that GCSFR, TF and VEGF: VEGFR expression as well as their co-expression might influence breast cancer biology, and may increase thromboembolic adverse events incidence.

hCG-dependent regulation of angiogenic factors in human granulosa lutein cells.

PubMed

Phan, B; Rakenius, A; Pietrowski, D; Bettendorf, H; Keck, C; Herr, D

2006-07-01

As prerequisite for development and maintenance of many diseases angiogenesis is of particular interest in medicine. Pathologic angiogenesis takes place in chronic arthritis, collagen diseases, arteriosclerosis, retinopathy associated with diabetes, and particularly in cancers. However, angiogenesis as a physiological process regularly occurs in the ovary. After ovulation the corpus luteum is formed by rapid vascularization of initially avascular granulosa lutein cell tissue. This process is regulated by gonadotropic hormones. In order to gain further insights in the regulatory mechanisms of angiogenesis in the ovary, we investigated these mechanisms in cell culture of human granulosa lutein cells. In particular, we determined the expression and production of several angiogenic factors including tissue inhibitor of matrix metalloproteinases-1 (TIMP-1), Leptin, connective tissue growth factor (CTGF), meningioma-associated complimentary DNA (Mac25), basic fibroblast growth factor (bFGF), and Midkine. In addition, we showed that human chorionic gonadotropin (hCG) has distinct effects on their expression and production. hCG enhances the expression and production of TIMP-1, whereas it downregulates the expression of CTGF and Mac25. Furthermore it decreases the expression of Leptin. Our results provide evidence that hCG determines growth and development of the corpus luteum by mediating angiogenic pathways in human granulosa lutein cells. Hence we describe a further approach to understand the regulation of angiogenesis in the ovary.

Monitoring of treatment with vitamin K antagonists: recombinant thromboplastins are more sensitive to factor VII than tissue-extract thromboplastins.

PubMed

Biedermann, J S; van den Besselaar, A M H P; de Maat, M P M; Leebeek, F W G; Kruip, M J H A

2017-03-01

Essentials Differences in sensitivity to factor VII (FVII) have been suggested between thromboplastins. FVII-induced International Normalized Ratio (INR) changes differ between commercial reagents. Recombinant human thromboplastins are more sensitive to FVII than tissue-extract thromboplastins. Thromboplastin choice may affect FVII-mediated INR stability. Background Differences regarding sensitivity to factor VII have been suggested for recombinant human and tissue-extract thromboplastins used for International Normalized Ratio (INR) measurement, but the evidence is scarce. Differences in FVII sensitivity are clinically relevant, as they can affect INR stability during treatment with vitamin K antagonists (VKAs). Objectives To determine whether commercial thromboplastins react differently to changes in FVII. Methods We studied the effect of addition of FVII on the INR in plasma by using three tissue-extract (Neoplastin C1+, Hepato Quick, and Thromborel S) and three recombinant human (Recombiplastin 2G, Innovin, and CoaguChek XS) thromboplastins. Three different concentrations of purified human FVII (0.006, 0.012 and 0.062 μg mL -1 plasma), or buffer, were added to five certified pooled plasmas of patients using VKAs (INR of 1.5-3.5). Changes in FVII activity were measured with two bioassays (Neoplastin and Recombiplastin), and relative INR changes were compared between reagents. Results After addition of 0.062 μg mL -1 FVII, FVII activity in the pooled plasmas increased by approximately 20% (Neoplastin) or 32% (Recombiplastin) relative to the activity in pooled normal plasma. All thromboplastins showed dose-dependent INR decreases. The relative INR change in the pooled plasmas significantly differed between the six thromboplastins. No differences were observed among recombinant or tissue-extract thromboplastins. Pooled results indicated that the FVII-induced INR change was greater for recombinant than for tissue-extract thromboplastins. Conclusions Differences regarding FVII sensitivity exist between various thromboplastins used for VKA monitoring. Recombinant human thromboplastins are more sensitive to FVII than tissue-extract thromboplastins. Therefore, thromboplastin choice may affect FVII-mediated INR stability. © 2017 International Society on Thrombosis and Haemostasis.

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.

Factors for C-Kit Expression in Cardiac Outgrowth Cells and Human Heart Tissue.

PubMed

Matsushita, Satoshi; Minematsu, Kazuo; Yamamoto, Taira; Inaba, Hirotaka; Kuwaki, Kenji; Shimada, Akie; Yokoyama, Yasutaka; Amano, Atsushi

2017-12-12

We determined the factors associated with the expression of c-kit in the heart and the proliferation of c-kit-positive (c-kit pos ) cardiac stem cells among the outgrowth cells cultured from human cardiac explants.Samples of the right atrium (RA), left atrium (LA), and left ventricle obtained from patients during open-heart surgery were processed for cell culture of outgrowth cells and tissue analysis. The total number of growing cells and the population of c-kit pos cells were measured and compared with c-kit expression in native tissues and characteristics of the patients according to the region of the heart.We analyzed 452 samples from 334 patients. Atrial fibrillation (AF) in the patients reduced the number of outgrowth cells from the RA and LA, and aging was a co-factor for the LA. The c-kit pos population from the RA was associated with serum brain natriuretic peptide (BNP). C-kit expression in native tissue was also associated with BNP expression. However, we observed no relationship in expression between outgrowth cells and native tissue. In addition, the RA tissue provided the highest number of c-kit pos cells, and the left ventricle provided the lowest.C-kit was weakly expressed in response to damage. In addition, no correlation between outgrowth cells and native tissue was found for c-kit expression.

Human tissue models in cancer research: looking beyond the mouse.

PubMed

Jackson, Samuel J; Thomas, Gareth J

2017-08-01

Mouse models, including patient-derived xenograft mice, are widely used to address questions in cancer research. However, there are documented flaws in these models that can result in the misrepresentation of human tumour biology and limit the suitability of the model for translational research. A coordinated effort to promote the more widespread development and use of 'non-animal human tissue' models could provide a clinically relevant platform for many cancer studies, maximising the opportunities presented by human tissue resources such as biobanks. A number of key factors limit the wide adoption of non-animal human tissue models in cancer research, including deficiencies in the infrastructure and the technical tools required to collect, transport, store and maintain human tissue for lab use. Another obstacle is the long-standing cultural reliance on animal models, which can make researchers resistant to change, often because of concerns about historical data compatibility and losing ground in a competitive environment while new approaches are embedded in lab practice. There are a wide range of initiatives that aim to address these issues by facilitating data sharing and promoting collaborations between organisations and researchers who work with human tissue. The importance of coordinating biobanks and introducing quality standards is gaining momentum. There is an exciting opportunity to transform cancer drug discovery by optimising the use of human tissue and reducing the reliance on potentially less predictive animal models. © 2017. Published by The Company of Biologists Ltd.

Identification of the IGF-1 processing product human Ec/rodent Eb peptide in various tissues: Evidence for its differential regulation after exercise-induced muscle damage in humans.

PubMed

Vassilakos, George; Philippou, Anastassios; Koutsilieris, Michael

2017-02-01

Insulin-like growth factor-1 (IGF-1) is a pleiotropic factor expressed in various tissues and plays a critical role in skeletal muscle physiology. Alternative splicing of the IGF-1 gene gives rise to different precursor polypeptides (isoforms) which could undergo post-translational cleavage, generating the common mature IGF-1 peptide and different carboxyl terminal extension (E-) peptides, with the fate of the latter being, so far, unknown. The objective if this study was to identify the IGF-1Ec forms or processing product(s), other than mature IGF-1, generated in different human and rodent tissues and particularly in human skeletal muscle after exercise-induced damage. Protein lysates from a wide range of human and rodent tissues were immunoblotted with a rabbit anti-human Ec polyclonal antibody raised against the last 24 amino acids of the C-terminal of the Ec peptide. This antibody can recognize the Ec peptide, both as part of IGF-1Ec and alone, and also the corresponding rodent forms, due to the high homology that the human Ec shares with the rodent Eb. We were able to confirm, for the first time, that the human Ec peptide and its rodent homologous Eb peptide are produced simultaneously with their precursor protein (pro-IGF-1Ec/Eb) in vivo, in a wide range of tissues (e.g. muscle, liver, heart). Proprotein convertase furin digestion of human muscle and liver protein lysates confirmed that the higher molecular form, pro-IGF-1Ec, can be cleaved to produce the free Ec peptide. Furthermore, initial evidence is provided that Ec peptide is differentially regulated during the process of muscle regeneration after exercise-induced damage in humans. The findings of this study possibly imply that the post-translational modification of the IGF-1Ec pro-peptide may regulate the bioavailability and activity of the processing product(s). Copyright © 2016. Published by Elsevier Ltd.

Human acellular cartilage matrix powders as a biological scaffold for cartilage tissue engineering with synovium-derived mesenchymal stem cells.

PubMed

Chang, Chih-Hung; Chen, Chia-Chun; Liao, Cheng-Hao; Lin, Feng-Huei; Hsu, Yuan-Ming; Fang, Hsu-Wei

2014-07-01

In our previous study, we found that cartilage fragments from osteoarthritic knee promoted chondrogenesis of mesenchymal stem cells. In this study, we further transformed the cartilage tissues into acellular cartilage matrix (ACM) and explored the feasibility of using ACM as a biological scaffold. Nonworn parts of cartilage tissues were obtained during total knee arthroplasty (TKA) surgery and were successfully fabricated into ACM powders. The ACM powders and human synovium-derived mesenchymal stem cells (SMSCs) were mixed into collagen gel for in vitro culture. Histological results showed a synergistic effect of ACM powders and chondrogenic growth factors in the formation of engineered cartilage. The findings of real-time polymerase chain reaction (PCR) suggested that ACM powders had the potential of promoting type II collagen gene expression in the growth factors-absent environment. Moreover, with growth factors induction, the ACM powders could reduce the hypertrophy in chondrogenesis of SMSCs. In summary, ACM powders could serve as a functional scaffold that benefited the chondrogenesis of SMSCs for cartilage tissue engineering. © 2013 Wiley Periodicals, Inc.

Therapeutic modulation of growth factors and cytokines in regenerative medicine.

PubMed

Ioannidou, Effie

2006-01-01

Regeneration that takes place in the human body is limited throughout life. Therefore, when organs are irreparably damaged, they are usually replaced with an artificial device or donor organ. The term "regenerative medicine" covers the restoration or replacement of cells, tissues, and organs. Stem cells play a major role in regenerative medicine by providing the way to repopulate organs damaged by disease. Stem cells have the ability to self renew and to regenerate cells of diverse lineages within the tissue in which they reside. Stem cells could originate from embryos or adult tissues. Growth factors are proteins that may act locally or systemically to affect the growth of cells in several ways. Various cell activities, including division, are influenced by growth factors. Cytokines are a family of low-molecular-weight proteins that are produced by numerous cell types and are responsible for regulating the immune response, inflammation, tissue remodeling and cellular differentiation. Target cells of growth factors and cytokines are mesenchymal, epithelial and endothelial cells. These molecules frequently have overlapping activities and can act in an autocrine or paracrine fashion. A complex network of growth factors and cytokines guides cellular differentiation and regeneration in all organs and tissues. The aim of this paper is to review the role of growth factors and cytokines in different organs or systems and explore their therapeutic application in regenerative medicine. The role of stem cells combined with growth factors and cytokines in the regeneration of vascular and hematopoietic, neural, skeletal, pancreatic, periodontal, and mucosal tissue is reviewed. There is evidence that supports the use of growth factors and cytokines in the treatment of neurological diseases, diabetes, cardiovascular disease, periodontal disease, cancer and its complication, oral mucositis. After solving the ethical issues and establishing clear and reasonable regulations, regenerative medicine through stem cell application combined with specific growth factors and cytokines will have great potential in curing a variety of human diseases.

Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells.

PubMed

Sawada, Keigo; Takedachi, Masahide; Yamamoto, Satomi; Morimoto, Chiaki; Ozasa, Masao; Iwayama, Tomoaki; Lee, Chun Man; Okura, Hanayuki; Matsuyama, Akifumi; Kitamura, Masahiro; Murakami, Shinya

2015-08-14

Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response. ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. Copyright © 2015 Elsevier Inc. All rights reserved.

Characterization of In Vitro Engineered Human Adipose Tissues: Relevant Adipokine Secretion and Impact of TNF-α

PubMed Central

Aubin, Kim; Safoine, Meryem; Proulx, Maryse; Audet-Casgrain, Marie-Alice; Côté, Jean-François; Têtu, Félix-André; Roy, Alphonse; Fradette, Julie

2015-01-01

Representative modelling of human adipose tissue functions is central to metabolic research. Tridimensional models able to recreate human adipogenesis in a physiological tissue-like context in vitro are still scarce. We describe the engineering of white adipose tissues reconstructed from their cultured adipose-derived stromal precursor cells. We hypothesize that these reconstructed tissues can recapitulate key functions of AT under basal and pro-inflammatory conditions. These tissues, featuring human adipocytes surrounded by stroma, were stable and metabolically active in long-term cultures (at least 11 weeks). Secretion of major adipokines and growth factors by the reconstructed tissues was determined and compared to media conditioned by human native fat explants. Interestingly, the secretory profiles of the reconstructed adipose tissues indicated an abundant production of leptin, PAI-1 and angiopoietin-1 proteins, while higher HGF levels were detected for the human fat explants. We next demonstrated the responsiveness of the tissues to the pro-inflammatory stimulus TNF-α, as reflected by modulation of MCP-1, NGF and HGF secretion, while VEGF and leptin protein expression did not vary. TNF-α exposure induced changes in gene expression for adipocyte metabolism-associated mRNAs such as SLC2A4, FASN and LIPE, as well as for genes implicated in NF-κB activation. Finally, this model was customized to feature adipocytes representative of progressive stages of differentiation, thereby allowing investigations using newly differentiated or more mature adipocytes. In conclusion, we produced tridimensional tissues engineered in vitro that are able to recapitulate key characteristics of subcutaneous white adipose tissue. These tissues are produced from human cells and their neo-synthesized matrix elements without exogenous or synthetic biomaterials. Therefore, they represent unique tools to investigate the effects of pharmacologically active products on human stromal cells, extracellular matrix and differentiated adipocytes, in addition to compounds modulating adipogenesis from precursor cells. PMID:26367137

Biology and pathological implications of brown adipose tissue: promises and caveats for the control of obesity and its associated complications.

PubMed

Tapia, Pablo; Fernández-Galilea, Marta; Robledo, Fermín; Mardones, Pablo; Galgani, José E; Cortés, Víctor A

2018-05-01

The discovery of metabolically active brown adipose tissue (BAT) in adult humans has fuelled the research of diverse aspects of this previously neglected tissue. BAT is solely present in mammals and its clearest physiological role is non-shivering thermogenesis, owing to the capacity of brown adipocytes to dissipate metabolic energy as heat. Recently, a number of other possible functions have been proposed, including direct regulation of glucose and lipid homeostasis and the secretion of a number of factors with diverse regulatory actions. Herein, we review recent advances in general biological knowledge of BAT and discuss the possible implications of this tissue in human metabolic health. In particular, we confront the claimed thermogenic potential of BAT for human energy balance and body mass regulation, mostly based on animal studies, with the most recent quantifications of human BAT. © 2017 Cambridge Philosophical Society.

From the Cover: Cell-replacement therapy for diabetes: Generating functional insulin-producing tissue from adult human liver cells

NASA Astrophysics Data System (ADS)

Sapir, Tamar; Shternhall, Keren; Meivar-Levy, Irit; Blumenfeld, Tamar; Cohen, Hamutal; Skutelsky, Ehud; Eventov-Friedman, Smadar; Barshack, Iris; Goldberg, Iris; Pri-Chen, Sarah; Ben-Dor, Lya; Polak-Charcon, Sylvie; Karasik, Avraham; Shimon, Ilan; Mor, Eytan; Ferber, Sarah

2005-05-01

Shortage in tissue availability from cadaver donors and the need for life-long immunosuppression severely restrict the large-scale application of cell-replacement therapy for diabetic patients. This study suggests the potential use of adult human liver as alternate tissue for autologous beta-cell-replacement therapy. By using pancreatic and duodenal homeobox gene 1 (PDX-1) and soluble factors, we induced a comprehensive developmental shift of adult human liver cells into functional insulin-producing cells. PDX-1-treated human liver cells express insulin, store it in defined granules, and secrete the hormone in a glucose-regulated manner. When transplanted under the renal capsule of diabetic, immunodeficient mice, the cells ameliorated hyperglycemia for prolonged periods of time. Inducing developmental redirection of adult liver offers the potential of a cell-replacement therapy for diabetics by allowing the patient to be the donor of his own insulin-producing tissue. pancreas | transdifferentiation

Initial Biochemical Characterization of Cells Derived from Human Periodontium and Their In vitro Response to Platelet-Derived Growth Factor, Epidermal Growth Factor and Transforming Growth Factor-Beta

DTIC Science & Technology

1988-05-01

Periodontal disease is characterized by a loss of connective tissue...obtained for bone cells and fibroblasts. • " S,. O. ipr’ 0 II. LITERATURE REVIEW A . Periodontal Regeneration Periodontal disease is characterized by a ...fracture are felt to involve a similar sequence of cellular events. Since periodontal disease also involves the loss of soft tissue structures, such

Human Umbilical Cord Mesenchymal Stem Cells Reduce Fibrosis of Bleomycin-Induced Lung Injury

PubMed Central

Moodley, Yuben; Atienza, Daniel; Manuelpillai, Ursula; Samuel, Chrishan S.; Tchongue, Jorge; Ilancheran, Sivakami; Boyd, Richard; Trounson, Alan

2009-01-01

Acute respiratory distress syndrome is characterized by loss of lung tissue as a result of inflammation and fibrosis. Augmenting tissue repair by the use of mesenchymal stem cells may be an important advance in treating this condition. We evaluated the role of term human umbilical cord cells derived from Wharton’s jelly with a phenotype consistent with mesenchymal stem cells (uMSCs) in the treatment of a bleomycin-induced mouse model of lung injury. uMSCs were administered systemically, and lungs were harvested at 7, 14, and 28 days post-bleomycin. Injected uMSCs were located in the lung 2 weeks later only in areas of inflammation and fibrosis but not in healthy lung tissue. The administration of uMSCs reduced inflammation and inhibited the expression of transforming growth factor-β, interferon-γ, and the proinflammatory cytokines macrophage migratory inhibitory factor and tumor necrosis factor-α. Collagen concentration in the lung was significantly reduced by uMSC treatment, which may have been a consequence of the simultaneous reduction in Smad2 phosphorylation (transforming growth factor-β activity). uMSCs also increased matrix metalloproteinase-2 levels and reduced their endogenous inhibitors, tissue inhibitors of matrix metalloproteinases, favoring a pro-degradative milieu following collagen deposition. Notably, injected human lung fibroblasts did not influence either collagen or matrix metalloproteinase levels in the lung. The results of this study suggest that uMSCs have antifibrotic properties and may augment lung repair if used to treat acute respiratory distress syndrome. PMID:19497992

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

The Role of Nerve Growth Factor (NGF) and Its Precursor Forms in Oral Wound Healing

PubMed Central

Schenck, Karl; Schreurs, Olav; Hayashi, Katsuhiko; Helgeland, Kristen

2017-01-01

Nerve growth factor (NGF) and its different precursor forms are secreted into human saliva by salivary glands and are also produced by an array of cells in the tissues of the oral cavity. The major forms of NGF in human saliva are forms of pro-nerve growth factor (pro-NGF) and not mature NGF. The NGF receptors tropomyosin-related kinase A (TrkA) and p75 neurotrophin receptor (p75NTR) are widely expressed on cells in the soft tissues of the human oral cavity, including keratinocytes, endothelial cells, fibroblasts and leukocytes, and in ductal and acinar cells of all types of salivary glands. In vitro models show that NGF can contribute at most stages in the oral wound healing process: restitution, cell survival, apoptosis, cellular proliferation, inflammation, angiogenesis and tissue remodeling. NGF may therefore take part in the effective wound healing in the oral cavity that occurs with little scarring. As pro-NGF forms appear to be the major form of NGF in human saliva, efforts should be made to study its function, specifically in the process of wound healing. In addition, animal and clinical studies should be initiated to examine if topical application of pro-NGF or NGF can be a therapy for chronic oral ulcerations and wounds. PMID:28208669

The Role of Nerve Growth Factor (NGF) and Its Precursor Forms in Oral Wound Healing.

PubMed

Schenck, Karl; Schreurs, Olav; Hayashi, Katsuhiko; Helgeland, Kristen

2017-02-11

Nerve growth factor (NGF) and its different precursor forms are secreted into human saliva by salivary glands and are also produced by an array of cells in the tissues of the oral cavity. The major forms of NGF in human saliva are forms of pro-nerve growth factor (pro-NGF) and not mature NGF. The NGF receptors tropomyosin-related kinase A (TrkA) and p75 neurotrophin receptor (p75 NTR ) are widely expressed on cells in the soft tissues of the human oral cavity, including keratinocytes, endothelial cells, fibroblasts and leukocytes, and in ductal and acinar cells of all types of salivary glands. In vitro models show that NGF can contribute at most stages in the oral wound healing process: restitution, cell survival, apoptosis, cellular proliferation, inflammation, angiogenesis and tissue remodeling. NGF may therefore take part in the effective wound healing in the oral cavity that occurs with little scarring. As pro-NGF forms appear to be the major form of NGF in human saliva, efforts should be made to study its function, specifically in the process of wound healing. In addition, animal and clinical studies should be initiated to examine if topical application of pro-NGF or NGF can be a therapy for chronic oral ulcerations and wounds.

Inhibition of connective tissue growth factor (CTGF/CCN2) expression decreases the survival and myogenic differentiation of human rhabdomyosarcoma cells.

PubMed

Croci, Stefania; Landuzzi, Lorena; Astolfi, Annalisa; Nicoletti, Giordano; Rosolen, Angelo; Sartori, Francesca; Follo, Matilde Y; Oliver, Noelynn; De Giovanni, Carla; Nanni, Patrizia; Lollini, Pier-Luigi

2004-03-01

Connective tissue growth factor (CTGF/CCN2), a cysteine-rich protein of the CCN (Cyr61, CTGF, Nov) family of genes, emerged from a microarray screen of genes expressed by human rhabdomyosarcoma cells. Rhabdomyosarcoma is a soft tissue sarcoma of childhood deriving from skeletal muscle cells. In this study, we investigated the role of CTGF in rhabdomyosarcoma. Human rhabdomyosarcoma cells of the embryonal (RD/12, RD/18, CCA) and the alveolar histotype (RMZ-RC2, SJ-RH4, SJ-RH30), rhabdomyosarcoma tumor specimens, and normal skeletal muscle cells expressed CTGF. To determine the function of CTGF, we treated rhabdomyosarcoma cells with a CTGF antisense oligonucleotide or with a CTGF small interfering RNA (siRNA). Both treatments inhibited rhabdomyosarcoma cell growth, suggesting the existence of a new autocrine loop based on CTGF. CTGF antisense oligonucleotide-mediated growth inhibition was specifically due to a significant increase in apoptosis, whereas cell proliferation was unchanged. CTGF antisense oligonucleotide induced a strong decrease in the level of myogenic differentiation of rhabdomyosarcoma cells, whereas the addition of recombinant CTGF significantly increased the proportion of myosin-positive cells. CTGF emerges as a survival and differentiation factor and could be a new therapeutic target in human rhabdomyosarcoma.

Biological properties of dehydrated human amnion/chorion composite graft: implications for chronic wound healing.

PubMed

Koob, Thomas J; Rennert, Robert; Zabek, Nicole; Massee, Michelle; Lim, Jeremy J; Temenoff, Johnna S; Li, William W; Gurtner, Geoffrey

2013-10-01

Human amnion/chorion tissue derived from the placenta is rich in cytokines and growth factors known to promote wound healing; however, preservation of the biological activities of therapeutic allografts during processing remains a challenge. In this study, PURION® (MiMedx, Marietta, GA) processed dehydrated human amnion/chorion tissue allografts (dHACM, EpiFix®, MiMedx) were evaluated for the presence of growth factors, interleukins (ILs) and tissue inhibitors of metalloproteinases (TIMPs). Enzyme-linked immunosorbent assays (ELISA) were performed on samples of dHACM and showed quantifiable levels of the following growth factors: platelet-derived growth factor-AA (PDGF-AA), PDGF-BB, transforming growth factor α (TGFα), TGFβ1, basic fibroblast growth factor (bFGF), epidermal growth factor (EGF), placental growth factor (PLGF) and granulocyte colony-stimulating factor (GCSF). The ELISA assays also confirmed the presence of IL-4, 6, 8 and 10, and TIMP 1, 2 and 4. Moreover, the relative elution of growth factors into saline from the allograft ranged from 4% to 62%, indicating that there are bound and unbound fractions of these compounds within the allograft. dHACM retained biological activities that cause human dermal fibroblast proliferation and migration of human mesenchymal stem cells (MSCs) in vitro. An in vivo mouse model showed that dHACM when tested in a skin flap model caused mesenchymal progenitor cell recruitment to the site of implantation. The results from both the in vitro and in vivo experiments clearly established that dHACM contains one or more soluble factors capable of stimulating MSC migration and recruitment. In summary, PURION® processed dHACM retains its biological activities related to wound healing, including the potential to positively affect four distinct and pivotal physiological processes intimately involved in wound healing: cell proliferation, inflammation, metalloproteinase activity and recruitment of progenitor cells. This suggests a paracrine mechanism of action for dHACM when used for wound healing applications. ©2013 The Authors. International Wound Journal published by John Wiley & Sons Ltd and Medicalhelplines.com Inc.

Optical properties of human colon tissues in the 350 – 2500 nm spectral range

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

Bashkatov, A N; Genina, E A; Kochubey, V I

2014-08-31

We present the optical characteristics of the mucosa and submucosa of human colon tissue. The experiments are performed in vitro using a LAMBDA 950 spectrophotometer in the 350 – 2500 nm spectral range. The absorption and scattering coefficients and the scattering anisotropy factor are calculated based on the measured diffuse reflectance and total and collimated transmittance spectra using the inverse Monte Carlo method. (laser biophotonics)

Recent Advances in Cartilage Tissue Engineering: From the Choice of Cell Sources to the Use of Bioreactors

NASA Astrophysics Data System (ADS)

Martin, Ivan; Démarteau, Olivier; Braccini, Alessandra

Grafting engineered cartilage tissues represents a promising approach for the repair of joint injuries. Recent animal experiments have demonstrated that tissues engineered by culturing chondrocytes on 3D scaffolds in bioreactors provide functional templates for orderly repair of large osteochondral lesions. To date, however, a reproducible generation of uniform cartilage tissues of predefined size starting from adult human cells has not been achieved. In this paper we review some of the recent advances and challenges ahead in the identification of appropriate (i) cell sources, (ii) bioactive factors, (iii) 3D scaffolds and (iv) bioreactors for human cartilage tissue engineering. We also present an example of how integrated efforts in these different areas can help addressing fundamental questions and advancing the field of cartilage tissue engineering towards clinical use. The presented experiment demonstrates that human nasal chondrocytes are responsive to dynamic loading and thus could be further investigated as a cell source for implantation in a joint environment.

Toll-Like Receptor and Accessory Molecule mRNA Expression in Humans and Mice as Well as in Murine Autoimmunity, Transient Inflammation, and Progressive Fibrosis

PubMed Central

Ramaiah, Santhosh Kumar Vankayala; Günthner, Roman; Lech, Maciej; Anders, Hans-Joachim

2013-01-01

The cell type-, organ-, and species-specific expression of the Toll-like receptors (TLRs) are well described, but little is known about the respective expression profiles of their accessory molecules. We therefore determined the mRNA expression levels of LBP, MD2, CD36, CD14, granulin, HMGB1, LL37, GRP94, UNC93b1, TRIL, PRAT4A, AP3B1, AEP and the respective TLRs in human and mouse solid organs. Humans and mice displayed significant differences between their respective mRNA expression patterns of these factors. In addition, the expression profiles in transient tissue inflammation upon renal ischemia-reperfusion injury, in spleens and kidneys from mice with lupus-like systemic autoimmunity, and in progressive tissue fibrosis upon unilateral ureteral obstruction were studied. Several TLR co-factors were specifically regulated during the different phases of these disease entities, suggesting a functional involvement in the disease process. Thus, the organ- and species-specific expression patterns need to be considered in the design and interpretation of studies related to TLR-mediated innate immunity, which seems to be involved in the tissue injury phase, in the phase of tissue regeneration, and in progressive tissue remodelling. PMID:23803655

The Role of T-Cell Leukemia Translocation-Associated Gene Protein in Human Tumorigenesis and Osteoclastogenesis

PubMed Central

Kotake, Shigeru; Yago, Toru; Kawamoto, Manabu; Nanke, Yuki

2012-01-01

Synovial tissues of patients with rheumatoid arthritis (RA) include factors regulating bone resorption, such as receptor activator NF-κB ligand (RANKL), TNF-α, IL-6, IL-17, and IFN-γ. However, in addition to these cytokines, other factors expressed in synovial tissues may play a role in regulating bone resorption. In 2009, we demonstrated that novel peptides from T-cell leukemia translocation-associated gene (TCTA) protein expressed in synovial tissues from patients with RA inhibit human osteoclastogenesis, preventing cellular fusion via the interaction between TCTA protein and a putative counterpart molecule. Only a few studies on the role of TCTA protein have been reported. Genomic Southern blots demonstrated a reduced TCTA signal in three of four small cell lung cancer cell lines, suggesting the loss of one of the two copies of the gene. In the current paper, we reviewed the roles of TCTA protein in lung cancer cell lines and human osteoclastogenesis. PMID:22174563

Tissue-specific DNA methylation is conserved across human, mouse, and rat, and driven by primary sequence conservation.

PubMed

Zhou, Jia; Sears, Renee L; Xing, Xiaoyun; Zhang, Bo; Li, Daofeng; Rockweiler, Nicole B; Jang, Hyo Sik; Choudhary, Mayank N K; Lee, Hyung Joo; Lowdon, Rebecca F; Arand, Jason; Tabers, Brianne; Gu, C Charles; Cicero, Theodore J; Wang, Ting

2017-09-12

Uncovering mechanisms of epigenome evolution is an essential step towards understanding the evolution of different cellular phenotypes. While studies have confirmed DNA methylation as a conserved epigenetic mechanism in mammalian development, little is known about the conservation of tissue-specific genome-wide DNA methylation patterns. Using a comparative epigenomics approach, we identified and compared the tissue-specific DNA methylation patterns of rat against those of mouse and human across three shared tissue types. We confirmed that tissue-specific differentially methylated regions are strongly associated with tissue-specific regulatory elements. Comparisons between species revealed that at a minimum 11-37% of tissue-specific DNA methylation patterns are conserved, a phenomenon that we define as epigenetic conservation. Conserved DNA methylation is accompanied by conservation of other epigenetic marks including histone modifications. Although a significant amount of locus-specific methylation is epigenetically conserved, the majority of tissue-specific DNA methylation is not conserved across the species and tissue types that we investigated. Examination of the genetic underpinning of epigenetic conservation suggests that primary sequence conservation is a driving force behind epigenetic conservation. In contrast, evolutionary dynamics of tissue-specific DNA methylation are best explained by the maintenance or turnover of binding sites for important transcription factors. Our study extends the limited literature of comparative epigenomics and suggests a new paradigm for epigenetic conservation without genetic conservation through analysis of transcription factor binding sites.

Effect of Hemodilution on Coagulation and Recombinant Factor VIIa Efficacy in Human Blood In Vitro

DTIC Science & Technology

2011-11-01

thrombasthenia.12 In trauma, when a blood vessel is injured, tissue factor on subendothelial pericytes is exposed and binds to endogenous FVII ...a more complex effect on coagulation than simply dilution of any single coagulation factor like FVII or fibrinogen (Fig. 1). It is interesting to note...ORIGINAL ARTICLE Effect of Hemodilution on Coagulation and Recombinant Factor VIIa Efficacy in Human Blood In Vitro Daniel N. Darlington, PhD, Angel

Treatment of Ebola Virus Infection With a Recombinant Inhibitor of Factor Vlla/Tissue Factor: A Study in Rhesus Monkeys

DTIC Science & Technology

2003-12-13

ameliorate the effects of Ebola haemorrhagic fever . Here, we tested the notion that blockade of fVIIa/tissue factor is beneficial after infection with...Ebola virus. Methods We used a rhesus macaque model of Ebola haemorrhagic fever , which produces near 100% mortality. We administered recombinant...severe haemorrhagic fever in primates.1,2 Acute mortality caused by the Zaire species of Ebola virus has been about 80% in outbreaks in human beings1

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection

PubMed Central

Gordon, Claire L.; Thome, Joseph J.C.; Igarashi, Suzu

2017-01-01

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. PMID:28130404

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection.

PubMed

Gordon, Claire L; Miron, Michelle; Thome, Joseph J C; Matsuoka, Nobuhide; Weiner, Joshua; Rak, Michael A; Igarashi, Suzu; Granot, Tomer; Lerner, Harvey; Goodrum, Felicia; Farber, Donna L

2017-03-06

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. @Gordon et al.

Development of human nervous tissue upon differentiation of embryonic stem cells in three-dimensional culture.

PubMed

Preynat-Seauve, Olivier; Suter, David M; Tirefort, Diderik; Turchi, Laurent; Virolle, Thierry; Chneiweiss, Herve; Foti, Michelangelo; Lobrinus, Johannes-Alexander; Stoppini, Luc; Feki, Anis; Dubois-Dauphin, Michel; Krause, Karl Heinz

2009-03-01

Researches on neural differentiation using embryonic stem cells (ESC) require analysis of neurogenesis in conditions mimicking physiological cellular interactions as closely as possible. In this study, we report an air-liquid interface-based culture of human ESC. This culture system allows three-dimensional cell expansion and neural differentiation in the absence of added growth factors. Over a 3-month period, a macroscopically visible, compact tissue developed. Histological coloration revealed a dense neural-like neural tissue including immature tubular structures. Electron microscopy, immunochemistry, and electrophysiological recordings demonstrated a dense network of neurons, astrocytes, and oligodendrocytes able to propagate signals. Within this tissue, tubular structures were niches of cells resembling germinal layers of human fetal brain. Indeed, the tissue contained abundant proliferating cells expressing markers of neural progenitors. Finally, the capacity to generate neural tissues on air-liquid interface differed for different ESC lines, confirming variations of their neurogenic potential. In conclusion, this study demonstrates in vitro engineering of a human neural-like tissue with an organization that bears resemblance to early developing brain. As opposed to previously described methods, this differentiation (a) allows three-dimensional organization, (b) yields dense interconnected neural tissue with structurally and functionally distinct areas, and (c) is spontaneously guided by endogenous developmental cues.

Expression and localization of endocrine gland-derived vascular endothelial growth factor (EG-VEGF) in human pancreas and pancreatic adenocarcinoma.

PubMed

Morales, Angélica; Vilchis, Felipe; Chávez, Bertha; Chan, Carlos; Robles-Díaz, Guillermo; Díaz-Sánchez, Vicente

2007-10-01

Endocrine gland-derived vascular endothelial growth factor (EG-VEGF) was recently identified as the first tissue-specific angiogenic molecule. EG-VEGF (the gene product of PROK-1) appears to be expressed exclusively in steroid-producing organs such as the ovary, testis, adrenals and placenta. Since the human pancreatic cells retain steroidogenic activity, in the present study we ascertained whether this angiogenic factor is expressed in normal pancreas and pancreatic adenocarcinoma. Tissue samples from normal males (n=5), normal females (n=5) and from surgically resected adenocarcinomas (n=2) were processed for RT-PCR and immunohistochemical studies. Results from semi-quantitative analysis by RT-PCR suggest a distinct expression level for EG-VEGF in the different tissue samples. The relative amount of EG-VEGF mRNA in pancreas was more abundant in female adenocarcinoma (0.89) followed by male adenocarcinoma (0.71), than normal female (0.64) and normal male (0.38). The expression of mRNA for EG-VEGF in normal tissue was significantly higher in females than in males. All samples examined showed specific immunostaining for EG-VEGF. In male preparations, the positive labeling was localized predominantly within the pancreatic islets while in female preparations the main staining was detected towards the exocrine portion. Specific immunolabeling was also observed in endothelial cells of pancreatic blood vessels. Our data provide evidence that the human pancreas expresses the EG-VEGF, a highly specific mitogen which regulates proliferation and differentiation of the vascular endothelium. The significance of this finding could be interpreted as either, EG-VEGF is not exclusive of endocrine organs, or the pancreas should be considered as a functional steroidogenic tissue. The extent of the expression of EG-VEGF appears to have a dimorphic pattern in normal and tumoral pancreatic tissue.

QUANTITATIVE TOXICOPROTEOMIC ANALYSIS OF CARCINOGEN-TREATED ANIMAL TISSUES AND HUMAN CELLS FOR HUMAN HEALTH RISK ASSESSMENT

EPA Science Inventory

Humans are exposed to a variety of environmental toxicants, and this together with a large number of interacting factors can contribute to an individual's risk for health. To understand the toxic mechanisms and/or modes of action for human health risk assessment, molecular charac...

Coordinate tropic hormone regulation of mRNAs for insulin-like growth factor II and the cholesterol side-chain-cleavage enzyme, P450scc [corrected], in human steroidogenic tissues.

PubMed Central

Voutilainen, R; Miller, W L

1987-01-01

Insulin-like growth factors (IGFs) are single-chain polypeptides important for cell proliferation and growth. IGFs are produced in several tissues, suggesting that they function in a paracrine or autocrine fashion as well as functioning as endocrine hormones. We studied the hormonal regulation of IGF-I and IGF-II mRNA in human steroidogenic tissues. In cultured human ovarian granulosa cells, follicle-stimulating hormone, human chorionic gonadotropin, and dibutyryl cAMP increased IGF-II mRNA, but corticotropin [adrenocorticotropic hormone (ACTH)], chorionic somatomammotropin, growth hormone, prolactin, dexamethasone, estradiol, and progesterone had no effect. In cultured human fetal adrenal cells, ACTH and dibutyryl cAMP increased IGF-II mRNA accumulation, but human chorionic gonadotropin and angiotensin II did not. The same five size species of IGF-II mRNA were detected in transfer blots of RNA from granulosa cells and fetal adrenal cells, and all of these increased after hormonal stimuli. Dibutyryl cAMP also increased IGF-II mRNA accumulation in cultured human placental cells. Accumulation of mRNA for the cholesterol side-chain-cleavage monooxygenase [P450scc [corrected]; cholesterol, reduced-adrenal-ferredoxin:oxygen oxidoreductase (side-chain-cleaving), EC 1.14.15.6] was regulated in parallel with IGF-II mRNA in all these steroidogenic tissues. IGF-I mRNA was not detected in transfer blots of these RNAs, and the minimal amounts detected in dot blots showed no detectable change after any of the hormonal stimuli studied. The data indicate that the IGF-II gene is expressed in human steroidogenic tissues and is regulated by cAMP. These data suggest that IGF-II may act in an autocrine or paracrine fashion to stimulate the adrenal and gonadal growth stimulated by ACTH and gonadotropins, respectively. Images PMID:3031644

Host‐related factors explaining interindividual variability of carotenoid bioavailability and tissue concentrations in humans

PubMed Central

Desmarchelier, Charles; Dragsted, Lars O.; Nielsen, Charlotte S.; Stahl, Wilhelm; Rühl, Ralph; Keijer, Jaap; Borel, Patrick

2017-01-01

Carotenoid dietary intake and their endogenous levels have been associated with a decreased risk of several chronic diseases. There are indications that carotenoid bioavailability depends, in addition to the food matrix, on host factors. These include diseases (e.g. colitis), life‐style habits (e.g. smoking), gender and age, as well as genetic variations including single nucleotide polymorphisms that govern carotenoid metabolism. These are expected to explain interindividual differences that contribute to carotenoid uptake, distribution, metabolism and excretion, and therefore possibly also their association with disease risk. For instance, digestion enzymes fostering micellization (PNLIP, CES), expression of uptake/efflux transporters (SR‐BI, CD36, NPC1L1), cleavage enzymes (BCO1/2), intracellular transporters (FABP2), secretion into chylomicrons (APOB, MTTP), carotenoid metabolism in the blood and liver (LPL, APO C/E, LDLR), and distribution to target tissues such as adipose tissue or macula (GSTP1, StARD3) depend on the activity of these proteins. In addition, human microbiota, e.g. via altering bile‐acid concentrations, may play a role in carotenoid bioavailability. In order to comprehend individual, variable responses to these compounds, an improved knowledge on intra‐/interindividual factors determining carotenoid bioavailability, including tissue distribution, is required. Here, we highlight the current knowledge on factors that may explain such intra‐/interindividual differences. PMID:28101967

Identification of Extracellular Matrix Components and Biological Factors in Micronized Dehydrated Human Amnion/Chorion Membrane

PubMed Central

Lei, Jennifer; Priddy, Lauren B.; Lim, Jeremy J.; Massee, Michelle; Koob, Thomas J.

2017-01-01

Objective: The use of bioactive extracellular matrix (ECM) grafts such as amniotic membranes is an attractive treatment option for enhancing wound repair. In this study, the concentrations, activity, and distribution of matrix components, growth factors, proteases, and inhibitors were evaluated in PURION® Processed, micronized, dehydrated human amnion/chorion membrane (dHACM; MiMedx Group, Inc.). Approach: ECM components in dHACM tissue were assessed by using immunohistochemical staining, and growth factors, cytokines, proteases, and inhibitors were quantified by using single and multiplex ELISAs. The activities of proteases that were native to the tissue were determined via gelatin zymography and EnzChek® activity assay. Results: dHACM tissue contained the ECM components collagens I and IV, hyaluronic acid, heparin sulfate proteoglycans, fibronectin, and laminin. In addition, numerous growth factors, cytokines, chemokines, proteases, and protease inhibitors that are known to play a role in the wound-healing process were quantified in dHACM. Though matrix metalloproteinases (MMPs) were present in dHACM tissues, inhibitors of MMPs overwhelmingly outnumbered the MMP enzymes by an overall molar ratio of 28:1. Protease activity assays revealed that the MMPs in the tissue existed primarily either in their latent form or complexed with inhibitors. Innovation: This is the first study to characterize components that function in wound healing, including inhibitor and protease content and activity, in micronized dHACM. Conclusion: A variety of matrix components and growth factors, as well as proteases and their inhibitors, were identified in micronized dHACM, providing a better understanding of how micronized dHACM tissue can be used to effectively promote wound repair. PMID:28224047

Identification of Extracellular Matrix Components and Biological Factors in Micronized Dehydrated Human Amnion/Chorion Membrane.

PubMed

Lei, Jennifer; Priddy, Lauren B; Lim, Jeremy J; Massee, Michelle; Koob, Thomas J

2017-02-01

Objective: The use of bioactive extracellular matrix (ECM) grafts such as amniotic membranes is an attractive treatment option for enhancing wound repair. In this study, the concentrations, activity, and distribution of matrix components, growth factors, proteases, and inhibitors were evaluated in PURION ® Processed, micronized, dehydrated human amnion/chorion membrane (dHACM; MiMedx Group, Inc.). Approach: ECM components in dHACM tissue were assessed by using immunohistochemical staining, and growth factors, cytokines, proteases, and inhibitors were quantified by using single and multiplex ELISAs. The activities of proteases that were native to the tissue were determined via gelatin zymography and EnzChek ® activity assay. Results: dHACM tissue contained the ECM components collagens I and IV, hyaluronic acid, heparin sulfate proteoglycans, fibronectin, and laminin. In addition, numerous growth factors, cytokines, chemokines, proteases, and protease inhibitors that are known to play a role in the wound-healing process were quantified in dHACM. Though matrix metalloproteinases (MMPs) were present in dHACM tissues, inhibitors of MMPs overwhelmingly outnumbered the MMP enzymes by an overall molar ratio of 28:1. Protease activity assays revealed that the MMPs in the tissue existed primarily either in their latent form or complexed with inhibitors. Innovation: This is the first study to characterize components that function in wound healing, including inhibitor and protease content and activity, in micronized dHACM. Conclusion: A variety of matrix components and growth factors, as well as proteases and their inhibitors, were identified in micronized dHACM, providing a better understanding of how micronized dHACM tissue can be used to effectively promote wound repair.

Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells

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

Sawada, Keigo; Takedachi, Masahide, E-mail: takedati@dent.osaka-u.ac.jp; Yamamoto, Satomi

Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response.more » ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. - Highlights: • ADMPC-derived humoral factors stimulate cytodifferentiation of HPDLs. • ADMPCs secret growth factors including IGFBP6, VEGF and HGF. • IGFBP6 is involved in the promotion effect of ADMPC-CM on HPDL cytodifferentiation.« less

Expression of Folliculogenesis-Related Genes in Vitrified Human Ovarian Tissue after Two Weeks In Vitro Culture.

PubMed

Shams Mofarahe, Zahra; Salehnia, Mojdeh; Ghaffari Novin, Marefat; Ghorbanmehr, Nassim; Fesharaki, Mohammad Gholami

2017-01-01

This study was designed to evaluate the effects of vitrification and in vitro culture of human ovarian tissue on the expression of oocytic and follicular cell-related genes. In this experimental study, ovarian tissue samples were obtained from eight transsexual women. Samples were cut into small fragments and were then assigned to vitrified and non-vitrified groups. In each group, some tissue fragments were divided into un-cultured and cultured (in α-MEM medium for 2 weeks) subgroups. The normality of follicles was assessed by morphological observation under a light microscope using hematoxylin and eosin (H&E) staining. Expression levels of factor in the germ line alpha ( FIGLA ), KIT ligand ( KL ), growth differentiation factor 9 ( GDF-9 ) and follicle stimulating hormone receptor ( FSHR ) genes were quantified in both groups by real-time reverse transcriptase polymerase chain reaction (RT-PCR) at the beginning and the end of culture. The percentage of normal follicles was similar between non-cultured vitrified and non-vitrified groups (P>0.05), however, cultured tissues had significantly fewer normal follicles than non-cultured tissues in both vitrified and non-vitrified groups (P<0.05). In both cultured groups the rate of primary and secondary follicles was significantly higher than non-cultured tissues (P<0.05). The expression of all examined genes was not significantly altered in both non-cultured groups. Whiles, in comparison with cultured tissues non-cultured tissues, the expression of FIGLA gene was significantly decreased, KL gene was not changed, GDF-9 and FSHR genes was significantly increased (P<0.05). Human ovarian vitrification following in vitro culture has no impairing effects on follicle normality and development and expression of related-genes. However, in vitro culture condition has deleterious effects on normality of follicles.

RNA Extraction from Animal and Human's Cancerous Tissues: Does Tissue Matter?

PubMed

Samadani, Ali Akbar; Nikbakhsh, Novin; Fattahi, Sadegh; Pourbagher, Roghayeh; Aghajanpour Mir, Seyyed Mohsen; Mousavi Kani, Narges; Abedian, Zeinab; Akhavan-Niaki, Haleh

2015-01-01

The reliability of gene expression profiling, based technologies and methods to find transcriptional differences representative of the original samples is influenced by the quality of the extracted RNA. Hence, RNA extraction is the first step to investigate the gene expression and its function. Consequently, the quality of extracted RNA is really significant. Correspondingly, this research was accomplished to optimize the RNA extraction methods and compare the amounts of tissue or quality of tissue. Relatively, the cancerous tissue of human stomach in fresh and frozen conditions and also the mouse fresh tissue were studied. Some factors like the amount of samples, efficacy differences of diverse extraction buffers (TriPure, Trizol) and also the efficacy of b-mercaptoethanol were compared and investigated. The results indicated that the less amount (1-2 mg) compared to other amounts (2-5 mg, 5-15 mg) yielded the best quality and the RNA bands (5S, 18S, 28S) were observed perfectly. Relatively, comparing and measuring some kinds of buffers (Trizol, TriPure) indicated no difference in RNA extraction quality. The last investigated factor was the effect of b- mercaptoethanol which was used along with TriPure to remove the RNAse. Conclusively, no effective impression was observed.

Comparison of the behavior of normal factor IX and the factor IX Bm variant Hilo in the prothrombin time test using tissue factors from bovine, human, and rabbit sources.

PubMed

Lefkowitz, J B; Monroe, D M; Kasper, C K; Roberts, H R

1993-07-01

A subset of hemophilia B patients have a prolonged bovine-brain prothrombin time. These CRM+ patients are classified as having hemophilia Bm. The prolongation of the prothrombin time has been reported only with bovine brain (referred to as ox brain in some literature) as the source of thromboplastin; prothrombin times determined with thromboplastin from rabbit brain or human brain are not reported to be prolonged. Factor IX from a hemophilia Bm patient (factor IX Hilo) was isolated. The activity of factor IX Hilo was compared to that of normal factor IX in prothrombin time assays when the thromboplastin source was of bovine, rabbit, or human origin. Factor IX, either normal or Hilo, prolonged a prothrombin time regardless of the tissue factor source. However, unless thromboplastin was from a bovine source, this prolongation required high concentrations of factor IX. Further, factor IX normal was as effective as factor IX Hilo in prolonging the prothrombin time when rabbit or human thromboplastin was used. With bovine thromboplastin, factor IX Hilo was significantly better than factor IX normal at prolonging the prothrombin time. The amount of prolongation was dependent on the amount of factor IX Hilo added. In addition, the prolongation was dependent on the concentration of factor X present in the sample. The prothrombin time changed as much as 20 seconds when the factor X concentration was varied from 50% to 150% to normal (fixed concentration of factor IX Hilo). These results demonstrate the difficulty of classifying the severity of a hemophilia Bm patient based on the bovine brain prothrombin time unless both the factor IX and factor X concentrations are known.

Strategies for reducing body fat mass: effects of liposuction and exercise on cardiovascular risk factors and adiposity

PubMed Central

Benatti, Fabiana Braga; Lira, Fábio Santos; Oyama, Lila Missae; do Nascimento, Cláudia Maria da Penha Oller; Lancha, Antonio Herbert

2011-01-01

Liposuction is the most popular aesthetic surgery performed in Brazil and worldwide. Evidence showing that adipose tissue is a metabolically active tissue has led to the suggestion that liposuction could be a viable method for improving metabolic profile through the immediate loss of adipose tissue. However, the immediate liposuction-induced increase in the proportion of visceral to subcutaneous adipose tissue could be detrimental to metabolism, because a high proportion of visceral to subcutaneous adipose tissue is associated with risk factors for cardiovascular disease. The results of studies investigating the effects of liposuction on the metabolic profile are inconsistent, however, with most studies reporting either no change or improvements in one or more cardiovascular risk factors. In addition, animal studies have demonstrated a compensatory growth of intact adipose tissue in response to lipectomy, although studies with humans have reported inconsistent results. Exercise training improves insulin sensitivity, inflammatory balance, lipid oxidation, and adipose tissue distribution; increases or preserves the fat-free mass; and increases total energy expenditure. Thus, liposuction and exercise appear to directly affect metabolism in similar ways, which suggests a possible interaction between these two strategies. To our knowledge, no studies have reported the associated effects of liposuction and exercise in humans. Nonetheless, one could suggest that exercise training associated with liposuction could attenuate or even block the possible compensatory fat deposition in intact depots or regrowth of the fat mass and exert an additive or even a synergistic effect to liposuction on improving insulin sensitivity and the inflammatory balance, resulting in an improvement of cardiovascular risk factors. Consequently, one could suggest that liposuction and exercise appear to be safe and effective strategies for either the treatment of metabolic disorders or aesthetic purposes. PMID:21779146

Fibroblast growth factors: key players in regeneration and tissue repair.

PubMed

Maddaluno, Luigi; Urwyler, Corinne; Werner, Sabine

2017-11-15

Tissue injury initiates a complex repair process, which in some organisms can lead to the complete regeneration of a tissue. In mammals, however, the repair of most organs is imperfect and results in scar formation. Both regeneration and repair are orchestrated by a highly coordinated interplay of different growth factors and cytokines. Among the key players are the fibroblast growth factors (FGFs), which control the migration, proliferation, differentiation and survival of different cell types. In addition, FGFs influence the expression of other factors involved in the regenerative response. Here, we summarize current knowledge on the roles of endogenous FGFs in regeneration and repair in different organisms and in different tissues and organs. Gaining a better understanding of these FGF activities is important for appropriate modulation of FGF signaling after injury to prevent impaired healing and to promote organ regeneration in humans. © 2017. Published by The Company of Biologists Ltd.

Human Factors Validation of the AeroForm Tissue Expander System for Breast Reconstruction.

PubMed

Kelley, Kathy; Kim, Jennie

The tissue expansion process using traditional saline expanders is lengthy and uncomfortable. A new technology has been developed, providing a needle-free option implanted after a mastectomy, and is activated by a handheld remote control releasing small amounts (10 cc) of carbon dioxide from an internal reservoir. The expander is gradually filled with CO2 resulting in mechanical stretching of the overlying tissue. The AeroForm System has been evaluated in a series of clinical trials including a randomized, controlled U.S. study comparing the AeroForm System with saline expanders. Results demonstrated patients can safely and reliably dose and complete their expansions in half the time compared to saline expanders. A human factors validation study was conducted in 8 patients to evaluate whether patients could correctly use the device to complete their expansion at home. The sessions were recorded and data on performance, behavioral, and subjective measures were collected and analyzed and submitted to the FDA as part of the U.S. marketing approval. All 8 participants were successful in using the controller to deliver a simulated dose. Participants found the device easy to use and the training material provided adequate to understand use of the controller. For women who choose 2-stage breast reconstruction, a new safe and effective option is available for tissue expansion, offering a convenient and empowering alternative. The human factors validation study conducted confirmed the simplicity of the device and further validated that the device can be used safely and effectively for breast tissue expansion.

Autopsy tissues as biological monitors of human exposure to environmental pollutants. A case study: Concentrations of metals and PCDD/Fs in subjects living near a hazardous waste incinerator.

PubMed

Domingo, José L; García, Francisco; Nadal, Martí; Schuhmacher, Marta

2017-04-01

Human biomonitoring is of tremendous importance to prevent potential adverse effects derived from human exposure to chemicals. Blood and urine are among the biological monitors more frequently used. However, biological matrices such as breast milk, hair, nails, saliva, feces, teeth, and expired air are also often used. In addition, and focused mainly on long-term exposure, adipose tissue and other human tissues like bone, liver, brain or kidney, are also used as biological monitors of certain substances, especially for long-term biomonitoring. However, for this kind of tissues sampling is always a limiting factor. In this paper, we have examined the role of autopsy tissues as biological monitors of human exposure to environmental pollutants. For it, we have used a case study conducted near a hazardous waste incinerator (HWI) in Catalonia (Spain), in which the concentrations of metals and polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs), have been periodically determined in autopsy tissues of subjects living in the area under potential influence of the facility. This case study does not show advantages -in comparison to other appropriate biomonitors such as blood- in using autopsy tissues in the monitoring of long-term exposure to metals and PCDD/Fs. Copyright © 2017 Elsevier Inc. All rights reserved.

Direct Reprogramming of Human Amniotic Fluid Stem Cells by OCT4 and Application in Repairing of Cerebral Ischemia Damage

PubMed Central

Qin, Mingde; Chen, Ruihua; Li, Hong; Liang, Hansi; Xue, Qun; Li, Fang; Chen, Ying; Zhang, Xueguang

2016-01-01

Amniotic fluid stem cells (AFSCs) are a type of fetal stem cell whose stemness encompasses both embryonic and adult stem cells, suggesting that they may be easily and efficiently reprogrammed into induced pluripotent stem cells (iPSCs). To further simplify the reprogramming process, the creation of AFSC-derived iPSCs using a single factor is desirable. Here we report the generation of one-factor human AFSC-iPSCs (AiPSCs) from human AFSCs by ectopic expression of the transcription factor OCT4. Just like human embryonic stem cells, AiPSCs exhibited similar epigenetic status, global gene expression profiles, teratoma formation and in vitro & in vivo pluripotency. Our results indicate that the OCT4 is necessary and sufficient to directly reprogram human AFSCs into pluripotent AiPSCs. Moreover, reflecting the similar memory characteristics of AFSCs and neural stem cells, we show that AiPSC membrane-derived vesicles (MVs) repair cerebral ischemia damage. We anticipate that the successful generation of one-factor AiPSCs will facilitate the creation of patient-specific pluripotent stem cells without the need for transgenic expression of oncogenes. Moreover, MVs from tissue-specific AiPSCs have potential in tissue repair, representing a novel application of iPSCs. PMID:27019637

Vertical ridge augmentation using an equine bone and collagen block infused with recombinant human platelet-derived growth factor-BB: a randomized single-masked histologic study in non-human primates.

PubMed

Nevins, Myron; Al Hezaimi, Khalid; Schupbach, Peter; Karimbux, Nadeem; Kim, David M

2012-07-01

This study tests the effectiveness of hydroxyapatite and collagen bone blocks of equine origin (eHAC), infused with recombinant human platelet-derived growth factor-BB (rhPDGF-BB), to augment localized posterior mandibular defects in non-human primates (Papio hamadryas). Bilateral critical-sized defects simulating severe atrophy were created at the time of the posterior teeth extraction. Test and control blocks (without growth factor) were randomly grafted into the respective sites in each non-human primate. All sites exhibited vertical ridge augmentation, with physiologic hard- and soft-tissue integration of the blocks when clinical and histologic examinations were done at 4 months after the vertical ridge augmentation procedure. There was a clear, although non-significant, tendency to increased regeneration in the test sites. As in the first two preclinical studies in this series using canines, experimental eHAC blocks infused with rhPDGF-BB proved to be a predictable and technically viable method to predictably regenerate bone and soft tissue in critical-sized defects. This investigation supplies additional evidence that eHAC blocks infused with rhPDGF-BB growth factor is a predictable and technically feasible option for vertical augmentation of severely resorbed ridges.

Higher Numbers of T-Bet+ Tumor-Infiltrating Lymphocytes Associate with Better Survival in Human Epithelial Ovarian Cancer.

PubMed

Xu, Yun; Chen, Lujun; Xu, Bin; Xiong, Yuqi; Yang, Min; Rui, Xiaohui; Shi, Liangrong; Wu, Changping; Jiang, Jingting; Lu, Binfeng

2017-01-01

T-bet, a member of the T-box family of transcription factors, is a key marker of type I immune response within the tumor microenvironment, and has been previously reported by us to serve as an important prognostic indicator for human gastric cancer patients and a potential biomarker for immunotherapy. In the present study, we aimed to assess the clinical significance and prognostic value of T-bet+ tumor-infiltrating lymphocytes in human epithelial ovarian cancer. The immunohistochemistry was used to analyze the infiltration density of T-bet+ lymphoid cells in human epithelial ovarian cancer tissues, and the flow cytometry analysis was used to further analyze the presence of T-bet+ tumor-infiltrating lymphocytes subgroups in cancer tissues. Our immunohistochemistry analysis showed increased number of T-bet+ lymphoid cells in the human epithelial ovarian cancer tissues, and the flow cytometry analysis further demonstrated the presence of T-bet+ tumor-infiltrating lymphocytes subgroups including CD4+ , CD8+ T cells and NK cells. In addition, we also observed a significant association of T-bet+ tumor-infiltrating lymphocytes density in the tumor nest of cancer with not only serum CA125 levels but also with distant metastasis. However no association was observed with other characteristics like patients' age, pathological type, FIGO stage, tumor site and tumor size. Furthermore, the survival analysis showed that higher density of T-bet+ tumor-infiltrating lymphocytes both in tumor nest and tumor stroma of cancer tissues was significantly associated with better patient survival. In addition, the density of T-bet+ tumor-infiltrating lymphocytes in tumor nest appeared to be an independent risk factor for predicting patients' postoperative prognoses. Our data indicated that the key transcription factor T-bet might play an important role in the type I immune cells mediated antitumor response, and the density of T-bet+ lymphocytes in human epithelial ovarian cancer tissues could serve as a prognostic predictor for ovarian cancer patients. © 2017 The Author(s)Published by S. Karger AG, Basel.

Impact of static magnetic fields on human myoblast cell cultures.

PubMed

Stern-Straeter, Jens; Bonaterra, Gabriel Alejandro; Kassner, Stefan S; Faber, Anne; Sauter, Alexander; Schulz, Johannes D; Hörmann, Karl; Kinscherf, Ralf; Goessler, Ulrich Reinhart

2011-12-01

Treatment of skeletal muscle loss due to trauma or tumor ablation therapy still lacks a suitable clinical approach. Creation of functional muscle tissue in vitro using the differentiation potential of human satellite cells (myoblasts) is a promising new research field called tissue engineering. Strong differentiation stimuli, which can induce formation of myofibers after cell expansion, have to be identified and evaluated in order to create sufficient amounts of neo-tissue. The objective of this study was to determine the influence of static magnetic fields (SMF) on human satellite cell cultures as one of the preferred stem cell sources in skeletal muscle tissue engineering. Experiments were performed using human satellite cells with and without SMF stimulation after incubation with a culture medium containing low [differentiation medium (DM)] or high [growth medium (GM)] concentrations of growth factors. Proliferation analysis using the alamarBlue assay revealed no significant influence of SMF on cell division. Real-time RT-PCR of the following marker genes was investigated: myogenic factor 5 (MYF5), myogenic differentiation antigen 1 (MYOD1), myogenin (MYOG), skeletal muscle α1 actin (ACTA1), and embryonic (MYH3), perinatal (MYH8) and adult (MYH1) skeletal muscle myosin heavy chain. We detected an influence on marker gene expression by SMF in terms of a down-regulation of the marker genes in cell cultures treated with SMF and DM, but not in cell cultures treated with SMF and GM. Immunocytochemical investigations using antibodies directed against the differentiation markers confirmed the gene expression results and showed an enhancement of maturation after stimulation with GM and SMF. Additional calculation of the fusion index also revealed an increase in myotube formation in cell cultures treated with SMF and GM. Our findings show that the effect of SMF on the process of differentiation depends on the growth factor concentration in the culture medium in human satellite cultures. SMF alone enhances the maturation of human satellite cells treated with GM, but not satellite cells that were additionally stimulated with serum cessation. Therefore, further investigations are necessary before consideration of SMF for skeletal muscle tissue engineering approaches.

Porcine endothelium induces DNA-histone complex formation in human whole blood: a harmful effect of histone on coagulation and endothelial activation.

PubMed

Yoo, Hyun Ju; Kim, Ji-Eun; Gu, Ja Yoon; Lee, Sae Bom; Lee, Hyun Joo; Hwang, Ho Young; Hwang, Yoohwa; Kim, Young Tae; Kim, Hyun Kyung

2016-11-01

Neutrophils play a role in xenograft rejection. When neutrophils are stimulated, they eject the DNA-histone complex into the extracellular space, called neutrophil extracellular traps (NET). We investigated whether NET formation actively occurs in the xenograft and contributes to coagulation and endothelial activation. Human whole blood was incubated with porcine aortic endothelial cells (pEC) from wild-type or α1,3-galactosyltransferase gene-knockout (GTKO) pigs. In the supernatant plasma from human blood, the level of the DNA-histone complex was measured by ELISA, and thrombin generation was measured using a calibrated automated thrombogram. Histone-induced tissue factor and adhesion molecule expression were measured by flow cytometry. pEC from both wild-type and GTKO pigs significantly induced DNA-histone complex formation in human whole blood. The DNA-histone complex produced shortened the thrombin generation time and clotting time. Histone alone dose-dependently induced tissue factor and adhesion molecule expression in pEC. Aurintricarboxylic acid pretreatment partially inhibited pEC-induced DNA-histone complex formation. DNA-histone complex actively generated upon xenotransplantation is a novel target to inhibit coagulation and endothelial activation. To prevent tissue factor and adhesion molecule expression, a strategy to block soluble histone may be required in xenotransplantation. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Immunohistochemical Localization of Periostin in Human Gingiva

PubMed Central

Cobo, T.; Obaya, A.; Cal, S.; Solares, L.; Cabo, R.; Vega, J.A.; Cobo, J.

2015-01-01

The periostin is a matricellular protein expressed in collagen-rich tissues including some dental and periodontal tissues where it is regulated by mechanical forces, growth factors and cytokines. Interestingly the expression of this protein has been found modified in different gingival pathologies although the expression of periostin in normal human gingiva was never investigated. Here we used Western blot and double immunofluorescence coupled to laser-confocal microscopy to investigated the occurrence and distribution of periostin in different segments of the human gingival in healthy subjects. By Western blot a protein band with an estimated molecular mass of 94 kDa was observed. Periostin was localized at the epithelial-connective tissue junction, or among the fibers of the periodontal ligament, and never co-localized with cytokeratin or vimentin thus suggesting it is an extracellular protein. These results demonstrate the occurrence of periostin in adult human gingiva; its localization suggests a role in the bidirectional interactions between the connective tissue and the epithelial cells, and therefore in the physiopathological conditions in which these interactions are altered. PMID:26428890

Fluorescence spectroscopy using excitation and emission matrix for quantification of tissue native fluorophores and cancer diagnosis

NASA Astrophysics Data System (ADS)

Wu, Binlin; Gayen, S. K.; Xu, M.

2014-03-01

Native fluorescence spectrum of normal and cancerous human prostate tissues is studied to distinguish between normal and cancerous tissues, and cancerous tissues at different cancer grade. The tissue samples were obtained from Cooperative Human Tissue Network (CHTN) and National Disease Research Interchange(NDRI). An excitation and emission matrix (EEM) was generated for each tissue sample by acquiring native fluorescence spectrum of the sample using multiple excitation wavelengths. The non-negative matrix factorization algorithm was used to generate fluorescence EEMs that correspond to the fluorophores in biological tissues, including tryptophan, collagen, elastin, nicotinamide adenine dinucleotide (NADH), flavin adenine dinucleotide (FAD) and the background paraffin. We hypothesize that, as a consequence of metabolic changes associated with the development of cancer, the concentrations of NADH and FAD are different in normal and cancerous tissues, and also different for different cancer grades. We used the ratio of the abundances of FAD and NADH to distinguish between normal and cancerous tissues, and the tissue cancer grade. The FAD-to-NADH ratio was found to be the highest for normal tissue and decreased as the cancer grade increased.

The Human Microbiome and Skin and Soft-Tissue Infections

DTIC Science & Technology

2015-09-23

254   Cell harvesting, RNA extraction, and RT-PCR ...................................................... 257   Primer extension and 5...contribution of the human microbiota to human health is not surprising given that the number of human-associated bacterial cells far exceeds the 37.5 trillion... cells of the human body by a factor of ~10 (27; 266). Additionally, the human 13 Figure 1. Microbiome and disease-related PubMed

Impaired coordination between signaling pathways is revealed in human colorectal cancer using single-cell mass cytometry of archival tissue blocks

PubMed Central

Simmons, Alan J.; Scurrah, Cherie’ R.; McKinley, Eliot T.; Herring, Charles A.; Irish, Jonathan M.; Washington, Mary K.; Coffey, Robert J.; Lau, Ken S.

2016-01-01

Cellular heterogeneity poses a significant challenge to understanding tissue level phenotypes and confounds conventional bulk analyses. To facilitate the analysis of signaling at the single-cell level in human tissues, we applied mass cytometry using CyTOF (Cytometry Time-of-Flight) to formalin-fixed paraffin-embedded (FFPE) normal and diseased intestinal specimens. We developed and validated a technique called FFPE-DISSECT (Disaggregation for Intracellular Signaling in Single Epithelial Cells from Tissue), a single-cell approach for characterizing native signaling states from embedded solid tissue samples. We applied FFPE-DISSECT coupled to mass cytometry and found differential signaling by tumor necrosis factor α (TNF-α) in intestinal enterocytes, goblet cells and enteroendocrine cells, implicating the role of the downstream RAS-RAF-MEK-ERK signaling pathway in dictating goblet cell identity. In addition, application of FFPE-DISSECT, mass cytometry, and data-driven computational analyses to human colon specimens confirmed reduced differentiation in colorectal cancer (CRC) compared to normal colon, and revealed quantitative increases in inter- and intra-tissue heterogeneity in CRC with regards to the modular regulation of signaling pathways. Specifically, modular co-regulation of the kinases P38 and ERK, the translation regulator 4EBP1, and the transcription factor CREB in the proliferative compartment of the normal colon was loss in CRC, as evidenced by their impaired coordination over samplings of single cells in tissue. Our data suggest that this single-cell approach, applied in conjunction with genomic annotation, such as microsatellite instability and mutations in KRAS and BRAF, allows rapid and detailed characterization of cellular heterogeneity from clinical repositories of embedded human tissues. FFPE-DISSECT coupled of mass cytometry can be used for deriving cellular landscapes from archived patient samples, beyond CRC, and as a high resolution tool for disease characterization and subtyping. PMID:27729552

The use of platelet-rich fibrin combined with periodontal ligament and jaw bone mesenchymal stem cell sheets for periodontal tissue engineering.

PubMed

Wang, Zhong-Shan; Feng, Zhi-Hong; Wu, Guo-Feng; Bai, Shi-Zhu; Dong, Yan; Chen, Fa-Ming; Zhao, Yi-Min

2016-06-21

Periodontal regeneration involves the restoration of at least three unique tissues: cementum, periodontal ligament tissue (PDL) and alveolar bone tissue. Here, we first isolated human PDL stem cells (PDLSCs) and jaw bone mesenchymal stem cells (JBMSCs). These cells were then induced to form cell sheets using an ascorbic acid-rich approach, and the cell sheet properties, including morphology, thickness and gene expression profile, were compared. Platelet-rich fibrin (PRF) derived from human venous blood was then fabricated into bioabsorbable fibrin scaffolds containing various growth factors. Finally, the in vivo potential of a cell-material construct based on PDLSC sheets, PRF scaffolds and JBMSC sheets to form periodontal tissue was assessed in a nude mouse model. In this model, PDLSC sheet/PRF/JBMSC sheet composites were placed in a simulated periodontal space comprising human treated dentin matrix (TDM) and hydroxyapatite (HA)/tricalcium phosphate (TCP) frameworks. Eight weeks after implantation, the PDLSC sheets tended to develop into PDL-like tissues, while the JBMSC sheets tended to produce predominantly bone-like tissues. In addition, the PDLSC sheet/PRF/JBMSC sheet composites generated periodontal tissue-like structures containing PDL- and bone-like tissues. Further improvements in this cell transplantation design may have the potential to provide an effective approach for future periodontal tissue regeneration.

Complement factor H: spatial and temporal expression and localization in the eye.

PubMed

Mandal, Md Nawajes A; Ayyagari, Radha

2006-09-01

Complement factor H (CFH) is a component of the mammalian complement system, which regulates the alternative pathway of complement activation and protects the host cell from inappropriate complement activation. CFH is a key regulator of innate immunity, and CFH deficiency leads to membranoproliferative glomerulonephritis type II. A variation in human CFH, Y402H, has been shown to be associated with an increased risk for age-related macular degeneration. The authors describe studies on the spatial and temporal expression of the CFH gene and localization of this protein in ocular tissues to gain insight into its role in the eye. CFH expression in human and mouse tissues was studied by quantitative RT-PCR and Western blot analysis, and localization of CFH was studied by immunohistochemical analysis followed by fluorescence microscopy. In human and mouse, CFH expression was found to be similar to the highest level of expression in the liver. In ocular tissue, CFH was detected in the distalmost optic nerve (3 mm) cut from the scleral surface of the eyeball, sclera, RPE-choroid, retina, lens, and ciliary body. In mouse, Cfh expression was observed from early embryonic stages, and in the eye its expression increased with age. A significant level of CFH expression is maintained in different ocular tissues during development and aging. Sustained high levels of CFH expression in eye tissues suggest that this protein may play a role in protecting these tissues from indiscriminate complement activation and inflammatory insult.

The effect of Setarud (IMOD(TM)) on angiogenesis in transplanted human ovarian tissue to nude mice.

PubMed

Hormozi, Maryam; Talebi, Saeed; Khorram Khorshid, Hamid Reza; Zarnani, Amir-Hassan; Kamali, Koorosh; Jeddi-Tehrani, Mahmood; Soltangoraee, Haleh; Akhondi, Mohammad Mehdi

2015-10-01

One of the promising methods in fertility preservation among women with cancer is cryopreservation of ovarian cortex but there are many drawbacks such as apoptosis and considerable reduction of follicular density in the transplanted ovary. One solution to reduce ischemic damage is enhancing angiogenesis after transplantation of ovarian cortex tissue. The aim of this study was to investigate the effect of Setarud, on angiogenesis in transplanted human ovarian tissue. In this case control study, twenty four nude mice were implanted subcutaneously, with human ovarian tissues, from four women. The mice were randomly divided into two groups (n=12): the experimental group was treated with Setarud, while control group received only vehicle. Each group was divided into three subgroups (n=4) based on the graft recovery days post transplantation (PT). The transplanted fragments were removed on days 2, 7, and 30 PT and the expression of Angiopoietin-1, Angiopoietin-2, and Vascular endothelial growth factor at both gene and protein levels and vascular density were studied in the grafted ovarian tissues. On the 2(nd) and 7(th) day PT, the level of Angiopoietin-1 gene expression in case group was significantly lower than that in control group, while the opposite results were obtained for Angiopoietin-2 and Vascular endothelial growth factor. These results were also confirmed at the protein level. The density of vessels in Setarud group elevated significantly on day 7 PT compared to pre-treatment state. Our results showed that administration of Setarud may stimulates angiogenesis in transplanted human ovarian tissues, although further researches are needed before a clear judgment is made.

The effect of Setarud (IMODTM) on angiogenesis in transplanted human ovarian tissue to nude mice

PubMed Central

Hormozi, Maryam; Talebi, Saeed; Khorram Khorshid, Hamid Reza; Zarnani, Amir-Hassan; Kamali, Koorosh; Jeddi-Tehrani, Mahmood; Soltangoraee, Haleh; Akhondi, Mohammad Mehdi

2015-01-01

Background: One of the promising methods in fertility preservation among women with cancer is cryopreservation of ovarian cortex but there are many drawbacks such as apoptosis and considerable reduction of follicular density in the transplanted ovary. One solution to reduce ischemic damage is enhancing angiogenesis after transplantation of ovarian cortex tissue. Objective: The aim of this study was to investigate the effect of Setarud, on angiogenesis in transplanted human ovarian tissue. Materials and Methods: In this case control study, twenty four nude mice were implanted subcutaneously, with human ovarian tissues, from four women. The mice were randomly divided into two groups (n=12): the experimental group was treated with Setarud, while control group received only vehicle. Each group was divided into three subgroups (n=4) based on the graft recovery days post transplantation (PT). The transplanted fragments were removed on days 2, 7, and 30 PT and the expression of Angiopoietin-1, Angiopoietin-2, and Vascular endothelial growth factor at both gene and protein levels and vascular density were studied in the grafted ovarian tissues. Results: On the 2nd and 7th day PT, the level of Angiopoietin-1 gene expression in case group was significantly lower than that in control group, while the opposite results were obtained for Angiopoietin-2 and Vascular endothelial growth factor. These results were also confirmed at the protein level. The density of vessels in Setarud group elevated significantly on day 7 PT compared to pre-treatment state. Conclusion: Our results showed that administration of Setarud may stimulates angiogenesis in transplanted human ovarian tissues, although further researches are needed before a clear judgment is made. PMID:26644788

Guidelines for preventing transmission of human immunodeficiency virus through transplantation of human tissue and organs. Centers for Disease Control and Prevention.

PubMed

1994-05-20

Although previous recommendations for preventing transmission of human immunodeficiency virus (HIV) through transplantation of human tissue and organs have markedly reduced the risk for this type of transmission, a case of HIV transmission from a screened, antibody-negative donor to several recipients raised questions about the need for additional federal oversight of transplantation of organs and tissues. A working group formed by the Public Health Service (PHS) in 1991 to address these issues concluded that further recommendations should be made to reduce the already low risk of HIV transmission by transplantation of organs and tissues. In revising these recommendations, the PHS sought assistance from public and private health professionals and representatives of transplant, public health, and other organizations. The revised guidelines address issues such as donor screening, testing, and exclusionary criteria; quarantine of tissue from living donors; inactivation or elimination of infectious organisms in organs and tissues before transplantation; timely detection, reporting, and tracking of potentially infected tissues, organs, and recipients; and recall of stored tissues from donors found after donation to have been infected. Factors considered in the development of these guidelines include differences between the screening of living and cadaveric donors; time constraints due to organ/tissue viability that may preclude performing certain screening procedures; differences in the risk of HIV transmission from various organs and tissues; differences between systems for procuring and distributing organs and tissues; the effect of screening practices on the limited availability of organs and some tissues; and the benefit of the transplant to the recipient.

[The gastrointestinal tract microbiom in connective tissue diseases].

PubMed

Krajewska-Włodarczyk, Magdalena

Factors such as genetics, the environment, infections, and the human body microbiota, mainly gastrointestinal tract microbiota may play a role in the pathogenesis of autoimmune disorders. There is an increasing evidence that suggest an association between gastrointestinal tract dysbiosis, and in particular gut dysbiosis, and connective tissue diseases but it still remains unclear whether alterations in the microbiome are a pathogenic cause or an effect of autoimmune disease. Given the strong variability and abundance of microbes living in close relation with human host, it becomes a difficult task to define what should be considered the normal or the favorable microbiome. Further studies are needed to establish how the human microbiome contributes to disease susceptibility, and to characterize the role of microbial diversity in the pathogenesis of connective tissue diseases and their clinical manifestations. The identification of dysbiosis specific for certain connective tissue diseases may help in the development of an individualized management for each patient. This review aims to summarize current data on the role of the gastrointestinal tract microbiome in connective tissue diseases.

Expression of connective tissue growth factor is a prognostic marker for patients with intrahepatic cholangiocarcinoma.

PubMed

Gardini, A; Corti, B; Fiorentino, M; Altimari, A; Ercolani, G; Grazi, G L; Pinna, A D; Grigioni, W F; D'Errico Grigioni, A

2005-04-01

Connective tissue growth factor is a member of the 'CCN' protein family. Consistent with its profibrotic properties, it is over-expressed in several human epithelial malignancies. We have retrospectively evaluated by immunohistochemistry the presence of connective tissue growth factor in archival tissues from 55 resected intrahepatic cholangiocarcinomas and compared its expression to the main pathological parameters, disease free and overall survival. Tumours were scored as high and low/absent expressers (> or =50%, 0-50% cells, respectively). Thirty-three of 55 cholangiocarcinomas (60%) were high and 22 (40%) low expressers. No significant correlation was found between connective tissue growth factor and tumour grade, tumour location, vascular and perineural invasion. Eighteen of 22 (82%) low/absent expressers and 12/33 (36%) high expressers had recurrence of disease (P=0.001). Low/absent expressers showed a poor disease free and overall survival compared with the higher expressers (P<0.001). Vascular invasion was related to tumour recurrence (P=0.025) and to decreased disease free survival (P<0.05). During proportional hazard regression analysis, only connective tissue growth factor was found to influence disease free survival (P=0.01). Expression of connective tissue growth factor is an independent prognostic indicator of both tumour recurrence and overall survival for intrahepatic cholangiocarcinoma patients regardless of tumour location, tumour grade, vascular and perineural invasion.

Performance of automated scoring of ER, PR, HER2, CK5/6 and EGFR in breast cancer tissue microarrays in the Breast Cancer Association Consortium

PubMed Central

Howat, William J; Blows, Fiona M; Provenzano, Elena; Brook, Mark N; Morris, Lorna; Gazinska, Patrycja; Johnson, Nicola; McDuffus, Leigh‐Anne; Miller, Jodi; Sawyer, Elinor J; Pinder, Sarah; van Deurzen, Carolien H M; Jones, Louise; Sironen, Reijo; Visscher, Daniel; Caldas, Carlos; Daley, Frances; Coulson, Penny; Broeks, Annegien; Sanders, Joyce; Wesseling, Jelle; Nevanlinna, Heli; Fagerholm, Rainer; Blomqvist, Carl; Heikkilä, Päivi; Ali, H Raza; Dawson, Sarah‐Jane; Figueroa, Jonine; Lissowska, Jolanta; Brinton, Louise; Mannermaa, Arto; Kataja, Vesa; Kosma, Veli‐Matti; Cox, Angela; Brock, Ian W; Cross, Simon S; Reed, Malcolm W; Couch, Fergus J; Olson, Janet E; Devillee, Peter; Mesker, Wilma E; Seyaneve, Caroline M; Hollestelle, Antoinette; Benitez, Javier; Perez, Jose Ignacio Arias; Menéndez, Primitiva; Bolla, Manjeet K; Easton, Douglas F; Schmidt, Marjanka K; Pharoah, Paul D; Sherman, Mark E

2014-01-01

Abstract Breast cancer risk factors and clinical outcomes vary by tumour marker expression. However, individual studies often lack the power required to assess these relationships, and large‐scale analyses are limited by the need for high throughput, standardized scoring methods. To address these limitations, we assessed whether automated image analysis of immunohistochemically stained tissue microarrays can permit rapid, standardized scoring of tumour markers from multiple studies. Tissue microarray sections prepared in nine studies containing 20 263 cores from 8267 breast cancers stained for two nuclear (oestrogen receptor, progesterone receptor), two membranous (human epidermal growth factor receptor 2 and epidermal growth factor receptor) and one cytoplasmic (cytokeratin 5/6) marker were scanned as digital images. Automated algorithms were used to score markers in tumour cells using the Ariol system. We compared automated scores against visual reads, and their associations with breast cancer survival. Approximately 65–70% of tissue microarray cores were satisfactory for scoring. Among satisfactory cores, agreement between dichotomous automated and visual scores was highest for oestrogen receptor (Kappa = 0.76), followed by human epidermal growth factor receptor 2 (Kappa = 0.69) and progesterone receptor (Kappa = 0.67). Automated quantitative scores for these markers were associated with hazard ratios for breast cancer mortality in a dose‐response manner. Considering visual scores of epidermal growth factor receptor or cytokeratin 5/6 as the reference, automated scoring achieved excellent negative predictive value (96–98%), but yielded many false positives (positive predictive value = 30–32%). For all markers, we observed substantial heterogeneity in automated scoring performance across tissue microarrays. Automated analysis is a potentially useful tool for large‐scale, quantitative scoring of immunohistochemically stained tissue microarrays available in consortia. However, continued optimization, rigorous marker‐specific quality control measures and standardization of tissue microarray designs, staining and scoring protocols is needed to enhance results. PMID:27499890

The effect of dietary carbohydrate on genes for fatty acid synthase and inflammatory cytokines in adipose tissues from lean and obese subjects.

PubMed

Hudgins, Lisa C; Baday, Aline; Hellerstein, Marc K; Parker, Thomas S; Levine, Daniel M; Seidman, Cynthia E; Neese, Richard A; Tremaroli, Jolanta D; Hirsch, Jules

2008-04-01

Hepatic de novo lipogenesis (DNL) is markedly stimulated in humans by low-fat diets enriched in simple sugars. However, the dietary responsiveness of the key enzyme controlling DNL in human adipose tissue, fatty acid synthase (FAS), is uncertain. Adipose tissue mRNA for FAS is increased in lean and obese subjects when hepatic DNL is elevated by a eucaloric, low-fat, high-sugar diet. Twelve lean and seven obese volunteers were given two eucaloric diets (10% vs. 30% fat; 75% vs. 55% carbohydrate; sugar/starch 60/40) each for 2 weeks by a random-order cross-over design. FAS mRNA in abdominal and gluteal adipose tissues was compared to hepatic DNL measured in serum by isotopic and nonisotopic methods. Adipose tissue mRNA for tumor necrosis factor-alpha and IL-6, which are inflammatory cytokines that modulate DNL, was also assayed. The low-fat high-sugar diet induced a 4-fold increase in maximum hepatic DNL (P<.001) but only a 1.3-fold increase in adipose tissue FAS mRNA (P=.029) and no change in cytokine mRNA. There was a borderline significant positive correlation between changes in FAS mRNA and hepatic DNL (P=.039). Compared to lean subjects, obese subjects had lower levels of FAS mRNA and higher levels of cytokine mRNA (P<.001). The results suggest that key elements of human adipose tissue DNL are less responsive to dietary carbohydrate than is hepatic DNL and may be regulated by diet-independent factors. Irrespective of diet, there is reduced expression of the FAS gene and increased expression of cytokine genes in adipose tissues of obese subjects.

FXIa and platelet polyphosphate as therapeutic targets during human blood clotting on collagen/tissue factor surfaces under flow

PubMed Central

Zhu, Shu; Travers, Richard J.; Morrissey, James H.

2015-01-01

Factor XIIa (FXIIa) and factor XIa (FXIa) contribute to thrombosis in animal models, whereas platelet-derived polyphosphate (polyP) may potentiate contact or thrombin-feedback pathways. The significance of these mediators in human blood under thrombotic flow conditions on tissue factor (TF) –bearing surfaces remains inadequately resolved. Human blood (corn trypsin inhibitor treated [4 μg/mL]) was tested by microfluidic assay for clotting on collagen/TF at TF surface concentration ([TF]wall) from ∼0.1 to 2 molecules per μm2. Anti-FXI antibodies (14E11 and O1A6) or polyP-binding protein (PPXbd) were used to block FXIIa-dependent FXI activation, FXIa-dependent factor IX (FIX) activation, or platelet-derived polyP, respectively. Fibrin formation was sensitive to 14E11 at 0 to 0.1 molecules per µm2 and sensitive to O1A6 at 0 to 0.2 molecules per µm2. However, neither antibody reduced fibrin generation at ∼2 molecules per µm2 when the extrinsic pathway became dominant. Interestingly, PPXbd reduced fibrin generation at low [TF]wall (0.1 molecules per µm2) but not at zero or high [TF]wall, suggesting a role for polyP distinct from FXIIa activation and requiring low extrinsic pathway participation. Regardless of [TF]wall, PPXbd enhanced fibrin sensitivity to tissue plasminogen activator and promoted clot retraction during fibrinolysis concomitant with an observed PPXbd-mediated reduction of fibrin fiber diameter. This is the first detection of endogenous polyP function in human blood under thrombotic flow conditions. When triggered by low [TF]wall, thrombosis may be druggable by contact pathway inhibition, although thrombolytic susceptibility may benefit from polyP antagonism regardless of [TF]wall. PMID:26136249

Delivery of growth factors for tissue regeneration and wound healing.

PubMed

Koria, Piyush

2012-06-01

Growth factors are soluble secreted proteins capable of affecting a variety of cellular processes important for tissue regeneration. Consequently, the self-healing capacity of patients can be augmented by artificially enhancing one or more processes important for healing through the application of growth factors. However, their application in clinics remains limited due to lack of robust delivery systems and biomaterial carriers. Interestingly, all clinically approved therapies involving growth factors utilize some sort of a biomaterial carrier for growth factor delivery. This suggests that biomaterial delivery systems are extremely important for successful usage of growth factors in regenerative medicine. This review outlines the role of growth factors in tissue regeneration, and their application in both pre-clinical animal models of regeneration and clinical trials is discussed. Additionally, current status of biomaterial substrates and sophisticated delivery systems such as nanoparticles for delivery of exogenous growth factors and peptides in humans are reviewed. Finally, issues and possible future research directions for growth factor therapy in regenerative medicine are discussed.

Requirement of Vascular Integrin α_vβ_3 for Angiogenesis

NASA Astrophysics Data System (ADS)

Brooks, Peter C.; Clark, Richard A. F.; Cheresh, David A.

1994-04-01

Angiogenesis depends on the adhesive interactions of vascular cells. The adhesion receptor integrin α_vβ_3 was identified as a marker of angiogenic vascular tissue. Integrin α_vβ_3 was expressed on blood vessels in human wound granulation tissue but not in normal skin, and it showed a fourfold increase in expression during angiogenesis on the chick chorioallantoic membrane. In the latter assay, a monoclonal antibody to α_vβ_3 blocked angiogenesis induced by basic fibroblast growth factor, tumor necrosis factor-α, and human melanoma fragments but had no effect on preexisting vessels. These findings suggest that α_vβ_3 may be a useful therapeutic target for diseases characterized by neovascularization.

[Effect of Leonurus Heterophyllus Sweet on tissue factor transcription and expression in human umbilical vein endothelial cells in vitro].

PubMed

Zheng, Lian; Fang, Chi-hua

2007-06-01

To investigate the effect of Leonurus Heterophyllus Sweet, (LHS) on tissue factor (TF) transcription and expression induced by thrombin in human umbilical vein endothelial cells (HUVECs). HUVECs were incubated with different concentrations of LHS and the TF mRNA expression was detected by reverse transcript-polymerase chain reaction (RT-PCR). LHS treatment of HUVECs at different concentrations and for different times resulted in significant differences in TF expression (Plt;0.01). The transcription of TF in LHS-treated cells was significantly different from that of the blank control group (Plt;0.01). LHS can decrease the expression of TF and intervene with TF transcription in HUVECs in vitro.

Opposite Effects of Soluble Factors Secreted by Adipose Tissue on Proliferating and Quiescent Osteosarcoma Cells.

PubMed

Avril, Pierre; Duteille, Franck; Ridel, Perrine; Heymann, Marie-Françoise; De Pinieux, Gonzague; Rédini, Françoise; Blanchard, Frédéric; Heymann, Dominique; Trichet, Valérie; Perrot, Pierre

2016-03-01

Autologous adipose tissue transfer may be performed for aesthetic needs following resection of osteosarcoma, the most frequent primary malignant tumor of bone, excluding myeloma. The safety of autologous adipose tissue transfer regarding the potential risk of cancer recurrence must be addressed. Adipose tissue injection was tested in a human osteosarcoma preclinical model induced by MNNG-HOS cells. Culture media without growth factors from fetal bovine serum were conditioned with adipose tissue samples and added to two osteosarcoma cell lines (MNNG-HOS and MG-63) that were cultured in monolayer or maintained in nonadherent spheres, favoring a proliferation or quiescent stage, respectively. Proliferation and cell cycle were analyzed. Adipose tissue injection increased local growth of osteosarcoma in mice but was not associated with aggravation of lung metastasis or osteolysis. Adipose tissue-derived soluble factors increased the in vitro proliferation of osteosarcoma cells up to 180 percent. Interleukin-6 and leptin were measured in higher concentrations in adipose tissue-conditioned medium than in osteosarcoma cell-conditioned medium, but the authors' results indicated that they were not implicated alone. Furthermore, adipose tissue-derived soluble factors did not favor a G0-to-G1 phase transition of MNNG-HOS cells in nonadherent oncospheres. This study indicates that adipose tissue-soluble factors activate osteosarcoma cell cycle from G1 to mitosis phases, but do not promote the transition from quiescent G0 to G1 phases. Autologous adipose tissue transfer may not be involved in the activation of dormant tumor cells or cancer stem cells.

Cell plasticity in wound healing: paracrine factors of M1/ M2 polarized macrophages influence the phenotypical state of dermal fibroblasts

PubMed Central

2013-01-01

Background Macrophages and fibroblasts are two major players in tissue repair and fibrosis. Despite the relevance of macrophages and fibroblasts in tissue homeostasis, remarkably little is known whether macrophages are able to influence the properties of fibroblasts. Here we investigated the role of paracrine factors secreted by classically activated (M1) and alternatively activated (M2) human macrophages on human dermal fibroblasts (HDFs). Results HDFs stimulated with paracrine factors from M1 macrophages showed a 10 to > 100-fold increase in the expression of the inflammatory cytokines IL6, CCL2 and CCL7 and the matrix metalloproteinases MMP1 and MMP3. This indicates that factors produced by M1 macrophages induce a fibroblast phenotype with pro-inflammatory and extracellular matrix (ECM) degrading properties. HDFs stimulated with paracrine factors secreted by M2 macrophages displayed an increased proliferation rate. Interestingly, the M1-activated pro-inflammatory fibroblasts downregulated, after exposure to paracrine factors produced by M2 macrophages or non-conditioned media, the inflammatory markers as well as MMPs and upregulated their collagen production. Conclusions Paracrine factors of M1 or M2 polarized macrophages induced different phenotypes of HDFs and the HDF phenotypes can in turn be reversed, pointing to a high dynamic plasticity of fibroblasts in the different phases of tissue repair. PMID:23601247

Heparanase enhances the generation of activated factor X in the presence of tissue factor and activated factor VII.

PubMed

Nadir, Yona; Brenner, Benjamin; Fux, Liat; Shafat, Itay; Attias, Judith; Vlodavsky, Israel

2010-11-01

Heparanase is an endo-β-D-glucuronidase dominantly involved in tumor metastasis and angiogenesis. Recently, we demonstrated that heparanase is involved in the regulation of the hemostatic system. Our hypothesis was that heparanase is directly involved in activation of the coagulation cascade. Activated factor X and thrombin were studied using chromogenic assays, immunoblotting and thromboelastography. Heparanase levels were measured by enzyme-linked immunosorbent assay. A potential direct interaction between tissue factor and heparanase was studied by co-immunoprecipitation and far-western assays. Interestingly, addition of heparanase to tissue factor and activated factor VII resulted in a 3- to 4-fold increase in activation of the coagulation cascade as shown by increased activated factor X and thrombin production. Culture medium of human embryonic kidney 293 cells over-expressing heparanase and its derivatives increased activated factor X levels in a non-enzymatic manner. When heparanase was added to pooled normal plasma, a 7- to 8-fold increase in activated factor X level was observed. Subsequently, we searched for clinical data supporting this newly identified role of heparanase. Plasma samples from 35 patients with acute leukemia at presentation and 20 healthy donors were studied for heparanase and activated factor X levels. A strong positive correlation was found between plasma heparanase and activated factor X levels (r=0.735, P=0.001). Unfractionated heparin and an inhibitor of activated factor X abolished the effect of heparanase, while tissue factor pathway inhibitor and tissue factor pathway inhibitor-2 only attenuated the procoagulant effect. Using co-immunoprecipitation and far-western analyses it was shown that heparanase interacts directly with tissue factor. Overall, our results support the notion that heparanase is a potential modulator of blood hemostasis, and suggest a novel mechanism by which heparanase increases the generation of activated factor X in the presence of tissue factor and activated factor VII.

[Review of influence of landing impact on human body (correction of boby) and its medical evaluation].

PubMed

Guo, Yao-yu; Tan, Cheng; Liu, Bing-kun; Jiang, Shi-zhong

2002-12-01

Landing impact is the dynamic factor that manned spaceship will inevitably meet after the mission has been completed, and impact force may cause damages to human tissues [correction of tissuses] and organs, even death. This paper described the characteristics of pathological and dynamic response of human body to landing impact, and discussed various related factors such as impact angle, fetters, design of cushion, harness and terrain condition. Medical evaluation of +Gx, +Gz, +/- Gy impacts were summarized.

Vaginal Lactobacillus Inhibits HIV-1 Replication in Human Tissues Ex Vivo

PubMed Central

Ñahui Palomino, Rogers A.; Zicari, Sonia; Vanpouille, Christophe; Vitali, Beatrice; Margolis, Leonid

2017-01-01

Lactobacillus species, which dominate vaginal microbiota of healthy reproductive-age women, lower the risks of sexually transmitted infections, including the risk of human immunodeficiency virus (HIV) acquisition. The exact mechanisms of this protection remain to be understood. Here, we investigated these mechanisms in the context of human cervico-vaginal and lymphoid tissues ex vivo. We found that all six Lactobacillus strains tested in these systems significantly suppressed HIV type-1 (HIV-1) infection. We identified at least three factors that mediated this suppression: (i) Acidification of the medium. The pH of the undiluted medium conditioned by lactobacilli was between 3.8 and 4.6. Acidification of the culture medium with hydrochloric acid (HCl) to this pH in control experiments was sufficient to abrogate HIV-1 replication. However, the pH of the Lactobacillus-conditioned medium (CM) diluted fivefold, which reached ∼6.9, was also suppressive for HIV-1 infection, while in control experiments HIV-1 infection was not abrogated when the pH of the medium was brought to 6.9 through the use of HCl. This suggested the existence of other factors responsible for HIV-1 inhibition by lactobacilli. (ii) Lactic acid. There was a correlation between the concentration of lactic acid in the Lactobacillus-CM and its ability to suppress HIV-1 infection in human tissues ex vivo. Addition of lactic acid isomers D and L to tissue culture medium at the concentration that corresponded to their amount released by lactobacilli resulted in HIV-1 inhibition. Isomer L was produced in higher quantities than isomer D and was mostly responsible for HIV-1 inhibition. These results indicate that lactic acid, in particular its L-isomer, inhibits HIV-1 independently of lowering of the pH. (iii) Virucidal effect. Incubation of HIV-1 in Lactobacillus-CM significantly suppressed viral infectivity for human tissues ex vivo. Finally, lactobacilli adsorb HIV-1, serving as a sink decreasing the number of free virions. In summary, we found that lactobacilli inhibit HIV-1 replication in human tissue ex vivo by multiple mechanisms. Further studies are needed to evaluate the potential of altering the spectra of vaginal microbiota as an effective strategy to enhance vaginal health. Human tissues ex vivo may serve as a test system for these strategies. PMID:28579980

Properties of dehydrated human amnion/chorion composite grafts: Implications for wound repair and soft tissue regeneration.

PubMed

Koob, Thomas J; Lim, Jeremy J; Massee, Michelle; Zabek, Nicole; Denozière, Guilhem

2014-08-01

PURION(®) processed dehydrated human amnion/chorion membrane (dHACM; MiMedx Group, Marietta, GA) tissue products were analyzed for the effectiveness of the PURION(®) process in retaining the native composition of the amniotic membrane and preserving bioactivity in the resulting products. dHACM was analyzed for extracellular matrix (ECM) composition through histological staining and for growth factor content via multiplex ELISA arrays. Bioactivity was assessed by evaluating endogenous growth factor production by human dermal fibroblasts in response to dHACM and for thermal stability by mechanical tests and in vitro cell proliferation assays. Histology of dHACM demonstrated preservation of the native amnion and chorion layers with intact, nonviable cells, collagen, proteoglycan, and elastic fibers distributed in the individual layers. An array of 36 cytokines known to regulate processes involved in inflammation and wound healing were identified in dHACM. When treated with dHACM extracts, bioactivity was demonstrated through an upregulation of basic fibroblast growth factor, granulocyte colony-stimulating factor, and placental growth factor biosynthesis, three growth factors involved in wound healing, by dermal fibroblasts in vitro. After conditioning at temperatures ranging from -78.7 to +73.5°C, dHACM retained its tensile strength and ability to promote proliferation of dermal fibroblasts in vitro. Elution experiments demonstrated a soluble fraction of growth factors that eluted from the tissue and another fraction sequestered within the matrix. The PURION(®) process retains the native composition of ECM and signaling molecules and preserves bioactivity. The array of cytokines preserved in dHACM are in part responsible for its therapeutic efficacy in treating chronic wounds by orchestrating a "symphony of signals" to promote healing. © 2014 Wiley Periodicals, Inc.

Engineered cartilaginous tubes for tracheal tissue replacement via self-assembly and fusion of human mesenchymal stem cell constructs.

PubMed

Dikina, Anna D; Strobel, Hannah A; Lai, Bradley P; Rolle, Marsha W; Alsberg, Eben

2015-06-01

There is a critical need to engineer a neotrachea because currently there are no long-term treatments for tracheal stenoses affecting large portions of the airway. In this work, a modular tracheal tissue replacement strategy was developed. High-cell density, scaffold-free human mesenchymal stem cell-derived cartilaginous rings and tubes were successfully generated through employment of custom designed culture wells and a ring-to-tube assembly system. Furthermore, incorporation of transforming growth factor-β1-delivering gelatin microspheres into the engineered tissues enhanced chondrogenesis with regard to tissue size and matrix production and distribution in the ring- and tube-shaped constructs, as well as luminal rigidity of the tubes. Importantly, all engineered tissues had similar or improved biomechanical properties compared to rat tracheas, which suggests they could be transplanted into a small animal model for airway defects. The modular, bottom up approach used to grow stem cell-based cartilaginous tubes in this report is a promising platform to engineer complex organs (e.g., trachea), with control over tissue size and geometry, and has the potential to be used to generate autologous tissue implants for human clinical applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

Biochemical Fractionation and Stable Isotope Dilution Liquid Chromatography-mass Spectrometry for Targeted and Microdomain-specific Protein Quantification in Human Postmortem Brain Tissue*

PubMed Central

MacDonald, Matthew L.; Ciccimaro, Eugene; Prakash, Amol; Banerjee, Anamika; Seeholzer, Steven H.; Blair, Ian A.; Hahn, Chang-Gyu

2012-01-01

Synaptic architecture and its adaptive changes require numerous molecular events that are both highly ordered and complex. A majority of neuropsychiatric illnesses are complex trait disorders, in which multiple etiologic factors converge at the synapse via many signaling pathways. Investigating the protein composition of synaptic microdomains from human patient brain tissues will yield valuable insights into the interactions of risk genes in many disorders. These types of studies in postmortem tissues have been limited by the lack of proper study paradigms. Thus, it is necessary not only to develop strategies to quantify protein and post-translational modifications at the synapse, but also to rigorously validate them for use in postmortem human brain tissues. In this study we describe the development of a liquid chromatography-selected reaction monitoring method, using a stable isotope-labeled neuronal proteome standard prepared from the brain tissue of a stable isotope-labeled mouse, for the multiplexed quantification of target synaptic proteins in mammalian samples. Additionally, we report the use of this method to validate a biochemical approach for the preparation of synaptic microdomain enrichments from human postmortem prefrontal cortex. Our data demonstrate that a targeted mass spectrometry approach with a true neuronal proteome standard facilitates accurate and precise quantification of over 100 synaptic proteins in mammalian samples, with the potential to quantify over 1000 proteins. Using this method, we found that protein enrichments in subcellular fractions prepared from human postmortem brain tissue were strikingly similar to those prepared from fresh mouse brain tissue. These findings demonstrate that biochemical fractionation methods paired with targeted proteomic strategies can be used in human brain tissues, with important implications for the study of neuropsychiatric disease. PMID:22942359

Use of ex vivo and in vitro cultures of the human respiratory tract to study the tropism and host responses of highly pathogenic avian influenza A (H5N1) and other influenza viruses.

PubMed

Chan, Renee W Y; Chan, Michael C W; Nicholls, John M; Malik Peiris, J S

2013-12-05

The tropism of influenza viruses for the human respiratory tract is a key determinant of host-range, and consequently, of pathogenesis and transmission. Insights can be obtained from clinical and autopsy studies of human disease and relevant animal models. Ex vivo cultures of the human respiratory tract and in vitro cultures of primary human cells can provide complementary information provided they are physiologically comparable in relevant characteristics to human tissues in vivo, e.g. virus receptor distribution, state of differentiation. We review different experimental models for their physiological relevance and summarize available data using these cultures in relation to highly pathogenic avian influenza H5N1, in comparison where relevant, with other influenza viruses. Transformed continuous cell-lines often differ in important ways to the corresponding tissues in vivo. The state of differentiation of primary human cells (respiratory epithelium, macrophages) can markedly affect virus tropism and host responses. Ex vivo cultures of human respiratory tissues provide a close resemblance to tissues in vivo and may be used to risk assess animal viruses for pandemic threat. Physiological factors (age, inflammation) can markedly affect virus receptor expression and virus tropism. Taken together with data from clinical studies on infected humans and relevant animal models, data from ex vivo and in vitro cultures of human tissues and cells can provide insights into virus transmission and pathogenesis and may provide understanding that leads to novel therapeutic interventions. Copyright © 2013 Elsevier B.V. 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.

Inorganic particles in human tissues and their association with neoplastic disease

PubMed Central

Langer, Arthur M.

1974-01-01

An increased gastrointestinal cancer risk is associated with occupational exposure to asbestos fiber. Examination of tissues obtained from extrapulmonary organs of exposed workmen demonstrates the presence of asbestos fibers and bodies. The amount of fiber present in these tissues is many magnitudes less than encountered in the lung tissues from the same individuals. Ingestion of asbestos fiber in some environmental instances may approach in magnitude the amount resulting from occupational exposure. Disease factors are discussed. PMID:4470940

mRNA expression levels of hypoxia-induced and stem cell-associated genes in human glioblastoma.

PubMed

Bache, Matthias; Rot, Swetlana; Keßler, Jacqueline; Güttler, Antje; Wichmann, Henri; Greither, Thomas; Wach, Sven; Taubert, Helge; Söling, Ariane; Bilkenroth, Udo; Kappler, Matthias; Vordermark, Dirk

2015-06-01

The roles of hypoxia-induced and stem cell-associated genes in the development of malignancy and tumour progression are well known. However, there are a limited number of studies analysing the impact of mRNA expression levels of hypoxia-induced and stem cell-associated genes in the tissues of brain tumours and glioblastoma patients. In this study, tumour tissues from patients with glioblastoma multiforme and tumour adjacent tissues were analysed. We investigated mRNA expression levels of hypoxia-inducible factor-1α (HIF-1α), hypoxia-inducible factor-2α (HIF-2α), carbonic anhydrase 9 (CA9), vascular endothelial growth factor (VEGF), glucose transporter-1 (GLUT-1) and osteopontin (OPN), and stem cell-associated genes survivin, epidermal growth factor receptor (EGFR), human telomerase reverse transcriptase (hTERT), Nanog and octamer binding transcription factor 4 (OCT4) using quantitative real-time polymerase chain reaction (qRT-PCR). Our data revealed higher mRNA expression levels of hypoxia-induced and stem cell-associated genes in tumour tissue than levels in the tumour adjacent tissues in patients with glioblastoma multiforme. A strong positive correlation between the mRNA expression levels of HIF-2α, CA9, VEGF, GLUT-1 and OPN suggests a specific hypoxia-associated profile of mRNA expression in glioblastoma multiforme. Additionally, the results indicate the role of stem-cell-related genes in tumour hypoxia. Kaplan-Maier analysis revealed that high mRNA expression levels of hypoxia-induced markers showed a trend towards shorter overall survival in glioblastoma patients (P=0.061). Our data suggest that mRNA expression levels of hypoxia-induced genes are important tumour markers in patients with glioblastoma multiforme.

Insulin-like growth factor-I and growth differentiation factor-5 promote the formation of tissue-engineered human nasal septal cartilage.

PubMed

Alexander, Thomas H; Sage, August B; Chen, Albert C; Schumacher, Barbara L; Shelton, Elliot; Masuda, Koichi; Sah, Robert L; Watson, Deborah

2010-10-01

Tissue engineering of human nasal septal chondrocytes offers the potential to create large quantities of autologous material for use in reconstructive surgery of the head and neck. Culture with recombinant human growth factors may improve the biochemical and biomechanical properties of engineered tissue. The objectives of this study were to (1) perform a high-throughput screen to assess multiple combinations of growth factors and (2) perform more detailed testing of candidates identified in part I. In part I, human nasal septal chondrocytes from three donors were expanded in monolayer with pooled human serum (HS). Cells were then embedded in alginate beads for 2 weeks of culture in medium supplemented with 2% or 10% HS and 1 of 90 different growth factor combinations. Combinations of insulin-like growth factor-I (IGF-1), bone morphogenetic protein (BMP)-2, BMP-7, BMP-13, growth differentiation factor-5 (GDF-5), transforming growth factor β (TGFβ)-2, insulin, and dexamethasone were evaluated. Glycosaminoglycan (GAG) accumulation was measured. A combination of IGF-1 and GDF-5 was selected for further testing based on the results of part I. Chondrocytes from four donors underwent expansion followed by three-dimensional alginate culture for 2 weeks in medium supplemented with 2% or 10% HS with or without IGF-1 and GDF-5. Chondrocytes and their associated matrix were then recovered and cultured for 4 weeks in 12 mm transwells in medium supplemented with 2% or 10% HS with or without IGF-1 and GDF-5 (the same medium used for alginate culture). Biochemical and biomechanical properties of the neocartilage were measured. In part I, GAG accumulation was highest for growth factor combinations including both IGF-1 and GDF-5. In part II, the addition of IGF-1 and GDF-5 to 2% HS resulted in a 12-fold increase in construct thickness compared with 2% HS alone (p < 0.0001). GAG and type II collagen accumulation was significantly higher with IGF-1 and GDF-5. Confined compression modulus was greatest with 2% HS, IGF-1, and GDF-5. Supplementation of medium with IGF-1 and GDF-5 during creation of neocartilage constructs results in increased accumulation of GAG and type II collagen and improved biomechanical properties compared with constructs created without the growth factors.

Tissue regeneration in dentistry: Can salamanders provide insight?

PubMed

Sader, F; Denis, J-F; Roy, S

2018-05-01

The ability to regenerate damaged tissues would be of tremendous benefit for medicine and dentistry. Unfortunately, humans are unable to regenerate tissues such as teeth and fingers or to repair injured spinal cord. With an aging population, health problems are more prominent and dentistry is no exception as loss of bone tissue in the orofacial sphere from periodontal disease is on the rise. Humans can repair oral soft tissues exceptionally well; however, hard tissues, such as bone and teeth, are devoid of the ability to repair well or at all. Fortunately, Mother Nature has solved nearly every problem that we would like to solve for our own benefit and tissue regeneration is no exception. By studying animals that can regenerate, like Axolotls (Mexican salamander), we hope to find ways to stimulate regeneration in humans. We will discuss the role of the transforming growth factor beta cytokines as they are central to wound healing in humans and regeneration in Axolotls. We will also compare wound healing in humans (skin and oral mucosa) to Axolotl skin wound healing and limb regeneration. Finally, we will address the problem of bone regeneration and present results in salamanders which indicate that in order to regenerate bone you need to recruit non-bone cells. Fundamental research, such as the work being performed in animals that can regenerate, offers insight to help understand why some treatments are successful while others fail when it comes to specific tissues such as bones. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Human fetal enterocytes in vitro: modulation of the phenotype by extracellular matrix.

PubMed Central

Sanderson, I R; Ezzell, R M; Kedinger, M; Erlanger, M; Xu, Z X; Pringault, E; Leon-Robine, S; Louvard, D; Walker, W A

1996-01-01

The differentiation of small intestinal epithelial cells may require stimulation by microenvironmental factors in vivo. In this study, the effects of mesenchymal and luminal elements in nonmalignant epithelia] cells isolated from the human fetus were studied in vitro. Enterocytes from the human fetus were cultured and microenvironmental factors were added in stages, each stage more closely approximating the microenvironment in vivo. Four stages were examined: epithelial cells derived on plastic from intestinal culture and grown as a cell clone, the same cells grown on connective tissue support, primary epithelial explants grown on fibroblasts with a laminin base, and primary epithelial explants grown on fibroblasts and laminin with n-butyrate added to the incubation medium. The epithelial cell clone dedifferentiated when grown on plastic; however, the cells expressed cytokeratins and villin as evidence of their epithelial cell origin. Human connective tissue matrix from Engelbreth-Holm-Swarm sarcoma cells (Matrigel) modulated their phenotype: alkaline phosphatase activity increased, microvilli developed on their apical surface, and the profile of insulin-like growth factor binding proteins resembled that secreted by differentiated enterocytes. Epithelial cells taken directly from the human fetus as primary cultures and grown as explants on fibroblasts and laminin expressed greater specific enzyme activities in brush border membrane fractions than the cell clone. These activities were enhanced by the luminal molecule sodium butyrate. Thus the sequential addition of connective tissue and luminal molecules to nonmalignant epithelia] cells in vitro induces a spectrum of changes in the epithelial cell phenotype toward full differentiation. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:8755542

Soft tissue wound healing at teeth, dental implants and the edentulous ridge when using barrier membranes, growth and differentiation factors and soft tissue substitutes.

PubMed

Vignoletti, Fabio; Nunez, Javier; Sanz, Mariano

2014-04-01

To review the biological processes of wound healing following periodontal and periimplant plastic surgery when different technologies are used in a) the coverage of root and implant dehiscences, b) the augmentation of keratinized tissue (KT) and c) the augmentation of soft tissue volume. An electronic search from The National Library of Medicine (MEDLINE-PubMed) was performed: English articles with research focus in oral soft tissue regeneration, providing histological outcomes, either from animal experimental studies or human biopsy material were included. Barrier membranes, enamel matrix derivatives, growth factors, allogeneic and xenogeneic soft tissue substitutes have been used in soft tissue regeneration demonstrating different degrees of regeneration. In root coverage, these technologies were able to improve new attachment, although none has shown complete regeneration. In KT augmentation, tissue-engineered allogenic products and xenogeneic collagen matrixes demonstrated integration within the host connective tissue and promotion of keratinization. In soft tissue augmentation and peri-implant plastic surgery there are no histological data currently available. Soft tissue substitutes, growth differentiation factors demonstrated promising histological results in terms of soft tissue regeneration and keratinization, whereas there is a need for further studies to prove their added value in soft tissue augmentation. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Three-Dimensional Rotating Wall Vessel-Derived Cell Culture Models for Studying Virus-Host Interactions

PubMed Central

Gardner, Jameson K.; Herbst-Kralovetz, Melissa M.

2016-01-01

The key to better understanding complex virus-host interactions is the utilization of robust three-dimensional (3D) human cell cultures that effectively recapitulate native tissue architecture and model the microenvironment. A lack of physiologically-relevant animal models for many viruses has limited the elucidation of factors that influence viral pathogenesis and of complex host immune mechanisms. Conventional monolayer cell cultures may support viral infection, but are unable to form the tissue structures and complex microenvironments that mimic host physiology and, therefore, limiting their translational utility. The rotating wall vessel (RWV) bioreactor was designed by the National Aeronautics and Space Administration (NASA) to model microgravity and was later found to more accurately reproduce features of human tissue in vivo. Cells grown in RWV bioreactors develop in a low fluid-shear environment, which enables cells to form complex 3D tissue-like aggregates. A wide variety of human tissues (from neuronal to vaginal tissue) have been grown in RWV bioreactors and have been shown to support productive viral infection and physiological meaningful host responses. The in vivo-like characteristics and cellular features of the human 3D RWV-derived aggregates make them ideal model systems to effectively recapitulate pathophysiology and host responses necessary to conduct rigorous basic science, preclinical and translational studies. PMID:27834891

Malondialdehyde-Derived Epitopes In Human Skin Result From Acute Exposure To Solar UV And Occur In Nonmelanoma Skin Cancer Tissue

PubMed Central

Williams, Joshua D.; Bermudez, Yira; Park, Sophia L.; Stratton, Steven P.; Uchida, Koji; Hurst, Craig A.; Wondrak, Georg T.

2014-01-01

Cutaneous exposure to solar ultraviolet radiation (UVR) is a causative factor in photoaging and photocarcinogenesis. In human skin, oxidative stress is widely considered a key mechanism underlying the detrimental effects of acute and chronic UVR exposure. The lipid peroxidation product malondialdehyde (MDA) accumulates in tissue under conditions of increased oxidative stress, and the occurrence of MDA-derived protein epitopes, including dihydropyridine-lysine (DHP), has recently been substantiated in human skin. Here we demonstrate for the first time that acute exposure to sub-apoptogenic doses of solar simulated UV light (SSL) causes the formation of free MDA and protein-bound MDA-derived epitopes in cultured human HaCaT keratinocytes and healthy human skin. Immunohistochemical staining revealed that acute exposure to SSL is sufficient to cause an almost twenty-fold increase in general MDA- and specific DHP-epitope content in human skin. When compared to dose-matched solar simulated UVA, complete SSL was more efficient generating both free MDA and MDA-derived epitopes. Subsequent tissue microarray (TMA) analysis revealed the prevalence of MDA- and DHP-epitopes in nonmelanoma skin cancer (NMSC). In squamous cell carcinoma tissue, both MDA- and DHP-epitopes were increased more than three-fold as compared to adjacent normal tissue. Taken together, these date demonstrate the occurrence of MDA-derived epitopes in both solar UVR-exposed healthy human skin and NMSC TMA tissue; however, the potential utility of these epitopes as novel biomarkers of cutaneous photodamage and a functional role in the process of skin photocarcinogenesis remain to be explored. PMID:24584085

Perianal implantation of bioengineered human internal anal sphincter constructs intrinsically innervated with human neural progenitor cells.

PubMed

Raghavan, Shreya; Miyasaka, Eiichi A; Gilmont, Robert R; Somara, Sita; Teitelbaum, Daniel H; Bitar, Khalil N

2014-04-01

The internal anal sphincter (IAS) is a major contributing factor to pressure within the anal canal and is required for maintenance of rectoanal continence. IAS damage or weakening results in fecal incontinence. We have demonstrated that bioengineered, intrinsically innervated, human IAS tissue replacements possess key aspects of IAS physiology, such as the generation of spontaneous basal tone and contraction/relaxation in response to neurotransmitters. The objective of this study is to demonstrate the feasibility of implantation of bioengineered IAS constructs in the perianal region of athymic rats. Human IAS tissue constructs were bioengineered from isolated human IAS circular smooth muscle cells and human enteric neuronal progenitor cells. After maturation of the bioengineered constructs in culture, they were implanted operatively into the perianal region of athymic rats. Platelet-derived growth factor was delivered to the implanted constructs through a microosmotic pump. Implanted constructs were retrieved from the animals 4 weeks postimplantation. Animals tolerated the implantation well, and there were no early postoperative complications. Normal stooling was observed during the implantation period. At harvest, implanted constructs were adherent to the perirectal rat tissue and appeared healthy and pink. Immunohistochemical analysis revealed neovascularization. Implanted smooth muscle cells maintained contractile phenotype. Bioengineered constructs responded in vitro in a tissue chamber to neuronally evoked relaxation in response to electrical field stimulation and vasoactive intestinal peptide, indicating the preservation of neuronal networks. Our results indicate that bioengineered innervated IAS constructs can be used to augment IAS function in an animal model. This is a regenerative medicine based therapy for fecal incontinence that would directly address the dysfunction of the IAS muscle. Copyright © 2014 Mosby, Inc. All rights reserved.

A genome-wide interactome of DNA-associated proteins in the human liver.

PubMed

Ramaker, Ryne C; Savic, Daniel; Hardigan, Andrew A; Newberry, Kimberly; Cooper, Gregory M; Myers, Richard M; Cooper, Sara J

2017-11-01

Large-scale efforts like the ENCODE Project have made tremendous progress in cataloging the genomic binding patterns of DNA-associated proteins (DAPs), such as transcription factors (TFs). However, most chromatin immunoprecipitation-sequencing (ChIP-seq) analyses have focused on a few immortalized cell lines whose activities and physiology differ in important ways from endogenous cells and tissues. Consequently, binding data from primary human tissue are essential to improving our understanding of in vivo gene regulation. Here, we identify and analyze more than 440,000 binding sites using ChIP-seq data for 20 DAPs in two human liver tissue samples. We integrated binding data with transcriptome and phased WGS data to investigate allelic DAP interactions and the impact of heterozygous sequence variation on the expression of neighboring genes. Our tissue-based data set exhibits binding patterns more consistent with liver biology than cell lines, and we describe uses of these data to better prioritize impactful noncoding variation. Collectively, our rich data set offers novel insights into genome function in human liver tissue and provides a valuable resource for assessing disease-related disruptions. © 2017 Ramaker et al.; Published by Cold Spring Harbor Laboratory Press.

Influence of postmortem time on the outcome of blood cultures among cadaveric tissue donors.

PubMed

Saegeman, V; Verhaegen, J; Lismont, D; Verduyckt, B; De Rijdt, T; Ectors, N

2009-02-01

Tissue banks provide tissues of human cadaver donors for transplantation. The maximal time limit for tissue retrieval has been set at 24 h postmortem. This study aimed at evaluating the evidence for this limit from a microbiological point of view. The delay of growth in postmortem blood cultures, the identification of the species isolated and clinical/environmental factors were investigated among 100 potential tissue donors. No significant difference was found in the rate of donors with grown blood cultures within (25/65=38%) compared with after (24/65=37%) 24 h of death. Coagulase-negative staphylococci and gastro-intestinal microorganisms were isolated within and after 24 h of death. Two factors--antimicrobial therapy and "delay before body cooling"--were significantly inversely related with donors' blood culture results. From a microbiological point of view, there is no evidence for avoiding tissue retrieval among donors after 24 h of death.

The use of platelet-rich fibrin combined with periodontal ligament and jaw bone mesenchymal stem cell sheets for periodontal tissue engineering

PubMed Central

Wang, Zhong-Shan; Feng, Zhi-Hong; Wu, Guo-Feng; Bai, Shi-Zhu; Dong, Yan; Chen, Fa-Ming; Zhao, Yi-Min

2016-01-01

Periodontal regeneration involves the restoration of at least three unique tissues: cementum, periodontal ligament tissue (PDL) and alveolar bone tissue. Here, we first isolated human PDL stem cells (PDLSCs) and jaw bone mesenchymal stem cells (JBMSCs). These cells were then induced to form cell sheets using an ascorbic acid-rich approach, and the cell sheet properties, including morphology, thickness and gene expression profile, were compared. Platelet-rich fibrin (PRF) derived from human venous blood was then fabricated into bioabsorbable fibrin scaffolds containing various growth factors. Finally, the in vivo potential of a cell-material construct based on PDLSC sheets, PRF scaffolds and JBMSC sheets to form periodontal tissue was assessed in a nude mouse model. In this model, PDLSC sheet/PRF/JBMSC sheet composites were placed in a simulated periodontal space comprising human treated dentin matrix (TDM) and hydroxyapatite (HA)/tricalcium phosphate (TCP) frameworks. Eight weeks after implantation, the PDLSC sheets tended to develop into PDL-like tissues, while the JBMSC sheets tended to produce predominantly bone-like tissues. In addition, the PDLSC sheet/PRF/JBMSC sheet composites generated periodontal tissue-like structures containing PDL- and bone-like tissues. Further improvements in this cell transplantation design may have the potential to provide an effective approach for future periodontal tissue regeneration. PMID:27324079

Tissue Factor-Factor VII Complex As a Key Regulator of Ovarian Cancer Phenotypes.

PubMed

Koizume, Shiro; Miyagi, Yohei

2015-01-01

Tissue factor (TF) is an integral membrane protein widely expressed in normal human cells. Blood coagulation factor VII (fVII) is a key enzyme in the extrinsic coagulation cascade that is predominantly secreted by hepatocytes and released into the bloodstream. The TF-fVII complex is aberrantly expressed on the surface of cancer cells, including ovarian cancer cells. This procoagulant complex can initiate intracellular signaling mechanisms, resulting in malignant phenotypes. Cancer tissues are chronically exposed to hypoxia. TF and fVII can be induced in response to hypoxia in ovarian cancer cells at the gene expression level, leading to the autonomous production of the TF-fVII complex. Here, we discuss the roles of the TF-fVII complex in the induction of malignant phenotypes in ovarian cancer cells. The hypoxic nature of ovarian cancer tissues and the roles of TF expression in endometriosis are discussed. Arguments will be extended to potential strategies to treat ovarian cancers based on our current knowledge of TF-fVII function.

Fell-Muir lecture: connective tissue growth factor (CCN2) – a pernicious and pleiotropic player in the development of kidney fibrosis

PubMed Central

Mason, Roger M

2013-01-01

Connective tissue growth factor (CTGF, CCN2) is a member of the CCN family of matricellular proteins. It interacts with many other proteins, including plasma membrane proteins, modulating cell function. It is expressed at low levels in normal adult kidney cells but is increased in kidney diseases, playing important roles in inflammation and in the development of glomerular and interstitial fibrosis in chronic disease. This review reports the evidence for its expression in human and animal models of chronic kidney disease and summarizes data showing that anti-CTGF therapy can successfully attenuate fibrotic changes in several such models, suggesting that therapies targeting CTGF and events downstream of it in renal cells may be useful for the treatment of human kidney fibrosis. Connective tissue growth factor stimulates the development of fibrosis in the kidney in many ways including activating cells to increase extracellular matrix synthesis, inducing cell cycle arrest and hypertrophy, and prolonging survival of activated cells. The relationship between CTGF and the pro-fibrotic factor TGFβ is examined and mechanisms by which CTGF promotes signalling by the latter are discussed. No specific cellular receptors for CTGF have been discovered but it interacts with and activates several plasma membrane proteins including low-density lipoprotein receptor-related protein (LRP)-1, LRP-6, tropomyosin-related kinase A, integrins and heparan sulphate proteoglycans. Intracellular signalling and downstream events triggered by such interactions are reviewed. Finally, the relationships between CTGF and several anti-fibrotic factors, such as bone morphogenetic factor-4 (BMP4), BMP7, hepatocyte growth factor, CCN3 and Oncostatin M, are discussed. These may determine whether injured tissue heals or progresses to fibrosis. PMID:23110747

Analysis of the N-glycans of recombinant human Factor IX purified from transgenic pig milk

PubMed Central

Gil, Geun-Cheol; Velander, William H; Van Cott, Kevin E

2008-01-01

Glycosylation of recombinant proteins is of particular importance because it can play significant roles in the clinical properties of the glycoprotein. In this work, the N-glycan structures of recombinant human Factor IX (tg-FIX) produced in the transgenic pig mammary gland were determined. The majority of the N-glycans of transgenic pig-derived Factor IX (tg-FIX) are complex, bi-antennary with one or two terminal N-acetylneuraminic acid (Neu5Ac) moieties. We also found that the N-glycan structures of tg-FIX produced in the porcine mammary epithelial cells differed with respect to N-glycans from glycoproteins produced in other porcine tissues. tg-FIX contains no detectable Neu5Gc, the sialic acid commonly found in porcine glycoproteins produced in other tissues. Additionally, we were unable to detect glycans in tg-FIX that have a terminal Galα(1,3)Gal disaccharide sequence, which is strongly antigenic in humans. The N-glycan structures of tg-FIX are also compared to the published N-glycan structures of recombinant human glycoproteins produced in other transgenic animal species. While tg-FIX contains only complex structures, antithrombin III (goat), C1 inhibitor (rabbit), and lactoferrin (cow) have both high mannose and complex structures. Collectively, these data represent a beginning point for the future investigation of species-specific and tissue/cell-specific differences in N-glycan structures among animals used for transgenic animal bioreactors. PMID:18456721

[Fluorides in the human bones – selected issues].

PubMed

Palczewska-Komsa, Mirona; Kalisińska, Elżbieta; Stogiera, Anna; Szmidt, Monika

Long -term intake of luoride leads to skeletal luorosis. The toxicity of luoride, not only for the human body, but also the entire ecosystem makes it necessary to constantly monitor their content in the environment. Accordingly, there is a need to control the level of luorides (F⁻) in humans, particularly in bone tissue, which relects long -term accumulation of these compounds. The aim of the study was to determine the concentration of luoride in the human bones depending on biological factors and environmental conditions on the basis of the published literature. Given the importance of bone tissue as the main reservoir of luoride ions is an important issue to continue to monitor the concentration of F⁻ in this tissue, particularly for people living in the polluted environment luorine compounds. There are numerous works on concentrations of this element in human bones in world literature which proves the great interest in the subject. It should be underlined the need for further study of this issue for people living in different regions of Poland.

High expression of arachidonate 15-lipoxygenase and proinflammatory markers in human ischemic heart tissue

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

Magnusson, Lisa U.; Lundqvist, Annika; Asp, Julia

Highlights: Black-Right-Pointing-Pointer We found a 17-fold upregulation of ALOX15 in the ischemic heart. Black-Right-Pointing-Pointer Incubation of human muscle cells in hypoxia showed a 22-fold upregulation of ALOX15. Black-Right-Pointing-Pointer We observed increased levels of proinflammatory markers in ischemic heart tissue. Black-Right-Pointing-Pointer Suggesting a link between ischemia and inflammation in ischemic heart biopsies. -- Abstract: A common feature of the ischemic heart and atherosclerotic plaques is the presence of hypoxia (insufficient levels of oxygen in the tissue). Hypoxia has pronounced effects on almost every aspect of cell physiology, and the nuclear transcription factor hypoxia inducible factor-1{alpha} (HIF-1{alpha}) regulates adaptive responses to lowmore » concentrations of oxygen in mammalian cells. In our recent work, we observed that hypoxia increases the proinflammatory enzyme arachidonate 15-lipoxygenase (ALOX15B) in human carotid plaques. ALOX15 has recently been shown to be present in the human myocardium, but the effect of ischemia on its expression has not been investigated. Here we test the hypothesis that ischemia of the heart leads to increased expression of ALOX15, and found an almost 2-fold increase in HIF-1{alpha} mRNA expression and a 17-fold upregulation of ALOX15 mRNA expression in the ischemic heart biopsies from patients undergoing coronary bypass surgery compared with non ischemic heart tissue. To investigate the effect of low oxygen concentration on ALOX15 we incubated human vascular muscle cells in hypoxia and showed that expression of ALOX15 increased 22-fold compared with cells incubated in normoxic conditions. We also observed increased mRNA levels of proinflammatory markers in ischemic heart tissue compared with non-ischemic controls. In summary, we demonstrate increased ALOX15 in human ischemic heart biopsies. Furthermore we demonstrate that hypoxia increases ALOX15 in human muscle cells. Our results yield important insights into the underlying association between hypoxia and inflammation in the human ischemic heart disease.« less

Expansion and redifferentiation of chondrocytes from osteoarthritic cartilage: cells for human cartilage tissue engineering.

PubMed

Hsieh-Bonassera, Nancy D; Wu, Iwen; Lin, Jonathan K; Schumacher, Barbara L; Chen, Albert C; Masuda, Koichi; Bugbee, William D; Sah, Robert L

2009-11-01

To determine if selected culture conditions enhance the expansion and redifferentiation of chondrocytes isolated from human osteoarthritic cartilage with yields appropriate for creation of constructs for treatment of joint-scale cartilage defects, damage, or osteoarthritis. Chondrocytes isolated from osteoarthritic cartilage were analyzed to determine the effects of medium supplement on cell expansion in monolayer and then cell redifferentiation in alginate beads. Expansion was assessed as cell number estimated from DNA, growth rate, and day of maximal growth. Redifferentiation was evaluated quantitatively from proteoglycan and collagen type II content, and qualitatively by histology and immunohistochemistry. Using either serum or a growth factor cocktail (TFP: transforming growth factor beta1, fibroblast growth factor 2, and platelet-derived growth factor type bb), cell growth rate in monolayer was increased to 5.5x that of corresponding conditions without TFP, and cell number increased 100-fold within 17 days. In subsequent alginate bead culture with human serum or transforming growth factor beta1 and insulin-transferrin-selenium-linoleic acid-bovine serum albumin, redifferentiation was enhanced with increased proteoglycan and collagen type II production. Effects of human serum were dose dependent, and 5% or higher induced formation of chondron-like structures with abundant proteoglycan-rich matrix. Chondrocytes from osteoarthritic cartilage can be stimulated to undergo 100-fold expansion and then redifferentiation, suggesting that they may be useful as a cell source for joint-scale cartilage tissue engineering.

The human tissue-biomaterial interface: a role for PPARγ-dependent glucocorticoid receptor activation in regulating the CD163+ M2 macrophage phenotype.

PubMed

Bullers, Samuel J; Baker, Simon C; Ingham, Eileen; Southgate, Jennifer

2014-09-01

In vivo studies of implanted acellular biological scaffolds in experimental animals have shown constructive remodeling mediated by anti-inflammatory macrophages. Little is known about the human macrophage response to such biomaterials, or the nature of the signaling mechanisms that govern the macrophage phenotype in this environment. The cellular events at the interface of a tissue and implanted decellularized biomaterial were examined by establishing a novel ex vivo tissue culture model in which surgically excised human urinary tract tissue was combined with porcine acellular bladder matrix (PABM). Evaluation of the tissue-biomaterial interface showed a time-dependent infiltration of the biomaterial by CD68(+) CD80(-) macrophages. The migration of CD68(+) cells from the tissue to the interface was accompanied by maturation to a CD163(hi) phenotype, suggesting that factor(s) associated with the biomaterial or the wound edge was/were responsible for the active recruitment and polarization of local macrophages. Glucocorticoid receptor (GR) and peroxisome proliferator activated receptor gamma (PPARγ) signaling was investigated as candidate pathways for integrating inflammatory responses; both showed intense nuclear labeling in interface macrophages. GR and PPARγ activation polarized peripheral blood-derived macrophages from a default M1 (CD80(+)) toward an M2 (CD163(+)) phenotype, but PPARγ signaling predominated, as its antagonism blocked any GR-mediated effect. Seeding on PABM was effective at polarizing peripheral blood-derived macrophages from a default CD80(+) phenotype on glass to a CD80(-) phenotype, with intense nuclear localization of PPARγ. These results endorse in vivo observations that the infiltration of decellularized biological scaffolds, exemplified here by PABM, is pioneered by macrophages. Thus, it appears that natural factors present in PABM are involved in the active recruitment and polarization of macrophages to a CD163(+) phenotype, with activation of PPARγ identified as the candidate pathway. The harnessing of these natural matrix-associated factors may be useful in enhancing the integration of synthetic and other natural biomaterials by polarizing macrophage activation toward an M2 regulatory phenotype.

Platelet lysate-based pro-angiogenic nanocoatings.

PubMed

Oliveira, Sara M; Pirraco, Rogério P; Marques, Alexandra P; Santo, Vítor E; Gomes, Manuela E; Reis, Rui L; Mano, João F

2016-03-01

Human platelet lysate (PL) is a cost-effective and human source of autologous multiple and potent pro-angiogenic factors, such as vascular endothelial growth factor A (VEGF A), fibroblast growth factor b (FGF b) and angiopoietin-1. Nanocoatings previously characterized were prepared by layer-by-layer assembling incorporating PL with marine-origin polysaccharides and were shown to activate human umbilical vein endothelial cells (HUVECs). Within 20 h of incubation, the more sulfated coatings induced the HUVECS to the form tube-like structures accompanied by an increased expression of angiogenic-associated genes, such as angiopoietin-1 and VEGF A. This may be a cost-effective approach to modify 2D/3D constructs to instruct angiogenic cells towards the formation of neo-vascularization, driven by multiple and synergistic stimulations from the PL combined with sulfated polysaccharides. The presence, or fast induction, of a stable and mature vasculature inside 3D constructs is crucial for new tissue formation and its viability. This has been one of the major tissue engineering challenges, limiting the dimensions of efficient tissue constructs. Many approaches based on cells, growth factors, 3D bioprinting and channel incorporation have been proposed. Herein, we explored a versatile technique, layer-by-layer assembling in combination with platelet lysate (PL), that is a cost-effective source of many potent pro-angiogenic proteins and growth factors. Results suggest that the combination of PL with sulfated polyelectrolytes might be used to introduce interfaces onto 2D/3D constructs with potential to induce the formation of cell-based tubular structures. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Two-way learning with one-way supervision for gene expression data.

PubMed

Wong, Monica H T; Mutch, David M; McNicholas, Paul D

2017-03-04

A family of parsimonious Gaussian mixture models for the biclustering of gene expression data is introduced. Biclustering is accommodated by adopting a mixture of factor analyzers model with a binary, row-stochastic factor loadings matrix. This particular form of factor loadings matrix results in a block-diagonal covariance matrix, which is a useful property in gene expression analyses, specifically in biomarker discovery scenarios where blood can potentially act as a surrogate tissue for other less accessible tissues. Prior knowledge of the factor loadings matrix is useful in this application and is reflected in the one-way supervised nature of the algorithm. Additionally, the factor loadings matrix can be assumed to be constant across all components because of the relationship desired between the various types of tissue samples. Parameter estimates are obtained through a variant of the expectation-maximization algorithm and the best-fitting model is selected using the Bayesian information criterion. The family of models is demonstrated using simulated data and two real microarray data sets. The first real data set is from a rat study that investigated the influence of diabetes on gene expression in different tissues. The second real data set is from a human transcriptomics study that focused on blood and immune tissues. The microarray data sets illustrate the biclustering family's performance in biomarker discovery involving peripheral blood as surrogate biopsy material. The simulation studies indicate that the algorithm identifies the correct biclusters, most optimally when the number of observation clusters is known. Moreover, the biclustering algorithm identified biclusters comprised of biologically meaningful data related to insulin resistance and immune function in the rat and human real data sets, respectively. Initial results using real data show that this biclustering technique provides a novel approach for biomarker discovery by enabling blood to be used as a surrogate for hard-to-obtain tissues.

Expression of ribosomopathy genes during Xenopus tropicalis embryogenesis.

PubMed

Robson, Andrew; Owens, Nick D L; Baserga, Susan J; Khokha, Mustafa K; Griffin, John N

2016-10-26

Because ribosomes are ubiquitously required for protein production, it was long assumed that any inherited defect in ribosome manufacture would be embryonically lethal. However, several human congenital diseases have been found to be associated with mutations in ribosome biogenesis factors. Surprisingly, despite the global requirement for ribosomes, these "ribosomopathies" are characterized by distinct and tissue specific phenotypes. The reasons for such tissue proclivity in ribosomopathies remain mysterious but may include differential expression of ribosome biogenesis factors in distinct tissues. Here we use in situ hybridization of labeled antisense mRNA probes and ultra high temporal resolution RNA-Seq data to examine and compare expression of 13 disease associated ribosome biogenesis factors at six key stages in Xenopus tropicalis development. Rather than being ubiquitously expressed during development, mRNAs of all examined ribosome biogenesis factors were highly enriched in specific tissues, including the cranial neural crest and ventral blood islands. Interestingly, expression of ribosome biogenesis factors demonstrates clear differences in timing, transcript number and tissue localization. Ribosome biogenesis factor expression is more spatiotemporally regulated during embryonic development than previously expected and correlates closely with many of the common ribosomopathy phenotypes. Our findings provide information on the dynamic use of ribosome production machinery components during development and advance our understanding of their roles in disease.

Bio-Imaging of Colorectal Cancer Models Using Near Infrared Labeled Epidermal Growth Factor

PubMed Central

Cohen, Gadi; Lecht, Shimon; Arien-Zakay, Hadar; Ettinger, Keren; Amsalem, Orit; Oron-Herman, Mor; Yavin, Eylon; Prus, Diana; Benita, Simon; Nissan, Aviram; Lazarovici, Philip

2012-01-01

Novel strategies that target the epidermal growth factor receptor (EGFR) have led to the clinical development of monoclonal antibodies, which treat metastatic colorectal cancer (mCRC) but only subgroups of patients with increased wild type KRAS and EGFR gene copy, respond to these agents. Furthermore, resistance to EGFR blockade inevitably occurred, making future therapy difficult. Novel bio-imaging (BOI) methods may assist in quantization of EGFR in mCRC tissue thus complementing the immunohistochemistry methodology, in guiding the future treatment of these patients. The aim of the present study was to explore the usefulness of near infrared-labeled EGF (EGF-NIR) for bio-imaging of CRC using in vitro and in vivo orthotopic tumor CRC models and ex vivo human CRC tissues. We describe the preparation and characterization of EGF-NIR and investigate binding, using BOI of a panel of CRC cell culture models resembling heterogeneity of human CRC tissues. EGF-NIR was specifically and selectively bound by EGFR expressing CRC cells, the intensity of EGF-NIR signal to background ratio (SBR) reflected EGFR levels, dose-response and time course imaging experiments provided optimal conditions for quantization of EGFR levels by BOI. EGF-NIR imaging of mice with HT-29 orthotopic CRC tumor indicated that EGF-NIR is more slowly cleared from the tumor and the highest SBR between tumor and normal adjacent tissue was achieved two days post-injection. Furthermore, images of dissected tissues demonstrated accumulation of EGF-NIR in the tumor and liver. EGF-NIR specifically and strongly labeled EGFR positive human CRC tissues while adjacent CRC tissue and EGFR negative tissues expressed weak NIR signals. This study emphasizes the use of EGF-NIR for preclinical studies. Combined with other methods, EGF-NIR could provide an additional bio-imaging specific tool in the standardization of measurements of EGFR expression in CRC tissues. PMID:23144978

Bio-imaging of colorectal cancer models using near infrared labeled epidermal growth factor.

PubMed

Cohen, Gadi; Lecht, Shimon; Arien-Zakay, Hadar; Ettinger, Keren; Amsalem, Orit; Oron-Herman, Mor; Yavin, Eylon; Prus, Diana; Benita, Simon; Nissan, Aviram; Lazarovici, Philip

2012-01-01

Novel strategies that target the epidermal growth factor receptor (EGFR) have led to the clinical development of monoclonal antibodies, which treat metastatic colorectal cancer (mCRC) but only subgroups of patients with increased wild type KRAS and EGFR gene copy, respond to these agents. Furthermore, resistance to EGFR blockade inevitably occurred, making future therapy difficult. Novel bio-imaging (BOI) methods may assist in quantization of EGFR in mCRC tissue thus complementing the immunohistochemistry methodology, in guiding the future treatment of these patients. The aim of the present study was to explore the usefulness of near infrared-labeled EGF (EGF-NIR) for bio-imaging of CRC using in vitro and in vivo orthotopic tumor CRC models and ex vivo human CRC tissues. We describe the preparation and characterization of EGF-NIR and investigate binding, using BOI of a panel of CRC cell culture models resembling heterogeneity of human CRC tissues. EGF-NIR was specifically and selectively bound by EGFR expressing CRC cells, the intensity of EGF-NIR signal to background ratio (SBR) reflected EGFR levels, dose-response and time course imaging experiments provided optimal conditions for quantization of EGFR levels by BOI. EGF-NIR imaging of mice with HT-29 orthotopic CRC tumor indicated that EGF-NIR is more slowly cleared from the tumor and the highest SBR between tumor and normal adjacent tissue was achieved two days post-injection. Furthermore, images of dissected tissues demonstrated accumulation of EGF-NIR in the tumor and liver. EGF-NIR specifically and strongly labeled EGFR positive human CRC tissues while adjacent CRC tissue and EGFR negative tissues expressed weak NIR signals. This study emphasizes the use of EGF-NIR for preclinical studies. Combined with other methods, EGF-NIR could provide an additional bio-imaging specific tool in the standardization of measurements of EGFR expression in CRC tissues.

Advancing osteochondral tissue engineering: bone morphogenetic protein, transforming growth factor, and fibroblast growth factor signaling drive ordered differentiation of periosteal cells resulting in stable cartilage and bone formation in vivo.

PubMed

Mendes, L F; Katagiri, H; Tam, W L; Chai, Y C; Geris, L; Roberts, S J; Luyten, F P

2018-02-21

Chondrogenic mesenchymal stem cells (MSCs) have not yet been used to address the clinical demands of large osteochondral joint surface defects. In this study, self-assembling tissue intermediates (TIs) derived from human periosteum-derived stem/progenitor cells (hPDCs) were generated and validated for stable cartilage formation in vivo using two different animal models. hPDCs were aggregated and cultured in the presence of a novel growth factor (GF) cocktail comprising of transforming growth factor (TGF)-β1, bone morphogenetic protein (BMP)2, growth differentiation factor (GDF)5, BMP6, and fibroblast growth factor (FGF)2. Quantitative polymerase chain reaction (PCR) and immunohistochemistry were used to study in vitro differentiation. Aggregates were then implanted ectopically in nude mice and orthotopically in critical-size osteochondral defects in nude rats and evaluated by microcomputed tomography (µCT) and immunohistochemistry. Gene expression analysis after 28 days of in vitro culture revealed the expression of early and late chondrogenic markers and a significant upregulation of NOGGIN as compared to human articular chondrocytes (hACs). Histological examination revealed a bilayered structure comprising of chondrocytes at different stages of maturity. Ectopically, TIs generated both bone and mineralized cartilage at 8 weeks after implantation. Osteochondral defects treated with TIs displayed glycosaminoglycan (GAG) production, type-II collagen, and lubricin expression. Immunostaining for human nuclei protein suggested that hPDCs contributed to both subchondral bone and articular cartilage repair. Our data indicate that in vitro derived osteochondral-like tissues can be generated from hPDCs, which are capable of producing bone and cartilage ectopically and behave orthotopically as osteochondral units.

Lemongrass (Cymbopogon flexuosus) essential oil demonstrated anti-inflammatory effect in pre-inflamed human dermal fibroblasts.

PubMed

Han, Xuesheng; Parker, Tory L

2017-06-01

Lemongrass ( Cymbopogon flexuosus ) essential oil (LEO), which has citral as its main component, has exhibited anti-inflammatory effect in both animal and human cells. In this study, we evaluated the anti-inflammatory activity of a commercially available LEO in pre-inflamed human dermal fibroblasts. We first studied the impact of LEO on 17 protein biomarkers that are critically associated with inflammation and tissue remodeling. LEO significantly inhibited production of the inflammatory biomarkers vascular cell adhesion molecule 1 (VCAM-1), interferon gamma-induced protein 10 (IP-10), interferon-inducible T-cell alpha chemoattractant (I-TAC), and monokine induced by gamma interferon (MIG); decreased levels of the tissue remodeling biomarkers collagen-I and III, epidermal growth factor receptor (EGFR), and plasminogen activator inhibitor (PAI-1); and inhibited the immunomodulatory biomarker macrophage colony-stimulating factor (M-CSF). Furthermore, we studied the impact of LEO on genome-wide gene expression profiles. LEO significantly modulated global gene expression and robustly impacted signaling pathways, many of which are critical for inflammation and tissue remodeling processes. This study provides the first evidence of the anti-inflammatory activity of LEO in human skin cells and indicates that it is a good therapeutic candidate for treating inflammatory conditions of the skin.

Breast Cancer Cell Colonization of the Human Bone Marrow Adipose Tissue Niche.

PubMed

Templeton, Zach S; Lie, Wen-Rong; Wang, Weiqi; Rosenberg-Hasson, Yael; Alluri, Rajiv V; Tamaresis, John S; Bachmann, Michael H; Lee, Kitty; Maloney, William J; Contag, Christopher H; King, Bonnie L

2015-12-01

Bone is a preferred site of breast cancer metastasis, suggesting the presence of tissue-specific features that attract and promote the outgrowth of breast cancer cells. We sought to identify parameters of human bone tissue associated with breast cancer cell osteotropism and colonization in the metastatic niche. Migration and colonization patterns of MDA-MB-231-fLuc-EGFP (luciferase-enhanced green fluorescence protein) and MCF-7-fLuc-EGFP breast cancer cells were studied in co-culture with cancellous bone tissue fragments isolated from 14 hip arthroplasties. Breast cancer cell migration into tissues and toward tissue-conditioned medium was measured in Transwell migration chambers using bioluminescence imaging and analyzed as a function of secreted factors measured by multiplex immunoassay. Patterns of breast cancer cell colonization were evaluated with fluorescence microscopy and immunohistochemistry. Enhanced MDA-MB-231-fLuc-EGFP breast cancer cell migration to bone-conditioned versus control medium was observed in 12/14 specimens (P = .0014) and correlated significantly with increasing levels of the adipokines/cytokines leptin (P = .006) and IL-1β (P = .001) in univariate and multivariate regression analyses. Fluorescence microscopy and immunohistochemistry of fragments underscored the extreme adiposity of adult human bone tissues and revealed extensive breast cancer cell colonization within the marrow adipose tissue compartment. Our results show that breast cancer cells migrate to human bone tissue-conditioned medium in association with increasing levels of leptin and IL-1β, and colonize the bone marrow adipose tissue compartment of cultured fragments. Bone marrow adipose tissue and its molecular signals may be important but understudied components of the breast cancer metastatic niche. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Cell biology and biotechnology research for exploration of the Moon and Mars

NASA Astrophysics Data System (ADS)

Pellis, N.; North, R.

Health risks generated by human long exposure to radiation, microgravity, and unknown factors in the planetary environment are the major unresolved issues for human space exploration. A complete characterization of human and other biological systems adaptation processes to long-duration space missions is necessary for the development of countermeasures. The utilization of cell and engineered tissue cultures in space research and exploration complements research in human, animal, and plant subjects. We can bring a small number of humans, animals, or plants to the ISS, Moon, and Mars. However, we can investigate millions of their cells during these missions. Furthermore, many experiments can not be performed on humans, e.g. radiation exposure, cardiac muscle. Cells from critical tissues and tissue constructs per se are excellent subjects for experiments that address underlying mechanisms important to countermeasures. The development of cell tissue engineered for replacement, implantation of biomaterial to induce tissue regeneration (e.g. absorbable collagen matrix for guiding tissue regeneration in periodontal surgery), and immunoisolation (e.g. biopolymer coating on transplanted tissues to ward off immunological rejection) are good examples of cell research and biotechnology applications. NASA Cell Biology and Biotechnology research include Bone/Muscle and Cardiovascular cell culture and tissue engineering; Environmental Health and Life Support Systems; Immune System; Radiation; Gravity Thresholds ; and Advanced Biotechnology Development to increase the understanding of animal and plant cell adaptive behavior when exposed to space, and to advance technologies that facilitates exploration. Cell systems can be used to investigate processes related to food, microbial proliferation, waste management, biofilms and biomaterials. The NASA Cell Science Program has the advantage of conducting research in microgravity based on significantly small resources, and the ability to conduct experiments in the early phase of the development of requirements for exploration. Supporting the NASA concept of stepping stones, we believe that ground based, International Space Station, robotic and satellite missions offer the ideal environment to perform experiments and secure answers necessary for human exploration.

[Insulin-like growth factor-1 (IGF-1) - structure and the role in the human body].

PubMed

Filus, Alicja; Zdrojewicz, Zygmunt

2015-01-01

In the recent years, managed to broadly explore the structure and role of insulin-like growth factors type 1 and 2 (IGF1 I 2). They belong to the structure of polypeptide hormones homologous to proinsulin. They are characterized by a wide range of activities. IGF-1 is a key mediator of most tissue effects of growth hormone (GH). In addition to effects on growth processes of the body, is also an important factor for cell homeostasis, is subject to both endocrine and tissue-specific auto- and paracrine regulation. In this paper, the current, general knowledge on the structure, function and mechanism of biological effects of IGF-1 in the human body was presented. Attention was also drawn to the directions of use of IGf-1 in the treatment of other diseases than the diseases of the hypothalamic-pituitary and growth disorders in children. © Polish Society for Pediatric Endocrinology and Diabetology.

A convex optimization approach for identification of human tissue-specific interactomes.

PubMed

Mohammadi, Shahin; Grama, Ananth

2016-06-15

Analysis of organism-specific interactomes has yielded novel insights into cellular function and coordination, understanding of pathology, and identification of markers and drug targets. Genes, however, can exhibit varying levels of cell type specificity in their expression, and their coordinated expression manifests in tissue-specific function and pathology. Tissue-specific/tissue-selective interaction mechanisms have significant applications in drug discovery, as they are more likely to reveal drug targets. Furthermore, tissue-specific transcription factors (tsTFs) are significantly implicated in human disease, including cancers. Finally, disease genes and protein complexes have the tendency to be differentially expressed in tissues in which defects cause pathology. These observations motivate the construction of refined tissue-specific interactomes from organism-specific interactomes. We present a novel technique for constructing human tissue-specific interactomes. Using a variety of validation tests (Edge Set Enrichment Analysis, Gene Ontology Enrichment, Disease-Gene Subnetwork Compactness), we show that our proposed approach significantly outperforms state-of-the-art techniques. Finally, using case studies of Alzheimer's and Parkinson's diseases, we show that tissue-specific interactomes derived from our study can be used to construct pathways implicated in pathology and demonstrate the use of these pathways in identifying novel targets. http://www.cs.purdue.edu/homes/mohammas/projects/ActPro.html mohammadi@purdue.edu. © The Author 2016. Published by Oxford University Press.

Coupled Analysis of In Vitro and Histology Tissue Samples to Quantify Structure-Function Relationship

PubMed Central

Acar, Evrim; Plopper, George E.; Yener, Bülent

2012-01-01

The structure/function relationship is fundamental to our understanding of biological systems at all levels, and drives most, if not all, techniques for detecting, diagnosing, and treating disease. However, at the tissue level of biological complexity we encounter a gap in the structure/function relationship: having accumulated an extraordinary amount of detailed information about biological tissues at the cellular and subcellular level, we cannot assemble it in a way that explains the correspondingly complex biological functions these structures perform. To help close this information gap we define here several quantitative temperospatial features that link tissue structure to its corresponding biological function. Both histological images of human tissue samples and fluorescence images of three-dimensional cultures of human cells are used to compare the accuracy of in vitro culture models with their corresponding human tissues. To the best of our knowledge, there is no prior work on a quantitative comparison of histology and in vitro samples. Features are calculated from graph theoretical representations of tissue structures and the data are analyzed in the form of matrices and higher-order tensors using matrix and tensor factorization methods, with a goal of differentiating between cancerous and healthy states of brain, breast, and bone tissues. We also show that our techniques can differentiate between the structural organization of native tissues and their corresponding in vitro engineered cell culture models. PMID:22479315

Inspiration from heart development: Biomimetic development of functional human cardiac organoids.

PubMed

Richards, Dylan J; Coyle, Robert C; Tan, Yu; Jia, Jia; Wong, Kerri; Toomer, Katelynn; Menick, Donald R; Mei, Ying

2017-10-01

Recent progress in human organoids has provided 3D tissue systems to model human development, diseases, as well as develop cell delivery systems for regenerative therapies. While direct differentiation of human embryoid bodies holds great promise for cardiac organoid production, intramyocardial cell organization during heart development provides biological foundation to fabricate human cardiac organoids with defined cell types. Inspired by the intramyocardial organization events in coronary vasculogenesis, where a diverse, yet defined, mixture of cardiac cell types self-organizes into functional myocardium in the absence of blood flow, we have developed a defined method to produce scaffold-free human cardiac organoids that structurally and functionally resembled the lumenized vascular network in the developing myocardium, supported hiPSC-CM development and possessed fundamental cardiac tissue-level functions. In particular, this development-driven strategy offers a robust, tunable system to examine the contributions of individual cell types, matrix materials and additional factors for developmental insight, biomimetic matrix composition to advance biomaterial design, tissue/organ-level drug screening, and cell therapy for heart repair. Copyright © 2017 Elsevier Ltd. All rights reserved.

Biological characterization of soft tissue sarcomas.

PubMed

Hayashi, Takuma; Horiuchi, Akiko; Sano, Kenji; Kanai, Yae; Yaegashi, Nobuo; Aburatani, Hiroyuki; Konishi, Ikuo

2015-12-01

Soft tissue sarcomas are neoplastic malignancies that typically arise in tissues of mesenchymal origin. The identification of novel molecular mechanisms leading to mesenchymal transformation and the establishment of new therapies and diagnostic biomarker has been hampered by several critical factors. First, malignant soft tissue sarcomas are rarely observed in the clinic with fewer than 15,000 newly cases diagnosed each year in the United States. Another complicating factor is that soft tissue sarcomas are extremely heterogeneous as they arise in a multitude of tissues from many different cell lineages. The scarcity of clinical materials coupled with its inherent heterogeneity creates a challenging experimental environment for clinicians and scientists. Faced with these challenges, there has been extremely limited advancement in clinical treatment options available to patients as compared to other malignant tumours. In order to glean insight into the pathobiology of soft tissue sarcomas, scientists are now using mouse models whose genomes have been specifically tailored to carry gene deletions, gene amplifications, and somatic mutations commonly observed in human soft tissue sarcomas. The use of these model organisms has been successful in increasing our knowledge and understanding of how alterations in relevant oncogenic and/or tumour suppressive signal cascades, i.e., interferon-γ (IFN-γ), tumour protein 53 (TP53) and/or retinoblastoma (RB) pathway directly impact sarcomagenesis. It is the goal of many in the physiological community that the use of several mouse models will serve as powerful in vivo tools for further understanding of sarcomagenesis and potentially identify new diagnostic biomarker and therapeutic strategies against human soft tissue sarcomas.

Negative regulatory role of PI3-kinase in TNF-induced tumor necrosis.

PubMed

Matschurat, Susanne; Blum, Sabine; Mitnacht-Kraus, Rita; Dijkman, Henry B P M; Kanal, Levent; De Waal, Robert M W; Clauss, Matthias

2003-10-20

Tissue factor is the prime initiator of blood coagulation. Expression of tissue factor in tumor endothelial cells leads to thrombus formation, occlusion of vessels and development of hemorrhagic infarctions in the tumor tissue, often followed by regression of the tumor. Tumor cells produce endogenous vascular endothelial growth factor (VEGF), which sensitizes endothelial cells for systemically administered tumor necrosis factor alpha (TNF alpha) and synergistically enhances the TNF-induced expression of tissue factor. We have analyzed the pathways involved in the induction of tissue factor in human umbilical cord vein endothelial cells (HUVECs) after combined stimulation with TNF and VEGF. By using specific low molecular weight inhibitors, we demonstrated that protein kinase C (PKC), p44/42 and p38 mitogen-activated protein (MAP) kinases, and stress-activated protein kinase (JNK) are essentially involved in the induction of tissue factor. In contrast, the application of wortmannin, an inhibitor of phosphatidylinositol 3 (PI3)-kinase, led to strongly enhanced expression of tissue factor in TNF- and VEGF-treated cells, implicating a negative regulatory role for PI3-kinase. In vivo, the application of wortmannin promoted the formation of TNF-induced hemorrhages and intratumoral necroses in murine meth A tumors. The co-injection of wortmannin lowered the effective dose of applied TNF. Therefore, it is conceivable that the treatment of TNF-sensitive tumors with a combination of TNF and wortmannin will ensure the selective damage of the tumor endothelium and minimize the risk of systemic toxicity of TNF. TNF-treatment in combination with specific inhibition of PI3-kinase is a novel concept in anti-cancer therapy. Copyright 2003 Wiley-Liss, Inc.

Root cementum may modulate gene expression during periodontal regeneration: a preliminary study in humans.

PubMed

Gonçalves, Patricia F; Lima, Liana L; Sallum, Enilson A; Casati, Márcio Z; Nociti, Francisco H

2008-02-01

Previous data demonstrated that root cementum may affect periodontal regeneration. As such, this study aimed to explore further possible mechanisms involved in this process by investigating in humans whether root cementum modulates gene expression in the regenerating tissue formed under membrane-protected intrabony defects. Thirty subjects with deep intrabony defects (> or =5 mm; 2- or 3-wall) were selected and assigned to the control or test group. The control group received scaling and root planing with the removal of granulation tissue and root cementum; the test group underwent removal of granulation tissue and soft microbial deposits by cleaning the root surface with a microbrush and saline solution, aiming at cementum preservation. Guided tissue regeneration (GTR) was applied to both groups. Twenty-one days later, the newly formed tissue under the membrane was assessed for the expression of the following genes: alkaline phosphatase (ALP), osteopontin (OPN), osteocalcin (OCN), platelet-derived growth factor-alpha (PDGFA), bone sialoprotein (BSP), and basic fibroblast growth factor (bFGF). Data analysis demonstrated that mRNA levels for PDGFA, BSP, and bFGF were higher in the sites where root cementum was kept in place compared to the sites where root cementum was removed completely as part of the periodontal therapy (P <0.05); in contrast, OCN levels were lower (P <0.05). No difference for ALP or OPN was observed between the control and test groups (P >0.05). Root cementum may modulate the expression of growth and mineral-associated factors during periodontal regeneration.

CCR8 Signaling Influences Toll-Like Receptor 4 Responses in Human Macrophages in Inflammatory Diseases ▿

PubMed Central

Kvist Reimer, Martina; Brange, Charlotte; Rosendahl, Alexander

2011-01-01

CCR8 immunity is generally associated with Th2 responses in allergic diseases. In this study, we demonstrate for the first time a pronounced attenuated influx of macrophages in ovalbumin (OVA)-challenged CCR8 knockout mice. To explore whether macrophages in human inflamed lung tissue also were CCR8 positive, human lung tissue from patients with chronic obstructive pulmonary disease (COPD) was evaluated. Indeed, CCR8 expression was pronounced in invading monocytes/macrophages from lungs of patients with Global Initiative for Obstructive Lung Disease (GOLD) stage IV COPD. Given this expression pattern, the functional role of CCR8 on human macrophages was evaluated in vitro. Human peripheral blood monocytes expressed low levels of CCR8, while macrophage colony-stimulating factor (M-CSF)-derived human macrophages expressed significantly elevated surface levels of CCR8. Importantly, CCL1 directly regulated the expression of CD18 and CD49b and hence influenced the adhesion capacity of human macrophages. CCL1 drives chemotaxis in M-CSF-derived macrophages, and this could be completely inhibited by lipopolysaccharide (LPS). Whereas both CCL1 and LPS monotreatment inhibited spontaneous superoxide release in macrophages, CCL1 significantly induced superoxide release in the presence of LPS in a dose-dependent manner. Finally, CCL1 induced production of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) and could inhibit LPS-induced cytokine production in a dose-dependent manner. Our data demonstrate, for the first time, the presence of CCR8 on inflammatory macrophages in human COPD lung tissue. Importantly, the functional data from human macrophages suggest a potential cross talk between the CCR8 and the Toll-like receptor 4 (TLR4) pathways, both of which are present in COPD patients. PMID:21976223

CCR8 signaling influences Toll-like receptor 4 responses in human macrophages in inflammatory diseases.

PubMed

Reimer, Martina Kvist; Brange, Charlotte; Rosendahl, Alexander

2011-12-01

CCR8 immunity is generally associated with Th2 responses in allergic diseases. In this study, we demonstrate for the first time a pronounced attenuated influx of macrophages in ovalbumin (OVA)-challenged CCR8 knockout mice. To explore whether macrophages in human inflamed lung tissue also were CCR8 positive, human lung tissue from patients with chronic obstructive pulmonary disease (COPD) was evaluated. Indeed, CCR8 expression was pronounced in invading monocytes/macrophages from lungs of patients with Global Initiative for Obstructive Lung Disease (GOLD) stage IV COPD. Given this expression pattern, the functional role of CCR8 on human macrophages was evaluated in vitro. Human peripheral blood monocytes expressed low levels of CCR8, while macrophage colony-stimulating factor (M-CSF)-derived human macrophages expressed significantly elevated surface levels of CCR8. Importantly, CCL1 directly regulated the expression of CD18 and CD49b and hence influenced the adhesion capacity of human macrophages. CCL1 drives chemotaxis in M-CSF-derived macrophages, and this could be completely inhibited by lipopolysaccharide (LPS). Whereas both CCL1 and LPS monotreatment inhibited spontaneous superoxide release in macrophages, CCL1 significantly induced superoxide release in the presence of LPS in a dose-dependent manner. Finally, CCL1 induced production of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) and could inhibit LPS-induced cytokine production in a dose-dependent manner. Our data demonstrate, for the first time, the presence of CCR8 on inflammatory macrophages in human COPD lung tissue. Importantly, the functional data from human macrophages suggest a potential cross talk between the CCR8 and the Toll-like receptor 4 (TLR4) pathways, both of which are present in COPD patients.

MO-F-CAMPUS-I-03: Tissue Equivalent Material Phantom to Test and Optimize Coherent Scatter Imaging for Tumor Classification

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

Albanese, K; Morris, R; Lakshmanan, M

Purpose: To accurately model different breast geometries using a tissue equivalent phantom, and to classify these tissues in a coherent x-ray scatter imaging system. Methods: A breast phantom has been designed to assess the capability of coded aperture coherent x-ray scatter imaging system to classify different types of breast tissue (adipose, fibroglandular, tumor). The tissue-equivalent phantom was modeled as a hollow plastic cylinder containing multiple cylindrical and spherical inserts that can be positioned, rearranged, or removed to model different breast geometries. Each enclosure can be filled with a tissue-equivalent material and excised human tumors. In this study, beef and lard,more » placed inside 2-mm diameter plastic Nalgene containers, were used as surrogates for fibroglandular and adipose tissue, respectively. The phantom was imaged at 125 kVp, 40 mA for 10 seconds each with a 1-mm pencil beam. The raw data were reconstructed using a model-based reconstruction algorithm and yielded the location and form factor, or momentum transfer (q) spectrum of the materials that were imaged. The measured material form factors were then compared to the ground truth measurements acquired by x-ray diffraction (XRD) imaging. Results: The tissue equivalent phantom was found to accurately model different types of breast tissue by qualitatively comparing our measured form factors to those of adipose and fibroglandular tissue from literature. Our imaging system has been able to define the location and composition of the various materials in the phantom. Conclusion: This work introduces a new tissue equivalent phantom for testing and optimization of our coherent scatter imaging system for material classification. In future studies, the phantom will enable the use of a variety of materials including excised human tissue specimens in evaluating and optimizing our imaging system using pencil- and fan-beam geometries. United States Department of Homeland Security Duke University Medical Center - Department of Radiology Carl E Ravin Advanced Imaging Laboratories Duke University Medical Physics Graduate Program.« less

In vitro control of human bone marrow stromal cells for bone tissue engineering.

PubMed

Anselme, Karine; Broux, Odile; Noel, Benoit; Bouxin, Bertrand; Bascoulergue, Gerard; Dudermel, Anne-France; Bianchi, Fabien; Jeanfils, Joseph; Hardouin, Pierre

2002-12-01

For the clinical application of cultured human mesenchymal stem cells (MSCs), cells must have minimal contact with fetal calf serum (FCS) because it might be a potential vector for contamination by adventitious agents. The use of human plasma and serum for clinical applications also continues to give rise to considerable concerns with respect to the transmission of known and unknown human infectious agents. With the objective of clinical applications of cultured human MSCs, we tested the ability of autologous plasma, AB human serum, FCS, and artificial serum substitutes containing animal-derived proteins (Ultroser G) or vegetable-derived proteins (Prolifix S6) to permit their growth and differentiation in vitro. To conserve as much autologous plasma as possible, we attempted to mix it at decreasing concentrations with the serum substitute containing vegetable-derived mitogenic factors. Under control conditions, by day 10 all the fibroblast colony-forming units (CFU-Fs) were alkaline phosphatase (ALP) positive. However, their number and size were highly variable among donors. Better CFU-F formation was obtained with Ultroser G, and with human AB serum and autologous plasma mixed at, respectively, 5 and 1% with Prolifix S6. The effects of these mixtures on CFU-F formation demonstrate synergy, with the human serum or plasma supplying the factors that favor differentiation of MSCs while Prolifix S6 supplies the mitogenic factors. Finally, we demonstrated the possibility of controlling human MSC growth and differentiation in vitro. Notably, by means of a minimal quantity of human serum or human plasma mixed with a new serum substitute containing vegetable-derived proteins, we displayed growth and differentiation of human MSCs comparable to that obtained with FCS or serum substitutes containing animal-derived proteins. These results will have crucial significance for future applications of cultured human MSCs in bone tissue engineering.

Innate Immunity and Breast Milk.

PubMed

Cacho, Nicole Theresa; Lawrence, Robert M

2017-01-01

Human milk is a dynamic source of nutrients and bioactive factors; unique in providing for the human infant's optimal growth and development. The growing infant's immune system has a number of developmental immune deficiencies placing the infant at increased risk of infection. This review focuses on how human milk directly contributes to the infant's innate immunity. Remarkable new findings clarify the multifunctional nature of human milk bioactive components. New research techniques have expanded our understanding of the potential for human milk's effect on the infant that will never be possible with milk formulas. Human milk microbiome directly shapes the infant's intestinal microbiome, while the human milk oligosaccharides drive the growth of these microbes within the gut. New techniques such as genomics, metabolomics, proteomics, and glycomics are being used to describe this symbiotic relationship. An expanded role for antimicrobial proteins/peptides within human milk in innate immune protection is described. The unique milieu of enhanced immune protection with diminished inflammation results from a complex interaction of anti-inflammatory and antioxidative factors provided by human milk to the intestine. New data support the concept of mucosal-associated lymphoid tissue and its contribution to the cellular content of human milk. Human milk stem cells (hMSCs) have recently been discovered. Their direct role in the infant for repair and regeneration is being investigated. The existence of these hMSCs could prove to be an easily harvested source of multilineage stem cells for the study of cancer and tissue regeneration. As the infant's gastrointestinal tract and immune system develop, there is a comparable transition in human milk over time to provide fewer immune factors and more calories and nutrients for growth. Each of these new findings opens the door to future studies of human milk and its effect on the innate immune system and the developing infant.

Human basic fibroblast growth factor fused with Kringle4 peptide binds to a fibrin scaffold and enhances angiogenesis.

PubMed

Zhao, Wenxue; Han, Qianqian; Lin, Hang; Sun, Wenjie; Gao, Yuan; Zhao, Yannan; Wang, Bin; Wang, Xia; Chen, Bing; Xiao, Zhifeng; Dai, Jianwu

2009-05-01

Appropriate three-dimensional (3D) scaffolds and signal molecules could accelerate tissue regeneration and wound repair. In this work, we targeted human basic fibroblast growth factor (bFGF), a potent angiogenic factor, to a fibrin scaffold to improve therapeutic angiogenesis. We fused bFGF to the Kringle4 domain (K4), a fibrin-binding peptide from human plasminogen, to endow bFGF with specific fibrin-binding ability. The recombinant K4bFGF bound specifically to the fibrin scaffold so that K4bFGF was delivered in a site-specific manner, and the fibrin scaffold provided 3D support for cell migration and proliferation. Subcutaneous implantation of the fibrin scaffolds bound with K4bFGF but not with bFGF induced neovascularization. Immunohistochemical analysis showed significantly more proliferation cells in the fibrin scaffolds incorporated with K4bFGF than in those with bFGF. Moreover, the regenerative tissues were integrated well with the fibrin scaffolds, suggesting its good biocompatibility. In summary, targeted delivery of K4bFGF could potentially improve therapeutic angiogenesis.

Malondialdehyde-derived epitopes in human skin result from acute exposure to solar UV and occur in nonmelanoma skin cancer tissue.

PubMed

Williams, Joshua D; Bermudez, Yira; Park, Sophia L; Stratton, Steven P; Uchida, Koji; Hurst, Craig A; Wondrak, Georg T

2014-03-05

Cutaneous exposure to solar ultraviolet radiation (UVR) is a causative factor in photoaging and photocarcinogenesis. In human skin, oxidative stress is widely considered a key mechanism underlying the detrimental effects of acute and chronic UVR exposure. The lipid peroxidation product malondialdehyde (MDA) accumulates in tissue under conditions of increased oxidative stress, and the occurrence of MDA-derived protein epitopes, including dihydropyridine-lysine (DHP), has recently been substantiated in human skin. Here we demonstrate for the first time that acute exposure to sub-apoptogenic doses of solar simulated UV light (SSL) causes the formation of free MDA and protein-bound MDA-derived epitopes in cultured human HaCaT keratinocytes and healthy human skin. Immunohistochemical staining revealed that acute exposure to SSL is sufficient to cause an almost twenty-fold increase in general MDA- and specific DHP-epitope content in human skin. When compared to dose-matched solar simulated UVA, complete SSL was more efficient generating both free MDA and MDA-derived epitopes. Subsequent tissue microarray (TMA) analysis revealed the prevalence of MDA- and DHP-epitopes in nonmelanoma skin cancer (NMSC). In squamous cell carcinoma tissue, both MDA- and DHP-epitopes were increased more than threefold as compared to adjacent normal tissue. Taken together, these date demonstrate the occurrence of MDA-derived epitopes in both solar UVR-exposed healthy human skin and NMSC TMA tissue; however, the potential utility of these epitopes as novel biomarkers of cutaneous photodamage and a functional role in the process of skin photocarcinogenesis remain to be explored. Copyright © 2014 Elsevier B.V. All rights reserved.

Tissue-engineered human bioartificial muscles expressing a foreign recombinant protein for gene therapy

NASA Technical Reports Server (NTRS)

Powell, C.; Shansky, J.; Del Tatto, M.; Forman, D. E.; Hennessey, J.; Sullivan, K.; Zielinski, B. A.; Vandenburgh, H. H.

1999-01-01

Murine skeletal muscle cells transduced with foreign genes and tissue engineered in vitro into bioartificial muscles (BAMs) are capable of long-term delivery of soluble growth factors when implanted into syngeneic mice (Vandenburgh et al., 1996b). With the goal of developing a therapeutic cell-based protein delivery system for humans, similar genetic tissue-engineering techniques were designed for human skeletal muscle stem cells. Stem cell myoblasts were isolated, cloned, and expanded in vitro from biopsied healthy adult (mean age, 42 +/- 2 years), and elderly congestive heart failure patient (mean age, 76 +/- 1 years) skeletal muscle. Total cell yield varied widely between biopsies (50 to 672 per 100 mg of tissue, N = 10), but was not significantly different between the two patient groups. Percent myoblasts per biopsy (73 +/- 6%), number of myoblast doublings prior to senescence in vitro (37 +/- 2), and myoblast doubling time (27 +/- 1 hr) were also not significantly different between the two patient groups. Fusion kinetics of the myoblasts were similar for the two groups after 20-22 doublings (74 +/- 2% myoblast fusion) when the biopsy samples had been expanded to 1 to 2 billion muscle cells, a number acceptable for human gene therapy use. The myoblasts from the two groups could be equally transduced ex vivo with replication-deficient retroviral expression vectors to secrete 0.5 to 2 microg of a foreign protein (recombinant human growth hormone, rhGH)/10(6) cells/day, and tissue engineered into human BAMs containing parallel arrays of differentiated, postmitotic myofibers. This work suggests that autologous human skeletal myoblasts from a potential patient population can be isolated, genetically modified to secrete foreign proteins, and tissue engineered into implantable living protein secretory devices for therapeutic use.

Effect of Fixatives and Tissue Processing on the Content and Integrity of Nucleic Acids

PubMed Central

Srinivasan, Mythily; Sedmak, Daniel; Jewell, Scott

2002-01-01

Clinical and molecular medicines are undergoing a revolution based on the accelerated advances in biotechnology such as DNA microarrays and proteomics. Answers to fundamental questions such as how does the DNA sequence differ between individuals and what makes one individual more prone for a certain disease are eagerly being sought in this postgenomic era. Several government and nonprofit organizations provide the researchers access to human tissues for molecular studies. The tissues procured by the different organizations may differ with respect to fixation and processing parameters that may affect significantly the molecular profile of the tissues. It is imperative that a prospective investigator be aware of the potential contributing factors before designing a project. The purpose of this review is to provide an overview of the methods of human tissue acquisition, fixation, and preservation. In addition, the parameters of procurement and fixation that affect the quality of the tissues at the molecular level are discussed. PMID:12466110

Factors Affecting the Use of Human Tissues in Biomedical Research: Implications in the Design and Operation of a Biorepository.

PubMed

Atherton, Daniel S; Sexton, Katherine C; Otali, Dennis; Bell, Walter C; Grizzle, William E

2016-01-01

The availability of high-quality human tissues is necessary to advance medical research. Although there are inherent and induced limitations on the use of human tissues in research, biorepositories play critical roles in minimizing the effects of such limitations. Specifically, the optimal utilization of tissues in research requires tissues to be diagnosed accurately, and the actual specimens provided to investigators must be carefully described (i.e., there must be quality control of each aliquot of the tissue provided for research, including a description of any damage to tissues). Tissues also should be collected, processed, stored, and distributed (i.e., handled) uniformly under a rigorous quality management system (QMS). Frequently, tissues are distributed to investigators by tissue banks which have collected, processed, and stored them by standard operating procedures (SOPs). Alternatively, tissues for research may be handled via SOPs that are modified to the specific requirements of investigators (i.e., using a prospective biorepository model). The primary goal of any type of biorepository should be to ensure its specimens are of high quality and are utilized appropriately in research; however, approaches may vary based on the tissues available and requested. For example, extraction of specific molecules (e.g., microRNA) to study molecular characteristics of a tissue may require less clinical annotation than tissues that are utilized to identify how the molecular expression might be used to clarify a clinical outcome of a disease or the response to a specific therapy. This review focuses on the limitations of the use of tissues in research and how the design and operations of a tissue biorepository can minimize some of these limitations.

C-terminal peptides of tissue factor pathway inhibitor are novel host defense molecules.

PubMed

Papareddy, Praveen; Kalle, Martina; Kasetty, Gopinath; Mörgelin, Matthias; Rydengård, Victoria; Albiger, Barbara; Lundqvist, Katarina; Malmsten, Martin; Schmidtchen, Artur

2010-09-03

Tissue factor pathway inhibitor (TFPI) inhibits tissue factor-induced coagulation, but may, via its C terminus, also modulate cell surface, heparin, and lipopolysaccharide interactions as well as participate in growth inhibition. Here we show that C-terminal TFPI peptide sequences are antimicrobial against the gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa, gram-positive Bacillus subtilis and Staphylococcus aureus, as well as the fungi Candida albicans and Candida parapsilosis. Fluorescence studies of peptide-treated bacteria, paired with analysis of peptide effects on liposomes, showed that the peptides exerted membrane-breaking effects similar to those seen for the "classic" human antimicrobial peptide LL-37. The killing of E. coli, but not P. aeruginosa, by the C-terminal peptide GGLIKTKRKRKKQRVKIAYEEIFVKNM (GGL27), was enhanced in human plasma and largely abolished in heat-inactivated plasma, a phenomenon linked to generation of antimicrobial C3a and activation of the classic pathway of complement activation. Furthermore, GGL27 displayed anti-endotoxic effects in vitro and in vivo in a mouse model of LPS shock. Importantly, TFPI was found to be expressed in the basal layers of normal epidermis, and was markedly up-regulated in acute skin wounds as well as wound edges of chronic leg ulcers. Furthermore, C-terminal fragments of TFPI were associated with bacteria present in human chronic leg ulcers. These findings suggest a new role for TFPI in cutaneous defense against infections.

The molecular functions of hepatocyte nuclear factors - In and beyond the liver.

PubMed

Lau, Hwee Hui; Ng, Natasha Hui Jin; Loo, Larry Sai Weng; Jasmen, Joanita Binte; Teo, Adrian Kee Keong

2018-05-01

The hepatocyte nuclear factors (HNFs) namely HNF1α/β, FOXA1/2/3, HNF4α/γ and ONECUT1/2 are expressed in a variety of tissues and organs, including the liver, pancreas and kidney. The spatial and temporal manner of HNF expression regulates embryonic development and subsequently the development of multiple tissues during adulthood. Though the HNFs were initially identified individually based on their roles in the liver, numerous studies have now revealed that the HNFs cross-regulate one another and exhibit synergistic relationships in the regulation of tissue development and function. The complex HNF transcriptional regulatory networks have largely been elucidated in rodent models, but less so in human biological systems. Several heterozygous mutations in these HNFs were found to cause diseases in humans but not in rodents, suggesting clear species-specific differences in mutational mechanisms that remain to be uncovered. In this review, we compare and contrast the expression patterns of the HNFs, the HNF cross-regulatory networks and how these liver-enriched transcription factors serve multiple functions in the liver and beyond, extending our focus to the pancreas and kidney. We also summarise the insights gained from both human and rodent studies of mutations in several HNFs that are known to lead to different disease conditions. Copyright © 2017 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Transforming growth factor beta mediates the progesterone suppression of an epithelial metalloproteinase by adjacent stroma in the human endometrium.

PubMed Central

Bruner, K L; Rodgers, W H; Gold, L I; Korc, M; Hargrove, J T; Matrisian, L M; Osteen, K G

1995-01-01

Unlike most normal adult tissues, cyclic growth and tissue remodeling occur within the uterine endometrium throughout the reproductive years. The matrix metalloproteinases (MMPs), a family of structurally related enzymes that degrade specific components of the extracellular matrix are thought to be the physiologically relevant mediators of extracellular matrix composition and turnover. Our laboratory has identified MMPs of the stromelysin family in the cycling human endometrium, implicating these enzymes in mediating the extensive remodeling that occurs in this tissue. While the stromelysins are expressed in vivo during proliferation-associated remodeling and menstruation-associated endometrial breakdown, none of the stromelysins are expressed during the progesterone-dominated secretory phase of the cycle. Our in vitro studies of isolated cell types have confirmed progesterone suppression of stromal MMPs, but a stromal-derived paracrine factor was found necessary for suppression of the epithelial-specific MMP matrilysin. In this report, we demonstrate that transforming growth factor beta (TGF-beta) is produced by endometrial stroma in response to progesterone and can suppress expression of epithelial matrilysin independent of progesterone. Additionally, we find that an antibody directed against the mammalian isoforms of TGF-beta abolishes progesterone suppression of matrilysin in stromal-epithelial cocultures, implicating TGF-beta as the principal mediator of matrilysin suppression in the human endometrium. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:7638197

In vitro 3D regeneration-like growth of human patient brain tissue.

PubMed

Tang-Schomer, M D; Wu, W B; Kaplan, D L; Bookland, M J

2018-05-01

In vitro culture of primary neurons is widely adapted with embryonic but not mature brain tissue. Here, we extended a previously developed bioengineered three-dimensional (3D) embryonic brain tissue model to resected normal patient brain tissue in an attempt to regenerate human neurons in vitro. Single cells and small sized (diameter < 100 μm) spheroids from dissociated brain tissue were seeded into 3D silk fibroin-based scaffolds, with or without collagen or Matrigel, and compared with two-dimensional cultures and scaffold-free suspension cultures. Changes of cell phenotypes (neuronal, astroglial, neural progenitor, and neuroepithelial) were quantified with flow cytometry and analyzed with a new method of statistical analysis specifically designed for percentage comparison. Compared with a complete lack of viable cells in conventional neuronal cell culture condition, supplements of vascular endothelial growth factor-containing pro-endothelial cell condition led to regenerative growth of neurons and astroglial cells from "normal" human brain tissue of epilepsy surgical patients. This process involved delayed expansion of Nestin+ neural progenitor cells, emergence of TUJ1+ immature neurons, and Vimentin+ neuroepithelium-like cell sheet formation in prolonged cultures (14 weeks). Micro-tissue spheroids, but not single cells, supported the brain tissue growth, suggesting importance of preserving native cell-cell interactions. The presence of 3D scaffold, but not hydrogel, allowed for Vimentin+ cell expansion, indicating a different growth mechanism than pluripotent cell-based brain organoid formation. The slow and delayed process implied an origin of quiescent neural precursors in the neocortex tissue. Further optimization of the 3D tissue model with primary human brain cells could provide personalized brain disease models. Copyright © 2018 John Wiley & Sons, Ltd.

Aptamer-conjugated PEGylated quantum dots targeting epidermal growth factor receptor variant III for fluorescence imaging of glioma.

PubMed

Tang, Jiaze; Huang, Ning; Zhang, Xiang; Zhou, Tao; Tan, Ying; Pi, Jiangli; Pi, Li; Cheng, Si; Zheng, Huzhi; Cheng, Yuan

2017-01-01

The extent of resection is a significant prognostic factor in glioma patients. However, the maximum safe resection level is difficult to determine due to the inherent infiltrative character of tumors. Recently, fluorescence-guided surgery has emerged as a new technique that allows safe resection of glioma. In this study, we constructed a new kind of quantum dot (QD)-labeled aptamer (QD-Apt) nanoprobe by conjugating aptamer 32 (A32) to the QDs surface, which can specially bind to the tumors. A32 is a single-stranded DNA capable of binding to the epidermal growth factor receptor variant III (EGFRvIII) specially distributed on the surface of glioma cells. To detect the expression of EGFRvIII in human brain tissues, 120 specimens, including 110 glioma tissues and 10 normal brain tissues, were examined by immunohistochemistry, and the results showed that the rate of positive expression of EGFRvIII in the glioma tissues was 41.82%, and 0.00% in normal brain tissues. Besides, the physiochemical properties of QD-Apt nanoparticles (NPs) were thoroughly characterized. Biocompatibility of the NPs was evaluated, and the results suggested that the QD-Apt was nontoxic in vivo and vitro. Furthermore, the use of the QD-Apt in labeling glioma cell lines and human brain glioma tissues, and target gliomas in situ was also investigated. We found that not only could QD-Apt specially bind to the U87-EGFRvIII glioma cells but also bind to human glioma tissues in vitro. Fluorescence imaging in vivo with orthotopic glioma model mice bearing U87-EGFRvIII showed that QD-Apt could penetrate the blood-brain barrier and then selectively accumulate in the tumors through binding to EGFRvIII, and consequently, generate a strong fluorescence, which contributed to the margins of gliomas that were visualized clearly, and thus, help the surgeons realize the maximum safe resection of glioma. In addition, QD-Apt can also be applied in preoperative diagnosis and postoperative examination of glioma. Therefore, these achievements facilitate the use of tumor-targeted fluorescence imaging in the diagnosis, surgical resection, and postoperative examination of glioma.

Aptamer-conjugated PEGylated quantum dots targeting epidermal growth factor receptor variant III for fluorescence imaging of glioma

PubMed Central

Tang, Jiaze; Huang, Ning; Zhang, Xiang; Zhou, Tao; Tan, Ying; Pi, Jiangli; Pi, Li; Cheng, Si; Zheng, Huzhi; Cheng, Yuan

2017-01-01

The extent of resection is a significant prognostic factor in glioma patients. However, the maximum safe resection level is difficult to determine due to the inherent infiltrative character of tumors. Recently, fluorescence-guided surgery has emerged as a new technique that allows safe resection of glioma. In this study, we constructed a new kind of quantum dot (QD)-labeled aptamer (QD-Apt) nanoprobe by conjugating aptamer 32 (A32) to the QDs surface, which can specially bind to the tumors. A32 is a single-stranded DNA capable of binding to the epidermal growth factor receptor variant III (EGFRvIII) specially distributed on the surface of glioma cells. To detect the expression of EGFRvIII in human brain tissues, 120 specimens, including 110 glioma tissues and 10 normal brain tissues, were examined by immunohistochemistry, and the results showed that the rate of positive expression of EGFRvIII in the glioma tissues was 41.82%, and 0.00% in normal brain tissues. Besides, the physiochemical properties of QD-Apt nanoparticles (NPs) were thoroughly characterized. Biocompatibility of the NPs was evaluated, and the results suggested that the QD-Apt was nontoxic in vivo and vitro. Furthermore, the use of the QD-Apt in labeling glioma cell lines and human brain glioma tissues, and target gliomas in situ was also investigated. We found that not only could QD-Apt specially bind to the U87-EGFRvIII glioma cells but also bind to human glioma tissues in vitro. Fluorescence imaging in vivo with orthotopic glioma model mice bearing U87-EGFRvIII showed that QD-Apt could penetrate the blood–brain barrier and then selectively accumulate in the tumors through binding to EGFRvIII, and consequently, generate a strong fluorescence, which contributed to the margins of gliomas that were visualized clearly, and thus, help the surgeons realize the maximum safe resection of glioma. In addition, QD-Apt can also be applied in preoperative diagnosis and postoperative examination of glioma. Therefore, these achievements facilitate the use of tumor-targeted fluorescence imaging in the diagnosis, surgical resection, and postoperative examination of glioma. PMID:28579776

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.

Human papilloma virus: a new risk factor in a subset of head and neck cancers.

PubMed

Bisht, Manisha; Bist, Sampan Singh

2011-01-01

Head and neck cancer is the sixth most common malignancy worldwide. Tobacco smoking and alcohol consumption are two well known behavioral risk factors associated with head and neck cancer. Recently, evidence is mounting that infection with human papilloma virus, most commonly human papilloma virus-16 is responsible for a subset of head and neck squamous cell carcinoma especially tumors of tonsillar origin. The molecular pathway used by human papilloma virus to trigger malignant transformation of tissue is different from that of other well known risk factors, i.e. smoking and alcohol, associated with squamous cell carcinoma. Apparently, these subsets of patients with human papilloma virus positive tumor are more likely to have a better prognosis than human papilloma virus negative tumor. Considering this fact, the human papilloma virus infection should be determined in all oropharyngeal cancers since it can have a major impact on the decision making process of the treatment.

Structure and Function of the Splice Variants of TMPRSS2-ERG, a Prevalent Genomic Alteration in Prostate Cancer

DTIC Science & Technology

2011-09-01

the ETS family of transcription factors showing diverse expression patterns in human tissues (Turner and Watson, 2008). ERG, similar to other...and adult mouse tissues . Most striking of these observations was highly selective and abundant expression of erg protein in endothelial cells of...mouse tissues . We for the first time clarified that endogenous ERG was not expressed in normal mouse prostate epithelium (Mohamed et al., 2010

The tumor secretory factor ZAG promotes white adipose tissue browning and energy wasting.

PubMed

Elattar, Sawsan; Dimri, Manali; Satyanarayana, Ande

2018-03-23

Cachexia is a complex tissue-wasting syndrome characterized by inflammation, hypermetabolism, increased energy expenditure, and anorexia. Browning of white adipose tissue (WAT) is one of the significant factors that contribute to energy wasting in cachexia. By utilizing a cell implantation model, we demonstrate here that the lipid mobilizing factor zinc-α 2 -glycoprotein (ZAG) induces WAT browning in mice. Increased circulating levels of ZAG not only induced lipolysis in adipose tissues but also caused robust browning in WAT. Stimulating WAT progenitors with ZAG recombinant protein or expression of ZAG in mouse embryonic fibroblasts (MEFs) strongly enhanced brown-like differentiation. At the molecular level, ZAG stimulated peroxisome proliferator-activated receptor γ (PPARγ) and early B cell factor 2 expression and promoted their recruitment to the PR/SET domain 16 (Prdm16) promoter, leading to enhanced expression of Prdm16, which determines brown cell fate. In brown adipose tissue, ZAG stimulated the expression of PPARγ and PPARγ coactivator 1α and promoted recruitment of PPARγ to the uncoupling protein 1 (Ucp1) promoter, leading to increased expression of Ucp1. Overall, our results reveal a novel function of ZAG in WAT browning and highlight the targeting of ZAG as a potential therapeutic application in humans with cachexia.-Elattar, S., Dimri, M., Satyanarayana, A. The tumor secretory factor ZAG promotes white adipose tissue browning and energy wasting.

Adenovirus-mediated transfer of hepatocyte growth factor gene to human dental pulp stem cells under good manufacturing practice improves their potential for periodontal regeneration in swine.

PubMed

Cao, Yu; Liu, Zhenhai; Xie, Yilin; Hu, Jingchao; Wang, Hua; Fan, Zhipeng; Zhang, Chunmei; Wang, Jingsong; Wu, Chu-Tse; Wang, Songlin

2015-12-15

Periodontitis is one of the most widespread infectious diseases in humans. We previously promoted significant periodontal tissue regeneration in swine models with the transplantation of autologous periodontal ligament stem cells (PDLSCs) and PDLSC sheet. We also promoted periodontal tissue regeneration in a rat model with a local injection of allogeneic bone marrow mesenchymal stem cells. The purpose of the present study is to investigate the roles of the hepatocyte growth factor (HGF) and human dental pulp stem cells (DPSCs) in periodontal tissue regeneration in swine. In the present study, we transferred an adenovirus that carried HGF gene into human DPSCs (HGF-hDPSCs) under good manufacturing practice (GMP) conditions. These cells were then transplanted into a swine model for periodontal regeneration. Twenty miniature pigs were used to generate periodontitis with bone defect of 5 mm in width, 7 mm in length, and 3 mm in depth. After 12 weeks, clinical, radiological, quantitative and histological assessment of regenerated periodontal tissues was performed to compare periodontal regeneration in swine treated with cell implantation. Our study showed that injecting HGF-hDPSCs into this large animal model could significantly improve periodontal bone regeneration and soft tissue healing. A hDPSC or HGF-hDPSC sheet showed superior periodontal tissue regeneration compared to the injection of dissociated cells. However, the sheets required surgical placement; thus, they were suitable for surgically-managed periodontitis treatments. The adenovirus-mediated transfer of the HGF gene markedly decreased hDPSC apoptosis in a hypoxic environment or in serum-free medium, and it increased blood vessel regeneration. This study indicated that HGF-hDPSCs produced under GMP conditions significantly improved periodontal bone regeneration in swine; thus, this method represents a potential clinical application for periodontal regeneration.

Tissue factor-expressing monocytes inhibit fibrinolysis through a TAFI-mediated mechanism, and make clots resistant to heparins

PubMed Central

Semeraro, Fabrizio; Ammollo, Concetta T.; Semeraro, Nicola; Colucci, Mario

2009-01-01

Background Thrombin is the main activator of the fibrinolysis inhibitor TAFI (thrombin activatable fibrinolysis inhibitor) and heightened clotting activation is believed to impair fibrinolysis through the increase of thrombin activatable fibrinolysis inhibitor activation. However, the enhancement of thrombin generation by soluble tissue factor was reported to have no effect on plasma fibrinolysis and it is not known whether the same is true for cell-associated tissue factor. The aim of this study was to evaluate the effect of tissue factor-expressing monocytes on plasma fibrinolysis in vitro. Design and Methods Tissue factor expression by human blood mononuclear cells (MNC) and monocytes was induced by LPS stimulation. Fibrinolysis was spectrophotometrically evaluated by measuring the lysis time of plasma clots containing LPS-stimulated or control cells and a low concentration of exogenous tissue plasminogen activator. Results LPS-stimulated MNC (LPS-MNC) prolonged fibrinolysis time as compared to unstimulated MNC (C-MNC) in contact-inhibited but not in normal citrated plasma. A significantly prolonged lysis time was observed using as few as 30 activated cells/μL. Fibrinolysis was also impaired when clots were generated on adherent LPS-stimulated monocytes. The antifibrinolytic effect of LPS-MNC or LPS-monocytes was abolished by an anti-tissue factor antibody, by an antibody preventing thrombin-mediated thrombin activatable fibrinolysis inhibitor activation, and by a TAFIa inhibitor (PTCI). Assays of thrombin and TAFIa in contact-inhibited plasma confirmed the greater generation of these enzymes in the presence of LPS-MNC. Finally, the profibrinolytic effect of unfractionated heparin and enoxaparin was markedly lower (~50%) in the presence of LPS-MNC than in the presence of a thromboplastin preparation displaying an identical tissue factor activity. Conclusions Our data indicate that LPS-stimulated monocytes inhibit fibrinolysis through a tissue factor-mediated enhancement of thrombin activatable fibrinolysis inhibitor activation and make clots resistant to the profibrinolytic activity of heparins, thus providing an additional mechanism whereby tissue factor-expressing monocytes/macrophages may favor fibrin accumulation and diminish the antithrombotic efficacy of heparins. PMID:19377079

The clinical use of regenerative therapy in COPD

PubMed Central

Lipsi, Roberto; Rogliani, Paola; Calzetta, Luigino; Segreti, Andrea; Cazzola, Mario

2014-01-01

Regenerative or stem cell therapy is an emerging field of treatment based on stimulation of endogenous resident stem cells or administration of exogenous stem cells to treat diseases or injury and to replace malfunctioning or damaged tissues. Current evidence suggests that in the lung, these cells may participate in tissue homeostasis and regeneration after injury. Animal and human studies have demonstrated that tissue-specific stem cells and bone marrow-derived cells contribute to lung tissue regeneration and protection, and thus administration of exogenous stem/progenitor cells or humoral factors responsible for the activation of endogenous stem/progenitor cells may be a potent next-generation therapy for chronic obstructive pulmonary disease. The use of bone marrow-derived stem cells could allow repairing and regenerate the damaged tissue present in chronic obstructive pulmonary disease by means of their engraftment into the lung. Another approach could be the stimulation of resident stem cells by means of humoral factors or photobiostimulation. PMID:25548520

High Molecular Weight Fibroblast Growth Factor-2 in the Human Heart Is a Potential Target for Prevention of Cardiac Remodeling

PubMed Central

Santiago, Jon-Jon; McNaughton, Leslie J.; Koleini, Navid; Ma, Xin; Bestvater, Brian; Nickel, Barbara E.; Fandrich, Robert R.; Wigle, Jeffrey T.; Freed, Darren H.; Arora, Rakesh C.; Kardami, Elissavet

2014-01-01

Fibroblast growth factor 2 (FGF-2) is a multifunctional protein synthesized as high (Hi-) and low (Lo-) molecular weight isoforms. Studies using rodent models showed that Hi- and Lo-FGF-2 exert distinct biological activities: after myocardial infarction, rat Lo-FGF-2, but not Hi-FGF-2, promoted sustained cardioprotection and angiogenesis, while Hi-FGF-2, but not Lo-FGF-2, promoted myocardial hypertrophy and reduced contractile function. Because there is no information regarding Hi-FGF-2 in human myocardium, we undertook to investigate expression, regulation, secretion and potential tissue remodeling-associated activities of human cardiac (atrial) Hi-FGF-2. Human patient-derived atrial tissue extracts, as well as pericardial fluid, contained Hi-FGF-2 isoforms, comprising, respectively, 53%(±20 SD) and 68% (±25 SD) of total FGF-2, assessed by western blotting. Human atrial tissue-derived primary myofibroblasts (hMFs) expressed and secreted predominantly Hi-FGF-2, at about 80% of total. Angiotensin II (Ang II) up-regulated Hi-FGF-2 in hMFs, via activation of both type 1 and type 2 Ang II receptors; the ERK pathway; and matrix metalloprotease-2. Treatment of hMFs with neutralizing antibodies selective for human Hi-FGF-2 (neu-AbHi-FGF-2) reduced accumulation of proteins associated with fibroblast-to-myofibroblast conversion and fibrosis, including α-smooth muscle actin, extra-domain A fibronectin, and procollagen. Stimulation of hMFs with recombinant human Hi-FGF-2 was significantly more potent than Lo-FGF-2 in upregulating inflammation-associated proteins such as pro-interleukin-1β and plasminogen-activator-inhibitor-1. Culture media conditioned by hMFs promoted cardiomyocyte hypertrophy, an effect that was prevented by neu-AbHi-FGF-2 in vitro. In conclusion, we have documented that Hi-FGF-2 represents a substantial fraction of FGF-2 in human cardiac (atrial) tissue and in pericardial fluid, and have shown that human Hi-FGF-2, unlike Lo-FGF-2, promotes deleterious (pro-fibrotic, pro-inflammatory, and pro-hypertrophic) responses in vitro. Selective targeting of Hi-FGF-2 production may, therefore, reduce pathological remodelling in the human heart. PMID:24827991

High molecular weight fibroblast growth factor-2 in the human heart is a potential target for prevention of cardiac remodeling.

PubMed

Santiago, Jon-Jon; McNaughton, Leslie J; Koleini, Navid; Ma, Xin; Bestvater, Brian; Nickel, Barbara E; Fandrich, Robert R; Wigle, Jeffrey T; Freed, Darren H; Arora, Rakesh C; Kardami, Elissavet

2014-01-01

Fibroblast growth factor 2 (FGF-2) is a multifunctional protein synthesized as high (Hi-) and low (Lo-) molecular weight isoforms. Studies using rodent models showed that Hi- and Lo-FGF-2 exert distinct biological activities: after myocardial infarction, rat Lo-FGF-2, but not Hi-FGF-2, promoted sustained cardioprotection and angiogenesis, while Hi-FGF-2, but not Lo-FGF-2, promoted myocardial hypertrophy and reduced contractile function. Because there is no information regarding Hi-FGF-2 in human myocardium, we undertook to investigate expression, regulation, secretion and potential tissue remodeling-associated activities of human cardiac (atrial) Hi-FGF-2. Human patient-derived atrial tissue extracts, as well as pericardial fluid, contained Hi-FGF-2 isoforms, comprising, respectively, 53%(±20 SD) and 68% (±25 SD) of total FGF-2, assessed by western blotting. Human atrial tissue-derived primary myofibroblasts (hMFs) expressed and secreted predominantly Hi-FGF-2, at about 80% of total. Angiotensin II (Ang II) up-regulated Hi-FGF-2 in hMFs, via activation of both type 1 and type 2 Ang II receptors; the ERK pathway; and matrix metalloprotease-2. Treatment of hMFs with neutralizing antibodies selective for human Hi-FGF-2 (neu-AbHi-FGF-2) reduced accumulation of proteins associated with fibroblast-to-myofibroblast conversion and fibrosis, including α-smooth muscle actin, extra-domain A fibronectin, and procollagen. Stimulation of hMFs with recombinant human Hi-FGF-2 was significantly more potent than Lo-FGF-2 in upregulating inflammation-associated proteins such as pro-interleukin-1β and plasminogen-activator-inhibitor-1. Culture media conditioned by hMFs promoted cardiomyocyte hypertrophy, an effect that was prevented by neu-AbHi-FGF-2 in vitro. In conclusion, we have documented that Hi-FGF-2 represents a substantial fraction of FGF-2 in human cardiac (atrial) tissue and in pericardial fluid, and have shown that human Hi-FGF-2, unlike Lo-FGF-2, promotes deleterious (pro-fibrotic, pro-inflammatory, and pro-hypertrophic) responses in vitro. Selective targeting of Hi-FGF-2 production may, therefore, reduce pathological remodelling in the human heart.

Three-dimensional organotypic culture: experimental models of mammalian biology and disease.

PubMed

Shamir, Eliah R; Ewald, Andrew J

2014-10-01

Mammalian organs are challenging to study as they are fairly inaccessible to experimental manipulation and optical observation. Recent advances in three-dimensional (3D) culture techniques, coupled with the ability to independently manipulate genetic and microenvironmental factors, have enabled the real-time study of mammalian tissues. These systems have been used to visualize the cellular basis of epithelial morphogenesis, to test the roles of specific genes in regulating cell behaviours within epithelial tissues and to elucidate the contribution of microenvironmental factors to normal and disease processes. Collectively, these novel models can be used to answer fundamental biological questions and generate replacement human tissues, and they enable testing of novel therapeutic approaches, often using patient-derived cells.

Brown adipose tissue: The heat is on the heart.

PubMed

Thoonen, Robrecht; Hindle, Allyson G; Scherrer-Crosbie, Marielle

2016-06-01

The study of brown adipose tissue (BAT) has gained significant scientific interest since the discovery of functional BAT in adult humans. The thermogenic properties of BAT are well recognized; however, data generated in the last decade in both rodents and humans reveal therapeutic potential for BAT against metabolic disorders and obesity. Here we review the current literature in light of a potential role for BAT in beneficially mediating cardiovascular health. We focus mainly on BAT's actions in obesity, vascular tone, and glucose and lipid metabolism. Furthermore, we discuss the recently discovered endocrine factors that have a potential beneficial role in cardiovascular health. These BAT-secreted factors may have a favorable effect against cardiovascular risk either through their metabolic role or by directly affecting the heart. Copyright © 2016 the American Physiological Society.

Tissue factor-dependent coagulation activation by heme: A thromboelastometry study.

PubMed

de Souza, Gleice Regina; Hounkpe, Bidossessi Wilfried; Fiusa, Maiara Marx Luz; Colella, Marina Pereira; Annichino-Bizzacchi, Joyce M; Traina, Fabiola; Costa, Fernando Ferreira; De Paula, Erich Vinicius

2017-01-01

Heme has been characterized as potent trigger of inflammation. In hemostasis, although heme has been shown to both induce and inhibit different compartments of hemostasis, its net effect on the hemostatic balance, and the biological relevance of these effects remain to be determined. Herein we evaluated the effect of heme on hemostasis using a global assay able to generate clinically relevant data in several other complex hemostatic diseases. Citrated whole blood samples from healthy participants were stimulated by heme or vehicle and incubated for 4h at 37°C. Rotational thromboelastometry was immediately performed. The participation of tissue factor in coagulation activation was evaluated using inhibitory antibody. Heme was able of inducing ex vivo coagulation activation in whole blood, affecting predominantly parameters associated with the initial phases of clot formation. This activation effect was at least partially dependent on hematopoietic tissue factor, since the effects of heme were partially abrogated by the inhibition of human tissue factor. In conclusion, using a global hemostasis assay, our study confirmed that heme is able to activate coagulation in whole blood, in a tissue factor-dependent way. These findings could explain the disturbance in hemostatic balance observed in conditions associated with the release of heme such as sickle cell disease.

Activated Monocytes Enhance Platelet-Driven Contraction of Blood Clots via Tissue Factor Expression.

PubMed

Peshkova, Alina D; Le Minh, Giang; Tutwiler, Valerie; Andrianova, Izabella A; Weisel, John W; Litvinov, Rustem I

2017-07-11

Platelet-driven reduction in blood clot volume (clot contraction or retraction) has been implicated to play a role in hemostasis and thrombosis. Although these processes are often linked with inflammation, the role of inflammatory cells in contraction of blood clots and thrombi has not been investigated. The aim of this work was to study the influence of activated monocytes on clot contraction. The effects of monocytes were evaluated using a quantitative optical tracking methodology to follow volume changes in a blood clot formed in vitro. When a physiologically relevant number of isolated human monocytes pre-activated with phorbol-12-myristate-13-acetate (PMA) were added back into whole blood, the extent and rate of clot contraction were increased compared to addition of non-activated cells. Inhibition of tissue factor expression or its inactivation on the surface of PMA-treated monocytes reduced the extent and rate of clot contraction back to control levels with non-activated monocytes. On the contrary, addition of tissue factor enhanced clot contraction, mimicking the effects of tissue factor expressed on the activated monocytes. These data suggest that the inflammatory cells through their expression of tissue factor can directly affect hemostasis and thrombosis by modulating the size and density of intra- and extravascular clots and thrombi.

Choice of surrogate tissue influences neonatal EWAS findings.

PubMed

Lin, Xinyi; Teh, Ai Ling; Chen, Li; Lim, Ives Yubin; Tan, Pei Fang; MacIsaac, Julia L; Morin, Alexander M; Yap, Fabian; Tan, Kok Hian; Saw, Seang Mei; Lee, Yung Seng; Holbrook, Joanna D; Godfrey, Keith M; Meaney, Michael J; Kobor, Michael S; Chong, Yap Seng; Gluckman, Peter D; Karnani, Neerja

2017-12-05

Epigenomes are tissue specific and thus the choice of surrogate tissue can play a critical role in interpreting neonatal epigenome-wide association studies (EWAS) and in their extrapolation to target tissue. To develop a better understanding of the link between tissue specificity and neonatal EWAS, and the contributions of genotype and prenatal factors, we compared genome-wide DNA methylation of cord tissue and cord blood, two of the most accessible surrogate tissues at birth. In 295 neonates, DNA methylation was profiled using Infinium HumanMethylation450 beadchip arrays. Sites of inter-individual variability in DNA methylation were mapped and compared across the two surrogate tissues at birth, i.e., cord tissue and cord blood. To ascertain the similarity to target tissues, DNA methylation profiles of surrogate tissues were compared to 25 primary tissues/cell types mapped under the Epigenome Roadmap project. Tissue-specific influences of genotype on the variable CpGs were also analyzed. Finally, to interrogate the impact of the in utero environment, EWAS on 45 prenatal factors were performed and compared across the surrogate tissues. Neonatal EWAS results were tissue specific. In comparison to cord blood, cord tissue showed higher inter-individual variability in the epigenome, with a lower proportion of CpGs influenced by genotype. Both neonatal tissues were good surrogates for target tissues of mesodermal origin. They also showed distinct phenotypic associations, with effect sizes of the overlapping CpGs being in the same order of magnitude. The inter-relationship between genetics, prenatal factors and epigenetics is tissue specific, and requires careful consideration in designing and interpreting future neonatal EWAS. This birth cohort is a prospective observational study, designed to study the developmental origins of health and disease, and was retrospectively registered on 1 July 2010 under the identifier NCT01174875 .

Microbial Dysbiosis Is Associated with Human Breast Cancer

PubMed Central

Xuan, Caiyun; Shamonki, Jaime M.; Chung, Alice; DiNome, Maggie L.; Chung, Maureen; Sieling, Peter A.; Lee, Delphine J.

2014-01-01

Breast cancer affects one in eight women in their lifetime. Though diet, age and genetic predisposition are established risk factors, the majority of breast cancers have unknown etiology. The human microbiota refers to the collection of microbes inhabiting the human body. Imbalance in microbial communities, or microbial dysbiosis, has been implicated in various human diseases including obesity, diabetes, and colon cancer. Therefore, we investigated the potential role of microbiota in breast cancer by next-generation sequencing using breast tumor tissue and paired normal adjacent tissue from the same patient. In a qualitative survey of the breast microbiota DNA, we found that the bacterium Methylobacterium radiotolerans is relatively enriched in tumor tissue, while the bacterium Sphingomonas yanoikuyae is relatively enriched in paired normal tissue. The relative abundances of these two bacterial species were inversely correlated in paired normal breast tissue but not in tumor tissue, indicating that dysbiosis is associated with breast cancer. Furthermore, the total bacterial DNA load was reduced in tumor versus paired normal and healthy breast tissue as determined by quantitative PCR. Interestingly, bacterial DNA load correlated inversely with advanced disease, a finding that could have broad implications in diagnosis and staging of breast cancer. Lastly, we observed lower basal levels of antibacterial response gene expression in tumor versus healthy breast tissue. Taken together, these data indicate that microbial DNA is present in the breast and that bacteria or their components may influence the local immune microenvironment. Our findings suggest a previously unrecognized link between dysbiosis and breast cancer which has potential diagnostic and therapeutic implications. PMID:24421902

Bioassay of procoagulant albumin in human plasma.

PubMed

Grosset, A; Liu, L; Parker, C J; Rodgers, G M

1994-09-01

Procoagulant albumin (P-Al) is present in normal human plasma and increases monocyte and endothelial cell expression of tissue factor activity. To develop a bioassay for P-Al, we partially purified plasma from healthy volunteers and several patient groups using BaCl2 and (NH4)2SO4 precipitation. The samples were assayed for tissue factor (TF) inducing activity, expressed as a percentage increase compared to a serum-free media control. Over six months, the assay was reproducible in stored samples and in serial samples from normal volunteers. The plasma P-Al activities of 35 volunteers averaged 141 +/- 8.2% (SEM). There was no diurnal variation. There was no difference in the P-Al activity after a 12 hour fast and 2 hours after a large meal in 4 healthy volunteers. There was no increase in activity (r = 0.16) with the subject's age. The average activity from 16 poorly-controlled diabetics was 131 +/- 11% (SEM). No alteration in activity was seen with samples from patients with uremia, liver dysfunction, hemophilia, thrombotic events, or adenocarcinoma. These results indicate that P-Al activity can be bioassayed in individual patient samples; however, pathologic states associated with abnormal P-Al-induced tissue factor activity presently remain unidentified.

FXIa and platelet polyphosphate as therapeutic targets during human blood clotting on collagen/tissue factor surfaces under flow.

PubMed

Zhu, Shu; Travers, Richard J; Morrissey, James H; Diamond, Scott L

2015-09-17

Factor XIIa (FXIIa) and factor XIa (FXIa) contribute to thrombosis in animal models, whereas platelet-derived polyphosphate (polyP) may potentiate contact or thrombin-feedback pathways. The significance of these mediators in human blood under thrombotic flow conditions on tissue factor (TF) -bearing surfaces remains inadequately resolved. Human blood (corn trypsin inhibitor treated [4 μg/mL]) was tested by microfluidic assay for clotting on collagen/TF at TF surface concentration ([TF]wall) from ∼0.1 to 2 molecules per μm(2). Anti-FXI antibodies (14E11 and O1A6) or polyP-binding protein (PPXbd) were used to block FXIIa-dependent FXI activation, FXIa-dependent factor IX (FIX) activation, or platelet-derived polyP, respectively. Fibrin formation was sensitive to 14E11 at 0 to 0.1 molecules per µm(2) and sensitive to O1A6 at 0 to 0.2 molecules per µm(2). However, neither antibody reduced fibrin generation at ∼2 molecules per µm(2) when the extrinsic pathway became dominant. Interestingly, PPXbd reduced fibrin generation at low [TF]wall (0.1 molecules per µm(2)) but not at zero or high [TF]wall, suggesting a role for polyP distinct from FXIIa activation and requiring low extrinsic pathway participation. Regardless of [TF]wall, PPXbd enhanced fibrin sensitivity to tissue plasminogen activator and promoted clot retraction during fibrinolysis concomitant with an observed PPXbd-mediated reduction of fibrin fiber diameter. This is the first detection of endogenous polyP function in human blood under thrombotic flow conditions. When triggered by low [TF]wall, thrombosis may be druggable by contact pathway inhibition, although thrombolytic susceptibility may benefit from polyP antagonism regardless of [TF]wall. © 2015 by The American Society of Hematology.

Comparative Biology of Decellularized Lung Matrix: Implications of Species Mismatch in Regenerative Medicine

PubMed Central

Balestrini, Jenna L.; Gard, Ashley L.; Gerhold, Kristin A.; Wilcox, Elise C.; Liu, Angela; Schwan, Jonas; Le, Andrew V.; Baevova, Pavlina; Dimitrievska, Sashka; Zhao, Liping; Sundaram, Sumati; Sun, Huanxing; Rittié, Laure; Dyal, Rachel; Broekelmann, Tom J.; Mecham, Robert P.; Schwartz, Martin A.; Niklason, Laura E.; White, Eric S.

2016-01-01

Lung engineering is a promising technology, relying on re-seeding of either human or xenographic decellularized matrices with patient-derived pulmonary cells. Little is known about the species-specificity of decellularization in various models of lung regeneration, or if species dependent cell-matrix interactions exist within these systems. Therefore decellularized scaffolds were produced from rat, pig, primate and human lungs, and assessed by measuring residual DNA, mechanical properties, and key matrix proteins (collagen, elastin, glycosaminoglycans). To study intrinsic matrix biologic cues, human endothelial cells were seeded onto acellular slices and analyzed for markers of cell health and inflammation. Despite similar levels of collagen after decellularization, human and primate lungs were stiffer, contained more elastin, and retained fewer glycosaminoglycans than pig or rat lung scaffolds. Human endothelial cells seeded onto human and primate lung tissue demonstrated less expression of vascular cell adhesion molecule and activation of nuclear factor-κB compared to those seeded onto rodent or porcine tissue. Adhesion of endothelial cells was markedly enhanced on human and primate tissues. Our work suggests that species-dependent biologic cues intrinsic to lung extracellular matrix could have profound effects on attempts at lung regeneration. PMID:27344365

Nerve Growth Factor Effects on the Immune System

DTIC Science & Technology

1989-12-19

neuroblastoma cell iine SY5Y was also used in this study of NGF and NGFR. NGF treatment of SY5Y induces differentiation events that are similar to the effect of...Regino Perez-Polo. Nerve growth factor induced neurite outgrowth in clone derived from NGF insensitive -7- human neuroblastoma cell line . Int. J. Devl...as outlined, were to characterize NGF binding in different rodent and human lymphoid tissues and to screen possible NGFR bearing cell lines

Fibroblast growth factor-2 regulates the cell function of human dental pulp cells.

PubMed

Shimabukuro, Yoshio; Ueda, Maki; Ozasa, Masao; Anzai, Jun; Takedachi, Masahide; Yanagita, Manabu; Ito, Masako; Hashikawa, Tomoko; Yamada, Satoru; Murakami, Shinya

2009-11-01

Homeostasis and tissue repair of dentin-pulp complex are attributed to dental pulp tissue and several growth factors. Dental pulp cells play a pivotal role in homeostasis of dentin-pulp complex and tissue responses after tooth injury. Among these cytokines, fibroblast growth factor (FGF)-2 has multifunctional biologic activity and is known as a signaling molecule that induces tissue regeneration. In this study, we examined the effects of FGF-2 on growth, migration, and differentiation of human dental pulp cells (HDPC). HDPC were isolated from healthy dental pulp. Cellular response was investigated by [(3)H]-thymidine incorporation into DNA. Cytodifferentiation was examined by alkaline phosphatase (ALPase) assay and cytochemical staining of calcium by using alizarin red. Migratory activity was determined by counting the cells migrating into cleared area that had introduced with silicon block. FGF-2 activated HDPC growth and migration but suppressed ALPase activity and calcified nodule formation. Interestingly, HDPC, which had been pretreated with FGF-2, showed increased ALPase activity and calcified nodule formation when subsequently cultured without FGF-2. These results suggest that FGF-2 potentiates cell growth and accumulation of HDPC that notably did not disturb cytodifferentiation of the cells later. Thus, FGF-2 is a favorable candidate for pulp capping agent. These results provide new evidence for the possible involvement of FGF-2 not only in homeostasis but also in regeneration of dentin-pulp complex.

Flow cytometric analysis of normal and neoplastic mast cells: role in diagnosis and follow-up of mast cell disease.

PubMed

Escribano, Luis; Garcia Montero, Andres C; Núñez, Rosa; Orfao, Alberto

2006-08-01

Human mast cells (MCs) are directly derived from human pluripotent CD34+ stem and progenitor hematopoietic cells with stem cell factor being a critical growth factor supporting human MC proliferation, differentiation, and survival. Because of the advantages that flow cytometry offers (it allows rapid, objective, and sensitive multiparameter analysis of high numbers of cells from a sample, with information being provided on the basis of a single cell), it has become the method of choice in the past decade for immunophenotypic identification, enumeration, and characterization of human MCs in bone marrow and other tissue specimens.

Characterization of human myoblast differentiation for tissue-engineering purposes by quantitative gene expression analysis.

PubMed

Stern-Straeter, Jens; Bonaterra, Gabriel Alejandro; Kassner, Stefan S; Zügel, Stefanie; Hörmann, Karl; Kinscherf, Ralf; Goessler, Ulrich Reinhart

2011-08-01

Tissue engineering of skeletal muscle is an encouraging possibility for the treatment of muscle loss through the creation of functional muscle tissue in vitro from human stem cells. Currently, the preferred stem cells are primary, non-immunogenic satellite cells ( = myoblasts). The objective of this study was to determine the expression patterns of myogenic markers within the human satellite cell population during their differentiation into multinucleated myotubes for an accurate characterization of stem cell behaviour. Satellite cells were incubated (for 1, 4, 8, 12 or 16 days) with a culture medium containing either a low [ = differentiation medium (DM)] or high [ = growth medium (GM)] concentration of growth factors. Furthermore, we performed a quantitative gene expression analysis of well-defined differentiation makers: myogenic factor 5 (MYF5), myogenin (MYOG), skeletal muscle αactin1 (ACTA1), embryonic (MYH3), perinatal (MYH8) and adult skeletal muscle myosin heavy chain (MYH1). Additionally, the fusion indices of forming myotubes of MYH1, MYH8 and ACTA1 were calculated. We show that satellite cells incubated with DM expressed multiple characteriztic features of mature skeletal muscles, verified by time-dependent upregulation of MYOG, MYH1, MYH3, MYH8 and ACTA1. However, satellite cells incubated with GM did not reveal all morphological aspects of muscle differentiation. Immunocytochemical investigations with antibodies directed against the differentiation markers showed correlations between the gene expression and differentiation. Our data provide information about time-dependent gene expression of differentiation markers in human satellite cells, which can be used for maturation analyses in skeletal muscle tissue-engineering applications. Copyright © 2011 John Wiley & Sons, Ltd.

The Neurovascular Properties of Dental Stem Cells and Their Importance in Dental Tissue Engineering

PubMed Central

Ratajczak, Jessica; Bronckaers, Annelies; Dillen, Yörg; Gervois, Pascal; Vangansewinkel, Tim; Driesen, Ronald B.; Wolfs, Esther; Lambrichts, Ivo

2016-01-01

Within the field of tissue engineering, natural tissues are reconstructed by combining growth factors, stem cells, and different biomaterials to serve as a scaffold for novel tissue growth. As adequate vascularization and innervation are essential components for the viability of regenerated tissues, there is a high need for easily accessible stem cells that are capable of supporting these functions. Within the human tooth and its surrounding tissues, different stem cell populations can be distinguished, such as dental pulp stem cells, stem cells from human deciduous teeth, stem cells from the apical papilla, dental follicle stem cells, and periodontal ligament stem cells. Given their straightforward and relatively easy isolation from extracted third molars, dental stem cells (DSCs) have become an attractive source of mesenchymal-like stem cells. Over the past decade, there have been numerous studies supporting the angiogenic, neuroprotective, and neurotrophic effects of the DSC secretome. Together with their ability to differentiate into endothelial cells and neural cell types, this makes DSCs suitable candidates for dental tissue engineering and nerve injury repair. PMID:27688777

Factor VIII-associated antigen in human lymphatic endothelium.

PubMed

Nagle, R B; Witte, M H; Martinez, A P; Witte, C L; Hendrix, M J; Way, D; Reed, K

1987-03-01

Lymphatic vascular endothelium both on tissue section and in culture exhibits positivity for Factor VIII-associated antigen although staining is generally less intense and more spotty than in comparable blood vascular endothelium. Lymphatic endothelium also exhibits Weibel-Palade bodies. Neither marker, therefore, reliably distinguishes blood vascular endothelium from lymphatic endothelium.

Novel Replication-Competent Circular DNA Molecules from Healthy Cattle Serum and Milk and Multiple Sclerosis-Affected Human Brain Tissue

PubMed Central

Whitley, Corinna; Gunst, Karin; Müller, Hermann; Funk, Mathis; zur Hausen, Harald

2014-01-01

Epidemiological data point to the involvement of a cow milk factor in the etiology of multiple sclerosis (MS). Eleven circular DNA molecules closely related to transmissible spongiform encephalopathy (TSE)-associated isolate Sphinx 1.76 were isolated from healthy cattle serum, cow milk, and serum and brain tissue from MS patients. PMID:25169859

Genome-wide DNA methylation measurements in prostate tissues uncovers novel prostate cancer diagnostic biomarkers and transcription factor binding patterns.

PubMed

Kirby, Marie K; Ramaker, Ryne C; Roberts, Brian S; Lasseigne, Brittany N; Gunther, David S; Burwell, Todd C; Davis, Nicholas S; Gulzar, Zulfiqar G; Absher, Devin M; Cooper, Sara J; Brooks, James D; Myers, Richard M

2017-04-17

Current diagnostic tools for prostate cancer lack specificity and sensitivity for detecting very early lesions. DNA methylation is a stable genomic modification that is detectable in peripheral patient fluids such as urine and blood plasma that could serve as a non-invasive diagnostic biomarker for prostate cancer. We measured genome-wide DNA methylation patterns in 73 clinically annotated fresh-frozen prostate cancers and 63 benign-adjacent prostate tissues using the Illumina Infinium HumanMethylation450 BeadChip array. We overlaid the most significantly differentially methylated sites in the genome with transcription factor binding sites measured by the Encyclopedia of DNA Elements consortium. We used logistic regression and receiver operating characteristic curves to assess the performance of candidate diagnostic models. We identified methylation patterns that have a high predictive power for distinguishing malignant prostate tissue from benign-adjacent prostate tissue, and these methylation signatures were validated using data from The Cancer Genome Atlas Project. Furthermore, by overlaying ENCODE transcription factor binding data, we observed an enrichment of enhancer of zeste homolog 2 binding in gene regulatory regions with higher DNA methylation in malignant prostate tissues. DNA methylation patterns are greatly altered in prostate cancer tissue in comparison to benign-adjacent tissue. We have discovered patterns of DNA methylation marks that can distinguish prostate cancers with high specificity and sensitivity in multiple patient tissue cohorts, and we have identified transcription factors binding in these differentially methylated regions that may play important roles in prostate cancer development.

Analysis of linear energy transfers and quality factors of charged particles produced by spontaneous fission neutrons from 252Cf and 244Pu in the human body.

PubMed

Endo, Akira; Sato, Tatsuhiko

2013-04-01

Absorbed doses, linear energy transfers (LETs) and quality factors of secondary charged particles in organs and tissues, generated via the interactions of the spontaneous fission neutrons from (252)Cf and (244)Pu within the human body, were studied using the Particle and Heavy Ion Transport Code System (PHITS) coupled with the ICRP Reference Phantom. Both the absorbed doses and the quality factors in target organs generally decrease with increasing distance from the source organ. The analysis of LET distributions of secondary charged particles led to the identification of the relationship between LET spectra and target-source organ locations. A comparison between human body-averaged mean quality factors and fluence-averaged radiation weighting factors showed that the current numerical conventions for the radiation weighting factors of neutrons, updated in ICRP103, and the quality factors for internal exposure are valid.

THE EFFECT OF POLIOMYELITIS VIRUS ON HUMAN BRAIN CELLS IN TISSUE CULTURE

PubMed Central

Hogue, M. J.; McAllister, R.; Greene, A. E.; Coriell, L. L.

1955-01-01

Poliomyelitis virus I, Mahoney strain, affected human brain cells grown in tissue cultures usually causing death of the cells in 3 days. The neurons reacted in different ways to the virus, some died with their neurites extended, others contracted one or more of their neurites. Terminal bulbs were frequently formed at the tips of the neurites when they were being drawn into the cell body. The final contraction of the cell body and the change into a mass of granules were often very sudden. Vacuoles often developed in the neuron. There was no recovery. Astrocytes, oligodendroglia, and macrophages were affected by the virus but not as quickly as the neurons. The age of the tissue culture was not a factor when the cells were in good condition. The age of the individual donor of the brain tissue was a factor; the fetal brain cells appeared to be more sensitive to the virus than the adult brain cells. The fetal neurons often reacted ½ hour after inoculation while the adult neurons reacted more slowly, 2 to 24 hours after inoculation. All these changes seemed to be caused by virus infection because they were prevented by specific antiserum or by preheating the virus. PMID:14392238

Expression and clinical significance of connective tissue growth factor in thyroid carcinomas.

PubMed

Wang, Guimin; Zhang, Wei; Meng, Wei; Liu, Jia; Wang, Peisong; Lin, Shan; Xu, Liyan; Li, Enmin; Chen, Guang

2013-08-01

To examine expression of the connective tissue growth factor (CTGF) gene in human thyroid cancer and establish whether a correlation exists between the presence of CTGF protein and clinicopathological parameters of the disease. CTGF protein expression was investigated retrospectively by immunohistochemical analysis of CTGF protein levels in thyroid tumour tissue. Associations between immunohistochemical score and several clinicopathological parameters were examined. In total, 131 thyroid tissue specimens were included. High levels of CTGF protein were observed in papillary thyroid carcinoma tissue; benign thyroid tumour tissue scored negatively for CTGF protein. In papillary thyroid carcinoma, there was a significant relationship between high CTGF protein levels and Union for International Cancer Control disease stage III-IV, and presence of lymph node metastasis. In papillary thyroid carcinomas, CTGF protein levels were not significantly associated with sex or age. These findings suggest that the CTGF protein level is increased in papillary thyroid carcinoma cells compared with benign thyroid tumours. CTGF expression might play a role in the development of malignant tumours in the thyroid.

Residual Expression of the Reprogramming Factors Prevents Differentiation of iPSC Generated from Human Fibroblasts and Cord Blood CD34+ Progenitors

PubMed Central

Ramos-Mejía, Verónica; Montes, Rosa; Bueno, Clara; Ayllón, Verónica; Real, Pedro J.; Rodríguez, René; Menendez, Pablo

2012-01-01

Human induced pluripotent stem cells (hiPSC) have been generated from different tissues, with the age of the donor, tissue source and specific cell type influencing the reprogramming process. Reprogramming hematopoietic progenitors to hiPSC may provide a very useful cellular system for modelling blood diseases. We report the generation and complete characterization of hiPSCs from human neonatal fibroblasts and cord blood (CB)-derived CD34+ hematopoietic progenitors using a single polycistronic lentiviral vector containing an excisable cassette encoding the four reprogramming factors Oct4, Klf4, Sox2 and c-myc (OKSM). The ectopic expression of OKSM was fully silenced upon reprogramming in some hiPSC clones and was not reactivated upon differentiation, whereas other hiPSC clones failed to silence the transgene expression, independently of the cell type/tissue origin. When hiPSC were induced to differentiate towards hematopoietic and neural lineages those hiPSC which had silenced OKSM ectopic expression displayed good hematopoietic and early neuroectoderm differentiation potential. In contrast, those hiPSC which failed to switch off OKSM expression were unable to differentiate towards either lineage, suggesting that the residual expression of the reprogramming factors functions as a developmental brake impairing hiPSC differentiation. Successful adenovirus-based Cre-mediated excision of the provirus OKSM cassette in CB-derived CD34+ hiPSC with residual transgene expression resulted in transgene-free hiPSC clones with significantly improved differentiation capacity. Overall, our findings confirm that residual expression of reprogramming factors impairs hiPSC differentiation. PMID:22545141

Human endogenous retroviruses: nature, occurrence, and clinical implications in human disease.

PubMed Central

Urnovitz, H B; Murphy, W H

1996-01-01

Retroviral diagnostics have become standard in human laboratory medicine. While current emphasis is placed on the human exogenous viruses (human immunodeficiency virus and human T-cell leukemia virus), evidence implicating human endogenous retroviruses (HERVs) in various human disease entities continues to mount. Literature on the occurrence of HERVs in human tissues and cells was analyzed. Substantial evidence documents that retrovirus particles were clearly demonstrable in various tissues and cells in both health and disease and were abundant in the placenta and that their occurrence could be implicated in some of the reproductive diseases. The characteristics of HERVs are summarized, mechanisms of replication and regulation are outlined, and the consistent hormonal responsiveness of HERVs is noted. Clear evidence implicating HERV gene products as participants in glomerulonephritis in some cases of systemic lupus erythematosus is adduced. Data implicating HERVs as etiologic factors in reproductive diseases, in some of the autoimmune diseases, in some forms of rheumatoid arthritis and connective tissue disease, in psoriasis, and in some of the inflammatory neurologic diseases are reviewed. The current major needs are to improve methods for HERV detection, to identify the most appropriate HERV prototypes, and to develop diagnostic reagents so that the putative biologic and pathologic roles of HERVs can be better evaluated. PMID:8665478

Proteomic analysis of human dental cementum and alveolar bone.

PubMed

Salmon, Cristiane R; Tomazela, Daniela M; Ruiz, Karina Gonzales Silvério; Foster, Brian L; Paes Leme, Adriana Franco; Sallum, Enilson Antonio; Somerman, Martha J; Nociti, Francisco H

2013-10-08

Dental cementum (DC) is a bone-like tissue covering the tooth root and responsible for attaching the tooth to the alveolar bone (AB) via the periodontal ligament (PDL). Studies have unsuccessfully tried to identify factors specific to DC versus AB, in an effort to better understand DC development and regeneration. The present study aimed to use matched human DC and AB samples (n=7) to generate their proteomes for comparative analysis. Bone samples were harvested from tooth extraction sites, whereas DC samples were obtained from the apical root portion of extracted third molars. Samples were denatured, followed by protein extraction reduction, alkylation and digestion for analysis by nanoAcquity HPLC system and LTQ-FT Ultra. Data analysis demonstrated that a total of 318 proteins were identified in AB and DC. In addition to shared proteins between these tissues, 105 and 83 proteins exclusive to AB or DC were identified, respectively. This is the first report analyzing the proteomic composition of human DC matrix and identifying putative unique and enriched proteins in comparison to alveolar bone. These findings may provide novel insights into developmental differences between DC and AB, and identify candidate biomarkers that may lead to more efficient and predictable therapies for periodontal regeneration. Periodontal disease is a highly prevalent disease affecting the world population, which involves breakdown of the tooth supporting tissues, the periodontal ligament, alveolar bone, and dental cementum. The lack of knowledge on specific factors that differentiate alveolar bone and dental cementum limits the development of more efficient and predictable reconstructive therapies. In order to better understand cementum development and potentially identify factors to improve therapeutic outcomes, we took the unique approach of using matched patient samples of dental cementum and alveolar bone to generate and compare a proteome list for each tissue. A potential biomarker for dental cementum was identified, superoxide dismutase 3 (SOD3), which is found in cementum and cementum-associated cells in mouse, pig, and human tissues. These findings may provide novel insights into developmental differences between alveolar bone and dental cementum, and represent the basis for improved and more predictable therapies. © 2013.

Utilizing two-photon fluorescence and second harmonic generation microscopy to study human bone marrow mesenchymal stem cell morphogenesis in chitosan scaffold

NASA Astrophysics Data System (ADS)

Su, Ping-Jung; Huang, Chi-Hsiu; Huang, Yi-You; Lee, Hsuan-Sue; Dong, Chen-Yuan

2008-02-01

A major goal of tissue engineering is to cultivate the cartilage in vitro. One approach is to implant the human bone marrow mesenchymal stem cells into the three dimensional biocompatible and biodegradable material. Through the action of the chondrogenic factor TGF-β3, the stem cells can be induced to secrete collagen. In this study, mesenchymal stem cells are implanted on the chitosan scaffold and TGF-β3 was added to produce the cartilage tissue and TP autofluorescence and SHG microscopy was used to image the process of chondrogenesis. With additional development, multiphoton microscopy can be developed into an effective tool for evaluating the quality of tissue engineering products.

Increased COX-2 expression in epithelial and stromal cells of high mammographic density tissues and in a xenograft model of mammographic density.

PubMed

Chew, G L; Huo, C W; Huang, D; Hill, P; Cawson, J; Frazer, H; Hopper, J L; Haviv, I; Henderson, M A; Britt, K; Thompson, E W

2015-08-01

Mammographic density (MD) adjusted for age and body mass index is one of the strongest known risk factors for breast cancer. Given the high attributable risk of MD for breast cancer, chemoprevention with a safe and available agent that reduces MD and breast cancer risk would be beneficial. Cox-2 has been implicated in MD-related breast cancer risk, and was increased in stromal cells in high MD tissues in one study. Our study assessed differential Cox-2 expression in epithelial and stromal cells in paired samples of high and low MD human breast tissue, and in a validated xenograft biochamber model of MD. We also examined the effects of endocrine treatment upon Cox-2 expression in high and low MD tissues in the MD xenograft model. Paired high and low MD human breast tissue samples were immunostained for Cox-2, then assessed for differential expression and staining intensity in epithelial and stromal cells. High and low MD human breast tissues were separately maintained in biochambers in mice treated with Tamoxifen, oestrogen or placebo implants, then assessed for percentage Cox-2 staining in epithelial and stromal cells. Percentage Cox-2 staining was greater for both epithelial (p = 0.01) and stromal cells (p < 0.0001) of high compared with low MD breast tissues. In high MD biochamber tissues, percentage Cox-2 staining was greater in stromal cells of oestrogen-treated versus placebo-treated tissues (p = 0.05).

Human fallopian tube epithelium co-culture with murine ovarian follicles reveals crosstalk in the reproductive cycle.

PubMed

Zhu, Jie; Xu, Yuanming; Rashedi, Alexandra S; Pavone, Mary Ellen; Kim, J Julie; Woodruff, Teresa K; Burdette, Joanna E

2016-11-01

Do interactions between human fallopian tube epithelium and murine follicles occur during an artificial reproductive cycle in a co-culture system in vitro? In a co-culture system, human fallopian tissues responded to the menstrual cycle mimetic by changes in morphology and levels of secreted factors, and increasing murine corpus luteum progesterone secretion. The entire fallopian tube epithelium, including ciliated and secretory cells, can be regulated in the reproductive cycle. Currently, there are no in vitro culture models that can monitor fallopian tissues in real time in response to factors produced by the ovary. In addition, there are no reports on the impact of fallopian tissue on ovarian function during the menstrual cycle. Human fallopian tissue (n = 24) was obtained by routine hysterectomies from women (aged 26-50 years, mean age = 43.6) who had not undergone exogenous hormonal treatment for at least 3 months prior to surgery. CD1 female mice were used for ovarian follicle isolation. The human fallopian epithelium layers were either co-cultured with five murine multilayer secondary follicles (150-180 μm follicles, encapsulated in one alginate gel bead) for 15 days or received stepwise steroid hormone additions for 13 days. The fallopian tissue morphology and cilia beating rate, as measured by an Andor Spinning Disk Confocal, were investigated. Oviduct-specific glycoprotein 1 (OVGP1), human insulin-like growth factor 1 (hIGF1), vascular endothelial growth factor A (VEGF-A) and interleukin 8 (IL8) as biological functional markers were measured either by ELISA or western blot to indicate dynamic changes in the fallopian epithelium during the reproductive cycle generated by mouse follicles or by stepwise steroid hormone induction. Three or four patients in each experiment were recruited for replicates. Data were presented as mean ± SD and further analyzed using one-way ANOVA followed by Tukey's multiple comparisons test. The cultured fallopian tube epithelium responded to exogenous steroid hormone stimulation, as demonstrated by enhanced cilia beating rate (~25% increase, P = 0.04) and an increase in OVGP1 secretion (P = 0.02) in response to 1 nM estradiol (E 2 ) treatment when compared with 0.1 nM E 2 . Conversely, 10 nM progesterone plus 1 nM E 2 suppressed cilia beating rate by ~30% (P = 0.008), while OVGP1 secretion was suppressed by 0.1 nM E 2 plus 50 nM progesterone (P = 0.002 versus 1 nM E 2 alone). Human fallopian tube epithelium was co-cultured with murine secondary follicles to mimic the human menstrual cycle. OVGP1 and VEGF-A secretion from fallopian tissue was similar with stepwise hormone treatment and when cultured with murine follicles. However, the secretion patterns of hIGF1 and IL8 differed in the luteal phase when comparing steroid treatment with follicle co-culture. In co-culture, hIGF1 secretion was suppressed in the luteal versus follicular phase (P = 0.005) but stepwise hormone treatment had no effect on hIGF1. In co-culture, IL8 secretion was also suppressed on luteal phase day 15 (P = 0.013) versus follicular phase day 7, but IL8 secretion increased continuously under high E 2 /progesterone treatment (P = 0.003 for D13 versus D3). In the co-culture system, the corpus luteum continuously produced progesterone in the presence of fallopian tube tissue until Day 18 while, without fallopian tissue, progesterone started to drop from Day 13. One limitation of this study is that murine follicles were used to mimic the human menstrual cycle. However, although secretion patterns of peptide hormones such as inhibins and activins differ in mice and humans, the co-culture system used here did reveal interactions between the tissues that govern reproductive function. In vitro co-culture models of fallopian reproductive tissues with ovarian follicles can provide an important tool for understanding fertility and for uncovering the mechanisms responsible for reduced fertility. In addition, the role of oviductal secretions and how they influence ovarian function, such as the production of progesterone during the menstrual cycle, can be uncovered using this model. None. This work was funded by grants from the NIH (UH3TR001207), the American Cancer Society (RSG-12-230-01-TBG) and NIH (R01EB014806). The authors declare no competing financial interest. © The Author 2016. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Computational Identification of Tissue-Specific Splicing Regulatory Elements in Human Genes from RNA-Seq Data.

PubMed

Badr, Eman; ElHefnawi, Mahmoud; Heath, Lenwood S

2016-01-01

Alternative splicing is a vital process for regulating gene expression and promoting proteomic diversity. It plays a key role in tissue-specific expressed genes. This specificity is mainly regulated by splicing factors that bind to specific sequences called splicing regulatory elements (SREs). Here, we report a genome-wide analysis to study alternative splicing on multiple tissues, including brain, heart, liver, and muscle. We propose a pipeline to identify differential exons across tissues and hence tissue-specific SREs. In our pipeline, we utilize the DEXSeq package along with our previously reported algorithms. Utilizing the publicly available RNA-Seq data set from the Human BodyMap project, we identified 28,100 differentially used exons across the four tissues. We identified tissue-specific exonic splicing enhancers that overlap with various previously published experimental and computational databases. A complicated exonic enhancer regulatory network was revealed, where multiple exonic enhancers were found across multiple tissues while some were found only in specific tissues. Putative combinatorial exonic enhancers and silencers were discovered as well, which may be responsible for exon inclusion or exclusion across tissues. Some of the exonic enhancers are found to be co-occurring with multiple exonic silencers and vice versa, which demonstrates a complicated relationship between tissue-specific exonic enhancers and silencers.

Selective localization of oxytocin receptors and vasopressin 1a receptors in the human brainstem

PubMed Central

Freeman, Sara M.; Smith, Aaron L.; Goodman, Mark M.; Bales, Karen L.

2017-01-01

Intranasal oxytocin affects a suite of human social behaviors, including trust, eye contact, and emotion recognition. However, it is unclear where oxytocin receptors (OXTR) and the structurally related vasopressin 1a receptors (AVPR1a) are expressed in the human brain. We have previously described a reliable, pharmacologically informed receptor autoradiography protocol for visualizing these receptors in postmortem primate brain tissue. We used this technique in human brainstem tissue to identify the neural targets of oxytocin and vasopressin. To determine binding selectivity of the OXTR radioligand and AVPR1a radioligand, sections were incubated in four conditions: radioligand alone, radioligand with the selective AVPR1a competitor SR49059, and radioligand with a low or high concentration of the selective OXTR competitor ALS-II-69. We found selective OXTR binding in the spinal trigeminal nucleus, a conserved region of OXTR expression in all primate species investigated to date. We found selective AVPR1a binding in the nucleus prepositus, an area implicated in eye gaze stabilization. The tissue's postmortem interval was not correlated with either the specific or nonspecific binding of either radioligand, indicating that it will not likely be a factor in similar postmortem studies. This study provides critical data for future studies of OXTR and AVPR1a in human brain tissue. PMID:26911439

76 FR 787 - Notice of Availability of the Recommended Toxicity Equivalence Factors (TEFs) for Human Health...

Federal Register 2010, 2011, 2012, 2013, 2014

2011-01-06

... manufacturing, combustion, and metal processing. There is global contamination of air, soil and water with trace... food chain, and accumulate in the tissues of animals. Human exposures to these compounds occur primarily through eating contaminated foods. The health effects from exposures to dioxins and DLCs have been...

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.

Equid Herpesvirus Type 1 Activates Platelets

PubMed Central

Stokol, Tracy; Yeo, Wee Ming; Burnett, Deborah; DeAngelis, Nicole; Huang, Teng; Osterrieder, Nikolaus; Catalfamo, James

2015-01-01

Equid herpesvirus type 1 (EHV-1) causes outbreaks of abortion and neurological disease in horses. One of the main causes of these clinical syndromes is thrombosis in placental and spinal cord vessels, however the mechanism for thrombus formation is unknown. Platelets form part of the thrombus and amplify and propagate thrombin generation. Here, we tested the hypothesis that EHV-1 activates platelets. We found that two EHV-1 strains, RacL11 and Ab4 at 0.5 or higher plaque forming unit/cell, activate platelets within 10 minutes, causing α-granule secretion (surface P-selectin expression) and platelet microvesiculation (increased small events double positive for CD41 and Annexin V). Microvesiculation was more pronounced with the RacL11 strain. Virus-induced P-selectin expression required plasma and 1.0 mM exogenous calcium. P-selectin expression was abolished and microvesiculation was significantly reduced in factor VII- or X-deficient human plasma. Both P-selectin expression and microvesiculation were re-established in factor VII-deficient human plasma with added purified human factor VIIa (1 nM). A glycoprotein C-deficient mutant of the Ab4 strain activated platelets as effectively as non-mutated Ab4. P-selectin expression was abolished and microvesiculation was significantly reduced by preincubation of virus with a goat polyclonal anti-rabbit tissue factor antibody. Infectious virus could be retrieved from washed EHV-1-exposed platelets, suggesting a direct platelet-virus interaction. Our results indicate that EHV-1 activates equine platelets and that α-granule secretion is a consequence of virus-associated tissue factor triggering factor X activation and thrombin generation. Microvesiculation was only partly tissue factor and thrombin-dependent, suggesting the virus causes microvesiculation through other mechanisms, potentially through direct binding. These findings suggest that EHV-1-induced platelet activation could contribute to the thrombosis that occurs in clinically infected horses and provides a new mechanism by which viruses activate hemostasis. PMID:25905776

Mouse and human BAC transgenes recapitulate tissue-specific expression of the vitamin D receptor in mice and rescue the VDR-null phenotype.

PubMed

Lee, Seong Min; Bishop, Kathleen A; Goellner, Joseph J; O'Brien, Charles A; Pike, J Wesley

2014-06-01

The biological actions of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) are mediated by the vitamin D receptor (VDR), which is expressed in numerous target tissues in a cell type-selective manner. Recent studies using genomic analyses and recombineered bacterial artificial chromosomes (BACs) have defined the specific features of mouse and human VDR gene loci in vitro. In the current study, we introduced recombineered mouse and human VDR BACs as transgenes into mice and explored their expression capabilities in vivo. Individual transgenic mouse strains selectively expressed BAC-derived mouse or human VDR proteins in appropriate vitamin D target tissues, thereby recapitulating the tissue-specific expression of endogenous mouse VDR. The mouse VDR transgene was also regulated by 1,25(OH)2D3 and dibutyryl-cAMP. When crossed into a VDR-null mouse background, both transgenes restored wild-type basal as well as 1,25(OH)2D3-inducible gene expression patterns in the appropriate tissues. This maneuver resulted in the complete rescue of the aberrant phenotype noted in the VDR-null mouse, including systemic features associated with altered calcium and phosphorus homeostasis and disrupted production of parathyroid hormone and fibroblast growth factor 23, and abnormalities associated with the skeleton, kidney, parathyroid gland, and the skin. This study suggests that both mouse and human VDR transgenes are capable of recapitulating basal and regulated expression of the VDR in the appropriate mouse tissues and restore 1,25(OH)2D3 function. These results provide a baseline for further dissection of mechanisms integral to mouse and human VDR gene expression and offer the potential to explore the consequence of selective mutations in VDR proteins in vivo.

Induction of Tissue Factor Pathway Inhibitor 2 by hCG Regulates Periovulatory Gene Expression and Plasmin Activity.

PubMed

Puttabyatappa, Muraly; Al-Alem, Linah F; Zakerkish, Farnosh; Rosewell, Katherine L; Brännström, Mats; Curry, Thomas E

2017-01-01

Increased proteolytic activity is a key event that aids in breakdown of the follicular wall to permit oocyte release. How the protease activity is regulated is still unknown. We hypothesize that tissue factor pathway inhibitor 2 (TFPI2), a Kunitz-type serine protease inhibitor, plays a role in regulating periovulatory proteolytic activity as in other tissues. TFPI2 is secreted into the extracellular matrix (ECM) where it is postulated to regulate physiological ECM remodeling. The expression profile of TFPI2 during the periovulatory period was assessed utilizing a well-characterized human menstrual cycle model and a gonadotropin-primed rat model. Administration of an ovulatory dose of human chorionic gonadotropin (hCG) increased TFPI2 expression dramatically in human and rat granulosa and theca cells. This increase in Tfpi2 expression in rat granulosa cells required hCG-mediated epidermal growth factor, protein kinase A, mitogen-activated protein kinase (MAPK) 1/2, p38 MAPK and protease activated receptor 1-dependent cell signaling. A small interferingRNA-mediated knockdown of TFPI2 in rat granulosa cells resulted in increased plasmin activity in the granulosa cell conditioned media. Knockdown of TFPI2 also reduced expression of multiple genes including interleukin 6 (Il6) and amphiregulin (Areg). Overexpression of TFPI2 using an adenoviral vector partially restored the expression of Il6 and Areg in TFPI2 siRNA treated rat granulosa cells. These data support the hypothesis that TFPI2 is important for moderating plasmin activity and regulating granulosa cell gene expression during the periovulatory period. We, therefore, propose that through these actions, TFPI2 aids in the tissue remodeling taking place during follicular rupture and corpus luteum formation. Copyright © 2017 by the Endocrine Society.

Induction of Tissue Factor Pathway Inhibitor 2 by hCG Regulates Periovulatory Gene Expression and Plasmin Activity

PubMed Central

Puttabyatappa, Muraly; Al-Alem, Linah F.; Zakerkish, Farnosh; Rosewell, Katherine L.; Brännström, Mats

2017-01-01

Increased proteolytic activity is a key event that aids in breakdown of the follicular wall to permit oocyte release. How the protease activity is regulated is still unknown. We hypothesize that tissue factor pathway inhibitor 2 (TFPI2), a Kunitz-type serine protease inhibitor, plays a role in regulating periovulatory proteolytic activity as in other tissues. TFPI2 is secreted into the extracellular matrix (ECM) where it is postulated to regulate physiological ECM remodeling. The expression profile of TFPI2 during the periovulatory period was assessed utilizing a well-characterized human menstrual cycle model and a gonadotropin-primed rat model. Administration of an ovulatory dose of human chorionic gonadotropin (hCG) increased TFPI2 expression dramatically in human and rat granulosa and theca cells. This increase in Tfpi2 expression in rat granulosa cells required hCG-mediated epidermal growth factor, protein kinase A, mitogen-activated protein kinase (MAPK) 1/2, p38 MAPK and protease activated receptor 1-dependent cell signaling. A small interferingRNA-mediated knockdown of TFPI2 in rat granulosa cells resulted in increased plasmin activity in the granulosa cell conditioned media. Knockdown of TFPI2 also reduced expression of multiple genes including interleukin 6 (Il6) and amphiregulin (Areg). Overexpression of TFPI2 using an adenoviral vector partially restored the expression of Il6 and Areg in TFPI2 siRNA treated rat granulosa cells. These data support the hypothesis that TFPI2 is important for moderating plasmin activity and regulating granulosa cell gene expression during the periovulatory period. We, therefore, propose that through these actions, TFPI2 aids in the tissue remodeling taking place during follicular rupture and corpus luteum formation. PMID:27813674

Periodontal ligament stem/progenitor cells with protein-releasing scaffolds for cementum formation and integration on dentin surface.

PubMed

Cho, Hankyu; Tarafder, Solaiman; Fogge, Michael; Kao, Kristy; Lee, Chang H

2016-11-01

Purpose/Aim: Cementogenesis is a critical step in periodontal tissue regeneration given the essential role of cementum in anchoring teeth to the alveolar bone. This study is designed to achieve integrated cementum formation on the root surfaces of human teeth using growth factor-releasing scaffolds with periodontal ligament stem/progenitor cells (PDLSCs). Human PDLSCs were sorted by CD146 expression, and characterized using CFU-F assay and induced multi-lineage differentiation. Polycaprolactone scaffolds were fabricated using 3D printing, embedded with poly(lactic-co-glycolic acids) (PLGA) microspheres encapsulating connective tissue growth factor (CTGF), bone morphogenetic protein-2 (BMP-2), or bone morphogenetic protein-7 (BMP-7). After removing cementum on human tooth roots, PDLSC-seeded scaffolds were placed on the exposed dentin surface. After 6-week culture with cementogenic/osteogenic medium, cementum formation and integration were evaluated by histomorphometric analysis, immunofluorescence, and qRT-PCR. Periodontal ligament (PDL) cells sorted by CD146 and single-cell clones show a superior clonogenecity and multipotency as compared with heterogeneous populations. After 6 weeks, all the growth factor-delivered groups showed resurfacing of dentin with a newly formed cementum-like layer as compared with control. BMP-2 and BMP-7 showed de novo formation of tissue layers significantly thicker than all the other groups, whereas CTGF and BMP-7 resulted in significantly improved integration on the dentin surface. The de novo mineralized tissue layer seen in BMP-7-treated samples expressed cementum matrix protein 1 (CEMP1). Consistently, BMP-7 showed a significant increase in CEMP1 mRNA expression. Our findings represent important progress in stem cell-based cementum regeneration as an essential part of periodontium regeneration.

Localization of matrix metalloproteinases, (MMPs) their tissue inhibitors, and vascular endothelial growth factor (VEGF) in growth plates of children and adolescents indicates a role for MMPs in human postnatal growth and skeletal maturation.

PubMed

Haeusler, G; Walter, I; Helmreich, M; Egerbacher, M

2005-05-01

Numerous studies have focused on the expression, regulation, and biological significance of matrix metalloproteinases (MMPs) in the growth plate. Findings in mouse knockout models and in vitro data from various species indicate that MMPs not only degrade extracellular matrix components but may regulate the activity of local growth factors. In this study we investigated the presence, distribution, and activity of various MMPs and inhibitors, tissue transglutaminase (tTG or TG2) and vascular endothelial growth factor (VEGF) in the human child and adolescent growth plates by means of immunohistochemistry and gelatin zymography. Tissue was derived during orthopedic surgery (epiphysiodesis) in two prepubertal and four pubertal patients.MMP-2 and MMP-14 were present in reserve cell chondrocytes. MMP-14 was the most prominent MMP within all zones of the growth plate including proliferating chondrocytes. MMP-1 and MMP-13 (collagenases 1 and 3), MMP-9 (gelatinases B), MMP-10, and MMP-11 (stromelysins) and VEGF were positive in hypertrophic chondrocytes and osteoblasts. MMP-2 showed the same expression pattern but was negative in osteoblasts. Osteoclasts stained positive for MMP-9, MMP-2, and TG2. Tissue inhibitor of MMP (TIMP)-1 was present in all zones of the growth plate, osteoblasts, and osteoclasts; TIMP-2 was found in hypertrophic chondrocytes and osteoblasts. In summary, the presence of MMPs, TIMPs, TG2, and VEGF in our study indicated that the MMPs are relevant in growth plate physiology during the postnatal period in humans. The specific location of MMP expression within the growth plate may be the basis for further studies on the role of MMPs in the local regulation of chondrocyte differentiation, proliferation, and ossification at the chondroosseus junction.

Insulin-like growth factor I (IGF-1) Ec/Mechano Growth factor--a splice variant of IGF-1 within the growth plate.

PubMed

Schlegel, Werner; Raimann, Adalbert; Halbauer, Daniel; Scharmer, Daniela; Sagmeister, Susanne; Wessner, Barbara; Helmreich, Magdalena; Haeusler, Gabriele; Egerbacher, Monika

2013-01-01

Human insulin-like growth factor 1 Ec (IGF-1Ec), also called mechano growth factor (MGF), is a splice variant of insulin-like growth factor 1 (IGF-1), which has been shown in vitro as well as in vivo to induce growth and hypertrophy in mechanically stimulated or damaged muscle. Growth, hypertrophy and responses to mechanical stimulation are important reactions of cartilaginous tissues, especially those in growth plates. Therefore, we wanted to ascertain if MGF is expressed in growth plate cartilage and if it influences proliferation of chondrocytes, as it does in musculoskeletal tissues. MGF expression was analyzed in growth plate and control tissue samples from piglets aged 3 to 6 weeks. Furthermore, growth plate chondrocyte cell culture was used to evaluate the effects of the MGF peptide on proliferation. We showed that MGF is expressed in considerable amounts in the tissues evaluated. We found the MGF peptide to be primarily located in the cytoplasm, and in some instances, it was also found in the nucleus of the cells. Addition of MGF peptides was not associated with growth plate chondrocyte proliferation.

Insulin-Like Growth Factor I (IGF-1) Ec/Mechano Growth Factor – A Splice Variant of IGF-1 within the Growth Plate

PubMed Central

Schlegel, Werner; Raimann, Adalbert; Halbauer, Daniel; Scharmer, Daniela; Sagmeister, Susanne; Wessner, Barbara; Helmreich, Magdalena; Haeusler, Gabriele; Egerbacher, Monika

2013-01-01

Human insulin-like growth factor 1 Ec (IGF-1Ec), also called mechano growth factor (MGF), is a splice variant of insulin-like growth factor 1 (IGF-1), which has been shown in vitro as well as in vivo to induce growth and hypertrophy in mechanically stimulated or damaged muscle. Growth, hypertrophy and responses to mechanical stimulation are important reactions of cartilaginous tissues, especially those in growth plates. Therefore, we wanted to ascertain if MGF is expressed in growth plate cartilage and if it influences proliferation of chondrocytes, as it does in musculoskeletal tissues. MGF expression was analyzed in growth plate and control tissue samples from piglets aged 3 to 6 weeks. Furthermore, growth plate chondrocyte cell culture was used to evaluate the effects of the MGF peptide on proliferation. We showed that MGF is expressed in considerable amounts in the tissues evaluated. We found the MGF peptide to be primarily located in the cytoplasm, and in some instances, it was also found in the nucleus of the cells. Addition of MGF peptides was not associated with growth plate chondrocyte proliferation. PMID:24146828

Development of a 3D co-culture model using human stem ...

EPA Pesticide Factsheets

Morphogenetic tissue fusion is a critical and complex event in embryonic development and failure of this event leads to birth defects, such as cleft palate. Palatal fusion requires adhesion and subsequent dissolution of the medial epithelial layer of the mesenchymal palatal shelves, and is regulated by the growth factors EGF and TGFβ, and others, although the complete regulatory mechanism is not understood. Three dimensional (3D) organotypic models allow us to mimic the native architecture of human tissue to facilitate the study of tissue dynamics and their responses to developmental toxicants. Our goal was to develop and characterize a spheroidal model of palatal fusion to investigate the mechanisms regulating fusion with exposure to growth factors and chemicals in the ToxCast program known to disrupt this event. We present a spheroidal model using human umbilical-derived mesenchymal stem cells (hMSC) spheroid cores cultured for 13 days and then coated with MaxGel™ basement membrane and a layer of human progenitor epithelial keratinocytes (hPEK) (hMSC+hPEK spheroids). We characterized the growth, differentiation, proliferation and fusion activity of the model. Spheroid diameter was dependent on hMSC seeding density, size of the seeding wells, time in culture, and type of medium. hMSC spheroid growth was enhanced with osteogenic differentiation medium. Alkaline phosphatase activity in the hMSC spheroid, indicating osteogenic differentiation, increased in inte

A human osteoarthritis osteochondral organ culture model for cartilage tissue engineering.

PubMed

Yeung, P; Zhang, W; Wang, X N; Yan, C H; Chan, B P

2018-04-01

In vitro human osteoarthritis (OA)-mimicking models enabling pathophysiological studies and evaluation of emerging therapies such as cartilage tissue engineering are of great importance. We describe the development and characterization of a human OA osteochondral organ culture. We also apply this model for evaluation of the phenotype maintenance of a human MSC derived engineered cartilage, as an example of emerging therapeutics, under long term exposure to the OA-mimicking environment. We also test the sensitivity of the model to a series of external factors and a potential disease-modifying agent, in terms of chondrogenic phenotype maintenance of the engineered cartilage, under OA-mimicking environment. Excised joint tissues from total knee replacement surgeries were carved into numerous miniaturized and standardized osteochondral plugs for subsequent OA organ culture. The organ cultures were characterized in detail before being co-cultured with a tissue engineered cartilage. The chondrogenic phenotype of the tissue engineered cartilage co-cultured in long term up to 8 weeks under this OA-mimicking microenvironment was evaluated. Using the same co-culture model, we also screened for a number of biomimetic environmental factors, including oxygen tension, the presence of serum and the application of compression loading. Finally, we studied the effect of a matrix metalloprotease inhibitor, as an example of potential disease-modifying agents, on the co-cultured engineered cartilage. We demonstrate that cells in the OA organ culture were viable while both the typical chondrogenic phenotype and the characteristic OA phenotype were maintained for long period of time. We then demonstrate that upon co-culture with the OA-mimicking organ culture, the engineered cartilage initially exhibited a more fibrocartilage phenotype but progressively reverted back to the chondrogenic phenotype upon long term co-culture up to 8 weeks. The engineered cartilage was also found to be sensitive to all biomimetic environmental factors screened (oxygen tension, serum and compression). Moreover, under the effect of a MMP inhibitor, the chondrogenic phenotype of engineered cartilage was better maintained. We demonstrated the development of a human OA osteochondral organ culture and tested the feasibility and potential of using this model as an in vitro evaluation tool for emerging cartilage therapies. Copyright © 2018 Elsevier Ltd. All rights reserved.

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

PubMed Central

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

2015-01-01

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

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

PubMed

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

2015-12-01

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

Characterization of human myoblast cultures for tissue engineering.

PubMed

Stern-Straeter, Jens; Bran, Gregor; Riedel, Frank; Sauter, Alexander; Hörmann, Karl; Goessler, Ulrich Reinhart

2008-01-01

Skeletal muscle tissue engineering, a promising specialty, aims at the reconstruction of skeletal muscle loss. In vitro tissue engineering attempts to achieve this goal by creating differentiated, functional muscle tissue through a process in which stem cells are extracted from the patient, e.g. by muscle biopsies, expanded and differentiated in a controlled environment, and subsequently re-implanted. A prerequisite for this undertaking is the ability to cultivate and differentiate human skeletal muscle cell cultures. Evidently, optimal culture conditions must be investigated for later clinical utilization. We therefore analysed the proliferation of human cells in different environments and evaluated the differentiation potential of different culture media. It was shown that human myoblasts have a higher rate of proliferation in the alamarBlue assay when cultured on gelatin-coated culture flasks rather than polystyrene-coated flasks. We also demonstrated that myoblasts treated with a culture medium with a high concentration of growth factors [growth medium (GM)] showed a higher proliferation compared to cultures treated with a culture medium with lower amounts of growth factors [differentiation medium (DM)]. Differentiation of human myoblast cell cultures treated with GM and DM was analysed until day 16 and myogenesis was verified by expression of MyoD, myogenin, alpha-sarcomeric actin and myosin heavy chain by semi-quantitative RT-PCR. Immunohistochemical staining for desmin, Myf-5 and alpha-sarcomeric actin was performed to verify the myogenic phenotype of extracted satellite cells and to prove the maturation of cells. Cultures treated with DM showed positive staining for alpha-sarcomeric actin. Notably, markers of differentiation were also detected in cultures treated with GM, but there was no formation of myotubes. In the enzymatic assay of creatine phosphokinase, cultures treated with DM showed a higher activity, evidencing a higher degree of differentiation. In this study, we obtained detailed information regarding the cultivation and differentiation of human myoblast cultures in different environments. By exploring optimal culture conditions for skeletal muscle tissue engineering, we acquired culture data for comparison with other sources of stem cells in order to find the most applicable stem cell for focussed clinical usage.

The tumour-associated glycoprotein podoplanin is expressed in fibroblast-like synoviocytes of the hyperplastic synovial lining layer in rheumatoid arthritis

PubMed Central

2011-01-01

Introduction Activated fibroblast-like synoviocytes (FLSs) in rheumatoid arthritis (RA) share many characteristics with tumour cells and are key mediators of synovial tissue transformation and joint destruction. The glycoprotein podoplanin is upregulated in the invasive front of several human cancers and has been associated with epithelial-mesenchymal transition, increased cell migration and tissue invasion. The aim of this study was to investigate whether podoplanin is expressed in areas of synovial transformation in RA and especially in promigratory RA-FLS. Methods Podoplanin expression in human synovial tissue from 18 RA patients and nine osteoarthritis (OA) patients was assessed by immunohistochemistry and confirmed by Western blot analysis. The expression was related to markers of synoviocytes and myofibroblasts detected by using confocal immunofluoresence microscopy. Expression of podoplanin, with or without the addition of proinflammatory cytokines and growth factors, in primary human FLS was evaluated by using flow cytometry. Results Podoplanin was highly expressed in cadherin-11-positive cells throughout the synovial lining layer in RA. The expression was most pronounced in areas with lining layer hyperplasia and high matrix metalloproteinase 9 expression, where it coincided with upregulation of α-smooth muscle actin (α-sma). The synovium in OA was predominantly podoplanin-negative. Podoplanin was expressed in 50% of cultured primary FLSs, and the expression was increased by interleukin 1β, tumour necrosis factor α and transforming growth factor β receptor 1. Conclusions Here we show that podoplanin is highly expressed in FLSs of the invading synovial tissue in RA. The concomitant upregulation of α-sma and podoplanin in a subpopulation of FLSs indicates a myofibroblast phenotype. Proinflammatory mediators increased the podoplanin expression in cultured RA-FLS. We conclude that podoplanin might be involved in the synovial tissue transformation and increased migratory potential of activated FLSs in RA. PMID:21385358

Tissue engineering skeletal muscle for orthopaedic applications

NASA Technical Reports Server (NTRS)

Payumo, Francis C.; Kim, Hyun D.; Sherling, Michael A.; Smith, Lee P.; Powell, Courtney; Wang, Xiao; Keeping, Hugh S.; Valentini, Robert F.; Vandenburgh, Herman H.

2002-01-01

With current technology, tissue-engineered skeletal muscle analogues (bioartificial muscles) generate too little active force to be clinically useful in orthopaedic applications. They have been engineered genetically with numerous transgenes (growth hormone, insulinlike growth factor-1, erythropoietin, vascular endothelial growth factor), and have been shown to deliver these therapeutic proteins either locally or systemically for months in vivo. Bone morphogenetic proteins belonging to the transforming growth factor-beta superfamily are osteoinductive molecules that drive the differentiation pathway of mesenchymal cells toward the chondroblastic or osteoblastic lineage, and stimulate bone formation in vivo. To determine whether skeletal muscle cells endogenously expressing bone morphogenetic proteins might serve as a vehicle for systemic bone morphogenetic protein delivery in vivo, proliferating skeletal myoblasts (C2C12) were transduced with a replication defective retrovirus containing the gene for recombinant human bone morphogenetic protein-6 (C2BMP-6). The C2BMP-6 cells constitutively expressed recombinant human bone morphogenetic protein-6 and synthesized bioactive recombinant human bone morphogenetic protein-6, based on increased alkaline phosphatase activity in coincubated mesenchymal cells. C2BMP-6 cells did not secrete soluble, bioactive recombinant human bone morphogenetic protein-6, but retained the bioactivity in the cell layer. Therefore, genetically-engineered skeletal muscle cells might serve as a platform for long-term delivery of osteoinductive bone morphogenetic proteins locally.

A tissue dose-based comparative exposure assessment of manganese using physiologically based pharmacokinetic modeling—The importance of homeostatic control for an essential metal

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

Gentry, P. Robinan, E-mail: rgentry@ramboll.com

A physiologically-based pharmacokinetic (PBPK) model (Schroeter et al., 2011) was applied to simulate target tissue manganese (Mn) concentrations following occupational and environmental exposures. These estimates of target tissue Mn concentrations were compared to determine margins of safety (MOS) and to evaluate the biological relevance of applying safety factors to derive acceptable Mn air concentrations. Mn blood concentrations measured in occupational studies permitted verification of the human PBPK models, increasing confidence in the resulting estimates. Mn exposure was determined based on measured ambient air Mn concentrations and dietary data in Canada and the United States (US). Incorporating dietary and inhalation exposuresmore » into the models indicated that increases in target tissue concentrations above endogenous levels only begin to occur when humans are exposed to levels of Mn in ambient air (i.e. > 10 μg/m{sup 3}) that are far higher than those currently measured in Canada or the US. A MOS greater than three orders of magnitude was observed, indicating that current Mn air concentrations are far below concentrations that would be required to produce the target tissue Mn concentrations associated with subclinical neurological effects. This application of PBPK modeling for an essential element clearly demonstrates that the conventional application of default factors to “convert” an occupational exposure to an equivalent continuous environmental exposure, followed by the application of safety factors, is not appropriate in the case of Mn. PBPK modeling demonstrates that the relationship between ambient Mn exposures and dose-to-target tissue is not linear due to normal tissue background levels and homeostatic controls. - Highlights: • Manganese is an essential nutrient, adding complexity to its risk assessment. • Nonlinearities in biological processes are important for manganese risk assessment. • A PBPK model was used to estimate target tissue concentrations of manganese. • An MOS approach also considered target tissue concentrations for ambient exposures. • Relationships between ambient Mn exposures and dose-to-target tissue are not linear.« less

Implant materials generate different peri-implant inflammatory factors: poly-ether-ether-ketone promotes fibrosis and microtextured titanium promotes osteogenic factors.

PubMed

Olivares-Navarrete, Rene; Hyzy, Sharon L; Slosar, Paul J; Schneider, Jennifer M; Schwartz, Zvi; Boyan, Barbara D

2015-03-15

An in vitro study examining factors produced by human mesenchymal stem cells on spine implant materials. The aim of this study was to examine whether the inflammatory microenvironment generated by cells on titanium-aluminum-vanadium (Ti-alloy, TiAlV) surfaces is affected by surface microtexture and whether it differs from that generated on poly-ether-ether-ketone (PEEK). Histologically, implants fabricated from PEEK have a fibrous connective tissue surface interface whereas Ti-alloy implants demonstrate close approximation with surrounding bone. Ti-alloy surfaces with complex micron/submicron scale roughness promote osteoblastic differentiation and foster a specific cellular environment that favors bone formation whereas PEEK favors fibrous tissue formation. Human mesenchymal stem cells were cultured on tissue culture polystyrene, PEEK, smooth TiAlV, or macro-/micro-/nano-textured rough TiAlV (mmnTiAlV) disks. Osteoblastic differentiation and secreted inflammatory interleukins were assessed after 7 days. Fold changes in mRNAs for inflammation, necrosis, DNA damage, or apoptosis with respect to tissue culture polystyrene were measured by low-density polymerase chain reaction array. Data were analyzed by analysis of variance, followed by Bonferroni's correction of Student's t-test. Cells on PEEK upregulated mRNAs for chemokine ligand-2, interleukin (IL) 1β, IL6, IL8, and tumor necrosis factor. Cells grown on the mmnTiAlV had an 8-fold reduction in mRNAs for toll-like receptor-4. Cells grown on mmnTiAlV had reduced levels of proinflammatory interleukins. Cells on PEEK had higher mRNAs for factors strongly associated with cell death/apoptosis, whereas cells on mmnTiAlV exhibited reduced cytokine factor levels. All results were significant (P < 0.05). These results suggest that fibrous tissue around PEEK implants may be due to several factors: reduced osteoblastic differentiation of progenitor cells and production of an inflammatory environment that favors cell death via apoptosis and necrosis. Ti alloy surfaces with complex macro/micro/nanoscale roughness promote osteoblastic differentiation and foster a specific cellular environment that favors bone formation. N/A.

Engineered cartilage using primary chondrocytes cultured in a porous cartilage-derived matrix

PubMed Central

Cheng, Nai-Chen; Estes, Bradley T; Young, Tai-Horng; Guilak, Farshid

2011-01-01

Aim To investigate the cell growth, matrix accumulation and mechanical properties of neocartilage formed by human or porcine articular chondrocytes on a porous, porcine cartilage-derived matrix (CDM) for use in cartilage tissue engineering. Materials & methods We examined the physical properties, cell infiltration and matrix accumulation in different formulations of CDM and selected a CDM made of homogenized cartilage slurry as an appropriate scaffold for long-term culture of human and porcine articular chondrocytes. Results The CDM scaffold supported growth and proliferation of both human and porcine chondrocytes. Histology and immunohistochemistry showed abundant cartilage-specific macromolecule deposition at day 28. Human chondrocytes migrated throughout the CDM, showing a relatively homogeneous distribution of new tissue accumulation, whereas porcine chondrocytes tended to form a proteoglycan-rich layer primarily on the surfaces of the scaffold. Human chondrocyte-seeded scaffolds had a significantly lower aggregate modulus and hydraulic permeability at day 28. Conclusions These data show that a scaffold derived from native porcine articular cartilage can support neocartilage formation in the absence of exogenous growth factors. The overall characteristics and properties of the constructs depend on factors such as the concentration of CDM used, the porosity of the scaffold, and the species of chondrocytes. PMID:21175289

Advancing biomaterials of human origin for tissue engineering

PubMed Central

Chen, Fa-Ming; Liu, Xiaohua

2015-01-01

Biomaterials have played an increasingly prominent role in the success of biomedical devices and in the development of tissue engineering, which seeks to unlock the regenerative potential innate to human tissues/organs in a state of deterioration and to restore or reestablish normal bodily function. Advances in our understanding of regenerative biomaterials and their roles in new tissue formation can potentially open a new frontier in the fast-growing field of regenerative medicine. Taking inspiration from the role and multi-component construction of native extracellular matrices (ECMs) for cell accommodation, the synthetic biomaterials produced today routinely incorporate biologically active components to define an artificial in vivo milieu with complex and dynamic interactions that foster and regulate stem cells, similar to the events occurring in a natural cellular microenvironment. The range and degree of biomaterial sophistication have also dramatically increased as more knowledge has accumulated through materials science, matrix biology and tissue engineering. However, achieving clinical translation and commercial success requires regenerative biomaterials to be not only efficacious and safe but also cost-effective and convenient for use and production. Utilizing biomaterials of human origin as building blocks for therapeutic purposes has provided a facilitated approach that closely mimics the critical aspects of natural tissue with regard to its physical and chemical properties for the orchestration of wound healing and tissue regeneration. In addition to directly using tissue transfers and transplants for repair, new applications of human-derived biomaterials are now focusing on the use of naturally occurring biomacromolecules, decellularized ECM scaffolds and autologous preparations rich in growth factors/non-expanded stem cells to either target acceleration/magnification of the body's own repair capacity or use nature's paradigms to create new tissues for restoration. In particular, there is increasing interest in separating ECMs into simplified functional domains and/or biopolymeric assemblies so that these components/constituents can be discretely exploited and manipulated for the production of bioscaffolds and new biomimetic biomaterials. Here, following an overview of tissue auto-/allo-transplantation, we discuss the recent trends and advances as well as the challenges and future directions in the evolution and application of human-derived biomaterials for reconstructive surgery and tissue engineering. In particular, we focus on an exploration of the structural, mechanical, biochemical and biological information present in native human tissue for bioengineering applications and to provide inspiration for the design of future biomaterials. PMID:27022202

Differential expression of THOC1 and ALY mRNP biogenesis/export factors in human cancers.

PubMed

Domínguez-Sánchez, María S; Sáez, Carmen; Japón, Miguel A; Aguilera, Andrés; Luna, Rosa

2011-02-17

One key step in gene expression is the biogenesis of mRNA ribonucleoparticle complexes (mRNPs). Formation of the mRNP requires the participation of a number of conserved factors such as the THO complex. THO interacts physically and functionally with the Sub2/UAP56 RNA-dependent ATPase, and the Yra1/REF1/ALY RNA-binding protein linking transcription, mRNA export and genome integrity. Given the link between genome instability and cancer, we have performed a comparative analysis of the expression patterns of THOC1, a THO complex subunit, and ALY in tumor samples. The mRNA levels were measured by quantitative real-time PCR and hybridization of a tumor tissue cDNA array; and the protein levels and distribution by immunostaining of a custom tissue array containing a set of paraffin-embedded samples of different tumor and normal tissues followed by statistical analysis. We show that the expression of two mRNP factors, THOC1 and ALY are altered in several tumor tissues. THOC1 mRNA and protein levels are up-regulated in ovarian and lung tumors and down-regulated in those of testis and skin, whereas ALY is altered in a wide variety of tumors. In contrast to THOC1, ALY protein is highly detected in normal proliferative cells, but poorly in high-grade cancers. These results suggest a differential connection between tumorogenesis and the expression levels of human THO and ALY. This study opens the possibility of defining mRNP biogenesis factors as putative players in cell proliferation that could contribute to tumor development.

Functional roles of fibroblast growth factor receptors (FGFRs) signaling in human cancers.

PubMed

Tiong, Kai Hung; Mah, Li Yen; Leong, Chee-Onn

2013-12-01

The fibroblast growth factor receptors (FGFRs) regulate important biological processes including cell proliferation and differentiation during development and tissue repair. Over the past decades, numerous pathological conditions and developmental syndromes have emerged as a consequence of deregulation in the FGFRs signaling network. This review aims to provide an overview of FGFR family, their complex signaling pathways in tumorigenesis, and the current development and application of therapeutics targeting the FGFRs signaling for treatment of refractory human cancers.

Three-dimensional tissue assemblies: novel models for the study of Salmonella enterica serovar Typhimurium pathogenesis

NASA Technical Reports Server (NTRS)

Nickerson, C. A.; Goodwin, T. J.; Terlonge, J.; Ott, C. M.; Buchanan, K. L.; Uicker, W. C.; Emami, K.; LeBlanc, C. L.; Ramamurthy, R.; Clarke, M. S.;

Upregulated epidermal growth factor receptor expression following near-infrared irradiation simulating solar radiation in a three-dimensional reconstructed human corneal epithelial tissue culture model.

PubMed

Tanaka, Yohei; Nakayama, Jun

2016-01-01

Humans are increasingly exposed to near-infrared (NIR) radiation from both natural (eg, solar) and artificial (eg, electrical appliances) sources. Although the biological effects of sun and ultraviolet (UV) exposure have been extensively investigated, the biological effect of NIR radiation is still unclear. We previously reported that NIR as well as UV induces photoaging and standard UV-blocking materials, such as sunglasses, do not sufficiently block NIR. The objective of this study was to investigate changes in gene expression in three-dimensional reconstructed corneal epithelial tissue culture exposed to broad-spectrum NIR irradiation to simulate solar NIR radiation that reaches human tissues. DNA microarray and quantitative real-time polymerase chain reaction analysis were used to assess gene expression levels in a three-dimensional reconstructed corneal epithelial model composed of normal human corneal epithelial cells exposed to water-filtered broad-spectrum NIR irradiation with a contact cooling (20°C). The water-filter allowed 1,000-1,800 nm wavelengths and excluded 1,400-1,500 nm wavelengths. A DNA microarray with >62,000 different probes showed 25 and 150 genes that were up- or downregulated by at least fourfold and twofold, respectively, after NIR irradiation. In particular, epidermal growth factor receptor (EGFR) was upregulated by 19.4-fold relative to control cells. Quantitative real-time polymerase chain reaction analysis revealed that two variants of EGFR in human corneal epithelial tissue were also significantly upregulated after five rounds of 10 J/cm(2) irradiation (P<0.05). We found that NIR irradiation induced the upregulated expression of EGFR in human corneal cells. Since over half of the solar energy reaching the Earth is in the NIR region, which cannot be adequately blocked by eyewear and thus can induce eye damage with intensive or long-term exposure, protection from both UV and NIR radiation may prevent changes in gene expression and in turn eye damage.

Three-Dimensional Tissue Assemblies: Novel Models for the Study of Salmonella enterica Serovar Typhimurium Pathogenesis

PubMed Central

Nickerson, Cheryl A.; Goodwin, Thomas J.; Terlonge, Jacqueline; Ott, C. Mark; Buchanan, Kent L.; Uicker, William C.; Emami, Kamal; LeBlanc, Carly L.; Ramamurthy, Rajee; Clarke, Mark S.; Vanderburg, Charles R.; Hammond, Timothy; Pierson, Duane L.

2001-01-01

The lack of readily available experimental systems has limited knowledge pertaining to the development of Salmonella-induced gastroenteritis and diarrheal disease in humans. We used a novel low-shear stress cell culture system developed at the National Aeronautics and Space Administration in conjunction with cultivation of three-dimensional (3-D) aggregates of human intestinal tissue to study the infectivity of Salmonella enterica serovar Typhimurium for human intestinal epithelium. Immunohistochemical characterization and microscopic analysis of 3-D aggregates of the human intestinal epithelial cell line Int-407 revealed that the 3-D cells more accurately modeled human in vivo differentiated tissues than did conventional monolayer cultures of the same cells. Results from infectivity studies showed that Salmonella established infection of the 3-D cells in a much different manner than that observed for monolayers. Following the same time course of infection with Salmonella, 3-D Int-407 cells displayed minimal loss of structural integrity compared to that of Int-407 monolayers. Furthermore, Salmonella exhibited significantly lower abilities to adhere to, invade, and induce apoptosis of 3-D Int-407 cells than it did for infected Int-407 monolayers. Analysis of cytokine expression profiles of 3-D Int-407 cells and monolayers following infection with Salmonella revealed significant differences in expression of interleukin 1α (IL-1α), IL-1β, IL-6, IL-1Ra, and tumor necrosis factor alpha mRNAs between the two cultures. In addition, uninfected 3-D Int-407 cells constitutively expressed higher levels of transforming growth factor β1 mRNA and prostaglandin E2 than did uninfected Int-407 monolayers. By more accurately modeling many aspects of human in vivo tissues, the 3-D intestinal cell model generated in this study offers a novel approach for studying microbial infectivity from the perspective of the host-pathogen interaction. PMID:11598087

Microphysiological modeling of the reproductive tract: a fertile endeavor.

PubMed

Eddie, Sharon L; Kim, J Julie; Woodruff, Teresa K; Burdette, Joanna E

2014-09-01

Preclinical toxicity testing in animal models is a cornerstone of the drug development process, yet it is often unable to predict adverse effects and tolerability issues in human subjects. Species-specific responses to investigational drugs have led researchers to utilize human tissues and cells to better estimate human toxicity. Unfortunately, human cell-derived models are imperfect because toxicity is assessed in isolation, removed from the normal physiologic microenvironment. Microphysiological modeling often referred to as 'organ-on-a-chip' or 'human-on-a-chip' places human tissue into a microfluidic system that mimics the complexity of human in vivo physiology, thereby allowing for toxicity testing on several cell types, tissues, and organs within a more biologically relevant environment. Here we describe important concepts when developing a repro-on-a-chip model. The development of female and male reproductive microfluidic systems is critical to sex-based in vitro toxicity and drug testing. This review addresses the biological and physiological aspects of the male and female reproductive systems in vivo and what should be considered when designing a microphysiological human-on-a-chip model. Additionally, interactions between the reproductive tract and other systems are explored, focusing on the impact of factors and hormones produced by the reproductive tract and disease pathophysiology. © 2014 by the Society for Experimental Biology and Medicine.

Self-organized amniogenesis by human pluripotent stem cells in a biomimetic implantation-like niche

NASA Astrophysics Data System (ADS)

Shao, Yue; Taniguchi, Kenichiro; Gurdziel, Katherine; Townshend, Ryan F.; Xue, Xufeng; Yong, Koh Meng Aw; Sang, Jianming; Spence, Jason R.; Gumucio, Deborah L.; Fu, Jianping

2017-04-01

Amniogenesis--the development of amnion--is a critical developmental milestone for early human embryogenesis and successful pregnancy. However, human amniogenesis is poorly understood due to limited accessibility to peri-implantation embryos and a lack of in vitro models. Here we report an efficient biomaterial system to generate human amnion-like tissue in vitro through self-organized development of human pluripotent stem cells (hPSCs) in a bioengineered niche mimicking the in vivo implantation environment. We show that biophysical niche factors act as a switch to toggle hPSC self-renewal versus amniogenesis under self-renewal-permissive biochemical conditions. We identify a unique molecular signature of hPSC-derived amnion-like cells and show that endogenously activated BMP-SMAD signalling is required for the amnion-like tissue development by hPSCs. This study unveils the self-organizing and mechanosensitive nature of human amniogenesis and establishes the first hPSC-based model for investigating peri-implantation human amnion development, thereby helping advance human embryology and reproductive medicine.

21 CFR 1270.33 - Records, general requirements.

Code of Federal Regulations, 2013 CFR

2013-04-01

... assure freedom from risk factors for and clinical evidence of HIV infection, hepatitis B, and hepatitis C. (c) All human tissue processed or shipped prior to determination of donor suitability must be under...

21 CFR 1270.33 - Records, general requirements.

Code of Federal Regulations, 2012 CFR

2012-04-01

... assure freedom from risk factors for and clinical evidence of HIV infection, hepatitis B, and hepatitis C. (c) All human tissue processed or shipped prior to determination of donor suitability must be under...

21 CFR 1270.33 - Records, general requirements.

Code of Federal Regulations, 2014 CFR

2014-04-01

... assure freedom from risk factors for and clinical evidence of HIV infection, hepatitis B, and hepatitis C. (c) All human tissue processed or shipped prior to determination of donor suitability must be under...

The presence of pleiotrophin in the human intervertebral disc is associated with increased vascularization: an immunohistologic study.

PubMed

Johnson, William E B; Patterson, Angela M; Eisenstein, Stephen M; Roberts, Sally

2007-05-20

An immunohistological study of surgical specimens of human intervertebral disc. To examine the presence of pleiotrophin in diseased or damaged intervertebral disc tissue and the association between its presence and the extent of tissue vascularization and innervation. Increased levels of pleiotrophin, a growth and differentiation factor that is active in various pathophysiologic processes, including angiogenesis, has been associated with osteoarthritic changes of human articular cartilage. The association between pleiotrophin expression and pathologic conditions of the human intervertebral disc is unknown. Specimens of human lumbar intervertebral discs, obtained following surgical discectomy, were divided into 3 groups: non-degenerated discs (n = 7), degenerated discs (n = 6), and prolapsed discs (n = 11). Serial tissue sections of each specimen were immunostained to determine the presence of pleiotrophin, blood vessels (CD34-positive endothelial cells), and nerves (neurofilament 200 kDa [NF200]-positive nerve fibers). Pleiotrophin immunoreactivity was seen in disc cells, endothelial cells, and in the extracellular matrix in most specimens of intervertebral disc but was most prevalent in vascularized tissue in prolapsed discs. There was a significant correlation between the presence of pleiotrophin-positive disc cells and that of CD34-positive blood vessels. NF200-positive nerves were seen in vascularized areas of more degenerated discs, but nerves did not appear to codistribute with blood vessels or pleiotrophin positivity in prolapsed discs. Pleiotrophin is present in pathologic human intervertebral discs, and its prevalence and distribution suggest that it may play a role in neovascularization of diseased or damaged disc tissue.

Regulation of the human ascorbate transporter SVCT2 exon 1b gene by zinc-finger transcription factors

PubMed Central

Qiao, Huan; May, James M.

2011-01-01

The sodium-dependent vitamin C transporter (SVCT) 2 is crucial for ascorbate uptake in metabolically active and specialized tissues. The present study focused on the gene regulation of the SVCT2 exon 1b, which is ubiquitously expressed in human and mouse tissues. Although the human SVCT2 exon 1b promoter doesn’t contain a classical TATA-box, we found that it does contain a functional initiator (Inr) that binds YY1 and interacts with upstream Sp1/Sp3 elements in the proximal promoter region. These elements in turn play a critical role in regulating YY1-mediated transcription of the exon 1b gene. Formation of YY1/Sp complexes on the promoter is required for its optional function. YY1 with Sp1 or Sp3 synergistically enhanced exon 1b promoter activity as well as the endogenous SVCT2 protein expression. Further, in addition to Sp1/Sp3 both EGR-1 and -2 were detected in the protein complexes that bound the three GC boxes bearing overlapping binding sites for EGR/WT1 and Sp1/3. The EGR family factors, WT1 and MAZ were found to differentially regulate exon 1b promoter activity. These results show that differential occupancy of transcription factors on the GC-rich consensus sequences in SVCT2 exon 1b promoter contributes to the regulation of cell and tissue expression of SVCT2. PMID:21335086

Three-dimensional organotypic culture: experimental models of mammalian biology and disease

PubMed Central

Shamir, Eliah R.; Ewald, Andrew J.

2015-01-01

Mammalian organs are challenging to study as they are fairly inaccessible to experimental manipulation and optical observation. Recent advances in three-dimensional (3D) culture techniques, coupled with the ability to independently manipulate genetic and microenvironmental factors, have enabled the real-time study of mammalian tissues. These systems have been used to visualize the cellular basis of epithelial morphogenesis, to test the roles of specific genes in regulating cell behaviours within epithelial tissues and to elucidate the contribution of microenvironmental factors to normal and disease processes. Collectively, these novel models can be used to answer fundamental biological questions and generate replacement human tissues, and they enable testing of novel therapeutic approaches, often using patient-derived cells. PMID:25237826

Fabrication of novel high surface area mushroom gilled fibers and their effects on human adipose derived stem cells under pulsatile fluid flow for tissue engineering applications.

PubMed

Tuin, Stephen A; Pourdeyhimi, Behnam; Loboa, Elizabeth G

2016-05-01

The fabrication and characterization of novel high surface area hollow gilled fiber tissue engineering scaffolds via industrially relevant, scalable, repeatable, high speed, and economical nonwoven carding technology is described. Scaffolds were validated as tissue engineering scaffolds using human adipose derived stem cells (hASC) exposed to pulsatile fluid flow (PFF). The effects of fiber morphology on the proliferation and viability of hASC, as well as effects of varied magnitudes of shear stress applied via PFF on the expression of the early osteogenic gene marker runt related transcription factor 2 (RUNX2) were evaluated. Gilled fiber scaffolds led to a significant increase in proliferation of hASC after seven days in static culture, and exhibited fewer dead cells compared to pure PLA round fiber controls. Further, hASC-seeded scaffolds exposed to 3 and 6dyn/cm(2) resulted in significantly increased mRNA expression of RUNX2 after one hour of PFF in the absence of soluble osteogenic induction factors. This is the first study to describe a method for the fabrication of high surface area gilled fibers and scaffolds. The scalable manufacturing process and potential fabrication across multiple nonwoven and woven platforms makes them promising candidates for a variety of applications that require high surface area fibrous materials. We report here for the first time the successful fabrication of novel high surface area gilled fiber scaffolds for tissue engineering applications. Gilled fibers led to a significant increase in proliferation of human adipose derived stem cells after one week in culture, and a greater number of viable cells compared to round fiber controls. Further, in the absence of osteogenic induction factors, gilled fibers led to significantly increased mRNA expression of an early marker for osteogenesis after exposure to pulsatile fluid flow. This is the first study to describe gilled fiber fabrication and their potential for tissue engineering applications. The repeatable, industrially scalable, and versatile fabrication process makes them promising candidates for a variety of scaffold-based tissue engineering applications. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A Histologically Distinctive Interstitial Pneumonia Induced by Overexpression of the Interleukin 6, Transforming Growth Factor β1, or Platelet-Derived Growth Factor B Gene

NASA Astrophysics Data System (ADS)

Yoshida, Mitsuhiro; Sakuma, Junko; Hayashi, Seiji; Abe, Kin'ya; Saito, Izumu; Harada, Shizuko; Sakatani, Mitsunoir; Yamamoto, Satoru; Matsumoto, Norinao; Kaneda, Yasufumi; Kishmoto, Tadamitsu

1995-10-01

Interstitial pneumonia is characterized by alveolitis with resulting fibrosis of the interstitium. To determine the relevance of humoral factors in the pathogenesis of interstitial pneumonia, we introduced expression vectors into Wistar rats via the trachea to locally overexpress humoral factors in the lungs. Human interleukin (IL) 6 and IL-6 receptor genes induced lymphocytic alveolitis without marked fibroblast proliferation. In contrast, overexpression of human transforming growth factor β1 or human platelet-derived growth factor B gene induced only mild or apparent cellular infiltration in the alveoli, respectively. However, both factors induced significant proliferation of fibroblasts and deposition of collagen fibrils. These histopathologic changes induced by the transforming growth factor β1 and platelet-derived growth factor B gene are partly akin to those changes seen in lung tissues from patients with pulmonary fibrosis and markedly contrast with the changes induced by overexpression of the IL-6 and IL-6 receptor genes that mimics lymphocytic interstitial pneumonia.

The effect of AlloDerm® on the initiation and growth of human neovessels.

USDA-ARS?s Scientific Manuscript database

AlloDerm® is commonly employed for reconstruction of ablative soft tissue and mucosal defects following surgical resections. Although devoid of growth factors, AlloDerm® may serve as an adhesive matrix for binding of growth factors increasing local angiogenesis and wound healing. We hypothesized tha...

Expression profile of undifferentiated cell transcription factor 1 in normal and cancerous human epithelia.

PubMed

Mouallif, Mustapha; Albert, Adelin; Zeddou, Mustapha; Ennaji, My Mustapha; Delvenne, Philippe; Guenin, Samuel

2014-08-01

Undifferentiated cell Transcription Factor 1 (UTF1) is a chromatin-bound protein involved in stem cell differentiation. It was initially reported to be restricted to stem cells or germinal tissues. However, recent work suggests that UTF1 is also expressed in somatic cells and that its expression may increase during carcinogenesis. To further clarify the expression profile of UTF1, we evaluated UTF1 expression levels immunohistochemically in eight normal human epithelia (from breast, prostate, endometrium, bladder, colon, oesophagus, lung and kidney) and their corresponding tumours as well as in several epithelial cell lines. We showed UTF1 staining in normal and tumour epithelial tissues, but with varying intensities according to the tissue location. In vitro analyses also revealed that UTF1 is expressed in somatic epithelial cell lines even in the absence of Oct4A and Sox2, its two main known regulators. The comparison of UTF1 levels in normal and tumoral tissues revealed significant overexpression in endometrial and prostatic adenocarcinomas, whereas lower intensity of the staining was observed in renal and colic tumours, suggesting a potential tissue-specific function of UTF1. Altogether, these results highlight a potential dual role for UTF1, acting either as an oncogene or as a tumour suppressor depending on the tissue. These findings also question its role as a specific marker for stem cells. © 2014 The Authors. International Journal of Experimental Pathology © 2014 International Journal of Experimental Pathology.

Factors affecting the structure and maturation of human tissue engineered skeletal muscle.

PubMed

Martin, Neil R W; Passey, Samantha L; Player, Darren J; Khodabukus, Alastair; Ferguson, Richard A; Sharples, Adam P; Mudera, Vivek; Baar, Keith; Lewis, Mark P

2013-07-01

Tissue engineered skeletal muscle has great utility in experimental studies of physiology, clinical testing and its potential for transplantation to replace damaged tissue. Despite recent work in rodent tissue or cell lines, there is a paucity of literature concerned with the culture of human muscle derived cells (MDCs) in engineered constructs. Here we aimed to tissue engineer for the first time in the literature human skeletal muscle in self-assembling fibrin hydrogels and determine the effect of MDC seeding density and myogenic proportion on the structure and maturation of the constructs. Constructs seeded with 4 × 10(5) MDCs assembled to a greater extent than those at 1 × 10(5) or 2 × 10(5), and immunostaining revealed a higher fusion index and a higher density of myotubes within the constructs, showing greater structural semblance to in vivo tissue. These constructs primarily expressed perinatal and slow type I myosin heavy chain mRNA after 21 days in culture. In subsequent experiments MACS(®) technology was used to separate myogenic and non-myogenic cells from their heterogeneous parent population and these cells were seeded at varying myogenic (desmin +) proportions in fibrin based constructs. Only in the constructs seeded with 75% desmin + cells was there evidence of striations when immunostained for slow myosin heavy chain compared with constructs seeded with 10 or 50% desmin + cells. Overall, this work reveals the importance of cell number and myogenic proportions in tissue engineering human skeletal muscle with structural resemblance to in vivo tissue. Copyright © 2013 Elsevier Ltd. All rights reserved.

Human fallopian tube epithelium co-culture with murine ovarian follicles reveals crosstalk in the reproductive cycle

PubMed Central

Zhu, Jie; Xu, Yuanming; Rashedi, Alexandra S.; Pavone, Mary Ellen; Kim, J. Julie; Woodruff, Teresa K.; Burdette, Joanna E.

2016-01-01

Study question Do interactions between human fallopian tube epithelium and murine follicles occur during an artificial reproductive cycle in a co-culture system in vitro? Summary answer In a co-culture system, human fallopian tissues responded to the menstrual cycle mimetic by changes in morphology and levels of secreted factors, and increasing murine corpus luteum progesterone secretion. What is known already The entire fallopian tube epithelium, including ciliated and secretory cells, can be regulated in the reproductive cycle. Currently, there are no in vitro culture models that can monitor fallopian tissues in real time in response to factors produced by the ovary. In addition, there are no reports on the impact of fallopian tissue on ovarian function during the menstrual cycle. Study design, samples/materials, methods Human fallopian tissue (n = 24) was obtained by routine hysterectomies from women (aged 26–50 years, mean age = 43.6) who had not undergone exogenous hormonal treatment for at least 3 months prior to surgery. CD1 female mice were used for ovarian follicle isolation. The human fallopian epithelium layers were either co-cultured with five murine multilayer secondary follicles (150–180 μm follicles, encapsulated in one alginate gel bead) for 15 days or received stepwise steroid hormone additions for 13 days. The fallopian tissue morphology and cilia beating rate, as measured by an Andor Spinning Disk Confocal, were investigated. Oviduct-specific glycoprotein 1 (OVGP1), human insulin-like growth factor 1 (hIGF1), vascular endothelial growth factor A (VEGF-A) and interleukin 8 (IL8) as biological functional markers were measured either by ELISA or western blot to indicate dynamic changes in the fallopian epithelium during the reproductive cycle generated by mouse follicles or by stepwise steroid hormone induction. Three or four patients in each experiment were recruited for replicates. Data were presented as mean ± SD and further analyzed using one-way ANOVA followed by Tukey's multiple comparisons test. Main results and the role of chance The cultured fallopian tube epithelium responded to exogenous steroid hormone stimulation, as demonstrated by enhanced cilia beating rate (~25% increase, P = 0.04) and an increase in OVGP1 secretion (P = 0.02) in response to 1 nM estradiol (E2) treatment when compared with 0.1 nM E2. Conversely, 10 nM progesterone plus 1 nM E2 suppressed cilia beating rate by ~30% (P = 0.008), while OVGP1 secretion was suppressed by 0.1 nM E2 plus 50 nM progesterone (P = 0.002 versus 1 nM E2 alone). Human fallopian tube epithelium was co-cultured with murine secondary follicles to mimic the human menstrual cycle. OVGP1 and VEGF-A secretion from fallopian tissue was similar with stepwise hormone treatment and when cultured with murine follicles. However, the secretion patterns of hIGF1 and IL8 differed in the luteal phase when comparing steroid treatment with follicle co-culture. In co-culture, hIGF1 secretion was suppressed in the luteal versus follicular phase (P = 0.005) but stepwise hormone treatment had no effect on hIGF1. In co-culture, IL8 secretion was also suppressed on luteal phase day 15 (P = 0.013) versus follicular phase day 7, but IL8 secretion increased continuously under high E2/progesterone treatment (P = 0.003 for D13 versus D3). In the co-culture system, the corpus luteum continuously produced progesterone in the presence of fallopian tube tissue until Day 18 while, without fallopian tissue, progesterone started to drop from Day 13. Limitations, reasons for caution One limitation of this study is that murine follicles were used to mimic the human menstrual cycle. However, although secretion patterns of peptide hormones such as inhibins and activins differ in mice and humans, the co-culture system used here did reveal interactions between the tissues that govern reproductive function. Wider implications of the findings In vitro co-culture models of fallopian reproductive tissues with ovarian follicles can provide an important tool for understanding fertility and for uncovering the mechanisms responsible for reduced fertility. In addition, the role of oviductal secretions and how they influence ovarian function, such as the production of progesterone during the menstrual cycle, can be uncovered using this model. Large-scale data None. Study funding and competing interest(s) This work was funded by grants from the NIH (UH3TR001207), the American Cancer Society (RSG-12-230-01-TBG) and NIH (R01EB014806). The authors declare no competing financial interest. PMID:27542947

Clinicopathological significance of SLP-2 overexpression in human gallbladder cancer.

PubMed

Wang, Wei-Xin; Lin, Qing-Feng; Shen, Dong; Liu, Shao-Ping; Mao, Wei-Dong; Ma, Gui; Qi, Wei-Dong

2014-01-01

Several studies have indicated that overexpression of stomatin-like protein 2 (SLP-2) has been identified in several types of cancer. However, its role and clinical relevance in gallbladder cancer (GBC) is unknown. The purpose of this study was to reveal the prognostic significance of SLP-2 in GBC. The SLP-2 expression was examined at mRNA and protein levels by real-time quantitative polymerase chain reaction (qRT-PCR), and immunohistochemistry in GBC tissues and adjacent noncancerous tissues. Statistical analyses were applied to test the associations between SLP-2 expression, clinicopathologic factors, and prognosis. Immunohistochemistry and qRT-PCR showed that the protein and mRNA expression levels of SLP-2 were both significantly higher in GBC tissues than in adjacent noncancerous tissues. In addition, immunohistochemistry analysis showed that SLP-2 expression was significantly correlated with histological grade (P <0.001), pathologic T stage (P = 0.019), clinical stage (P = 0.001), and lymph node metastasis (P = 0.026). The Kaplan-Meier survival curves indicated that patients with high expression of SLP-2 had shorter overall survival than those with low expression (P <0.001). Meanwhile, the Cox multivariate analysis indicated that high expressions of SLP-2 were an independent prognostic factor for patients with GBC. These data showed that SLP-2 may play an important role in human GBC tumorigenesis, and SLP-2 might serve as a novel prognostic marker in human GBC.

The angiogenic factor CCN1 promotes adhesion and migration of circulating CD34+ progenitor cells: potential role in angiogenesis and endothelial regeneration.

PubMed

Grote, Karsten; Salguero, Gustavo; Ballmaier, Matthias; Dangers, Marc; Drexler, Helmut; Schieffer, Bernhard

2007-08-01

Tissue regeneration involves the formation of new blood vessels regulated by angiogenic factors. We reported recently that the expression of the angiogenic factor CCN1 is up-regulated under various pathophysiologic conditions within the cardiovascular system. Because CD34+ progenitor cells participate in cardiovascular tissue regeneration, we investigated whether CCN1-detected for the first time in human plasma-promotes the recruitment of CD34+ progenitor cells to endothelial cells, thereby enhancing endothelial proliferation and neovascularization. In this study, we demonstrated that CCN1 and supernatants from CCN1-stimulated human CD34+ progenitor cells promoted proliferation of endothelial cells and angiogenesis in vitro and in vivo. In addition, CCN1 induced migration and transendothelial migration of CD34+ cells and the release of multiple growth factors, chemokines, and matrix metalloproteinase-9 (MMP-9) from these cells. Moreover, the CCN1-specific integrins alpha(M)beta(2) and alpha(V)beta(3) are expressed on CD34+ cells and CCN1 stimulated integrin-dependent signaling. Furthermore, integrin antagonists (RGD-peptides) suppressed both binding of CCN1 to CD34+ cells and CCN1-induced adhesion of CD34+ cells to endothelial cells. These data suggest that CCN1 promotes integrin-dependent recruitment of CD34+ progenitor cells to endothelial cells, which may contribute to paracrine effects on angiogenesis and tissue regeneration.

Role of Angiogenesis in Endodontics: Contributions of Stem Cells and Proangiogenic and Antiangiogenic Factors to Dental Pulp Regeneration

PubMed Central

Saghiri, Mohammad Ali; Asatourian, Armen; Sorenson, Christine M.; Sheibani, Nader

2016-01-01

Introduction Dental pulp regeneration is a part of regenerative endodontics, which includes isolation, propagation, and re-transplantation of stem cells inside the prepared root canal space. The formation of new blood vessels through angiogenesis is mandatory to increase the survival rate of re-transplanted tissues. Angiogenesis is defined as the formation of new blood vessels from preexisting capillaries, which has great importance in pulp regeneration and homeostasis. Here the contribution of human dental pulp stem cells and proangiogenic and antiangiogenic factors to angiogenesis process and regeneration of dental pulp is reviewed. Methods A search was performed on the role of angiogenesis in dental pulp regeneration from January 2005 through April 2014. The recent aspects of the relationship between angiogenesis, human dental pulp stem cells, and proangiogenic and antiangiogenic factors in regeneration of dental pulp were assessed. Results Many studies have indicated an intimate relationship between angiogenesis and dental pulp regeneration. The contribution of stem cells and mechanical and chemical factors to dental pulp regeneration has been previously discussed. Conclusions Angiogenesis is an indispensable process during dental pulp regeneration. The survival of inflamed vital pulp and engineered transplanted pulp tissue are closely linked to the process of angiogenesis at sites of application. However, the detailed regulatory mechanisms involved in initiation and progression of angiogenesis in pulp tissue require investigation. PMID:25649306

A Prospective, Randomized, Double-blind, Split-face Clinical Trial Comparing the Efficacy of Two Topical Human Growth Factors for the Rejuvenation of the Aging Face

PubMed Central

Goldman, Mitchel P.

2017-01-01

Background: Cosmeceutical products represent an increasingly important therapeutic option for anti-aging and rejuvenation, either used alone or in combination with dermatologic surgical procedures. Among this group of products, topical growth factors have demonstrated efficacy in randomized, controlled clinical trials. However, comparisons between different products remain uncommon. Objective: The objective of this randomized, double-blind, split-face clinical trial was to compare two different topical growth factor formulations derived from either human fibroblasts or human adipose tissue derived mesenchymal stem cells. Methods: This was an institutional review board-approved, randomized, double-blind, split-face clinical trial involving 20 healthy subjects with moderate-to-severe facial wrinkling secondary to photodamage. One half of the face was randomized to receive topical human fibroblast growth factors and the other topical human mesenchymal stem cell growth factors. Treatment was continued for three months, and evaluations were performed in a double-blind fashion. Results: Both growth factor formulations achieved significant improvement in facial wrinkling. Blinded investigator and subject evaluations did not detect any significant differences between the two formulations in terms of efficacy, safety, or tolerability. Conclusion: Both human fibroblast growth factors and human mesenchymal stem cell growth factors are effective at facial rejuvenation. Topical growth factors represent a useful therapeutic modality. PMID:28670356

A Prospective, Randomized, Double-blind, Split-face Clinical Trial Comparing the Efficacy of Two Topical Human Growth Factors for the Rejuvenation of the Aging Face.

PubMed

Wu, Douglas C; Goldman, Mitchel P

2017-05-01

Background: Cosmeceutical products represent an increasingly important therapeutic option for anti-aging and rejuvenation, either used alone or in combination with dermatologic surgical procedures. Among this group of products, topical growth factors have demonstrated efficacy in randomized, controlled clinical trials. However, comparisons between different products remain uncommon. Objective: The objective of this randomized, double-blind, split-face clinical trial was to compare two different topical growth factor formulations derived from either human fibroblasts or human adipose tissue derived mesenchymal stem cells. Methods: This was an institutional review board-approved, randomized, double-blind, split-face clinical trial involving 20 healthy subjects with moderate-to-severe facial wrinkling secondary to photodamage. One half of the face was randomized to receive topical human fibroblast growth factors and the other topical human mesenchymal stem cell growth factors. Treatment was continued for three months, and evaluations were performed in a double-blind fashion. Results: Both growth factor formulations achieved significant improvement in facial wrinkling. Blinded investigator and subject evaluations did not detect any significant differences between the two formulations in terms of efficacy, safety, or tolerability. Conclusion: Both human fibroblast growth factors and human mesenchymal stem cell growth factors are effective at facial rejuvenation. Topical growth factors represent a useful therapeutic modality.

Multilayered epithelium in a rat model and human Barrett's esophagus: Similar expression patterns of transcription factors and differentiation markers

PubMed Central

Chen, Xiaoxin; Qin, Rong; Liu, Ba; Ma, Yan; Su, Yinghao; Yang, Chung S; Glickman, Jonathan N; Odze, Robert D; Shaheen, Nicholas J

2008-01-01

Background In rats, esophagogastroduodenal anastomosis (EGDA) without concomitant chemical carcinogen treatment leads to gastroesophageal reflux disease, multilayered epithelium (MLE, a presumed precursor in intestinal metaplasia), columnar-lined esophagus, dysplasia, and esophageal adenocarcinoma. Previously we have shown that columnar-lined esophagus in EGDA rats resembled human Barrett's esophagus (BE) in its morphology, mucin features and expression of differentiation markers (Lab. Invest. 2004;84:753–765). The purpose of this study was to compare the phenotype of rat MLE with human MLE, in order to gain insight into the nature of MLE and its potential role in the development of BE. Methods Serial sectioning was performed on tissue samples from 32 EGDA rats and 13 patients with established BE. Tissue sections were immunohistochemically stained for a variety of transcription factors and differentiation markers of esophageal squamous epithelium and intestinal columnar epithelium. Results We detected MLE in 56.3% (18/32) of EGDA rats, and in all human samples. As expected, both rat and human squamous epithelium, but not intestinal metaplasia, expressed squamous transcription factors and differentiation markers (p63, Sox2, CK14 and CK4) in all cases. Both rat and human intestinal metaplasia, but not squamous epithelium, expressed intestinal transcription factors and differentiation markers (Cdx2, GATA4, HNF1α, villin and Muc2) in all cases. Rat MLE shared expression patterns of Sox2, CK4, Cdx2, GATA4, villin and Muc2 with human MLE. However, p63 and CK14 were expressed in a higher proportion of rat MLE compared to humans. Conclusion These data indicate that rat MLE shares similar properties to human MLE in its expression pattern of these markers, not withstanding small differences, and support the concept that MLE may be a transitional stage in the metaplastic conversion of squamous to columnar epithelium in BE. PMID:18190713

Multilayered epithelium in a rat model and human Barrett's esophagus: similar expression patterns of transcription factors and differentiation markers.

PubMed

Chen, Xiaoxin; Qin, Rong; Liu, Ba; Ma, Yan; Su, Yinghao; Yang, Chung S; Glickman, Jonathan N; Odze, Robert D; Shaheen, Nicholas J

2008-01-11

In rats, esophagogastroduodenal anastomosis (EGDA) without concomitant chemical carcinogen treatment leads to gastroesophageal reflux disease, multilayered epithelium (MLE, a presumed precursor in intestinal metaplasia), columnar-lined esophagus, dysplasia, and esophageal adenocarcinoma. Previously we have shown that columnar-lined esophagus in EGDA rats resembled human Barrett's esophagus (BE) in its morphology, mucin features and expression of differentiation markers (Lab. Invest. 2004;84:753-765). The purpose of this study was to compare the phenotype of rat MLE with human MLE, in order to gain insight into the nature of MLE and its potential role in the development of BE. Serial sectioning was performed on tissue samples from 32 EGDA rats and 13 patients with established BE. Tissue sections were immunohistochemically stained for a variety of transcription factors and differentiation markers of esophageal squamous epithelium and intestinal columnar epithelium. We detected MLE in 56.3% (18/32) of EGDA rats, and in all human samples. As expected, both rat and human squamous epithelium, but not intestinal metaplasia, expressed squamous transcription factors and differentiation markers (p63, Sox2, CK14 and CK4) in all cases. Both rat and human intestinal metaplasia, but not squamous epithelium, expressed intestinal transcription factors and differentiation markers (Cdx2, GATA4, HNF1alpha, villin and Muc2) in all cases. Rat MLE shared expression patterns of Sox2, CK4, Cdx2, GATA4, villin and Muc2 with human MLE. However, p63 and CK14 were expressed in a higher proportion of rat MLE compared to humans. These data indicate that rat MLE shares similar properties to human MLE in its expression pattern of these markers, not withstanding small differences, and support the concept that MLE may be a transitional stage in the metaplastic conversion of squamous to columnar epithelium in BE.

A Six Months Exercise Intervention Influences the Genome-wide DNA Methylation Pattern in Human Adipose Tissue

PubMed Central

Rönn, Tina; Volkov, Petr; Davegårdh, Cajsa; Dayeh, Tasnim; Hall, Elin; Olsson, Anders H.; Nilsson, Emma; Tornberg, Åsa; Dekker Nitert, Marloes; Eriksson, Karl-Fredrik; Jones, Helena A.; Groop, Leif; Ling, Charlotte

2013-01-01

Epigenetic mechanisms are implicated in gene regulation and the development of different diseases. The epigenome differs between cell types and has until now only been characterized for a few human tissues. Environmental factors potentially alter the epigenome. Here we describe the genome-wide pattern of DNA methylation in human adipose tissue from 23 healthy men, with a previous low level of physical activity, before and after a six months exercise intervention. We also investigate the differences in adipose tissue DNA methylation between 31 individuals with or without a family history of type 2 diabetes. DNA methylation was analyzed using Infinium HumanMethylation450 BeadChip, an array containing 485,577 probes covering 99% RefSeq genes. Global DNA methylation changed and 17,975 individual CpG sites in 7,663 unique genes showed altered levels of DNA methylation after the exercise intervention (q<0.05). Differential mRNA expression was present in 1/3 of gene regions with altered DNA methylation, including RALBP1, HDAC4 and NCOR2 (q<0.05). Using a luciferase assay, we could show that increased DNA methylation in vitro of the RALBP1 promoter suppressed the transcriptional activity (p = 0.03). Moreover, 18 obesity and 21 type 2 diabetes candidate genes had CpG sites with differences in adipose tissue DNA methylation in response to exercise (q<0.05), including TCF7L2 (6 CpG sites) and KCNQ1 (10 CpG sites). A simultaneous change in mRNA expression was seen for 6 of those genes. To understand if genes that exhibit differential DNA methylation and mRNA expression in human adipose tissue in vivo affect adipocyte metabolism, we silenced Hdac4 and Ncor2 respectively in 3T3-L1 adipocytes, which resulted in increased lipogenesis both in the basal and insulin stimulated state. In conclusion, exercise induces genome-wide changes in DNA methylation in human adipose tissue, potentially affecting adipocyte metabolism. PMID:23825961

Extracellular acidification induces connective tissue growth factor production through proton-sensing receptor OGR1 in human airway smooth muscle cells

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

Matsuzaki, Shinichi; Ishizuka, Tamotsu, E-mail: tamotsui@showa.gunma-u.ac.jp; Yamada, Hidenori

Highlights: {yields} The involvement of extracellular acidification in airway remodeling was investigated. {yields} Extracellular acidification alone induced CTGF production in human ASMCs. {yields} Extracellular acidification enhanced TGF-{beta}-induced CTGF production in human ASMCs. {yields} Proton-sensing receptor OGR1 was involved in acidic pH-stimulated CTGF production. {yields} OGR1 may play an important role in airway remodeling in asthma. -- Abstract: Asthma is characterized by airway inflammation, hyper-responsiveness and remodeling. Extracellular acidification is known to be associated with severe asthma; however, the role of extracellular acidification in airway remodeling remains elusive. In the present study, the effects of acidification on the expression of connectivemore » tissue growth factor (CTGF), a critical factor involved in the formation of extracellular matrix proteins and hence airway remodeling, were examined in human airway smooth muscle cells (ASMCs). Acidic pH alone induced a substantial production of CTGF, and enhanced transforming growth factor (TGF)-{beta}-induced CTGF mRNA and protein expression. The extracellular acidic pH-induced effects were inhibited by knockdown of a proton-sensing ovarian cancer G-protein-coupled receptor (OGR1) with its specific small interfering RNA and by addition of the G{sub q/11} protein-specific inhibitor, YM-254890, or the inositol-1,4,5-trisphosphate (IP{sub 3}) receptor antagonist, 2-APB. In conclusion, extracellular acidification induces CTGF production through the OGR1/G{sub q/11} protein and inositol-1,4,5-trisphosphate-induced Ca{sup 2+} mobilization in human ASMCs.« less

Heterologous expression of Streptococcus mutans Cnm in Lactococcus lactis promotes intracellular invasion, adhesion to human cardiac tissues and virulence.

PubMed

Freires, Irlan A; Avilés-Reyes, Alejandro; Kitten, Todd; Simpson-Haidaris, P J; Swartz, Michael; Knight, Peter A; Rosalen, Pedro L; Lemos, José A; Abranches, Jacqueline

2017-01-02

In S. mutans, the expression of the surface glycoprotein Cnm mediates binding to extracellular matrix proteins, endothelial cell invasion and virulence in the Galleria mellonella invertebrate model. To further characterize Cnm as a virulence factor, the cnm gene from S. mutans strain OMZ175 was expressed in the non-pathogenic Lactococcus lactis NZ9800 using a nisin-inducible system. Despite the absence of the machinery necessary for Cnm glycosylation, Western blot and immunofluorescence microscopy analyses demonstrated that Cnm was effectively expressed and translocated to the cell wall of L. lactis. Similar to S. mutans, expression of Cnm in L. lactis enabled robust binding to collagen and laminin, invasion of human coronary artery endothelial cells and increased virulence in G. mellonella. Using an ex vivo human heart tissue colonization model, we showed that Cnm-positive strains of either S. mutans or L. lactis outcompete their Cnm-negative counterparts for tissue colonization. Finally, Cnm expression facilitated L. lactis adhesion and colonization in a rabbit model of infective endocarditis. Collectively, our results provide unequivocal evidence that binding to extracellular matrices mediated by Cnm is an important virulence attribute of S. mutans and confirm the usefulness of the L. lactis heterologous system for further characterization of bacterial virulence factors.

Mature brain-derived neurotrophic factor and its receptor TrkB are upregulated in human glioma tissues.

PubMed

Xiong, Jing; Zhou, L I; Lim, Yoon; Yang, Miao; Zhu, Yu-Hong; Li, Zhi-Wei; Fu, Deng-Li; Zhou, Xin-Fu

2015-07-01

There are two forms of brain-derived neurotrophic factor (BDNF), precursor of BDNF (proBDNF) and mature BDNF, which each exert opposing effects through two different transmembrane receptor signaling systems, consisting of p75 neurotrophin receptor (p75NTR) and tyrosine receptor kinase B (TrkB). Previous studies have demonstrated that proBDNF promotes cell death and inhibits the growth and migration of C6 glioma cells through p75NTR in vitro , while mature BDNF has opposite effects on C6 glioma cells. It is hypothesized that mature BDNF is essential in the development of malignancy in gliomas. However, histological data obtained in previous studies were unable distinguish mature BDNF from proBDNF due to the lack of specific antibodies. The present study investigated the expression of mature BDNF using a specific sheep monoclonal anti-mature BDNF antibody in 42 human glioma tissues of different grades and 10 control tissues. The correlation between mature BDNF and TrkB was analyzed. Mature BDNF expression was significantly increased in high-grade gliomas, and was positively correlated with the malignancy of the tumor and TrkB receptor expression. The present data have demonstrated that increased levels of mature BDNF contribute markedly to the development of malignancy of human gliomas through the primary BDNF receptor TrkB.

Purification of Growth Factor mRNA in Renal Tissues:bFGF-2, FGF-2, TGFα, and EGFR.

PubMed

Mydlo, J H

2001-01-01

Growth factors are polypeptides that induce cell mitogenicity, and thus play an important role in the etiology and progression of tumors (1). Fibroblast growth factors (FGF) constitute a family of structurally related polypeptides of 146 amino acids, which exhibit a wide spectrum of biologic activities, including angiogenesis or the formation of a vascular network. FGFs are mitogenic towards many mesodermal and ectodermal cell types, and can also induce and/or inhibit differentiation of cells (2). These heparin-binding factors are categorized as FGF-1 through FGF-10. Acidic FGF, or FGF-1, is found mostly in brain and other neural tissues. Basic FGF, or FGF- 2, a protein of 18 kDa mw, is one of the most ubiqitous growth factors. It is found in numerous benign and cancerous human and animal tissues, including kidney, prostate, and bladder (3-6). In some cases it has also been demonstrated to have potential as a tumor marker (7-11). One group reported greater recovery of both FGF-2 protein and FGF-2 mRNA from renal-cancer tissue compared to equal amounts of normal renal tissue (5). Furthermore, when purified FGF-2 from renal cell carcinoma (RCC) is added exogenously to other established renal tumorcell lines and endothelial cell lines, it demonstrates significant mitogenic activity (6). Thus, renal tumors may use FGF-2 in an autocrine manner to sustain themselves.

Comparison of fibrin clots derived from peripheral blood and bone marrow.

PubMed

Shoji, Takeshi; Nakasa, Tomoyuki; Yoshizuka, Masaaki; Yamasaki, Takuma; Yasunaga, Yuji; Adachi, Nobuo; Ochi, Mitsuo

2017-03-01

Autologous fibrin clots derived from peripheral blood (pb-fibrin clot) and bone marrow (bm-fibrin clot) are thought to be effective for tissue regeneration. However, there is no report detailing the amount of growth factors in pb-/bm-fibrin clot. In this study we evaluated the amount of growth factors in human pb-/bm-fibrin clot, and prove the validity of fibrin clot for clinical use. Human pb-/bm-fibrin clots were obtained during surgery. In the first experiment, enzyme-linked immunosorbent assay (ELISA) was performed for detecting the amount of vascular endothelial growth factor (VEGF), epidermal growth factor (EGF), insulin-like growth factor-1 (IGF-1), fibroblast growth factor basic (bFGF), hepatocyte growth factor (HGF), transforming growth factor-beta (TGF-β), platelet derived-growth factors-AB (PDGF-AB), and stromal cell-derived factor-1 (SDF-1). In the second experiment, the efficacy of fibrin clot on the osteogenic differentiation and fibroblast proliferation was evaluated. Pb-/bm-fibrin clots were incubated in human osteoblast derived from mesenchymal stromal cells (MSCs) or human skin fibroblast. Alizarin red staining and real-time PCR (COL1A1, RUNX2) were performed for the detection of osteogenic potential. Cell-growth assay (WST-8) and real-time PCR (COL1A1) were also performed for the detection of the potential of fibroblast proliferation. ELISA analysis revealed that the amount of VEGF, HGF, bFGF, IGF-1, and SDF-1 of bm-fibrin clot group is higher than that of pb-fibrin clot group with statistical differences. Besides, we confirmed that bm-fibrin clot has much potential for the osteogenic differentiation and fibroblast proliferation. The positive outcomes confirm the efficacy of pb-/bm-fibrin clot, and bm-fibrin clot was proved to have much potential for tissue regeneration compared with pb-fibrin clot. The current study showed the potential of a strategy for regenerative medicine using bm-fibrin clot.

Organochlorine residues in Baluchistan/Pakistan: Blood and fat concentrations in humans

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

Krawinkel, M.B.; Plehn, G.; Kruse, H.

1989-12-01

Organochlorine (OC)-residues are detected in measurable concentrations in various tissues of human beings because of the worldwide pollution of air, water, soil, and foods. The concentrations vary from region to region according to chemical, climatic, socio-economic, and geographic factors. Persisting pesticides used in agriculture are found in relevant concentrations in agriculture products, meat, and fish. As developing countries face economic pressure to increase their agricultural exports cheap pesticides are sometimes used without the precautions necessary to prevent contaminations of water and food. The authors conducted a small survey monitoring the OC-concentrations in human blood and fat tissue under the aimmore » to detect more recent as well as elder expositions.« less

Phenytoin activates Smad3 phosphorylation and periostin expression in drug-induced gingival enlargement.

PubMed

Kim, Shawna S; Nikoloudaki, Georgia; Darling, Mark; Rieder, Michael J; Hamilton, Douglas W

2018-06-19

Drug-induced gingival enlargement (DIGE) is a fibrotic condition associated with systemic administration of the anti-epileptic drug, phenytoin. We have previously demonstrated that periostin, which is transforming growth factor-beta (TGF-β) inducible gene, is upregulated in various fibrotic conditions including gingival enlargement associated with nifedipine. The objective of this study was to assess periostin expression in phenytoin-induced gingival enlargement (PIGE) tissues and to investigate the mechanisms underlying periostin expression. Human PIGE tissues were assessed using Masson's trichrome, with cell infiltration and changes in extracellular matrix composition characterized through labeling with antibodies to periostin, phospho-SMAD 3, TGF-β, as well as the macrophage markers CD68 and RM3/1. Using human gingival fibroblasts (HGFs) in vitro we examined the pathways through which phenytoin acts on fibroblasts. In PIGE tissues, which demonstrate altered collagen organization and increased inflammatory cell infiltration, periostin protein was increased compared with healthy tissues. p-SMAD2/3, the transcription factor associated with canonical TGF-β signaling, is localized to the nuclei in both gingival fibroblasts and oral epithelial cells in PIGE tissues, but not in healthy tissue. In vitro culture of HGFs with 15 and 30 μg/ml of phenytoin increased periostin protein levels, which correlated with p-SMAD3 phosphorylation. Inhibition of canonical TGF-β signaling with SB431542 significantly reduced phenytoin induction of SMAD3 phosphorylation and periostin expression in HGFs. Analysis of PIGE tissues showed a subset of CD68 stained macrophages were TGF-β positive and that RM1/3 regenerative macrophages were present in the tissues. Our results demonstrate that phenytoin up-regulates periostin in HGFs in a TGF-β-dependent manner.

Cigarette smoke condensate inhibits collagen gel contraction and prostaglandin E2 production in human gingival fibroblasts.

PubMed

Romero, A; Cáceres, M; Arancibia, R; Silva, D; Couve, E; Martínez, C; Martínez, J; Smith, P C

2015-06-01

Granulation tissue remodeling and myofibroblastic differentiation are critically important events during wound healing. Tobacco smoking has a detrimental effect in gingival tissue repair. However, studies evaluating the effects of cigarette smoke on these events are lacking. We used gingival fibroblasts cultured within free-floating and restrained collagen gels to simulate the initial and final steps of the granulation tissue phase during tissue repair. Collagen gel contraction was stimulated with serum or transforming growth factor-β1. Cigarette smoke condensate (CSC) was used to evaluate the effects of tobacco smoke on gel contraction. Protein levels of alpha-smooth muscle actin, β1 integrin, matrix metalloproteinase-3 and connective tissue growth factor were evaluated through Western blot. Prostaglandin E(2) (PGE(2)) levels were determined through ELISA. Actin organization was evaluated through confocal microscopy. CSC reduced collagen gel contraction induced by serum and transforming growth factor-β1 in restrained collagen gels. CSC also altered the development of actin stress fibers in fibroblasts cultured within restrained collagen gels. PGE(2) levels were strongly diminished by CSC in three-dimensional cell cultures. However, other proteins involved in granulation tissue remodeling and myofibroblastic differentiation such as alpha-smooth muscle actin, β1 integrin, matrix metalloproteinase-3 and connective tissue growth factor, were unmodified by CSC. CSC may alter the capacity of gingival fibroblasts to remodel and contract a collagen matrix. Inhibition of PGE(2) production and alterations of actin stress fibers in these cells may impair proper tissue maturation during wound healing in smokers. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Dynamic changes in high and low mammographic density human breast tissues maintained in murine tissue engineering chambers during various murine peripartum states and over time.

PubMed

Chew, G L; Huang, D; Huo, C W; Blick, T; Hill, P; Cawson, J; Frazer, H; Southey, M D; Hopper, J L; Henderson, M A; Haviv, I; Thompson, E W

2013-07-01

Mammographic density (MD) is a strong heritable risk factor for breast cancer, and may decrease with increasing parity. However, the biomolecular basis for MD-associated breast cancer remains unclear, and systemic hormonal effects on MD-associated risk is poorly understood. This study assessed the effect of murine peripartum states on high and low MD tissue maintained in a xenograft model of human MD. Method High and low MD human breast tissues were precisely sampled under radiographic guidance from prophylactic mastectomy specimens of women. The high and low MD tissues were maintained in separate vascularised biochambers in nulliparous or pregnant SCID mice for 4 weeks, or mice undergoing postpartum involution or lactation for three additional weeks. High and low MD biochamber material was harvested for histologic and radiographic comparisons during various murine peripartum states. High and low MD biochamber tissues in nulliparous mice were harvested at different timepoints for histologic and radiographic comparisons. Results High MD biochamber tissues had decreased stromal (p = 0.0027), increased adipose (p = 0.0003) and a trend to increased glandular tissue areas (p = 0.076) after murine postpartum involution. Stromal areas decreased (p = 0.042), while glandular (p = 0.001) and adipose areas (p = 0.009) increased in high MD biochamber tissues during lactation. A difference in radiographic density was observed in high (p = 0.0021) or low MD biochamber tissues (p = 0.004) between nulliparous, pregnant and involution groups. No differences in tissue composition were observed in high or low MD biochamber tissues maintained for different durations, although radiographic density increased over time. Conclusion High MD biochamber tissues had measurable histologic changes after postpartum involution or lactation. Alterations in radiographic density occurred in biochamber tissues between different peripartum states and over time. These findings demonstrate the dynamic nature of the human MD xenograft model, providing a platform for studying the biomolecular basis of MD-associated cancer risk.

Human papillomavirus infects placental trophoblast and Hofbauer cells, but appears not to play a causal role in miscarriage and preterm labor.

PubMed

Ambühl, Lea M M; Leonhard, Anne K; Widen Zakhary, Carina; Jørgensen, Annemette; Blaakaer, Jan; Dybkaer, Karen; Baandrup, Ulrik; Uldbjerg, Niels; Sørensen, Suzette

2017-10-01

Recently, an association between human papillomavirus infection and both spontaneous abortion and spontaneous preterm delivery was suggested. However, the reported human papillomavirus prevalence in pregnant women varies considerably and reliable conclusions are difficult. We aimed to investigate human papillomavirus infection in placental tissue of a Danish study cohort. Furthermore, we studied the cellular localization of human papillomavirus. In this prospective case-control study, placental tissue was analyzed for human papillomavirus infection by nested PCR in the following four study groups: full-term delivery (n = 103), spontaneous preterm delivery (n = 69), elective abortion (n = 54), and spontaneous abortion (n = 44). Moreover, human papillomavirus cellular target was identified using in situ hybridization. Human papillomavirus prevalence in placental tissue was 8.7% in full-term deliveries, 8.8% in spontaneous preterm deliveries, 10.9% in spontaneous abortions, and 20.4% in elective abortions. Twelve different human papillomavirus types were detected, and placental human papillomavirus infection was associated to a disease history of cervical cancer. Human papillomavirus DNA was identified in trophoblast cells, cells of the placental villi mesenchyme including Hofbauer cells, and in parts of the encasing endometrium. Placental human papillomavirus infections are not likely to constitute a risk factor for spontaneous preterm labor or spontaneous abortions in the Danish population, although an effect of human papillomavirus DNA in placental cells cannot be excluded. © 2017 Nordic Federation of Societies of Obstetrics and Gynecology.

Immunohistochemical profile of VEGF, TGF-β and PGE₂ in human pterygium and normal conjunctiva: experimental study and review of the literature.

PubMed

Bianchi, E; Scarinci, F; Grande, C; Plateroti, R; Plateroti, P; Plateroti, A M; Fumagalli, L; Capozzi, P; Feher, J; Artico, M

2012-01-01

Human pterygium is made up of chronic proliferative fibro-vascular tissue growing on the ocular surface. This disease exhibits both degenerative and hyperplastic properties. Some fibroangiogenic factors have recently been shown to play a potential role in fibrovascular diseases via the angiogenesis process. The aim of this study is to evaluate VEGF, TGF-β and PGE₂ expression in the epithelial, endothelial and stromal cells of human pterygium and normal conjunctiva in order to determine whether these factors participate in the development of pterygium. Ten specimens from patients with pterygium and two normal conjunctivas (cadavers) were analyzed by immunohistochemistry using specific antibodies against these growth factors. The technique used was ABC/HRP (Avidin complexed with biotinylated peroxidase). Immunoreactivity of VEGF was significantly increased in the epithelium, vascular endothelium and stromal cells in primary pterygium as compared with normal conjunctiva. A moderate expression of TGF-β in the pterygium was observed in the epithelial and stromal layers. On the contrary, immunolabeling of this growth factor in the human normal conjunctiva was weak. PGE₂ was strongly expressed in the epithelium of patients with pterygium, as in control conjunctival tissues, and the immunolabeling was moderate in the stroma from the same patients. Our results suggest that these growth factors may contribute to the progression of primary pterygium by increasing angiogenesis, thus leading to the formation of new blood vessels from the pre-existing vasculature. We conclude that VEGF, TGF-β and PGE₂ may be potential therapeutic targets in the treatment of this disease although proof of this evidence requires further studies.

Dynamic Proteomic Analysis of Pancreatic Mesenchyme Reveals Novel Factors That Enhance Human Embryonic Stem Cell to Pancreatic Cell Differentiation.

PubMed

Russ, Holger A; Landsman, Limor; Moss, Christopher L; Higdon, Roger; Greer, Renee L; Kaihara, Kelly; Salamon, Randy; Kolker, Eugene; Hebrok, Matthias

2016-01-01

Current approaches in human embryonic stem cell (hESC) to pancreatic beta cell differentiation have largely been based on knowledge gained from developmental studies of the epithelial pancreas, while the potential roles of other supporting tissue compartments have not been fully explored. One such tissue is the pancreatic mesenchyme that supports epithelial organogenesis throughout embryogenesis. We hypothesized that detailed characterization of the pancreatic mesenchyme might result in the identification of novel factors not used in current differentiation protocols. Supplementing existing hESC differentiation conditions with such factors might create a more comprehensive simulation of normal development in cell culture. To validate our hypothesis, we took advantage of a novel transgenic mouse model to isolate the pancreatic mesenchyme at distinct embryonic and postnatal stages for subsequent proteomic analysis. Refined sample preparation and analysis conditions across four embryonic and prenatal time points resulted in the identification of 21,498 peptides with high-confidence mapping to 1,502 proteins. Expression analysis of pancreata confirmed the presence of three potentially important factors in cell differentiation: Galectin-1 (LGALS1), Neuroplastin (NPTN), and the Laminin α-2 subunit (LAMA2). Two of the three factors (LGALS1 and LAMA2) increased expression of pancreatic progenitor transcript levels in a published hESC to beta cell differentiation protocol. In addition, LAMA2 partially blocks cell culture induced beta cell dedifferentiation. Summarily, we provide evidence that proteomic analysis of supporting tissues such as the pancreatic mesenchyme allows for the identification of potentially important factors guiding hESC to pancreas differentiation.

Dynamic Proteomic Analysis of Pancreatic Mesenchyme Reveals Novel Factors That Enhance Human Embryonic Stem Cell to Pancreatic Cell Differentiation

PubMed Central

Russ, Holger A.; Landsman, Limor; Moss, Christopher L.; Higdon, Roger; Greer, Renee L.; Kaihara, Kelly; Salamon, Randy; Kolker, Eugene; Hebrok, Matthias

2016-01-01

Current approaches in human embryonic stem cell (hESC) to pancreatic beta cell differentiation have largely been based on knowledge gained from developmental studies of the epithelial pancreas, while the potential roles of other supporting tissue compartments have not been fully explored. One such tissue is the pancreatic mesenchyme that supports epithelial organogenesis throughout embryogenesis. We hypothesized that detailed characterization of the pancreatic mesenchyme might result in the identification of novel factors not used in current differentiation protocols. Supplementing existing hESC differentiation conditions with such factors might create a more comprehensive simulation of normal development in cell culture. To validate our hypothesis, we took advantage of a novel transgenic mouse model to isolate the pancreatic mesenchyme at distinct embryonic and postnatal stages for subsequent proteomic analysis. Refined sample preparation and analysis conditions across four embryonic and prenatal time points resulted in the identification of 21,498 peptides with high-confidence mapping to 1,502 proteins. Expression analysis of pancreata confirmed the presence of three potentially important factors in cell differentiation: Galectin-1 (LGALS1), Neuroplastin (NPTN), and the Laminin α-2 subunit (LAMA2). Two of the three factors (LGALS1 and LAMA2) increased expression of pancreatic progenitor transcript levels in a published hESC to beta cell differentiation protocol. In addition, LAMA2 partially blocks cell culture induced beta cell dedifferentiation. Summarily, we provide evidence that proteomic analysis of supporting tissues such as the pancreatic mesenchyme allows for the identification of potentially important factors guiding hESC to pancreas differentiation. PMID:26681951

An evaluation of tyramide signal amplification and archived fixed and frozen tissue in microarray gene expression analysis

PubMed Central

Karsten, Stanislav L.; Van Deerlin, Vivianna M. D.; Sabatti, Chiara; Gill, Lisa H.; Geschwind, Daniel H.

2002-01-01

Archival formalin-fixed, paraffin-embedded and ethanol-fixed tissues represent a potentially invaluable resource for gene expression analysis, as they are the most widely available material for studies of human disease. Little data are available evaluating whether RNA obtained from fixed (archival) tissues could produce reliable and reproducible microarray expression data. Here we compare the use of RNA isolated from human archival tissues fixed in ethanol and formalin to frozen tissue in cDNA microarray experiments. Since an additional factor that can limit the utility of archival tissue is the often small quantities available, we also evaluate the use of the tyramide signal amplification method (TSA), which allows the use of small amounts of RNA. Detailed analysis indicates that TSA provides a consistent and reproducible signal amplification method for cDNA microarray analysis, across both arrays and the genes tested. Analysis of this method also highlights the importance of performing non-linear channel normalization and dye switching. Furthermore, archived, fixed specimens can perform well, but not surprisingly, produce more variable results than frozen tissues. Consistent results are more easily obtainable using ethanol-fixed tissues, whereas formalin-fixed tissue does not typically provide a useful substrate for cDNA synthesis and labeling. PMID:11788730

BNDF methylation in mothers and newborns is associated with maternal exposure to war trauma.

PubMed

Kertes, Darlene A; Bhatt, Samarth S; Kamin, Hayley S; Hughes, David A; Rodney, Nicole C; Mulligan, Connie J

2017-01-01

The BDNF gene codes for brain-derived neurotrophic factor, a growth factor involved in neural development, cell differentiation, and synaptic plasticity. Present in both the brain and periphery, BDNF plays critical roles throughout the body and is essential for placental and fetal development. Rodent studies show that early life stress, including prenatal stress, broadly alters BDNF methylation, with presumed changes in gene expression. No studies have assessed prenatal exposure to maternal traumatic stress and BDNF methylation in humans. This study examined associations of prenatal exposure to maternal stress and BDNF methylation at CpG sites across the BDNF gene. Among 24 mothers and newborns in the eastern Democratic Republic of Congo, a region with extreme conflict and violence to women, maternal experiences of war trauma and chronic stress were associated with BDNF methylation in umbilical cord blood, placental tissue, and maternal venous blood. Associations of maternal stress and BDNF methylation showed high tissue specificity. The majority of significant associations were observed in putative transcription factor binding regions. This is the first study in humans to examine BDNF methylation in relation to prenatal exposure to maternal stress in three tissues simultaneously and the first in any mammalian species to report associations of prenatal stress and BDNF methylation in placental tissue. The findings add to the growing body of evidence highlighting the importance of considering epigenetic effects when examining the impacts of trauma and stress, not only for adults but also for offspring exposed via effects transmitted before birth.

PEDF Is Associated with the Termination of Chondrocyte Phenotype and Catabolism of Cartilage Tissue.

PubMed

Klinger, P; Lukassen, S; Ferrazzi, F; Ekici, A B; Hotfiel, T; Swoboda, B; Aigner, T; Gelse, K

2017-01-01

Objective. To investigate the expression and target genes of pigment epithelium-derived factor (PEDF) in cartilage and chondrocytes, respectively. Methods. We analyzed the expression pattern of PEDF in different human cartilaginous tissues including articular cartilage, osteophytic cartilage, and fetal epiphyseal and growth plate cartilage, by immunohistochemistry and quantitative real-time (qRT) PCR. Transcriptome analysis after stimulation of human articular chondrocytes with rhPEDF was performed by RNA sequencing (RNA-Seq) and confirmed by qRT-PCR. Results. Immunohistochemically, PEDF could be detected in transient cartilaginous tissue that is prone to undergo endochondral ossification, including epiphyseal cartilage, growth plate cartilage, and osteophytic cartilage. In contrast, PEDF was hardly detected in healthy articular cartilage and in the superficial zone of epiphyses, regions that are characterized by a permanent stable chondrocyte phenotype. RNA-Seq analysis and qRT-PCR demonstrated that rhPEDF significantly induced the expression of a number of matrix-degrading factors including SAA1, MMP1, MMP3, and MMP13. Simultaneously, a number of cartilage-specific genes including COL2A1, COL9A2, COMP, and LECT were among the most significantly downregulated genes. Conclusions. PEDF represents a marker for transient cartilage during all neonatal and postnatal developmental stages and promotes the termination of cartilage tissue by upregulation of matrix-degrading factors and downregulation of cartilage-specific genes. These data provide the basis for novel strategies to stabilize the phenotype of articular cartilage and prevent its degradation.

Advances in Tissue Engineering Techniques for Articular Cartilage Repair

PubMed Central

Haleem, AM; Chu, CR

2010-01-01

The limited repair potential of human articular cartilage contributes to development of debilitating osteoarthritis and remains a great clinical challenge. This has led to evolution of cartilage treatment strategies from palliative to either reconstructive or reparative methods in an attempt to delay or “bridge the gap” to joint replacement. Further development of tissue engineering-based cartilage repair methods have been pursued to provide a more functional biological tissue. Currently, tissue engineering of articular cartilage has three cornerstones; a cell population capable of proliferation and differentiation into mature chondrocytes, a scaffold that can host these cells, provide a suitable environment for cellular functioning and serve as a sustained-release delivery vehicle of chondrogenic growth factors and thirdly, signaling molecules and growth factors that stimulate the cellular response and the production of a hyaline extracellular matrix (ECM). The aim of this review is to summarize advances in each of these three fields of tissue engineering with specific relevance to surgical techniques and technical notes. PMID:29430164

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

Assessment of stem cell differentiation based on genome-wide expression profiles.

PubMed

Godoy, Patricio; Schmidt-Heck, Wolfgang; Hellwig, Birte; Nell, Patrick; Feuerborn, David; Rahnenführer, Jörg; Kattler, Kathrin; Walter, Jörn; Blüthgen, Nils; Hengstler, Jan G

2018-07-05

In recent years, protocols have been established to differentiate stem and precursor cells into more mature cell types. However, progress in this field has been hampered by difficulties to assess the differentiation status of stem cell-derived cells in an unbiased manner. Here, we present an analysis pipeline based on published data and methods to quantify the degree of differentiation and to identify transcriptional control factors explaining differences from the intended target cells or tissues. The pipeline requires RNA-Seq or gene array data of the stem cell starting population, derived 'mature' cells and primary target cells or tissue. It consists of a principal component analysis to represent global expression changes and to identify possible problems of the dataset that require special attention, such as: batch effects; clustering techniques to identify gene groups with similar features; over-representation analysis to characterize biological motifs and transcriptional control factors of the identified gene clusters; and metagenes as well as gene regulatory networks for quantitative cell-type assessment and identification of influential transcription factors. Possibilities and limitations of the analysis pipeline are illustrated using the example of human embryonic stem cell and human induced pluripotent cells to generate 'hepatocyte-like cells'. The pipeline quantifies the degree of incomplete differentiation as well as remaining stemness and identifies unwanted features, such as colon- and fibroblast-associated gene clusters that are absent in real hepatocytes but typically induced by currently available differentiation protocols. Finally, transcription factors responsible for incomplete and unwanted differentiation are identified. The proposed method is widely applicable and allows an unbiased and quantitative assessment of stem cell-derived cells.This article is part of the theme issue 'Designer human tissue: coming to a lab near you'. © 2018 The Author(s).

Integrated approaches to spatiotemporally directing angiogenesis in host and engineered tissues.

PubMed

Kant, Rajeev J; Coulombe, Kareen L K

2018-03-15

The field of tissue engineering has turned towards biomimicry to solve the problem of tissue oxygenation and nutrient/waste exchange through the development of vasculature. Induction of angiogenesis and subsequent development of a vascular bed in engineered tissues is actively being pursued through combinations of physical and chemical cues, notably through the presentation of topographies and growth factors. Presenting angiogenic signals in a spatiotemporal fashion is beginning to generate improved vascular networks, which will allow for the creation of large and dense engineered tissues. This review provides a brief background on the cells, mechanisms, and molecules driving vascular development (including angiogenesis), followed by how biomaterials and growth factors can be used to direct vessel formation and maturation. Techniques to accomplish spatiotemporal control of vascularization include incorporation or encapsulation of growth factors, topographical engineering, and 3D bioprinting. The vascularization of engineered tissues and their application in angiogenic therapy in vivo is reviewed herein with an emphasis on the most densely vascularized tissue of the human body - the heart. Vascularization is vital to wound healing and tissue regeneration, and development of hierarchical networks enables efficient nutrient transfer. In tissue engineering, vascularization is necessary to support physiologically dense engineered tissues, and thus the field seeks to induce vascular formation using biomaterials and chemical signals to provide appropriate, pro-angiogenic signals for cells. This review critically examines the materials and techniques used to generate scaffolds with spatiotemporal cues to direct vascularization in engineered and host tissues in vitro and in vivo. Assessment of the field's progress is intended to inspire vascular applications across all forms of tissue engineering with a specific focus on highlighting the nuances of cardiac tissue engineering for the greater regenerative medicine community. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Regulatory aspects of tissue donation, banking and transplantation in India.

PubMed

Lobo Gajiwala, Astrid

2018-05-04

Amendments to India's Transplantation of Human Organs Act, 1994, have established the legality of tissue donation and transplantation from deceased donors and the conditions under which they are permitted. The amended Act, now known as The Transplantation of Human Organs and Tissues Act, 1994, seeks to prevent the commercialization of tissue donation and to guarantee the safety of indigenous allografts. Registration of tissue banks, compliance with national standards and the appointment of transplant co-ordinators in hospitals registered under the Act are now mandatory. A national registry and Regional and State networks for donation and transplantation of tissues have been introduced. Despite the amendments a few anomalies of the principal Act persist as some of the differences between tissue and organ donation and transplantation have been overlooked. These include the possibility of skin donation in locations other than hospitals; the donation of medical and surgical tissue residues which does not pose any risk to the living donor; the non-requirement for compatibility between donor and recipient; the delayed time factor between tissue donation and transplantation which makes identification of a recipient at the time of donation impossible; and the easy availability of alternatives to tissues which make waiting lists redundant for many tissues. Rules for the implementation of the amended Act were framed in 2014 but like the Act must be adopted by the State health assemblies to become universally applicable in the country.

Innate Immunity and Breast Milk

PubMed Central

Cacho, Nicole Theresa; Lawrence, Robert M.

2017-01-01

Human milk is a dynamic source of nutrients and bioactive factors; unique in providing for the human infant’s optimal growth and development. The growing infant’s immune system has a number of developmental immune deficiencies placing the infant at increased risk of infection. This review focuses on how human milk directly contributes to the infant’s innate immunity. Remarkable new findings clarify the multifunctional nature of human milk bioactive components. New research techniques have expanded our understanding of the potential for human milk’s effect on the infant that will never be possible with milk formulas. Human milk microbiome directly shapes the infant’s intestinal microbiome, while the human milk oligosaccharides drive the growth of these microbes within the gut. New techniques such as genomics, metabolomics, proteomics, and glycomics are being used to describe this symbiotic relationship. An expanded role for antimicrobial proteins/peptides within human milk in innate immune protection is described. The unique milieu of enhanced immune protection with diminished inflammation results from a complex interaction of anti-inflammatory and antioxidative factors provided by human milk to the intestine. New data support the concept of mucosal-associated lymphoid tissue and its contribution to the cellular content of human milk. Human milk stem cells (hMSCs) have recently been discovered. Their direct role in the infant for repair and regeneration is being investigated. The existence of these hMSCs could prove to be an easily harvested source of multilineage stem cells for the study of cancer and tissue regeneration. As the infant’s gastrointestinal tract and immune system develop, there is a comparable transition in human milk over time to provide fewer immune factors and more calories and nutrients for growth. Each of these new findings opens the door to future studies of human milk and its effect on the innate immune system and the developing infant. PMID:28611768

The effects of thyroid hormones on brown adipose tissue in humans: a PET-CT study.

PubMed

Zhang, Qiongyue; Miao, Qing; Ye, Hongying; Zhang, Zhaoyun; Zuo, Chuantao; Hua, Fengchun; Guan, Yihui; Li, Yiming

2014-09-01

Brown adipose tissue (BAT) is important for energy expenditure through thermogenesis, although its regulatory factors are not well known in humans. There is evidence suggesting that thyroid hormones affect BAT functions in some mammals, but the effects of thyroid hormones on BAT activity in humans are still unclear. The aim of this study was to investigate the effects of thyroid hormones on glucose metabolism of BAT and other organs in humans. Nine Graves' disease-caused hyperthyroid patients who were newly diagnosed and untreated were studied. Putative brown adipose tissue activity was determined by the integrated ¹⁸F-fluorodeoxyglucose (¹⁸F-FDG) positron-emission tomography and computed tomography (PET-CT). All hyperthyroid patients were treated with methimazole and had been monitored until their symptoms disappeared and thyroid hormone levels returned to normal. At the end, a second PET-CT scan was performed. The average follow-up period was 77 days. Meanwhile, compared with a group of seventy-five brown adipose tissue-negative controls, thyroid hormones of seventy-five BAT-positive healthy subjects were measured. Active brown adipose tissue was not present in any of the hyperthyroid patients. However, one patient with normalized thyroid function showed active BAT after therapy. The free T3 levels and free T4 levels were significantly lower in the 75 BAT-positive subjects than in the BAT-negative subjects. All hyperthyroid patients showed symmetrically increased uptake of fluorodeoxyglucose in skeletal muscles before treatment, whereas, the standardized uptake value was substantially decreased after treatment. Abnormally high circulating thyroid hormone levels may not increase brown adipose tissue activity, which may be limited by the increased obligatory thermogenesis of muscle in adult humans. Copyright © 2014 John Wiley & Sons, Ltd.

Metabolic Disposition of Osimertinib in Rats, Dogs, and Humans: Insights into a Drug Designed to Bind Covalently to a Cysteine Residue of Epidermal Growth Factor Receptor.

PubMed

Dickinson, Paul A; Cantarini, Mireille V; Collier, Jo; Frewer, Paul; Martin, Scott; Pickup, Kathryn; Ballard, Peter

2016-08-01

Preclinical and clinical studies were conducted to determine the metabolism and pharmacokinetics of osimertinib and key metabolites AZ5104 and AZ7550. Osimertinib was designed to covalently bind to epidermal growth factor receptors, allowing it to achieve nanomolar cellular potency (Finlay et al., 2014). Covalent binding was observed in incubations of radiolabeled osimertinib with human and rat hepatocytes, human and rat plasma, and human serum albumin. Osimertinib, AZ5104, and AZ7550 were predominantly metabolized by CYP3A. Seven metabolites were detected in human hepatocytes, also observed in rat or dog hepatocytes at similar or higher levels. After oral administration of radiolabeled osimertinib to rats, drug-related material was widely distributed, with the highest radioactivity concentrations measured at 6 hours postdose in most tissues; radioactivity was detectable in 42% of tissues 60 days postdose. Concentrations of [(14)C]-radioactivity in blood were lower than in most tissues. After the administration of a single oral dose of 20 mg of radiolabeled osimertinib to healthy male volunteers, ∼19% of the dose was recovered by 3 days postdose. At 84 days postdose, mean total radioactivity recovery was 14.2% and 67.8% of the dose in urine and feces. The most abundant metabolite identified in feces was AZ5104 (∼6% of dose). Osimertinib accounted for ∼1% of total radioactivity in the plasma of non-small cell lung cancer patients after 22 days of 80-mg osimertinib once-daily treatment; the most abundant circulatory metabolites were AZ7550 and AZ5104 (<10% of total osimertinib-related material). Osimertinib is extensively distributed and metabolized in humans and is eliminated primarily via the fecal route. Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.

FACS-based Isolation of Neural and Glioma Stem Cell Populations from Fresh Human Tissues Utilizing EGF Ligand

PubMed Central

Tome-Garcia, Jessica; Doetsch, Fiona; Tsankova, Nadejda M.

2018-01-01

Direct isolation of human neural and glioma stem cells from fresh tissues permits their biological study without prior culture and may capture novel aspects of their molecular phenotype in their native state. Recently, we demonstrated the ability to prospectively isolate stem cell populations from fresh human germinal matrix and glioblastoma samples, exploiting the ability of cells to bind the Epidermal Growth Factor (EGF) ligand in fluorescence-activated cell sorting (FACS). We demonstrated that FACS-isolated EGF-bound neural and glioblastoma populations encompass the sphere-forming colonies in vitro, and are capable of both self-renewal and multilineage differentiation. Here we describe in detail the purification methodology of EGF-bound (i.e., EGFR+) human neural and glioma cells with stem cell properties from fresh postmortem and surgical tissues. The ability to prospectively isolate stem cell populations using native ligand-binding ability opens new doors for understanding both normal and tumor cell biology in uncultured conditions, and is applicable for various downstream molecular sequencing studies at both population and single-cell resolution. PMID:29516026

The evolution and expression of the snaR family of small non-coding RNAs

PubMed Central

Parrott, Andrew M.; Tsai, Michael; Batchu, Priyanka; Ryan, Karen; Ozer, Harvey L.; Tian, Bin; Mathews, Michael B.

2011-01-01

We recently identified the snaR family of small non-coding RNAs that associate in vivo with the nuclear factor 90 (NF90/ILF3) protein. The major human species, snaR-A, is an RNA polymerase III transcript with restricted tissue distribution and orthologs in chimpanzee but not rhesus macaque or mouse. We report their expression in human tissues and their evolution in primates. snaR genes are exclusively in African Great Apes and some are unique to humans. Two novel families of snaR-related genetic elements were found in primates: CAS (catarrhine ancestor of snaR), limited to Old World Monkeys and apes; and ASR (Alu/snaR-related), present in all monkeys and apes. ASR and CAS appear to have spread by retrotransposition, whereas most snaR genes have spread by segmental duplication. snaR-A and snaR-G2 are differentially expressed in discrete regions of the human brain and other tissues, notably including testis. snaR-A is up-regulated in transformed and immortalized human cells, and is stably bound to ribosomes in HeLa cells. We infer that snaR evolved from the left monomer of the primate-specific Alu SINE family via ASR and CAS in conjunction with major primate speciation events, and suggest that snaRs participate in tissue- and species-specific regulation of cell growth and translation. PMID:20935053

IL-17 Promotes Angiogenic Factors IL-6, IL-8, and Vegf Production via Stat1 in Lung Adenocarcinoma.

PubMed

Huang, Qi; Duan, Limin; Qian, Xin; Fan, Jinshuo; Lv, Zhilei; Zhang, Xiuxiu; Han, Jieli; Wu, Feng; Guo, Mengfei; Hu, Guorong; Du, Jiao; Chen, Caiyun; Jin, Yang

2016-11-07

Inflammation and angiogenesis are two hallmarks of carcinoma. The proinflammatory cytokine interleukin-17 (IL-17) facilitates angiogenesis in lung cancer; however, the underlying mechanism is not fully understood. In this study, tumour microvessel density (MVD) was positively associated with IL-17, interleukin-6 (IL-6), interleukin-8 (IL-8), and vascular endothelial cell growth factor (VEGF) expression in human lung adenocarcinoma tissues, and it was increased in tumour tissues of A549-IL-17 cell-bearing nude mice. Importantly, positive correlations were also detected between IL-17 expression and IL-6, IL-8 and VEGF expression in human lung adenocarcinoma tissues. Furthermore, IL-6, IL-8 and VEGF production, as well as STAT1 phosphorylation, were increased in tumour tissues of A549-IL-17 cell-bearing nude mice in vivo and in A549 and H292 cells following IL-17 stimulation in vitro. In addition, STAT1 knockdown using an inhibitor and siRNA attenuated the IL-17-mediated increases in IL-6, IL-8 and VEGF expression in A549 and H292 cells. In conclusion, IL-17 may promote the production of the angiogenic inducers IL-6, IL-8 and VEGF via STAT1 signalling in lung adenocarcinoma.

Hepatic differentiation potential of commercially available human mesenchymal stem cells.

PubMed

Ong, Shin-Yeu; Dai, Hui; Leong, Kam W

2006-12-01

The ready availability and low immunogenicity of commercially available mesenchymal stem cells (MSC) render them a potential cell source for the development of therapeutic products. With cell source a major bottleneck in hepatic tissue engineering, we investigated whether commercially available human MSC (hMSC) can transdifferentiate into the hepatic lineage. Based on previous studies that find rapid gain of hepatic genes in bone marrow-derived stem cells cocultured with liver tissue, we used a similar approach to drive hepatic differentiation by coculturing the hMSC with rat livers treated or untreated with gadolinium chloride (GdCl(3)). After a 24-hour coculture period with liver tissue injured by GdCl(3) in a Transwell configuration, approximately 34% of the cells differentiated into albumin-expressing cells. Cocultured cells were subsequently maintained with growth factors to complete the hepatic differentiation. Cocultured cells expressed more hepatic gene markers, and had higher metabolic functions and P450 activity than cells that were only differentiated with growth factors. In conclusion, commercially available hMSC do show hepatic differentiation potential, and a liver microenvironment in culture can provide potent cues to accelerate and deepen the differentiation. The ability to generate hepatocyte-like cells from a commercially available cell source would find interesting applications in liver tissue engineering.

Increased circulating procoagulant and anticoagulant factors as TF and TFPI according to severity or infecting serotypes in human dengue infection.

PubMed

Leal de Azeredo, Elzinandes; Solórzano, Victor Edgar Fiestas; de Oliveira, Débora Batista; Marinho, Cintia Ferreira; de Souza, Luiz José; da Cunha, Rivaldo Venâncio; Damasco, Paulo Vieira; Kubelka, Claire Fernandes; de-Oliveira-Pinto, Luzia Maria

2017-01-01

Tissue Factor (TF) is the initiator of coagulation and Tissue Factor Inhibitor (TFPI) is the physiological inhibitor of the TF/FVIIa complex. Circulating levels of TF and TFPI were quantified in dengue patients and the relationships with disease severity and infecting serotype analysed. A significant decrease in TF and TPFI plasma levels was observed in mild DF patients compared with severe dengue. Furthermore, both factors were associated with haemorrhagic manifestations. Finally, TF levels were significantly increased in DENV-1/2 infected patients as compared with DENV-4. These findings suggest that activation of TF-pathway is an important component of DENV -related coagulation disorders. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Tissue Engineering Using Transfected Growth-Factor Genes

NASA Technical Reports Server (NTRS)

Madry, Henning; Langer, Robert S.; Freed, Lisa E.; Trippel, Stephen; Vunjak-Novakovic, Gordana

2005-01-01

A method of growing bioengineered tissues includes, as a major component, the use of mammalian cells that have been transfected with genes for secretion of regulator and growth-factor substances. In a typical application, one either seeds the cells onto an artificial matrix made of a synthetic or natural biocompatible material, or else one cultures the cells until they secrete a desired amount of an extracellular matrix. If such a bioengineered tissue construct is to be used for surgical replacement of injured tissue, then the cells should preferably be the patient s own cells or, if not, at least cells matched to the patient s cells according to a human-leucocyteantigen (HLA) test. The bioengineered tissue construct is typically implanted in the patient's injured natural tissue, wherein the growth-factor genes enhance metabolic functions that promote the in vitro development of functional tissue constructs and their integration with native tissues. If the matrix is biodegradable, then one of the results of metabolism could be absorption of the matrix and replacement of the matrix with tissue formed at least partly by the transfected cells. The method was developed for articular chondrocytes but can (at least in principle) be extended to a variety of cell types and biocompatible matrix materials, including ones that have been exploited in prior tissue-engineering methods. Examples of cell types include chondrocytes, hepatocytes, islet cells, nerve cells, muscle cells, other organ cells, bone- and cartilage-forming cells, epithelial and endothelial cells, connective- tissue stem cells, mesodermal stem cells, and cells of the liver and the pancreas. Cells can be obtained from cell-line cultures, biopsies, and tissue banks. Genes, molecules, or nucleic acids that secrete factors that influence the growth of cells, the production of extracellular matrix material, and other cell functions can be inserted in cells by any of a variety of standard transfection techniques.

Structural insights into humanization of anti-tissue factor antibody 10H10.

PubMed

Teplyakov, Alexey; Obmolova, Galina; Malia, Thomas J; Raghunathan, Gopalan; Martinez, Christian; Fransson, Johan; Edwards, Wilson; Connor, Judith; Husovsky, Matthew; Beck, Heena; Chi, Ellen; Fenton, Sandra; Zhou, Hong; Almagro, Juan Carlos; Gilliland, Gary L

Murine antibody 10H10 raised against human tissue factor is unique in that it blocks the signaling pathway, and thus inhibits angiogenesis and tumor growth without interfering with coagulation. As a potential therapeutic, the antibody was humanized in a two-step procedure. Antigen-binding loops were grafted onto selected human frameworks and the resulting chimeric antibody was subjected to affinity maturation by using phage display libraries. The results of humanization were analyzed from the structural perspective through comparison of the structure of a humanized variant with the parental mouse antibody. This analysis revealed several hot spots in the framework region that appear to affect antigen binding, and therefore should be considered in human germline selection. In addition, some positions in the Vernier zone, e.g., residue 71 in the heavy chain, that are traditionally thought to be crucial appear to tolerate amino acid substitutions without any effect on binding. Several humanized variants were produced using both short and long forms of complementarity-determining region (CDR) H2 following the difference in the Kabat and Martin definitions. Comparison of such pairs indicated consistently higher thermostability of the variants with short CDR H2. Analysis of the binding data in relation to the structures singled out the ImMunoGeneTics information system® germline IGHV1-2*01 as dubious owing to two potentially destabilizing mutations as compared to the other alleles of the same germline and to other human germlines.

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

Comparison of In Vitro- and Chorioallantoic Membrane (CAM)-Culture Systems for Cryopreserved Medulla-Contained Human Ovarian Tissue

PubMed Central

Isachenko, Vladimir; Mallmann, Peter; Petrunkina, Anna M.; Rahimi, Gohar; Nawroth, Frank; Hancke, Katharina; Felberbaum, Ricardo; Genze, Felicitas; Damjanoski, Ilija; Isachenko, Evgenia

2012-01-01

At present, there are three ways to determine effectively the quality of the cryopreservation procedure using ovarian tissue before the re-implantation treatment: evaluation of follicles after post-thawing xenotransplantation to SCID mouse, in-vitro culture in a large volume of culture medium under constant agitation and culture on embryonic chorio-allantoic membrane within a hen's eggs. The aim of this study was to compare the two methods, culture in vitro and culture on embryonic chorioallantoic membrane (CAM) of cryopreserved human ovarian medulla-contained and medulla-free cortex. Ovarian fragments were divided into small pieces (1.5–2.0×1.0–1.2×0.8–1.5) of two types, cortex with medulla and medulla-free cortex, frozen, thawed and randomly divided into the following four groups. Group 1: medulla-free cortex cultured in vitro for 8 days in large volume of medium with mechanical agitation, Group 2: medulla-containing cortex cultured in vitro, Group 3: medulla-free cortex cultured in CAM-system for 5 days, Group 4: medulla-containing cortex cultured in CAM-system. The efficacy of the tissue culture was evaluated by the development of follicles and by intensiveness of angiogenesis in the tissue (von Willebrand factor and Desmin). For Group 1, 2, 3 and 4, respectively 85%, 85%, 87% and 84% of the follicles were morphologically normal (P>0.1). The immunohistochemical analysis showed that angiogenesis detected by von Willebrand factor was lower in groups 1 and 3 (medulla-free cortex). Neo-vascularisation (by Desmin) was observed only in ovarian tissue of Group 4 (medulla-contained cortex after CAM-culture). It appears that the presence of medulla in ovarian pieces is beneficial for post-thaw development of cryopreserved human ovarian tissue. For medical practice it is recommended for evaluation of post-warming ovarian tissue to use the CAM-system as a valuable alternative to xenotransplantation and for cryopreservation of these tissues to prepare ovarian medulla-contained strips. PMID:22479331

Specific Accumulation of Tumor-Derived Adhesion Factor in Tumor Blood Vessels and in Capillary Tube-Like Structures of Cultured Vascular Endothelial Cells

NASA Astrophysics Data System (ADS)

Akaogi, Kotaro; Okabe, Yukie; Sato, Junji; Nagashima, Yoji; Yasumitsu, Hidetaro; Sugahara, Kazuyuki; Miyazaki, Kaoru

1996-08-01

Tumor-derived adhesion factor (TAF) was previously identified as a cell adhesion molecule secreted by human bladder carcinoma cell line EJ-1. To elucidate the physiological function of TAF, we examined its distribution in human normal and tumor tissues. Immunochemical staining with an anti-TAF monoclonal antibody showed that TAF was specifically accumulated in small blood vessels and capillaries within and adjacent to tumor nests, but not in those in normal tissues. Tumor blood vessel-specific staining of TAF was observed in various human cancers, such as esophagus, brain, lung, and stomach cancers. Double immunofluorescent staining showed apparent colocalization of TAF and type IV collagen in the vascular basement membrane. In vitro experiments demonstrated that TAF preferentially bound to type IV collagen among various extracellular matrix components tested. In cell culture experiments, TAF promoted adhesion of human umbilical vein endothelial cells to type IV collagen substrate and induced their morphological change. Furthermore, when the endothelial cells were induced to form capillary tube-like structures by type I collagen, TAF and type IV collagen were exclusively detected on the tubular structures. The capillary tube formation in vitro was prevented by heparin, which inhibited the binding of TAF to the endothelial cells. These results strongly suggest that TAF contributes to the organization of new capillary vessels in tumor tissues by modulating the interaction of endothelial cells with type IV collagen.

Characterization and classification of oral tissues using excitation and emission matrix: a statistical modeling approach

NASA Astrophysics Data System (ADS)

Kanniyappan, Udayakumar; Gnanatheepaminstein, Einstein; Prakasarao, Aruna; Dornadula, Koteeswaran; Singaravelu, Ganesan

2017-02-01

Cancer is one of the most common human threats around the world and diagnosis based on optical spectroscopy especially fluorescence technique has been established as the standard approach among scientist to explore the biochemical and morphological changes in tissues. In this regard, the present work aims to extract spectral signatures of the various fluorophores present in oral tissues using parallel factor analysis (PARAFAC). Subsequently, the statistical analysis also to be performed to show its diagnostic potential in distinguishing malignant, premalignant from normal oral tissues. Hence, the present study may lead to the possible and/or alternative tool for oral cancer diagnosis.

Altered Breast Development in Young Girls from an Agricultural Environment

PubMed Central

Guillette, Elizabeth A.; Conard, Craig; Lares, Fernando; Aguilar, Maria Guadalupe; McLachlan, John; Guillette, Louis J.

2006-01-01

In several human populations, the age at which female breast development begins is reported to have declined over the last five decades. Much debate has occurred over whether this reported decline has actually occurred and what factors contribute to it. However, geographical patterns reflecting earlier developmental onset in some human populations suggest environmental factors influence this phenomenon. These factors include interactions between genetic makeup, nutrition, and possible cumulative exposure to estrogens, both endogenous as well as environmental beginning during in utero development. We examined the onset of breast development in a group of peripubertal girls from the Yaqui Valley of Sonora, Mexico. We observed that girls from valley towns, areas using modern agricultural practices, exhibited larger breast fields than those of girls living in the foothills who exhibited similar stature [e.g., weight, height, body mass index (BMI)], and genetic background. Further, girls from valley towns displayed a poorly defined relationship between breast size and mammary gland development, whereas girls from the Yaqui foothills, where traditional ranching occurs, show a robust positive relationship between breast size and mammary size. The differences noted were obtained by a medically based exam involving morphometric analysis and palpation of tissues, in contrast to visual staging alone. In fact, use of the Tanner scale, involving visual staging of breast development for puberty, detected no differences between the study populations. Mammary tissue, determined by palpation, was absent in 18.5% of the girls living in agricultural areas, although palpable breast adipose tissue was present. No relationship was seen between mammary diameter and weight or BMI in either population. These data suggest that future in-depth studies examining mammary tissue growth and fat deposition in breast tissue are required if we are to understand environmental influences on these phenomena. PMID:16507474

Explant culture: An advantageous method for isolation of mesenchymal stem cells from human tissues.

PubMed

Hendijani, Fatemeh

2017-04-01

Mesenchymal stem cell (MSC) research progressively moves towards clinical phases. Accordingly, a wide range of different procedures were presented in the literature for MSC isolation from human tissues; however, there is not yet any close focus on the details to offer precise information for best method selection. Choosing a proper isolation method is a critical step in obtaining cells with optimal quality and yield in companion with clinical and economical considerations. In this concern, current review widely discusses advantages of omitting proteolysis step in isolation process and presence of tissue pieces in primary culture of MSCs, including removal of lytic stress on cells, reduction of in vivo to in vitro transition stress for migrated/isolated cells, reduction of price, processing time and labour, removal of viral contamination risk, and addition of supporting functions of extracellular matrix and released growth factors from tissue explant. In next sections, it provides an overall report of technical highlights and molecular events of explant culture method for isolation of MSCs from human tissues including adipose tissue, bone marrow, dental pulp, hair follicle, cornea, umbilical cord and placenta. Focusing on informative collection of molecular and methodological data about explant methods can make it easy for researchers to choose an optimal method for their experiments/clinical studies and also stimulate them to investigate and optimize more efficient procedures according to clinical and economical benefits. © 2017 John Wiley & Sons Ltd.

Isolation and characterization of adult human liver progenitors from ischemic liver tissue derived from therapeutic hepatectomies.

PubMed

Stachelscheid, Harald; Urbaniak, Thomas; Ring, Alexander; Spengler, Berlind; Gerlach, Jörg C; Zeilinger, Katrin

2009-07-01

Recent evidence suggests that progenitor cells in adult tissues and embryonic stem cells share a high resistance to hypoxia and ischemic stress. To study the ischemic resistance of adult liver progenitors, we characterized remaining viable cells in human liver tissue after cold ischemic treatment for 24-168 h, applied to the tissue before cell isolation. In vitro cultures of isolated cells showed a rapid decline of the number of different cell types with increasing ischemia length. After all ischemic periods, liver progenitor-like cells could be observed. The comparably small cells exhibited a low cytoplasm-to-nucleus ratio, formed densely packed colonies, and showed a hepatobiliary marker profile. The cells expressed epithelial cell adhesion molecule, epithelial-specific (CK8/18) and biliary-specific (CK7/19) cytokeratins, albumin, alpha-1-antitrypsin, cytochrome-P450 enzymes, as well as weak levels of hepatocyte nuclear factor-4 and gamma-glutamyl transferase, but not alpha-fetoprotein or Thy-1. In vitro survival and expansion was facilitated by coculture with mouse embryonic fibroblasts. Hepatic progenitor-like cells exhibit a high resistance to ischemic stress and can be isolated from human liver tissue after up to 7 days of ischemia. Ischemic liver tissue from various sources, thought to be unsuitable for cell isolation, may be considered as a prospective source of hepatic progenitor cells.

Immunohistochemical characterization of human olfactory tissue

PubMed Central

Holbrook, Eric H.; Wu, Enming; Curry, William T.; Lin, Derrick T.; Schwob, James E.

2011-01-01

Objectives/Hypothesis The pathophysiology underlying human olfactory disorders is poorly understood because biopsying the olfactory epithelium (OE) can be unrepresentative and extensive immunohistochemical analysis is lacking. Autopsy tissue enriches our grasp of normal and abnormal olfactory immunohistology and guides the sampling of the OE by biopsy. Furthermore, a comparison of the molecular phenotype of olfactory epithelial cells between rodents and humans will improve our ability to correlate human histopathology with olfactory dysfunction. Study Design An immunohistochemical analysis of human olfactory tissue using a comprehensive battery of proven antibodies. Methods Human olfactory mucosa obtained from 21 autopsy specimens was analyzed with immunohistochemistry. The position and extent of olfactory mucosa was assayed by staining whole mounts with neuronal markers. Sections of the OE were analyzed with an extensive group of antibodies directed against cytoskeletal proteins and transcription factors, as were surgical specimens from an esthesioneuroblastoma. Results Neuron-rich epithelium is always found inferior to the cribriform plate, even at advanced age, despite the interruptions in the neuroepithelial sheet caused by patchy respiratory metaplasia. The pattern of immunostaining with our antibody panel identifies two distinct types of basal cell progenitors in human OE similar to rodents. The panel also clarifies the complex composition of the esthesioneuroblastoma. Conclusion The extent of human olfactory mucosa at autopsy can easily be delineated as a function of age and neurological disease. The similarities in human vs. rodent OE will enable us to translate knowledge from experimental animals to humans and will extend our understanding of human olfactory pathophysiology. PMID:21792956

A short review on human exposure to and tissue distribution of per- and polyfluoroalkyl substances (PFASs).

PubMed

Jian, Jun-Meng; Chen, Da; Han, Fu-Juan; Guo, Ying; Zeng, Lixi; Lu, Xingwen; Wang, Fei

2018-09-15

PFASs are widely distributed in natural and living environment and can enter human bodies via different routes. Many studies have reported that PFASs may be associated with human diseases, such as urine acid and thyroid diseases. In this study, we reviewed PFAS levels in human bodies reported in past seven years, including blood, urine, milk, and tissues (hair and nails). Most studies focused on human blood. Blood type, spatiality, human age, and gender were found to have a strong relationship with PFAS levels in blood samples. The PFAS distribution in urine samples was reported to be associated with the chain length of PFASs and human gender. Urinary excretion was found to be an important pathway of PFAS elimination. PFAS levels in human milk might be affected by various factors, such as mothers' age, dietary habit, parity of mothers and the interval of interpregnancy. Data in hair and nails remain very limited, but these matrices offer a non-invasive approach to evaluate human exposure to PFASs. Copyright © 2018 Elsevier B.V. All rights reserved.

Msx homeobox gene family and craniofacial development.

PubMed

Alappat, Sylvia; Zhang, Zun Yi; Chen, Yi Ping

2003-12-01

Vertebrate Msx genes are unlinked, homeobox-containing genes that bear homology to the Drosophila muscle segment homeobox gene. These genes are expressed at multiple sites of tissue-tissue interactions during vertebrate embryonic development. Inductive interactions mediated by the Msx genes are essential for normal craniofacial, limb and ectodermal organ morphogenesis, and are also essential to survival in mice, as manifested by the phenotypic abnormalities shown in knockout mice and in humans. This review summarizes studies on the expression, regulation, and functional analysis of Msx genes that bear relevance to craniofacial development in humans and mice. Key words: Msx genes, craniofacial, tooth, cleft palate, suture, development, transcription factor, signaling molecule.

MACF1, versatility in tissue-specific function and in human disease.

PubMed

Hu, Lifang; Xiao, Yunyun; Xiong, Zhipeng; Zhao, Fan; Yin, Chong; Zhang, Yan; Su, Peihong; Li, Dijie; Chen, Zhihao; Ma, Xiaoli; Zhang, Ge; Qian, Airong

2017-09-01

Spectraplakins are a family of evolutionarily conserved gigantic proteins and play critical roles in many cytoskeleton-related processes. Microtubule actin crosslinking factor 1 (MACF1) is one of the most versatile spectraplakin with multiple isoforms. As a broadly expressed mammalian spectraplakin, MACF1 is important in maintaining normal functions of many tissues. The loss-of-function studies using knockout mouse models reveal the pivotal roles of MACF1 in embryo development, skin integrity maintenance, neural development, bone formation, and colonic paracellular permeability. Mutation in the human MACF1 gene causes a novel myopathy genetic disease. In addition, abnormal expression of MACF1 is associated with schizophrenia, Parkinson's disease, cancer and osteoporosis. This demonstrates the crucial roles of MACF1 in physiology and pathology. Here, we review the research advances of MACF1's roles in specific tissue and in human diseases, providing the perspectives of MACF1 for future studies. Copyright © 2017. Published by Elsevier Ltd.

Integrative analyses of conserved WNT clusters and their co-operative behaviour in human breast cancer

PubMed Central

Qurrat-ul-Ain; Seemab, Umair; Nawaz, Sulaman; Rashid, Sajid

2011-01-01

In human, WNT gene clusters are highly conserved at specie level and associated with carcinogenesis. Among them, WNT-10A and WNT-6 genes clustered in chromosome 2q35 are homologous to WNT-10B and WNT-1 located in chromosome 12q13, respectively. In an attempt to study co-regulation, the coordinated expression of these genes was monitored in human breast cancer tissues. As compared to normal tissue, both WNT-10A and WNT-10B genes exhibited lower expression while WNT-6 and WNT-1 showed increased expression in breast cancer tissues. The co-expression pattern was elaborated by detailed phylogenetic and syntenic analyses. Moreover, the intergenic and intragenic regions for these gene clusters were analyzed for studying the transcriptional regulation. In this context, adequate conserved binding sites for SOX and TCF family of transcriptional factors were observed. We propose that SOX9 and TCF4 may compete for binding at the promoters of WNT family genes thus regulating the disease phenotype. PMID:22355234

Human iPS cell-engineered cardiac tissue sheets with cardiomyocytes and vascular cells for cardiac regeneration

PubMed Central

Masumoto, Hidetoshi; Ikuno, Takeshi; Takeda, Masafumi; Fukushima, Hiroyuki; Marui, Akira; Katayama, Shiori; Shimizu, Tatsuya; Ikeda, Tadashi; Okano, Teruo; Sakata, Ryuzo; Yamashita, Jun K.

2014-01-01

To realize cardiac regeneration using human induced pluripotent stem cells (hiPSCs), strategies for cell preparation, tissue engineering and transplantation must be explored. Here we report a new protocol for the simultaneous induction of cardiomyocytes (CMs) and vascular cells [endothelial cells (ECs)/vascular mural cells (MCs)], and generate entirely hiPSC-engineered cardiovascular cell sheets, which showed advantageous therapeutic effects in infarcted hearts. The protocol adds to a previous differentiation protocol of CMs by using stage-specific supplementation of vascular endothelial cell growth factor for the additional induction of vascular cells. Using this cell sheet technology, we successfully generated physically integrated cardiac tissue sheets (hiPSC-CTSs). HiPSC-CTS transplantation to rat infarcted hearts significantly improved cardiac function. In addition to neovascularization, we confirmed that engrafted human cells mainly consisted of CMs in >40% of transplanted rats four weeks after transplantation. Thus, our HiPSC-CTSs show promise for cardiac regenerative therapy. PMID:25336194

A live zebrafish-based screening system for human nuclear receptor ligand and cofactor discovery.

PubMed

Tiefenbach, Jens; Moll, Pamela R; Nelson, Meryl R; Hu, Chun; Baev, Lilia; Kislinger, Thomas; Krause, Henry M

2010-03-22

Nuclear receptors (NRs) belong to a superfamily of transcription factors that regulate numerous homeostatic, metabolic and reproductive processes. Taken together with their modulation by small lipophilic molecules, they also represent an important and successful class of drug targets. Although many NRs have been targeted successfully, the majority have not, and one third are still orphans. Here we report the development of an in vivo GFP-based reporter system suitable for monitoring NR activities in all cells and tissues using live zebrafish (Danio rerio). The human NR fusion proteins used also contain a new affinity tag cassette allowing the purification of receptors with bound molecules from responsive tissues. We show that these constructs 1) respond as expected to endogenous zebrafish hormones and cofactors, 2) facilitate efficient receptor and cofactor purification, 3) respond robustly to NR hormones and drugs and 4) yield readily quantifiable signals. Transgenic lines representing the majority of human NRs have been established and are available for the investigation of tissue- and isoform-specific ligands and cofactors.

Injury-induced ctgfa directs glial bridging and spinal cord regeneration in zebrafish

PubMed Central

Mokalled, Mayssa H.; Patra, Chinmoy; Dickson, Amy L.; Endo, Toyokazu; Stainier, Didier Y. R.; Poss, Kenneth D.

2016-01-01

Unlike mammals, zebrafish efficiently regenerate functional nervous system tissue after major spinal cord injury. Whereas glial scarring presents a roadblock for mammalian spinal cord repair, glial cells in zebrafish form a bridge across severed spinal cord tissue and facilitate regeneration, a relatively unexplored process. Here, we performed a genome-wide profiling screen for secreted factors that are upregulated during zebrafish spinal cord regeneration. We find that connective tissue growth factor a (ctgfa) is induced in and around glial cells that participate in initial bridging events. Mutations in ctgfa disrupt spinal cord repair, while transgenic ctgfa overexpression and local human CTGF recombinant protein delivery accelerate bridging and functional regeneration. Our study reveals that CTGF is necessary and sufficient to stimulate glial bridging and natural spinal cord regeneration. PMID:27811277

Differential expression of THOC1 and ALY mRNP biogenesis/export factors in human cancers

PubMed Central

2011-01-01

Background One key step in gene expression is the biogenesis of mRNA ribonucleoparticle complexes (mRNPs). Formation of the mRNP requires the participation of a number of conserved factors such as the THO complex. THO interacts physically and functionally with the Sub2/UAP56 RNA-dependent ATPase, and the Yra1/REF1/ALY RNA-binding protein linking transcription, mRNA export and genome integrity. Given the link between genome instability and cancer, we have performed a comparative analysis of the expression patterns of THOC1, a THO complex subunit, and ALY in tumor samples. Methods The mRNA levels were measured by quantitative real-time PCR and hybridization of a tumor tissue cDNA array; and the protein levels and distribution by immunostaining of a custom tissue array containing a set of paraffin-embedded samples of different tumor and normal tissues followed by statistical analysis. Results We show that the expression of two mRNP factors, THOC1 and ALY are altered in several tumor tissues. THOC1 mRNA and protein levels are up-regulated in ovarian and lung tumors and down-regulated in those of testis and skin, whereas ALY is altered in a wide variety of tumors. In contrast to THOC1, ALY protein is highly detected in normal proliferative cells, but poorly in high-grade cancers. Conclusions These results suggest a differential connection between tumorogenesis and the expression levels of human THO and ALY. This study opens the possibility of defining mRNP biogenesis factors as putative players in cell proliferation that could contribute to tumor development. PMID:21329510

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

Recombinant Factor VIIa Reduces Rebleed Hemorrhage Volume in a Swine Aortotomy Model: A Randomized Double-Blinded Study

DTIC Science & Technology

2008-06-01

Bpops,[ a dose of 90 Hg kgj1 will produce serum concentrations of factor VII high enough to reduce hemorrhage by accelerating thrombin production and...interaction is species-specific, and human FVII seems to have only between 5% and 50% activity when exposed to porcine tissue factor (15). The results...Copyright @ 200 by the Shock Society. Unauthorized reproduction of this article is prohibited.8 RECOMBINANT FACTOR VIIA REDUCES REBLEED HEMORRHAGE

Differential Protein Expression Marks the Transition From Infection With Opisthorchis viverrini to Cholangiocarcinoma*

PubMed Central

Khoontawad, Jarinya; Pairojkul, Chawalit; Rucksaken, Rucksak; Pinlaor, Porntip; Wongkham, Chaisiri; Yongvanit, Puangrat; Pugkhem, Ake; Jones, Alun; Plieskatt, Jordan; Potriquet, Jeremy; Bethony, Jeffery; Pinlaor, Somchai; Mulvenna, Jason

2017-01-01

Parts of Southeast Asia have the highest incidence of intrahepatic cholangiocarcinoma (CCA) in the world because of infection by the liver fluke Opisthorchis viverrini (Ov). Ov-associated CCA is the culmination of chronic Ov-infection, with the persistent production of the growth factors and cytokines associated with persistent inflammation, which can endure for years in Ov-infected individuals prior to transitioning to CCA. Isobaric labeling and tandem mass spectrometry of liver tissue from a hamster model of CCA was used to compare protein expression profiles from inflammed tissue (Ovinfected but not cancerous) versus cancerous tissue (Ov-induced CCA). Immunohistochemistry and immunoblotting were used to verify dysregulated proteins in the animal model and in human tissue. We identified 154 dysregulated proteins that marked the transition from Ov-infection to Ov-induced CCA, i.e. proteins dysregulated during carcinogenesis but not Ov-infection. The verification of dysregulated proteins in resected liver tissue from humans with Ov-associated CCA showed the numerous parallels in protein dysregulation between human and animal models of Ov-induced CCA. To identify potential circulating markers for CCA, dysregulated proteins were compared with proteins isolated from exosomes secreted by a human CCA cell line (KKU055) and 27 proteins were identified as dysregulated in CCA and present in exosomes. These data form the basis of potential diagnostic biomarkers for human Ov-associated CCA. The profile of protein dysregulation observed during chronic Ovinfection and then in Ov-induced CCA provides insight into the etiology of an infection-induced inflammation-related cancer. PMID:28232516

Anti-inflammatory activity of clove (Eugenia caryophyllata) essential oil in human dermal fibroblasts.

PubMed

Han, Xuesheng; Parker, Tory L

2017-12-01

Clove (Eugenia caryophyllata Thunb. [Myrtaceae]) essential oil (CEO) has been shown to possess antimicrobial, antifungal, antiviral, antioxidant, anti-inflammatory and anticancer properties. However, few studies have focused on its topical use. We investigated the biological activity of a commercially available CEO in a human skin disease model. We evaluated the effect of CEO on 17 protein biomarkers that play critical roles in inflammation and tissue remodelling in a validated human dermal fibroblast system, which was designed to model chronic inflammation and fibrosis. Four concentrations of CEO (0.011, 0.0037, 0.0012, and 0.00041%, v/v) were studied. The effect of 0.011% CEO on genome-wide gene expression was also evaluated. CEO at a concentration of 0.011% showed robust antiproliferative effects on human dermal fibroblasts. It significantly inhibited the increased production of several proinflammatory biomarkers such as vascular cell adhesion molecule-1 (VCAM-1), interferon γ-induced protein 10 (IP-10), interferon-inducible T-cell α chemoattractant (I-TAC), and monokine induced by γ interferon (MIG). CEO also significantly inhibited tissue remodelling protein molecules, namely, collagen-I, collagen-III, macrophage colony-stimulating factor (M-CSF), and tissue inhibitor of metalloproteinase 2 (TIMP-2). Furthermore, it significantly modulated global gene expression and altered signalling pathways critical for inflammation, tissue remodelling, and cancer signalling processes. CEO significantly inhibited VCAM-1 and collagen III at both protein and gene expression levels. This study provides important evidence of CEO-induced anti-inflammatory and tissue remodelling activity in human dermal fibroblasts. This study also supports the anticancer properties of CEO and its major active component eugenol.

Human embryonic stem cell-derived mesodermal progenitors display substantially increased tissue formation compared to human mesenchymal stem cells under dynamic culture conditions in a packed bed/column bioreactor.

PubMed

de Peppo, Giuseppe Maria; Sladkova, Martina; Sjövall, Peter; Palmquist, Anders; Oudina, Karim; Hyllner, Johan; Thomsen, Peter; Petite, Hervé; Karlsson, Camilla

2013-01-01

Bone tissue engineering represents a promising strategy to obviate bone deficiencies, allowing the ex vivo construction of bone substitutes with unprecedented potential in the clinical practice. Considering that in the human body cells are constantly stimulated by chemical and mechanical stimuli, the use of bioreactor is emerging as an essential factor for providing the proper environment for the reproducible and large-scale production of the engineered substitutes. Human mesenchymal stem cells (hMSCs) are experimentally relevant cells but, regardless the encouraging results reported after culture under dynamic conditions in bioreactors, show important limitations for tissue engineering applications, especially considering their limited proliferative potential, loss of functionality following protracted expansion, and decline in cellular fitness associated with aging. On the other hand, we previously demonstrated that human embryonic stem cell-derived mesodermal progenitors (hES-MPs) hold great potential to provide a homogenous and unlimited source of cells for bone engineering applications. Based on prior scientific evidence using different types of stem cells, in the present study we hypothesized that dynamic culture of hES-MPs in a packed bed/column bioreactor had the potential to affect proliferation, expression of genes involved in osteogenic differentiation, and matrix mineralization, therefore resulting in increased bone-like tissue formation. The reported findings suggest that hES-MPs constitute a suitable alternative cell source to hMSCs and hold great potential for the construction of bone substitutes for tissue engineering applications in clinical settings.

A Novel Semi-Supervised Methodology for Extracting Tumor Type-Specific MRS Sources in Human Brain Data

PubMed Central

Ortega-Martorell, Sandra; Ruiz, Héctor; Vellido, Alfredo; Olier, Iván; Romero, Enrique; Julià-Sapé, Margarida; Martín, José D.; Jarman, Ian H.; Arús, Carles; Lisboa, Paulo J. G.

2013-01-01

Background The clinical investigation of human brain tumors often starts with a non-invasive imaging study, providing information about the tumor extent and location, but little insight into the biochemistry of the analyzed tissue. Magnetic Resonance Spectroscopy can complement imaging by supplying a metabolic fingerprint of the tissue. This study analyzes single-voxel magnetic resonance spectra, which represent signal information in the frequency domain. Given that a single voxel may contain a heterogeneous mix of tissues, signal source identification is a relevant challenge for the problem of tumor type classification from the spectroscopic signal. Methodology/Principal Findings Non-negative matrix factorization techniques have recently shown their potential for the identification of meaningful sources from brain tissue spectroscopy data. In this study, we use a convex variant of these methods that is capable of handling negatively-valued data and generating sources that can be interpreted as tumor class prototypes. A novel approach to convex non-negative matrix factorization is proposed, in which prior knowledge about class information is utilized in model optimization. Class-specific information is integrated into this semi-supervised process by setting the metric of a latent variable space where the matrix factorization is carried out. The reported experimental study comprises 196 cases from different tumor types drawn from two international, multi-center databases. The results indicate that the proposed approach outperforms a purely unsupervised process by achieving near perfect correlation of the extracted sources with the mean spectra of the tumor types. It also improves tissue type classification. Conclusions/Significance We show that source extraction by unsupervised matrix factorization benefits from the integration of the available class information, so operating in a semi-supervised learning manner, for discriminative source identification and brain tumor labeling from single-voxel spectroscopy data. We are confident that the proposed methodology has wider applicability for biomedical signal processing. PMID:24376744

Effect of chitosan and coagulation factors on the wound repair phenotype of bioengineered blood clots.

PubMed

Hoemann, Caroline D; Marchand, Catherine; Rivard, Georges-Etienne; El-Gabalawy, Hani; Poubelle, Patrice E

2017-11-01

Controlling the blood clot phenotype in a surgically prepared wound is an evolving concept in scaffold-guided tissue engineering. Here, we investigated the effect of added chitosan (80% or 95% Degree of Deacetylation, DDA) or coagulation factors (recombinant human Factor VIIa, Tissue Factor, thrombin) on inflammatory factors released by blood clots. We tested the hypothesis that 80% DDA chitosan specifically enhances leukotriene B 4 (LTB 4 ) production. Human or rabbit whole blood was combined with isotonic chitosan solutions, coagulation factors, or lipopolysaccharide, cultured in vitro at 37°C, and after 4hours the serum was assayed for LTB 4 or inflammatory factors. Only 80% DDA chitosan clots produced around 15-fold more LTB 4 over other clots including 95% DDA chitosan clots. All serum contained high levels of PDGF-BB and CXCL8. Normal clots produced very low type I cytokines compared to lipopolysaccharide clots, with even lower IL-6 and IL-12 and more CCL3/CCL4 produced by chitosan clots. Coagulation factors had no detectable effect on clot phenotype. Conclusion In blood clots from healthy individuals, 80% DDA chitosan has a unique influence of inducing more LTB 4 , a potent neutrophil chemoattractant, with similar production of PDGF-BB and CXCL8, and lower type I cytokines, compared to whole blood clots. Copyright © 2017 Elsevier B.V. All rights reserved.

Optical redox imaging indices discriminate human breast cancer from normal tissues

NASA Astrophysics Data System (ADS)

Xu, He N.; Tchou, Julia; Feng, Min; Zhao, Huaqing; Li, Lin Z.

2016-11-01

Our long-term goal was to investigate the potential of incorporating redox imaging technique as a breast cancer (BC) diagnosis component to increase the positive predictive value of suspicious imaging finding and to reduce unnecessary biopsies and overdiagnosis. We previously found that precancer and cancer tissues in animal models displayed abnormal mitochondrial redox state. We also revealed abnormal mitochondrial redox state in cancerous specimens from three BC patients. Here, we extend our study to include biopsies of 16 patients. Tissue aliquots were collected from both apparently normal and cancerous tissues from the affected cancer-bearing breasts shortly after surgical resection. All specimens were snap-frozen and scanned with the Chance redox scanner, i.e., the three-dimensional cryogenic NADH/Fp (reduced nicotinamide adenine dinucleotide/oxidized flavoproteins) fluorescence imager. We found both Fp and NADH in the cancerous tissues roughly tripled that in the normal tissues (p<0.05). The redox ratio Fp/(NADH + Fp) was ˜27% higher in the cancerous tissues (p<0.05). Additionally, Fp, or NADH, or the redox ratio alone could predict cancer with reasonable sensitivity and specificity. Our findings suggest that the optical redox imaging technique can provide parameters independent of clinical factors for discriminating cancer from noncancer breast tissues in human patients.

Optical redox imaging indices discriminate human breast cancer from normal tissues

PubMed Central

Xu, He N.; Tchou, Julia; Feng, Min; Zhao, Huaqing; Li, Lin Z.

2016-01-01

Abstract. Our long-term goal was to investigate the potential of incorporating redox imaging technique as a breast cancer (BC) diagnosis component to increase the positive predictive value of suspicious imaging finding and to reduce unnecessary biopsies and overdiagnosis. We previously found that precancer and cancer tissues in animal models displayed abnormal mitochondrial redox state. We also revealed abnormal mitochondrial redox state in cancerous specimens from three BC patients. Here, we extend our study to include biopsies of 16 patients. Tissue aliquots were collected from both apparently normal and cancerous tissues from the affected cancer-bearing breasts shortly after surgical resection. All specimens were snap-frozen and scanned with the Chance redox scanner, i.e., the three-dimensional cryogenic NADH/Fp (reduced nicotinamide adenine dinucleotide/oxidized flavoproteins) fluorescence imager. We found both Fp and NADH in the cancerous tissues roughly tripled that in the normal tissues (p<0.05). The redox ratio Fp/(NADH + Fp) was ∼27% higher in the cancerous tissues (p<0.05). Additionally, Fp, or NADH, or the redox ratio alone could predict cancer with reasonable sensitivity and specificity. Our findings suggest that the optical redox imaging technique can provide parameters independent of clinical factors for discriminating cancer from noncancer breast tissues in human patients. PMID:27896360

Human 'brite/beige' adipocytes develop from capillary networks, and their implantation improves metabolic homeostasis in mice.

PubMed

Min, So Yun; Kady, Jamie; Nam, Minwoo; Rojas-Rodriguez, Raziel; Berkenwald, Aaron; Kim, Jong Hun; Noh, Hye-Lim; Kim, Jason K; Cooper, Marcus P; Fitzgibbons, Timothy; Brehm, Michael A; Corvera, Silvia

2016-03-01

Uncoupling protein 1 (UCP1) is highly expressed in brown adipose tissue, where it generates heat by uncoupling electron transport from ATP production. UCP1 is also found outside classical brown adipose tissue depots, in adipocytes that are termed 'brite' (brown-in-white) or 'beige'. In humans, the presence of brite or beige (brite/beige) adipocytes is correlated with a lean, metabolically healthy phenotype, but whether a causal relationship exists is not clear. Here we report that human brite/beige adipocyte progenitors proliferate in response to pro-angiogenic factors, in association with expanding capillary networks. Adipocytes formed from these progenitors transform in response to adenylate cyclase activation from being UCP1 negative to being UCP1 positive, which is a defining feature of the beige/brite phenotype, while displaying uncoupled respiration. When implanted into normal chow-fed, or into high-fat diet (HFD)-fed, glucose-intolerant NOD-scid IL2rg(null) (NSG) mice, brite/beige adipocytes activated in vitro enhance systemic glucose tolerance. These adipocytes express neuroendocrine and secreted factors, including the pro-protein convertase PCSK1, which is strongly associated with human obesity. Pro-angiogenic conditions therefore drive the proliferation of human beige/brite adipocyte progenitors, and activated beige/brite adipocytes can affect systemic glucose homeostasis, potentially through a neuroendocrine mechanism.

Human ‘brite / beige’ adipocytes develop from capillary networks and their implantation improves metabolic homeostasis in mice

PubMed Central

Min, So Yun; Kady, Jamie; Nam, Minwoo; Rojas-Rodriguez, Raziel; Berkenwald, Aaron; Kim, Jong Hun; Noh, Hye-Lim; Kim, Jason K.; Cooper, Marcus P.; Fitzgibbons, Timothy; Brehm, Michael A.; Corvera, Silvia

2015-01-01

The uncoupling protein 1 (UCP1) is highly expressed in brown adipose tissue, where it generates heat by uncoupling electron transport from ATP production. UCP1 is also found outside classical brown adipose tissue depots1–4, in adipocytes termed ‘brite’ (brown-in-white) or ‘beige’. In humans, the presence of ‘brite/beige’ adipocytes correlates with a lean, metabolically healthy phenotype5–8, but whether a causal relationship exists is not clear. Here we report that human ‘brite/beige’ adipocyte progenitors proliferate in response to pro-angiogenic factors, in association with expanding capillary networks. Adipocytes formed from these progenitors transform from being UCP1-negative to UCP1-positive in response to adenylate cyclase activation, a defining feature of the ‘beige/brite’ phenotype, and display uncoupled respiration. When implanted into normal or high fat diet-fed, glucose intolerant NOD-scid IL2rgnull mice, activated ‘brite/beige’ adipocytes enhance systemic glucose tolerance. These adipocytes express neuroendocrine and secreted factors, including the pro-protein convertase PCSK1, which is strongly associated with human obesity. Thus, pro-angiogenic conditions drive proliferation of human ‘beige/brite’ adipocyte progenitors, and activated ‘beige/brite’ adipocytes can affect systemic glucose homeostasis, potentially through a neuroendocrine mechanism. PMID:26808348

PPAR-alpha agonist treatment increases trefoil factor family-3 expression and attenuates apoptosis in the liver tissue of bile duct-ligated rats.

PubMed

Karakan, Tarkan; Kerem, Mustafa; Cindoruk, Mehmet; Engin, Doruk; Alper, Murat; Akın, Okan

2013-01-01

Peroxisome proliferators-activated receptor alpha activation modulates cholesterol metabolism and suppresses bile acid synthesis. The trefoil factor family comprises mucin-associated proteins that increase the viscosity of mucins and help protect epithelial linings from insults. We evaluated the effect of short-term administration of fenofibrate, a peroxisome proliferators activated receptor alpha agonist, on trefoil factor family-3 expression, degree of apoptosis, generation of free radicals, and levels of proinflammatory cytokines in the liver tissue of bile duct-ligated rats. Forty male Wistar rats were randomly divided into four groups: 1 = sham operated, 2 = bile duct ligation, 3 = bile duct-ligated + vehicle (gum Arabic), and 4 = bile duct-ligated + fenofibrate (100 mg/kg/day). All rats were sacrificed on the 7 th day after obtaining blood samples and liver tissue. Liver function tests, tumor necrosis factor-alpha and interleukin 1 beta in serum, and trefoil factor family-3 mRNA expression, degree of apoptosis (TUNEL) and tissue malondialdehyde (malondialdehyde, end-product of lipid peroxidation by reactive oxygen species) in liver tissue were evaluated. Fenofibrate administration significantly reduced serum total bilirubin, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transferase, and tumor necrosis factor-alpha and interleukin-1β levels. Apoptosis and malondialdehyde were significantly reduced in the fenofibrate group. Trefoil factor family-3 expression increased with fenofibrate treatment in bile duct-ligated rats. The peroxisome proliferators-activated receptor alpha agonist fenofibrate significantly increased trefoil factor family-3 expression and decreased apoptosis and lipid peroxidation in the liver and attenuated serum levels of proinflammatory cytokines in bile duct-ligated rats. Further studies are needed to determine the protective role of fenofibrate in human cholestatic disorders.

The Effect of Cryopreserved Human Placental Tissues on Biofilm Formation of Wound-Associated Pathogens.

PubMed

Mao, Yong; Singh-Varma, Anya; Hoffman, Tyler; Dhall, Sandeep; Danilkovitch, Alla; Kohn, Joachim

2018-01-08

Biofilm, a community of bacteria, is tolerant to antimicrobial agents and ubiquitous in chronic wounds. In a chronic DFU (Diabetic Foot Ulcers) clinical trial, the use of a human cryopreserved viable amniotic membrane (CVAM) resulted in a high rate of wound closure and reduction of wound-related infections. Our previous study demonstrated that CVAM possesses intrinsic antimicrobial activity against a spectrum of wound-associated bacteria under planktonic culture conditions. In this study, we evaluated the effect of CVAM and cryopreserved viable umbilical tissue (CVUT) on biofilm formation of S. aureus and P. aeruginosa , the two most prominent pathogens associated with chronic wounds. Firstly, we showed that, like CVAM, CVUT released antibacterial activity against multiple bacterial pathogens and the devitalization of CVUT reduced its antibacterial activity. The biofilm formation was then measured using a high throughput method and an ex vivo porcine dermal tissue model. We demonstrate that the formation of biofilm was significantly reduced in the presence of CVAM- or CVUT-derived conditioned media compared to control assay medium. The formation of P. aeruginosa biofilm on CVAM-conditioned medium saturated porcine dermal tissues was reduced 97% compared with the biofilm formation on the control medium saturated dermal tissues. The formation of S. auerus biofilm on CVUT-conditioned medium saturated dermal tissues was reduced 72% compared with the biofilm formation on the control tissues. This study is the first to show that human cryopreserved viable placental tissues release factors that inhibit biofilm formation. Our results provide an explanation for the in vivo observation of their ability to support wound healing.

Increased bioactive lipids content in human subcutaneous and epicardial fat tissue correlates with insulin resistance.

PubMed

Błachnio-Zabielska, Agnieszka U; Baranowski, Marcin; Hirnle, Tomasz; Zabielski, Piotr; Lewczuk, Anna; Dmitruk, Iwona; Górski, Jan

2012-12-01

Obesity is a risk factor for metabolic diseases. Intramuscular lipid accumulation of ceramides, diacylglycerols, and long chain acyl-CoA is responsible for the induction of insulin resistance. These lipids are probably implicated in obesity-associated insulin resistance not only in skeletal muscle but also in fat tissue. Only few data are available about ceramide content in human subcutaneous adipose tissue. However, there are no data on DAG and LCACoA content in adipose tissue. The aim of our study was to measure the lipids content in human SAT and epicardial adipose tissue we sought to determine the bioactive lipids content by LC/MS/MS in fat tissue from lean non-diabetic, obese non-diabetic, and obese diabetic subjects and test whether the lipids correlate with HOMA-IR. We found, that total content of measured lipids was markedly higher in OND and OD subjects in both types of fat tissue (for all p < 0.001) as compared to LND group. In SAT we found positive correlation between HOMA-IR and C16:0-Cer (r = 0.79, p < 0.001) and between HOMA-IR and C16:0/18:2 DAG (r = 0.56, p < 0.001). In EAT we found a strong correlation between C16:0-CoA content and HOMA-IR (r = 0.73, p < 0.001). The study showed that in obese and obese diabetic patients, bioactive lipids content is greater in subcutaneous and epicardial fat tissue and the particular lipids content positively correlates with HOMA-IR.

3D Bioprinting and In Vitro Cardiovascular Tissue Modeling.

PubMed

Jang, Jinah

2017-08-18

Numerous microfabrication approaches have been developed to recapitulate morphologically and functionally organized tissue microarchitectures in vitro; however, the technical and operational limitations remain to be overcome. 3D printing technology facilitates the building of a construct containing biomaterials and cells in desired organizations and shapes that have physiologically relevant geometry, complexity, and micro-environmental cues. The selection of biomaterials for 3D printing is considered one of the most critical factors to achieve tissue function. It has been reported that some printable biomaterials, having extracellular matrix-like intrinsic microenvironment factors, were capable of regulating stem cell fate and phenotype. In particular, this technology can control the spatial positions of cells, and provide topological, chemical, and complex cues, allowing neovascularization and maturation in the engineered cardiovascular tissues. This review will delineate the state-of-the-art 3D bioprinting techniques in the field of cardiovascular tissue engineering and their applications in translational medicine. In addition, this review will describe 3D printing-based pre-vascularization technologies correlated with implementing blood perfusion throughout the engineered tissue equivalent. The described engineering method may offer a unique approach that results in the physiological mimicry of human cardiovascular tissues to aid in drug development and therapeutic approaches.

3D Bioprinting and In Vitro Cardiovascular Tissue Modeling

PubMed Central

Jang, Jinah

2017-01-01

Numerous microfabrication approaches have been developed to recapitulate morphologically and functionally organized tissue microarchitectures in vitro; however, the technical and operational limitations remain to be overcome. 3D printing technology facilitates the building of a construct containing biomaterials and cells in desired organizations and shapes that have physiologically relevant geometry, complexity, and micro-environmental cues. The selection of biomaterials for 3D printing is considered one of the most critical factors to achieve tissue function. It has been reported that some printable biomaterials, having extracellular matrix-like intrinsic microenvironment factors, were capable of regulating stem cell fate and phenotype. In particular, this technology can control the spatial positions of cells, and provide topological, chemical, and complex cues, allowing neovascularization and maturation in the engineered cardiovascular tissues. This review will delineate the state-of-the-art 3D bioprinting techniques in the field of cardiovascular tissue engineering and their applications in translational medicine. In addition, this review will describe 3D printing-based pre-vascularization technologies correlated with implementing blood perfusion throughout the engineered tissue equivalent. The described engineering method may offer a unique approach that results in the physiological mimicry of human cardiovascular tissues to aid in drug development and therapeutic approaches. PMID:28952550

Expression of the stem cell factor in fibroblasts, endothelial cells, and macrophages in periapical tissues in human chronic periapical diseases.

PubMed

Shen, S Q; Wang, R; Huang, S G

2017-03-08

Stem cell factor (SCF), an important stem cell cytokine, has multiple functions. Fibroblasts (FBs), mature mast cells, endothelial cells (ECs), and eosinophil granulocytes can produce SCF in the inflammatory process. Therefore, we aimed to observe SCF expression in FBs, ECs, and macrophages (MPs) in periapical tissues in human chronic periapical disease and investigate the effects of cells expressing SCF in pathogenesis of the disease. Healthy (N = 20), periapical cyst (N = 15), and periapical granuloma (N = 15) tissues were fixed in 10% formalin for 48 h, embedded in paraffin, and stained with hematoxylin and eosin to observe histological changes. SCF expression was observed in FBs, ECs, and MPs in periapical tissues by double immunofluorescence. CD334, CD31, and CD14 are specific markers of FBs, ECs, and MPs, respectively. Results showed that densities of CD334-SCF double-positive FBs, CD31-SCF double-positive ECs, and CD14-SCF double-positive MPs were significantly increased in periapical tissue groups (P < 0.01). There were no significant differences in CD334-SCF double-positive FB and CD31-SCF double-positive EC levels between the two periapical tissue groups (P > 0.05). CD14-SCF double-positive MP density was considerably higher in periapical granulomas than in cysts (P < 0.01). FB, EC, and MP levels were significantly high and densities of CD334-SCF double-positive FBs, CD31-SCF double-positive ECs, and CD14-SCF double-positive MPs improved considerably in chronic periapical tissues, suggesting that the cells might be related to occurrence, development, and pathogenesis of chronic periapical disease.

Development of an ex vivo cellular model of rheumatoid arthritis: critical role of CD14-positive monocyte/macrophages in the development of pannus tissue.

PubMed

Nozaki, Toshiko; Takahashi, Kyoko; Ishii, Osamu; Endo, Sachio; Hioki, Kyoji; Mori, Toshihito; Kikukawa, Tadahiro; Boumpas, Dimitrios T; Ozaki, Shoichi; Yamada, Hidehiro

2007-09-01

To establish an ex vivo cellular model of pannus, the aberrant overgrowth of human synovial tissue (ST). Inflammatory cells that infiltrated pannus tissue from patients with rheumatoid arthritis (RA) were collected without enzyme digestion, and designated as ST-derived inflammatory cells. Single-cell suspensions of ST-derived inflammatory cells were cultured in medium alone. Levels of cytokines produced in culture supernatants were measured using enzyme-linked immunosorbent assay kits. ST-derived inflammatory cells were transferred into the joints of immunodeficient mice to explore whether these cells could develop pannus. CD14 and CD2 cells were depleted by negative selection. Culture of ST-derived inflammatory cells from 92 of 111 patients with RA resulted in spontaneous reconstruction of inflammatory tissue in vitro within 4 weeks. Ex vivo tissue contained fibroblasts, macrophages, T cells, and tartrate-resistant acid phosphatase-positive multinucleated cells. On calcium phosphate-coated slides, ST-derived inflammatory cell cultures showed numerous resorption pits. ST-derived inflammatory cell cultures continuously produced matrix metalloproteinase 9 and proinflammatory cytokines associated with osteoclastogenesis, such as tumor necrosis factor alpha, interleukin-8, and macrophage colony-stimulating factor. More importantly, transferring ST-derived inflammatory cells into the joints of immunodeficient mice resulted in the development of pannus tissue and erosive joint lesions. Both in vitro development and in vivo development of pannus tissue by ST-derived inflammatory cells were inhibited by depleting CD14-positive, but not CD2-positive, cells from ST-derived inflammatory cells. These findings suggest that overgrowth of inflammatory cells from human rheumatoid synovium simulates the development of pannus. This may prove informative in the screening of potential antirheumatic drugs.

Increased expression of connective tissue growth factor (CTGF) in multiple organs after exposure of non-human primates (NHP) to lethal doses of radiation

PubMed Central

Zhang, Pei; Cui, Wanchang; Hankey, Kim G.; Gibbs, Allison M.; Smith, Cassandra P.; Taylor-Howell, Cheryl; Kearney, Sean R.; MacVittie, Thomas J.

2015-01-01

Exposure to sufficiently high doses of ionizing radiation is known to cause fibrosis in many different organs and tissues. Connective tissue growth factor (CTGF/CCN2), a member of the CCN family of matricellular proteins, plays an important role in the development of fibrosis in multiple organs. The aim of the present study was to quantify the gene and protein expression of CTGF in a variety of organs from non-human primates (NHP) that were previously exposed to potentially lethal doses of radiation. Tissues from non-irradiated NHP, and NHP exposed to whole thoracic lung irradiation (WTLI) or partial-body irradiation with 5% bone marrow sparing (PBI/BM5) were examined by real-time quantitative reverse transcription PCR, western blot, and immunohistochemistry. Expression of CTGF was elevated in the lung tissues of NHP exposed to WTLI relative to the lung tissues of the non-irradiated NHP. Increased expression of CTGF was also observed in multiple organs from NHP exposed to PBI/BM5 compared to non-irradiated NHP; these included the lung, kidney, spleen, thymus and liver. These irradiated organs also exhibited histological evidence of increased collagen deposition compared to the control tissues. There was significant correlation of CTGF expression with collagen deposition in the lung and spleen of NHP exposed to PBI/BM5. Significant correlations were observed between spleen and multiple organs on CTGF expression and collagen deposition respectively, suggesting possible crosstalk between spleen and other organs. Our data suggest that CTGF levels are increased in multiple organs after radiation exposure and that inflammatory cell infiltration may contribute to the elevated levels of CTGF in multiple organs. PMID:26425899

Connective tissue growth factor immunohistochemical expression is associated with gallbladder cancer progression.

PubMed

Garcia, Patricia; Leal, Pamela; Alvarez, Hector; Brebi, Priscilla; Ili, Carmen; Tapia, Oscar; Roa, Juan C

2013-02-01

Gallbladder cancer (GBC) is an aggressive neoplasia associated with late diagnosis, unsatisfactory treatment, and poor prognosis. Molecular mechanisms involved in GBC pathogenesis remain poorly understood. Connective tissue growth factor (CTGF) is thought to play a role in the pathologic processes and is overexpressed in several human cancers, including GBC. No information is available about CTGF expression in early stages of gallbladder carcinogenesis. Objective.- To evaluate the expression level of CTGF in benign and malignant lesions of gallbladder and its correlation with clinicopathologic features and GBC prognosis. Connective tissue growth factor protein was examined by immunohistochemistry on tissue microarrays containing tissue samples of chronic cholecystitis (n = 51), dysplasia (n = 15), and GBC (n = 169). The samples were scored according to intensity of staining as low/absent and high CTGF expressers. Statistical analysis was performed using the χ(2) test or Fisher exact probability test with a significance level of P < .05. Survival analysis was assessed by the Kaplan-Meier method and the log-rank test. Connective tissue growth factor expression showed a progressive increase from chronic cholecystitis to dysplasia and then to early and advanced carcinoma. Immunohistochemical expression (score ≥2) was significantly higher in advanced tumors, in comparison with chronic cholecystitis (P < .001) and dysplasia (P = .03). High levels of CTGF expression correlated with better survival (P = .04). Our results suggest a role for CTGF in GBC progression and a positive association with better prognosis. In addition, they underscore the importance of considering the involvement of inflammation on GBC development.

Therapeutic suppression of translation initiation factor eIF4E expression reduces tumor growth without toxicity

PubMed Central

Graff, Jeremy R.; Konicek, Bruce W.; Vincent, Thomas M.; Lynch, Rebecca L.; Monteith, David; Weir, Spring N.; Schwier, Phil; Capen, Andrew; Goode, Robin L.; Dowless, Michele S.; Chen, Yuefeng; Zhang, Hong; Sissons, Sean; Cox, Karen; McNulty, Ann M.; Parsons, Stephen H.; Wang, Tao; Sams, Lillian; Geeganage, Sandaruwan; Douglass, Larry E.; Neubauer, Blake Lee; Dean, Nicholas M.; Blanchard, Kerry; Shou, Jianyong; Stancato, Louis F.; Carter, Julia H.; Marcusson, Eric G.

2007-01-01

Expression of eukaryotic translation initiation factor 4E (eIF4E) is commonly elevated in human and experimental cancers, promoting angiogenesis and tumor growth. Elevated eIF4E levels selectively increase translation of growth factors important in malignancy (e.g., VEGF, cyclin D1) and is thereby an attractive anticancer therapeutic target. Yet to date, no eIF4E-specific therapy has been developed. Herein we report development of eIF4E-specific antisense oligonucleotides (ASOs) designed to have the necessary tissue stability and nuclease resistance required for systemic anticancer therapy. In mammalian cultured cells, these ASOs specifically targeted the eIF4E mRNA for destruction, repressing expression of eIF4E-regulated proteins (e.g., VEGF, cyclin D1, survivin, c-myc, Bcl-2), inducing apoptosis, and preventing endothelial cells from forming vessel-like structures. Most importantly, intravenous ASO administration selectively and significantly reduced eIF4E expression in human tumor xenografts, significantly suppressing tumor growth. Because these ASOs also target murine eIF4E, we assessed the impact of eIF4E reduction in normal tissues. Despite reducing eIF4E levels by 80% in mouse liver, eIF4E-specific ASO administration did not affect body weight, organ weight, or liver transaminase levels, thereby providing the first in vivo evidence that cancers may be more susceptible to eIF4E inhibition than normal tissues. These data have prompted eIF4E-specific ASO clinical trials for the treatment of human cancers. PMID:17786246

LIM Domain Only 2 Regulates Endothelial Proliferation, Angiogenesis, and Tissue Regeneration.

PubMed

Meng, Shu; Matrone, Gianfranco; Lv, Jie; Chen, Kaifu; Wong, Wing Tak; Cooke, John P

2016-10-06

LIM domain only 2 (LMO2, human gene) is a key transcription factor that regulates hematopoiesis and vascular development. However, its role in adult endothelial function has been incompletely characterized. In vitro loss- and gain-of-function studies on LMO2 were performed in human umbilical vein endothelial cells with lentiviral overexpression or short hairpin RNA knockdown (KD) of LMO2, respectively. LMO2 KD significantly impaired endothelial proliferation. LMO2 controls endothelial G1/S transition through transcriptional regulation of cyclin-dependent kinase 2 and 4 as determined by reverse transcription polymerase chain reaction (PCR), western blot, and chromatin immunoprecipitation, and also influences the expression of Cyclin D1 and Cyclin A1. LMO2 KD also impaired angiogenesis by reducing transforming growth factor-β (TGF-β) expression, whereas supplementation of exogenous TGF-β restored defective network formation in LMO2 KD human umbilical vein endothelial cells. In a zebrafish model of caudal fin regeneration, RT-PCR revealed that the lmo2 (zebrafish gene) gene was upregulated at day 5 postresection. The KD of lmo2 by vivo-morpholino injections in adult Tg(fli1:egfp) y1 zebrafish reduced 5-bromo-2'-deoxyuridine incorporation in endothelial cells, impaired neoangiogenesis in the resected caudal fin, and substantially delayed fin regeneration. The transcriptional factor LMO2 regulates endothelial proliferation and angiogenesis in vitro. Furthermore, LMO2 is required for angiogenesis and tissue healing in vivo. Thus, LMO2 is a critical determinant of vascular and tissue regeneration. © 2016 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell.

Impact of polyunsaturated and saturated fat overfeeding on the DNA-methylation pattern in human adipose tissue: a randomized controlled trial.

PubMed

Perfilyev, Alexander; Dahlman, Ingrid; Gillberg, Linn; Rosqvist, Fredrik; Iggman, David; Volkov, Petr; Nilsson, Emma; Risérus, Ulf; Ling, Charlotte

2017-04-01

Background: Dietary fat composition can affect ectopic lipid accumulation and, thereby, insulin resistance. Diets that are high in saturated fatty acids (SFAs) or polyunsaturated fatty acids (PUFAs) have different metabolic responses. Objective: We investigated whether the epigenome of human adipose tissue is affected differently by dietary fat composition and general overfeeding in a randomized trial. Design: We studied the effects of 7 wk of excessive SFA ( n = 17) or PUFA ( n = 14) intake (+750 kcal/d) on the DNA methylation of ∼450,000 sites in human subcutaneous adipose tissue. Both diets resulted in similar body weight increases. We also combined the data from the 2 groups to examine the overall effect of overfeeding on the DNA methylation in adipose tissue. Results: The DNA methylation of 4875 Cytosine-phosphate-guanine (CpG) sites was affected differently between the 2 diets. Furthermore, both the SFA and PUFA diets increased the mean degree of DNA methylation in adipose tissue, particularly in promoter regions. However, although the mean methylation was changed in 1797 genes [e.g., alpha-ketoglutarate dependent dioxygenase ( FTO ), interleukin 6 ( IL6 ), insulin receptor ( INSR ), neuronal growth regulator 1 ( NEGR1 ), and proopiomelanocortin ( POMC )] by PUFAs, only 125 genes [e.g., adiponectin, C1Q and collagen domain containing ( ADIPOQ )] were changed by SFA overfeeding. In addition, the SFA diet significantly altered the expression of 28 transcripts [e.g., acyl-CoA oxidase 1 ( ACOX1 ) and FAT atypical cadherin 1 ( FAT1 )], whereas the PUFA diet did not significantly affect gene expression. When the data from the 2 diet groups were combined, the mean methylation of 1444 genes, including fatty acid binding protein 1 ( FABP1 ), fatty acid binding protein 2 ( FABP2 ), melanocortin 2 receptor ( MC2R ), MC3R , PPARG coactivator 1 α ( PPARGC1A ), and tumor necrosis factor ( TNF ), was changed in adipose tissue by overfeeding. Moreover, the baseline DNA methylation of 12 CpG sites that was annotated to 9 genes [e.g., mitogen-activated protein kinase 7 ( MAPK7 ), melanin concentrating hormone receptor 1 ( MCHR1 ), and splicing factor SWAP homolog ( SFRS8 )] was associated with the degree of weight increase in response to extra energy intake. Conclusions: SFA overfeeding and PUFA overfeeding induce distinct epigenetic changes in human adipose tissue. In addition, we present data that suggest that baseline DNA methylation can predict weight increase in response to overfeeding in humans. This trial was registered at clinicaltrials.gov as NCT01427140. © 2017 American Society for Nutrition.

Overexpression of stromal cell-derived factor 1 and its receptor CXCR4 induces autocrine/paracrine cell proliferation in human pituitary adenomas.

PubMed

Barbieri, Federica; Bajetto, Adriana; Stumm, Ralf; Pattarozzi, Alessandra; Porcile, Carola; Zona, Gianluigi; Dorcaratto, Alessandra; Ravetti, Jean-Louis; Minuto, Francesco; Spaziante, Renato; Schettini, Gennaro; Ferone, Diego; Florio, Tullio

2008-08-15

Hypothalamic or locally produced growth factors and cytokines control pituitary development, functioning, and cell division. We evaluated the expression of the chemokine stromal cell-derived factor 1 (SDF1) and its receptor CXCR4 in human pituitary adenomas and normal pituitary tissues and their role in cell proliferation. The expression of SDF1 and CXCR4 in 65 human pituitary adenomas and 4 human normal pituitaries was determined by reverse transcription-PCR, immunohistochemistry, and confocal immunofluorescence. The proliferative effect of SDF1 was evaluated in eight fibroblast-free human pituitary adenoma cell cultures. CXCR4 mRNA was expressed in 92% of growth hormone (GH)-secreting pituitary adenomas (GHoma) and 81% of nonfunctioning pituitary adenomas (NFPA), whereas SDF1 was identified in 63% and 78% of GHomas and NFPAs, respectively. Immunostaining for CXCR4 and SDF1 showed a strong homogenous labeling in all tumoral cells in both GHomas and NFPAs. In normal tissues, CXCR4 and SDF1 were expressed only in a subset of anterior pituitary cells, with a lower expression of SDF1 compared with its cognate receptor. CXCR4 and SDF1 were not confined to a specific cell population in the anterior pituitary but colocalized with discrete subpopulations of GH-, prolactin-, and adrenocorticorticotropic hormone-secreting cells. Conversely, most of the SDF1-containing cells expressed CXCR4. In six of eight pituitary adenoma primary cultures, SDF1 induced a statistically significant increase in DNA synthesis that was prevented by the treatment with the CXCR4 antagonist AMD3100 or somatostatin. CXCR4 and SDF1 are overexpressed in human pituitary adenomas and CXCR4 activation may contribute to pituitary cell proliferation and, possibly, to adenoma development in humans.

Human hepatocyte growth factor promotes functional recovery in primates after spinal cord injury.

PubMed

Kitamura, Kazuya; Fujiyoshi, Kanehiro; Yamane, Jun-Ichi; Toyota, Fumika; Hikishima, Keigo; Nomura, Tatsuji; Funakoshi, Hiroshi; Nakamura, Toshikazu; Aoki, Masashi; Toyama, Yoshiaki; Okano, Hideyuki; Nakamura, Masaya

2011-01-01

Many therapeutic interventions for spinal cord injury (SCI) using neurotrophic factors have focused on reducing the area damaged by secondary, post-injury degeneration, to promote functional recovery. Hepatocyte growth factor (HGF), which is a potent mitogen for mature hepatocytes and a mediator of the inflammatory responses to tissue injury, was recently highlighted as a potent neurotrophic factor in the central nervous system. We previously reported that introducing exogenous HGF into the injured rodent spinal cord using a herpes simplex virus-1 vector significantly reduces the area of damaged tissue and promotes functional recovery. However, that study did not examine the therapeutic effects of administering HGF after injury, which is the most critical issue for clinical application. To translate this strategy to human treatment, we induced a contusive cervical SCI in the common marmoset, a primate, and then administered recombinant human HGF (rhHGF) intrathecally. Motor function was assessed using an original open field scoring system focusing on manual function, including reach-and-grasp performance and hand placement in walking. The intrathecal rhHGF preserved the corticospinal fibers and myelinated areas, thereby promoting functional recovery. In vivo magnetic resonance imaging showed significant preservation of the intact spinal cord parenchyma. rhHGF-treatment did not give rise to an abnormal outgrowth of calcitonin gene related peptide positive fibers compared to the control group, indicating that this treatment did not induce or exacerbate allodynia. This is the first study to report the efficacy of rhHGF for treating SCI in non-human primates. In addition, this is the first presentation of a novel scale for assessing neurological motor performance in non-human primates after contusive cervical SCI.

Keratinocyte growth factor expression in human gingival fibroblasts and stimulation of in vitro gene expression by retinoic acid.

PubMed

Mackenzie, I C; Gao, Z

2001-04-01

Keratinocyte growth factor (KGF) is a stromally derived growth factor of the fibroblast growth factor (FGF) family with paracrine effects targeted to influence the growth and differentiation of epithelia. Regional and temporal changes in KGF expression play important roles in the development and maintenance of epithelial structures and in epithelial wound healing. Differing patterns of expression of KGF by fibroblasts in the gingival region could therefore be related to the observed regional variation in the differentiation and behavior of gingival epithelia. The in vitro and in vivo patterns of expression of KGF mRNA in human gingival and periodontal fibroblasts were examined using reverse transcription polymerase chain reactions (RT-PCR) and in situ hybridization with digoxigenin-labeled riboprobes. The patterns observed for human gingiva were compared with those for human skin and for murine tissues. Gingival and periodontal fibroblasts showed expression of KGF transcripts in vitro, and the degree of expression was markedly influenced by the presence of retinoic acid, an agent known to influence patterns of epithelial differentiation. Sections of human and murine gingiva and skin showed regionally variable expression of transcripts with the cells expressing KGF in the subepithelial, rather than the deeper, connective tissues and periodontium. The results point to a role of KGF in the maintenance of normal growth and differentiation of gingival epithelia. A lack of KGF expression by periodontal fibroblasts in vivo is expected to hinder apical epithelial migration and thus stabilize the epithelial attachment. The effects of retinoic acid (RA) on KGF expression in vitro provide an indirect mechanism by which RA may regulate the growth and differentiation of gingival epithelia.

Human Hepatocyte Growth Factor Promotes Functional Recovery in Primates after Spinal Cord Injury

PubMed Central

Kitamura, Kazuya; Fujiyoshi, Kanehiro; Yamane, Jun-ichi; Toyota, Fumika; Hikishima, Keigo; Nomura, Tatsuji; Funakoshi, Hiroshi; Nakamura, Toshikazu; Aoki, Masashi; Toyama, Yoshiaki; Okano, Hideyuki; Nakamura, Masaya

2011-01-01

Many therapeutic interventions for spinal cord injury (SCI) using neurotrophic factors have focused on reducing the area damaged by secondary, post-injury degeneration, to promote functional recovery. Hepatocyte growth factor (HGF), which is a potent mitogen for mature hepatocytes and a mediator of the inflammatory responses to tissue injury, was recently highlighted as a potent neurotrophic factor in the central nervous system. We previously reported that introducing exogenous HGF into the injured rodent spinal cord using a herpes simplex virus-1 vector significantly reduces the area of damaged tissue and promotes functional recovery. However, that study did not examine the therapeutic effects of administering HGF after injury, which is the most critical issue for clinical application. To translate this strategy to human treatment, we induced a contusive cervical SCI in the common marmoset, a primate, and then administered recombinant human HGF (rhHGF) intrathecally. Motor function was assessed using an original open field scoring system focusing on manual function, including reach-and-grasp performance and hand placement in walking. The intrathecal rhHGF preserved the corticospinal fibers and myelinated areas, thereby promoting functional recovery. In vivo magnetic resonance imaging showed significant preservation of the intact spinal cord parenchyma. rhHGF-treatment did not give rise to an abnormal outgrowth of calcitonin gene related peptide positive fibers compared to the control group, indicating that this treatment did not induce or exacerbate allodynia. This is the first study to report the efficacy of rhHGF for treating SCI in non-human primates. In addition, this is the first presentation of a novel scale for assessing neurological motor performance in non-human primates after contusive cervical SCI. PMID:22140459

Exposure to transforming growth factor-β1 after basic fibroblast growth factor promotes the fibroblastic differentiation of human periodontal ligament stem/progenitor cell lines.

PubMed

Kono, Kiyomi; Maeda, Hidefumi; Fujii, Shinsuke; Tomokiyo, Atsushi; Yamamoto, Naohide; Wada, Naohisa; Monnouchi, Satoshi; Teramatsu, Yoko; Hamano, Sayuri; Koori, Katsuaki; Akamine, Akifumi

2013-05-01

Basic fibroblast growth factor (bFGF) is a cytokine that promotes the regeneration of the periodontium, the specialized tissues supporting the teeth. bFGF, does not, however, induce the synthesis of smooth muscle actin alpha 2 (ACTA2), type I collagen (COL1), or COL3, which are principal molecules in periodontal ligament (PDL) tissue, a component of the periodontium. We have suggested the feasibility of using transforming growth factor-β1 (TGFβ1) to induce fibroblastic differentiation of PDL stem/progenitor cells (PDLSCs). Here, we investigated the effect of the subsequent application of TGFβ1 after bFGF (bFGF/TGFβ1) on the differentiation of PDLSCs into fibroblastic cells. We first confirmed the expression of bFGF and TGFβ1 in rat PDL tissue and primary human PDL cells. Receptors for both bFGF and TGFβ1 were expressed in the human PDLSC lines 1-11 and 1-17. Exposure to bFGF for 2 days promoted vascular endothelial growth factor gene and protein expression in both cell lines and down-regulated the expression of ACTA2, COL1, and COL3 mRNA in both cell lines and the gene fibrillin 1 (FBN1) in cell line 1-11 alone. Furthermore, bFGF stimulated cell proliferation of these cell lines and significantly increased the number of cells in phase G2/M in the cell lines. Exposure to TGFβ1 for 2 days induced gene expression of ACTA2 and COL1 in both cell lines and FBN1 in cell line 1-11 alone. BFGF/TGFβ1 treatment significantly up-regulated ACTA2, COL1, and FBN1 expression as compared with the group treated with bFGF alone or the untreated control. This method might thus be useful for accelerating the generation and regeneration of functional periodontium.

Barley as a green factory for the production of functional Flt3 ligand.

PubMed

Erlendsson, Lýdur S; Muench, Marcus O; Hellman, Ulf; Hrafnkelsdóttir, Soffía M; Jonsson, Anders; Balmer, Yves; Mäntylä, Einar; Orvar, Björn L

2010-02-01

Biologically active recombinant human Flt3 ligand was expressed and isolated from transgenic barley seeds. Its expression is controlled by a tissue specific promoter that confines accumulation of the recombinant protein to the endosperm tissue of the seed. The recombinant Flt3 ligand variant expressed in the seeds contains an HQ-tag for affinity purification on immobilized metal ion affinity chromatography (IMAC) resin. The tagged protein was purified from seed extracts to near homogeneity using sequential chromatography on IMAC affinity resin and cation exchange resin. We also show that the recombinant Flt3 ligand protein undergoes posttranslational modifications: it is a glycoprotein containing alpha-1,3-fucose and alpha-1,2-xylose. The HQ-tagged Flt3 ligand variant exhibits comparable biological activity to commercial Flt3 ligand. This is the first report showing expression and accumulation of recombinant human growth factor in barley seeds with a yield of active protein similar to a bacterial expression system. The present results demonstrate that plant molecular farming is a viable approach for the bioproduction of human-derived growth factors.

The significance of folic acid, tissue iron stores, and tissue viability in determining iron uptake from serum by thyroid tissue slices

PubMed Central

Buchanan, W. M.

1971-01-01

This paper describes an attempt to measure in vitro iron uptake from serum by human thyroid slices and to relate the uptake to tissue iron stores, folic acid status, and tissue viability. It is an extension of work previously reported (Buchanan, 1969). Thyroids were obtained from patients undergoing partial thyroidectomy for colloid goitre and serum from clinically normal healthy adults. The haemoglobin, serum iron, and folic acid levels of both thyroid and serum donors were measured and thyroids examined histologically for the presence of stainable iron. Viable and non-viable tissue slices were incubated in sera treated with radioactive iron so as to produce high and normal levels of transferrin saturation. Iron was taken up both from sera with normal and high transferrin saturation but the amount was, in almost all cases, greater from the more highly saturated. The uptake by non-viable tissue was appreciable but did not vary to any great extent from one serum to the next, and was attributed to simple diffusion of ionic iron into the tissue. There was, however, marked variation in uptake from different sera by viable tissue. It was concluded therefore that viability is a factor affecting the uptake. As the variation in uptake by viable tissue incubated in a single serum was significantly less than tissue incubated in a number of different sera it was further concluded that there was also a factor in the serum itself affecting iron uptake. The nature of the factor was not elucidated but neither folic acid nor levels of iron stores appeared to influence uptake because no correlation was found between iron uptake and iron stores or folic acid. Images PMID:5556118

The Broad Spectrum of Human Natural Killer Cell Diversity.

PubMed

Freud, Aharon G; Mundy-Bosse, Bethany L; Yu, Jianhua; Caligiuri, Michael A

2017-11-21

Natural killer (NK) cells provide protection against infectious pathogens and cancer. For decades it has been appreciated that two major NK cell subsets (CD56 bright and CD56 dim ) exist in humans and have distinct anatomical localization patterns, phenotypes, and functions in immunity. In light of this traditional NK cell dichotomy, it is now clear that the spectrum of human NK cell diversity is much broader than originally appreciated as a result of variegated surface receptor, intracellular signaling molecule, and transcription factor expression; tissue-specific imprinting; and foreign antigen exposure. The recent discoveries of tissue-resident NK cell developmental intermediates, non-NK innate lymphoid cells, and the capacity for NK cells to adapt and differentiate into long-lived memory cells has added further complexity to this field. Here we review our current understanding of the breadth and generation of human NK cell diversity. Copyright © 2017 Elsevier Inc. All rights reserved.

microRNA-497 overexpression decreases proliferation, migration and invasion of human retinoblastoma cells via targeting vascular endothelial growth factor A

PubMed Central

Li, Jianjun; Zhang, Yinghui; Wang, Xiuchao; Zhao, Ruibo

2017-01-01

The expression level and roles of microRNA-497 (miR-497) have been frequently reported in previous studies on cancer. However, its expression, function and associated molecular mechanisms in retinoblastoma remain unknown. In the present study, miR-497 expression levels in human retinoblastoma tissues, normal retinal tissues and retinoblastoma cell lines were determined using reverse transcription-quantitative polymerase chain reaction. In addition, a Cell Counting Kit-8 assay, cell migration assay, cell invasion assay, western blot analysis and Dual-Luciferase reporter assay were used to explore the expression, functions and molecular mechanisms of miR-497 in human retinoblastoma. It was demonstrated that miR-497 was significantly downregulated in retinoblastoma tissues and cell lines compared with normal retinal tissues. Ectopic expression of miR-497 decreased the proliferation, migration and invasion of retinoblastoma cells. Furthermore, VEGFA was verified as a potential direct target of miR-497 in vitro. Taken together, the results indicate that miR-497 functions as a tumor suppressor in the carcinogenesis and progression of retinoblastoma via targeting VEGFA. miR-497 should be investigated as a potential therapeutic target for the treatment of retinoblastoma. PMID:28588740

Palatogenesis

PubMed Central

Levi, Benjamin; Brugman, Samantha; Wong, Victor W; Grova, Monica; Longaker, Michael T

2011-01-01

Cleft palate represents the second most common birth defect and carries substantial physiologic and social challenges for affected patients, as they often require multiple surgical interventions during their lifetime. A number of genes have been identified to be associated with the cleft palate phenotype, but etiology in the majority of cases remains elusive. In order to better understand cleft palate and both surgical and potential tissue engineering approaches for repair, we have performed an in-depth literature review into cleft palate development in humans and mice, as well as into molecular pathways underlying these pathologic developments. We summarize the multitude of pathways underlying cleft palate development, with the transforming growth factor β superfamily being the most commonly studied. Furthermore, while the majority of cleft palate studies are performed using a mouse model, studies focusing on tissue engineering have also focused heavily on mouse models. A paucity of human randomized controlled studies exists for cleft palate repair, and so far, tissue engineering approaches are limited. In this review, we discuss the development of the palate, explain the basic science behind normal and pathologic palate development in humans as well as mouse models and elaborate on how these studies may lead to future advances in palatal tissue engineering and cleft palate treatments. PMID:21964245

Platelet-Rich Plasma

PubMed Central

Cole, Brian J.; Seroyer, Shane T.; Filardo, Giuseppe; Bajaj, Sarvottam; Fortier, Lisa A.

2010-01-01

Context: Platelet-rich plasma (PRP) may affect soft tissue healing via growth factors released after platelet degranulation. Because of this potential benefit, clinicians have begun to inject PRP for the treatment of tendon, ligament, muscle, and cartilage injuries and early osteoarthritis. Evidence Acquisition: A PubMed search was performed for studies relating to PRP, growth factors, and soft tissue injuries from 1990 to 2010. Relevant references from these studies were also retrieved. Results: Soft tissue injury is a major source of disability that may often be complicated by prolonged and incomplete recovery. Numerous growth factors may potentiate the healing and regeneration of tendons and ligaments. The potential benefits of biologically enhanced healing processes have led to a recent interest in the use of PRP in orthopaedic sports medicine. There has been widespread anecdotal use of PRP for muscle strains, tendinopathy, and ligament injuries and as a surgical adjuvant to rotator cuff repair, anterior cruciate ligament reconstruction, and meniscal or labral repairs. Although the fascination with this emerging technology has led to a dramatic increase in its use, scientific data supporting this use are still in their infancy. Conclusions: The literature is replete with studies on the basic science of growth factors and their relation to the maintenance, proliferation, and regeneration of various tissues and tissue-derived cells. Despite the promising results of several animal studies, well-controlled human studies are lacking. PMID:23015939

Field and laboratory fish tissue accumulation of the anti-convulsant drug carbamazepine.

PubMed

Garcia, Santos N; Foster, Michael; Constantine, Lisa A; Huggett, Duane B

2012-10-01

Understanding the potential for human and veterinary pharmaceuticals to accumulate in the tissues of biota is a topic of increasing importance in the pharmaceutical risk assessment process. However, few data are available in the literature that compare the ability of laboratory bioconcentration studies to predict field tissue concentrations. To begin to address this data gap, bioconcentration factors (BCF) for carbamazepine (CBZ), a human anticonvulsant that modulates Na+ channels, were determined using laboratory experiments with Pimephales notatus and Ictalurus punctatus. These data were compared to field derived bioaccumulation factors (BAFs) for Oreochromis niloticus from the Denton, Texas Wastewater Treatment Plant. The 42 d kinetic BCFs (BCFk) for white muscle and liver of P. notatus were 1.9 and 4.6, respectively, while the white muscle, liver, brain, and plasma BCFk's of I. punctatus were 1.8, 1.5, 1.6, and 7.1, respectively. Field derived BAF values (2.5-3.8) for O. niloticus were similar to those derived in laboratory studies. Partitioning values between blood plasma and individual tissues were calculated for I. punctatus and O. niloticus, with the values indicating that tissue levels of carbamazepine are similar or slightly higher than plasma concentrations. Collectively these data suggest that the fish laboratory BCF and field derived BCF/BAF values for carbamazepine are similar and much lower than the European Union regulatory threshold of 2000 for designation of a "B" substance. Copyright © 2012 Elsevier Inc. All rights reserved.

Detection of hydroxyapatite in calcified cardiovascular tissues.

PubMed

Lee, Jae Sam; Morrisett, Joel D; Tung, Ching-Hsuan

2012-10-01

The objective of this study is to develop a method for selective detection of the calcific (hydroxyapatite) component in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues ex vivo. This method uses a novel optical molecular imaging contrast dye, Cy-HABP-19, to target calcified cells and tissues. A peptide that mimics the binding affinity of osteocalcin was used to label hydroxyapatite in vitro and ex vivo. Morphological changes in vascular smooth muscle cells were evaluated at an early stage of the mineralization process induced by extrinsic stimuli, osteogenic factors and a magnetic suspension cell culture. Hydroxyapatite components were detected in monolayers of these cells in the presence of osteogenic factors and a magnetic suspension environment. Atherosclerotic plaque contains multiple components including lipidic, fibrotic, thrombotic, and calcific materials. Using optical imaging and the Cy-HABP-19 molecular imaging probe, we demonstrated that hydroxyapatite components could be selectively distinguished from various calcium salts in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues, carotid endarterectomy samples and aortic valves, ex vivo. Hydroxyapatite deposits in cardiovascular tissues were selectively detected in the early stage of the calcification process using our Cy-HABP-19 probe. This new probe makes it possible to study the earliest events associated with vascular hydroxyapatite deposition at the cellular and molecular levels. This target-selective molecular imaging probe approach holds high potential for revealing early pathophysiological changes, leading to progression, regression, or stabilization of cardiovascular diseases. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Detection of Hydroxyapatite in Calcified Cardiovascular Tissues

PubMed Central

Lee, Jae Sam; Morrisett, Joel D.; Tung, Ching-Hsuan

2012-01-01

Objective The objective of this study is to develop a method for selective detection of the calcific (hydroxyapatite) component in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues ex vivo. This method uses a novel optical molecular imaging contrast dye, Cy-HABP-19, to target calcified cells and tissues. Methods A peptide that mimics the binding affinity of osteocalcin was used to label hydroxyapatite in vitro and ex vivo. Morphological changes in vascular smooth muscle cells were evaluated at an early stage of the mineralization process induced by extrinsic stimuli, osteogenic factors and a magnetic suspension cell culture. Hydroxyapatite components were detected in monolayers of these cells in the presence of osteogenic factors and a magnetic suspension environment. Results Atherosclerotic plaque contains multiple components including lipidic, fibrotic, thrombotic, and calcific materials. Using optical imaging and the Cy-HABP-19 molecular imaging probe, we demonstrated that hydroxyapatite components could be selectively distinguished from various calcium salts in human aortic smooth muscle cells in vitro and in calcified cardiovascular tissues, carotid endarterectomy samples and aortic valves, ex vivo. Conclusion Hydroxyapatite deposits in cardiovascular tissues were selectively detected in the early stage of the calcification process using our Cy-HABP-19 probe. This new probe makes it possible to study the earliest events associated with vascular hydroxyapatite deposition at the cellular and molecular levels. This target-selective molecular imaging probe approach holds high potential for revealing early pathophysiological changes, leading to progression, regression, or stabilization of cardiovascular diseases. PMID:22877867

Human cells and cell cultures: availability, authentication and future prospects.

PubMed

Hay, R J

1996-09-01

The availability of well characterized, viable human cells, tissues and cell lines along with pertinent data on the specific patient donors is a prerequisite for much current transplantation and biomedical research. In the USA, institutional and multi-center networks have been established for provision of primary human cells and tissues to qualified clinicians and research scientists. Monetary support derives from government, university, institutional and fee sources. Problems involved include concern for the rights and privacy of tissue donors, cultural reservations relating to tissue provision, the need for safe and expeditious transport, short term survival and limited supply, adequate correlation of patient data with samples provided, presence of infectious viruses and microorganisms, as well as state or government regulations regarding national or international shipping. The use of human cell lines with continuous or even somewhat limited doubling potentials overcomes many of the above difficulties. National cell banks have been established to provide reference lines for use by multiple investigators. Use of such cell lines assures improved research comparability both geographically and with time. Authentication procedures are critically important for all of these programs. Verification of tissue types and conditions is required through histological, biochemical and immunological assays. Tests for microbial and viral contaminants must be applied. In addition to such procedures utilized for tissues, with cell lines the banking agency must also verify species and where possible identity, properties and functions. The literature is replete with descriptions documenting incorrect identifications and infections of proliferating cell strains used for research. The availability of viable tissue through local sources and distribution agencies in the USA is becoming more commonplace even including full family participation and collection of related, detailed histories. Increased support for this developmental activity is needed, coupled with provision of blood and normal cells and cell lines from family members in many disease categories. Modern techniques, new and improved culture ware, serum-free media, reagents such as growth, adherence and transfer factors will permit isolation, propagation and wide spread distribution not only of human tumor cells but also normal and functional human cells of most renewing and expanding tissue types. Hybridization and immortalization techniques are enhancing this capability such that virtually all human cell types should be available for short or longer-term propagation and study in the foreseeable future.

An independent component analysis confounding factor correction framework for identifying broad impact expression quantitative trait loci

PubMed Central

Ju, Jin Hyun; Crystal, Ronald G.

2017-01-01

Genome-wide expression Quantitative Trait Loci (eQTL) studies in humans have provided numerous insights into the genetics of both gene expression and complex diseases. While the majority of eQTL identified in genome-wide analyses impact a single gene, eQTL that impact many genes are particularly valuable for network modeling and disease analysis. To enable the identification of such broad impact eQTL, we introduce CONFETI: Confounding Factor Estimation Through Independent component analysis. CONFETI is designed to address two conflicting issues when searching for broad impact eQTL: the need to account for non-genetic confounding factors that can lower the power of the analysis or produce broad impact eQTL false positives, and the tendency of methods that account for confounding factors to model broad impact eQTL as non-genetic variation. The key advance of the CONFETI framework is the use of Independent Component Analysis (ICA) to identify variation likely caused by broad impact eQTL when constructing the sample covariance matrix used for the random effect in a mixed model. We show that CONFETI has better performance than other mixed model confounding factor methods when considering broad impact eQTL recovery from synthetic data. We also used the CONFETI framework and these same confounding factor methods to identify eQTL that replicate between matched twin pair datasets in the Multiple Tissue Human Expression Resource (MuTHER), the Depression Genes Networks study (DGN), the Netherlands Study of Depression and Anxiety (NESDA), and multiple tissue types in the Genotype-Tissue Expression (GTEx) consortium. These analyses identified both cis-eQTL and trans-eQTL impacting individual genes, and CONFETI had better or comparable performance to other mixed model confounding factor analysis methods when identifying such eQTL. In these analyses, we were able to identify and replicate a few broad impact eQTL although the overall number was small even when applying CONFETI. In light of these results, we discuss the broad impact eQTL that have been previously reported from the analysis of human data and suggest that considerable caution should be exercised when making biological inferences based on these reported eQTL. PMID:28505156

An independent component analysis confounding factor correction framework for identifying broad impact expression quantitative trait loci.

PubMed

Ju, Jin Hyun; Shenoy, Sushila A; Crystal, Ronald G; Mezey, Jason G

2017-05-01

Genome-wide expression Quantitative Trait Loci (eQTL) studies in humans have provided numerous insights into the genetics of both gene expression and complex diseases. While the majority of eQTL identified in genome-wide analyses impact a single gene, eQTL that impact many genes are particularly valuable for network modeling and disease analysis. To enable the identification of such broad impact eQTL, we introduce CONFETI: Confounding Factor Estimation Through Independent component analysis. CONFETI is designed to address two conflicting issues when searching for broad impact eQTL: the need to account for non-genetic confounding factors that can lower the power of the analysis or produce broad impact eQTL false positives, and the tendency of methods that account for confounding factors to model broad impact eQTL as non-genetic variation. The key advance of the CONFETI framework is the use of Independent Component Analysis (ICA) to identify variation likely caused by broad impact eQTL when constructing the sample covariance matrix used for the random effect in a mixed model. We show that CONFETI has better performance than other mixed model confounding factor methods when considering broad impact eQTL recovery from synthetic data. We also used the CONFETI framework and these same confounding factor methods to identify eQTL that replicate between matched twin pair datasets in the Multiple Tissue Human Expression Resource (MuTHER), the Depression Genes Networks study (DGN), the Netherlands Study of Depression and Anxiety (NESDA), and multiple tissue types in the Genotype-Tissue Expression (GTEx) consortium. These analyses identified both cis-eQTL and trans-eQTL impacting individual genes, and CONFETI had better or comparable performance to other mixed model confounding factor analysis methods when identifying such eQTL. In these analyses, we were able to identify and replicate a few broad impact eQTL although the overall number was small even when applying CONFETI. In light of these results, we discuss the broad impact eQTL that have been previously reported from the analysis of human data and suggest that considerable caution should be exercised when making biological inferences based on these reported eQTL.

Haematopoietic stem and progenitor cells from human pluripotent stem cells

PubMed Central

Sugimura, Ryohichi; Jha, Deepak Kumar; Han, Areum; Soria-Valles, Clara; da Rocha, Edroaldo Lummertz; Lu, Yi-Fen; Goettel, Jeremy A.; Serrao, Erik; Rowe, R. Grant; Malleshaiah, Mohan; Wong, Irene; Sousa, Patricia; Zhu, Ted N.; Ditadi, Andrea; Keller, Gordon; Engelman, Alan N.; Snapper, Scott B.; Doulatov, Sergei; Daley, George Q.

2018-01-01

A variety of tissue lineages can be differentiated from pluripotent stem cells by mimicking embryonic development through stepwise exposure to morphogens, or by conversion of one differentiated cell type into another by enforced expression of master transcription factors. Here, to yield functional human haematopoietic stem cells, we perform morphogen-directed differentiation of human pluripotent stem cells into haemogenic endothelium followed by screening of 26 candidate haematopoietic stem-cell-specifying transcription factors for their capacity to promote multi-lineage haematopoietic engraftment in mouse hosts. We recover seven transcription factors (ERG, HOXA5, HOXA9, HOXA10, LCOR, RUNX1 and SPI1) that are sufficient to convert haemogenic endothelium into haematopoietic stem and progenitor cells that engraft myeloid, B and T cells in primary and secondary mouse recipients. Our combined approach of morphogen-driven differentiation and transcription-factor-mediated cell fate conversion produces haematopoietic stem and progenitor cells from pluripotent stem cells and holds promise for modelling haematopoietic disease in humanized mice and for therapeutic strategies in genetic blood disorders. PMID:28514439

Neuroinflammatory Mechanisms of Connective Tissue Fibrosis: Targeting Neurogenic and Mast Cell Contributions

PubMed Central

Monument, Michael J.; Hart, David A.; Salo, Paul T.; Befus, A. Dean; Hildebrand, Kevin A.

2015-01-01

Significance: The pathogenesis of fibrogenic wound and connective tissue healing is complex and incompletely understood. Common observations across a vast array of human and animal models of fibroproliferative conditions suggest neuroinflammatory mechanisms are important upstream fibrogenic events. Recent Advances: As detailed in this review, mast cell hyperplasia is a common observation in fibrotic tissue. Recent investigations in human and preclinical models of hypertrophic wound healing and post-traumatic joint fibrosis provides evidence that fibrogenesis is governed by a maladaptive neuropeptide-mast cell-myofibroblast signaling pathway. Critical Issues: The blockade and manipulation of these factors is providing promising evidence that if timed correctly, the fibrogenic process can be appropriately regulated. Clinically, abnormal fibrogenic healing responses are not ubiquitous to all patients and the identification of those at-risk remains an area of priority. Future Directions: Ultimately, an integrated appreciation of the common pathobiology shared by many fibrogenic connective tissue conditions may provide a scientific framework to facilitate the development of novel antifibrotic prevention and treatment strategies. PMID:25785237

Vasohibin 2 promotes epithelial-mesenchymal transition in human breast cancer via activation of transforming growth factor β 1 and hypoxia dependent repression of GATA-binding factor 3.

PubMed

Tu, Min; Li, Zhanjun; Liu, Xian; Lv, Nan; Xi, Chunhua; Lu, Zipeng; Wei, Jishu; Song, Guoxin; Chen, Jianmin; Guo, Feng; Jiang, Kuirong; Wang, Shui; Gao, Wentao; Miao, Yi

2017-03-01

Vasohibin 2 (VASH2) is identified as an angiogenic factor, and has been implicated in tumor angiogenesis, proliferation and epithelial-mesenchymal transition (EMT). To investigate the EMT role of VASH2 in breast cancer, we overexpressed or knocked down expression of VASH2 in human breast cancer cell lines. We observed that VASH2 induced EMT in vitro and in vivo. The transforming growth factor β1 (TGFβ1) pathway was activated by VASH2, and expression of a dominant negative TGFβ type II receptor could block VASH2-mediated EMT. In clinical breast cancer tissues VASH2 positively correlated with TGFβ1 expression, but negatively correlated with E-cadherin (a marker of EMT) expression. Under hypoxic conditions in vitro or in vivo, we found that down-regulation of estrogen receptor 1 (ESR1) in VASH2 overexpressing ESR1 positive cells suppressed E-cadherin. Correlation coefficient analysis indicated that VASH2 and ESR1 expression were negatively correlated in clinical human breast cancer tissues. Further study revealed that a transcription factor of ESR1, GATA-binding factor 3 (GATA3), was down-regulated by VASH2 under hypoxia or in vivo. These findings suggest that VASH2 drives breast cancer cells to undergo EMT by activation of the TGFβ1 pathway and hypoxia dependent repression GATA3-ESR1 pathway, leading to cancer metastasis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Increasing consistency of disease biomarker prediction across datasets.

PubMed

Chikina, Maria D; Sealfon, Stuart C

2014-01-01

Microarray studies with human subjects often have limited sample sizes which hampers the ability to detect reliable biomarkers associated with disease and motivates the need to aggregate data across studies. However, human gene expression measurements may be influenced by many non-random factors such as genetics, sample preparations, and tissue heterogeneity. These factors can contribute to a lack of agreement among related studies, limiting the utility of their aggregation. We show that it is feasible to carry out an automatic correction of individual datasets to reduce the effect of such 'latent variables' (without prior knowledge of the variables) in such a way that datasets addressing the same condition show better agreement once each is corrected. We build our approach on the method of surrogate variable analysis but we demonstrate that the original algorithm is unsuitable for the analysis of human tissue samples that are mixtures of different cell types. We propose a modification to SVA that is crucial to obtaining the improvement in agreement that we observe. We develop our method on a compendium of multiple sclerosis data and verify it on an independent compendium of Parkinson's disease datasets. In both cases, we show that our method is able to improve agreement across varying study designs, platforms, and tissues. This approach has the potential for wide applicability to any field where lack of inter-study agreement has been a concern.

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.

Storage and qualification of viable intact human amniotic graft and technology transfer to a tissue bank.

PubMed

Laurent, Romain; Nallet, Aurélie; Obert, Laurent; Nicod, Laurence; Gindraux, Florelle

2014-06-01

Human amniotic membrane (hAM) is known to have good potential to help the regeneration of tissue. It has been used for over 100 years in many medical disciplines because of its properties, namely a scaffold containing stem cells and growth factors, with low immunogenicity and anti-microbial, anti-inflammatory, anti-fibrotic and analgesic properties. In order to use this "boosted membrane" as an advanced therapeutic medicinal product for bone repair, we aimed to observe the influence of tissue culture and/or cryopreservation on cell viability and tissue structure, and secondly, to adapt to a tissue bank, identify easy processes to store hAM containing viable cells and to verify the quality of the graft before its release for use. To this end, we tested different published culture or cryopreservation storage conditions and cell viability assays. Tissue structure was evaluated by Giemsa staining and was compared to histological analysis. Preliminary results show no dramatic decrease in cell viability in cultured hAM as compared to cryopreserved hAM, but tissue structure alterations were observed with both storage conditions. Histological and immunohistochemical data highlight that tissue damage was associated with significantly modified protein expression, which could lead to a possible loss of differentiation potential. Finally, we report that trypan blue and Giemsa staining could constitute controls that are "materially and easily transferable" to a tissue bank.

Cyclic AMP-dependent modification of gonad-selective TAF(II)105 in a human ovarian granulosa cell line.

PubMed

Wu, Yimin; Lu, Yunzhe; Hu, Yanfen; Li, Rong

2005-11-01

In response to gonadotropins, the elevated level of intracellular-cyclic AMP (cAMP) in ovarian granulosa cells triggers an ordered activation of multiple ovarian genes, which in turn promotes various ovarian functions including folliculogenesis and steroidogenesis. Identification and characterization of transcription factors that control ovarian gene expression are pivotal to the understanding of the molecular basis of the tissue-specific gene regulation programs. The recent discovery of the mouse TATA binding protein (TBP)-associated factor 105 (TAF(II)105) as a gonad-selective transcriptional co-activator strongly suggests that general transcription factors such as TFIID may play a key role in regulating tissue-specific gene expression. Here we show that the human TAF(II)105 protein is preferentially expressed in ovarian granulosa cells. We also identified a novel TAF(II)105 mRNA isoform that results from alternative exon inclusion and is predicted to encode a dominant negative mutant of TAF(II)105. Following stimulation by the adenylyl cyclase activator forskolin, TAF(II)105 in granulosa cells undergoes rapid and transient phosphorylation that is dependent upon protein kinase A (PKA). Thus, our work suggests that pre-mRNA processing and post-translational modification represent two important regulatory steps for the gonad-specific functions of human TAF(II)105. Copyright 2005 Wiley-Liss, Inc.

Epidermal growth factor increases LRF/Pokemon expression in human prostate cancer cells.

PubMed

Aggarwal, Himanshu; Aggarwal, Anshu; Agrawal, Devendra K

2011-10-01

Leukemia/lymphoma related factor/POK erythroid myeloid ontogenic factor (LRF/Pokemon) is a member of the POK family of proteins that promotes oncogenesis in several forms of cancer. Recently, we found higher LRF expression in human breast and prostate carcinomas compared to the corresponding normal tissues. The aim of this study was to examine the regulation of LRF expression in human prostate cells. Epidermal growth factor (EGF) and its receptors mediate several tumorigenic cascades that regulate cell differentiation, proliferation, migration and survival of prostate cancer cells. There was significantly higher level of LRF expression in the nucleus of LNCaP and PC-3 cells than RWPE-1 cells. A significant increase in LRF expression was observed with increasing doses of EGF in more aggressive and androgen-sensitive prostate cancer cells suggesting that EGF signaling pathway is critical in upregulating the expression of LRF/Pokemon to promote oncogenesis. Copyright © 2011 Elsevier Inc. All rights reserved.

Epidermal Growth Factor Increases LRF/Pokemon Expression in Human Prostate Cancer Cells

PubMed Central

Aggarwal, Himanshu; Aggarwal, Anshu; Agrawal, Devendra K.

2011-01-01

Leukemia/lymphoma related factor/POK erythroid myeloid ontogenic factor (LRF/Pokemon) is a member of the POK family of proteins that promotes oncogenesis in several forms of cancer. Recently, we found higher LRF expression in human breast and prostate carcinomas compared to the corresponding normal tissues. The aim of this study was to examine the regulation of LRF expression in human prostate cells. Epidermal growth factor (EGF) and its receptors mediate several tumorigenic cascades that regulate cell differentiation, proliferation, migration and survival of prostate cancer cells. There was significantly higher level of LRF expression in the nucleus of LNCaP and PC-3 cells than RWPE-1 cells. A significant increase in LRF expression was observed with increasing doses of EGF in more aggressive and androgen-sensitive prostate cancer cells suggesting that EGF signaling pathway is critical in upregulating the expression of LRF/Pokemon to promote oncogenesis. PMID:21640721

Class I and II histone deacetylase expression in human chronic periodontitis gingival tissue.

PubMed

Cantley, M D; Dharmapatni, A A S S K; Algate, K; Crotti, T N; Bartold, P M; Haynes, D R

2016-04-01

Histone deacetylase inhibitors (HDACi) are being considered to treat chronic inflammatory diseases at low doses. Currently HDACi that are more specific are being developed to target particular HDACs; therefore, this study aimed to determine levels and distribution of class I and II HDAC in human gingival samples obtained from patients with chronic periodontitis. Gingival biopsies were obtained from patients with and without (mild inflammation, no bone loss) periodontitis. Total RNA was isolated for real-time quantitative polymerase chain reaction to determine expression of HDACs 1-10. Immunohistochemistry was used to determine protein distribution of HDACs 1, 5, 8 and 9. Factor VIII, CD3 and tartrate resistant acid phosphatase (TRAP) were detected in serial sections to identify blood vessels, lymphocytes, pre-osteoclasts and osteoclasts cells respectively. Tumour necrosis factor α (TNF-α) expression was also assessed. mRNA for HDAC 1, 5, 8 and 9 were significantly upregulated in chronic periodontitis gingival tissues compared to non-periodontitis samples (p < 0.05). Significantly higher HDAC 1 protein expression was observed in chronic periodontitis samples (p < 0.05), and was associated with CD3, TRAP and TNF-α-positive cells. HDAC 1, 5, 8 and 9 were expressed strongly by the factor VIII-positive microvasculature in the chronic periodontitis gingival tissues. HDAC 1, 5, 8 and 9 expression was higher in gingival tissues from patients with chronic periodontitis compared to non-periodontitis samples. Results suggest that these HDACs could therefore be targeted with specific acting HDACi. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

The role of granulocyte macrophage-colony stimulating factor in gastrointestinal immunity to salmonellosis.

PubMed

Coon, C; Beagley, K W; Bao, S

2009-08-01

Human Salmonella infection, in particular, typhoid fever is a highly infectious disease that remains a major public health problem causing significant morbidity and mortality. The outcome of these infections depends on the host's immune response, particularly the actions of granulocytes and macrophages. Using a mouse model of human typhoid fever, with Salmonella typhimurium infection of wild type and granulocyte macrophage-colony stimulating factor (GM-CSF) knock out mice we show a delay in the onset of immune-mediated tissue damage in the spleens and livers of GM-CSF(-/-) mice. Furthermore, GM-CSF(-/-) mice have a prolonged sequestration of S. typhimurium in affected tissues despite an increased production of F4/80+ effector cells. Moreover in the absence of GM-CSF, a decrease in pro-inflammatory cytokine expression of tumor necrosis factor-alpha, interleukin-12 (IL-12) and IL-18 was found, which may alter the host's immune response to infection. GM-CSF appears to play an important role in the pathogenesis of Salmonellosis, and may contribute significantly to the development of protective gastrointestinal mucosal immune responses against oral pathogens.

Connective tissue growth factor is a substrate of ADAM28

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

Mochizuki, Satsuki; Tanaka, Rena; Shimoda, Masayuki

2010-11-26

Research highlights: {yields} The hyper-variable region in the cysteine-rich domain of ADAM28 binds to C-terminal domain of CTGF. {yields} ADAM28 cleaves CTGF alone and CTGF in the CTGF/VEGF{sub 165} complex. {yields} CTGF digestion by ADAM28 releases biologically active VEGF{sub 165} from the complex. {yields} ADAM28, CTGF and VEGF{sub 165} are commonly co-expressed by carcinoma cells in human breast carcinoma tissues. {yields} These suggest that ADAM28 promotes VEGF{sub 165}-induced angiogenesis in the breast carcinomas by selective CTGF digestion in the CTGF/VEGF{sub 165} complex. -- Abstract: ADAM28, a member of the ADAM (a disintegrin and metalloproteinase) gene family, is over-expressed by carcinomamore » cells and the expression correlates with carcinoma cell proliferation and progression in human lung and breast carcinomas. However, information about substrates of ADAM28 is limited. We screened interacting molecules of ADAM28 in human lung cDNA library by yeast two-hybrid system and identified connective tissue growth factor (CTGF). Binding of CTGF to proADAM28 was demonstrated by yeast two-hybrid assay and protein binding assay. ADAM28 cleaved CTGF in dose- and time-dependent manners at the Ala{sup 181}-Tyr{sup 182} and Asp{sup 191}-Pro{sup 192} bonds in the hinge region of the molecule. ADAM28 selectively digested CTGF in the complex of CTGF and vascular endothelial growth factor{sub 165} (VEGF{sub 165}), releasing biologically active VEGF{sub 165} from the complex. RT-PCR and immunohistochemical analyses demonstrated that ADAM28, CTGF and VEGF are commonly co-expressed in the breast carcinoma tissues. These data provide the first evidence that CTGF is a novel substrate of ADAM28 and suggest that ADAM28 may promote VEGF{sub 165}-induced angiogenesis in the breast carcinomas by the CTGF digestion in the CTGF/VEGF{sub 165} complex.« less

Pre-Treatment of Human Mesenchymal Stem Cells With Inflammatory Factors or Hypoxia Does Not Influence Migration to Osteoarthritic Cartilage and Synovium.

PubMed

Leijs, Maarten J C; van Buul, Gerben M; Verhaar, Jan A N; Hoogduijn, Martin J; Bos, Pieter K; van Osch, Gerjo J V M

2017-04-01

Mesenchymal stem cells (MSCs) are promising candidates as a cell-based therapy for osteoarthritis (OA), although current results are modest. Pre-treatment of MSCs before application might improve their therapeutic efficacy. Pre-treatment of MSCs with inflammatory factors or hypoxia will improve their migration and adhesion capacities toward OA-affected tissues. Controlled laboratory study. We used real-time polymerase chain reaction to determine the effects of different fetal calf serum (FCS) batches, platelet lysate (PL), hypoxia, inflammatory factors, factors secreted by OA tissues, and OA synovial fluid (SF) on the expression of 12 genes encoding chemokine or adhesion receptors. Migration of MSCs toward factors secreted by OA tissues was studied in vitro, and attachment of injected MSCs was evaluated in vivo in healthy and OA knees of male Wistar rats. Different FCS batches, PL, or hypoxia did not influence the expression of the migration and adhesion receptor genes. Exposure to inflammatory factors altered the expression of CCR1, CCR4, CD44, PDGFRα, and PDGFRβ. MSCs migrated toward factors secreted by OA tissues in vitro. Neither pre-treatment with inflammatory factors nor the presence of OA influenced MSC migration in vitro or adhesion in vivo. Factors secreted by OA tissues increase MSC migration in vitro. In vivo, no difference in MSC adhesion was found between OA and healthy knees. Pre-treatment with inflammatory factors influenced the expression of migration and adhesion receptors of MSCs but not their migration in vitro or adhesion in vivo. To improve the therapeutic capacity of intra-articular injection of MSCs, they need to remain intra-articular for a longer period of time. Pre-treatment of MSCs with hypoxia or inflammatory factors did not increase the migration or adhesion capacity of MSCs and will therefore not likely prolong their intra-articular longevity. Alternative approaches to prolong the intra-articular presence of MSCs should be developed to increase the therapeutic effect of MSCs in OA.

Genetic compendium of 1511 human brains available through the UK Medical Research Council Brain Banks Network Resource.

PubMed

Keogh, Michael J; Wei, Wei; Wilson, Ian; Coxhead, Jon; Ryan, Sarah; Rollinson, Sara; Griffin, Helen; Kurzawa-Akanbi, Marzena; Santibanez-Koref, Mauro; Talbot, Kevin; Turner, Martin R; McKenzie, Chris-Anne; Troakes, Claire; Attems, Johannes; Smith, Colin; Al Sarraj, Safa; Morris, Chris M; Ansorge, Olaf; Pickering-Brown, Stuart; Ironside, James W; Chinnery, Patrick F

2017-01-01

Given the central role of genetic factors in the pathogenesis of common neurodegenerative disorders, it is critical that mechanistic studies in human tissue are interpreted in a genetically enlightened context. To address this, we performed exome sequencing and copy number variant analysis on 1511 frozen human brains with a diagnosis of Alzheimer's disease (AD, n = 289), frontotemporal dementia/amyotrophic lateral sclerosis (FTD/ALS, n = 252), Creutzfeldt-Jakob disease (CJD, n = 239), Parkinson's disease (PD, n = 39), dementia with Lewy bodies (DLB, n = 58), other neurodegenerative, vascular, or neurogenetic disorders (n = 266), and controls with no significant neuropathology (n = 368). Genomic DNA was extracted from brain tissue in all cases before exome sequencing (Illumina Nextera 62 Mb capture) with variants called by FreeBayes; copy number variant (CNV) analysis (Illumina HumanOmniExpress-12 BeadChip); C9orf72 repeat expansion detection; and APOE genotyping. Established or likely pathogenic heterozygous, compound heterozygous, or homozygous variants, together with the C9orf72 hexanucleotide repeat expansions and a copy number gain of APP, were found in 61 brains. In addition to known risk alleles in 349 brains (23.9% of 1461 undergoing exome sequencing), we saw an association between rare variants in GRN and DLB. Rare CNVs were found in <1.5% of brains, including copy number gains of PRPH that were overrepresented in AD. Clinical, pathological, and genetic data are available, enabling the retrieval of specific frozen brains through the UK Medical Research Council Brain Banks Network. This allows direct access to pathological and control human brain tissue based on an individual's genetic architecture, thus enabling the functional validation of known genetic risk factors and potentially pathogenic alleles identified in future studies. © 2017 Keogh et al.; Published by Cold Spring Harbor Laboratory Press.

Regulation of vascular endothelial growth factor-C by tumor necrosis factor-α in the conjunctiva and pterygium.

PubMed

Dong, Yoko; Kase, Satoru; Dong, Zhenyu; Fukuhara, Junichi; Tagawa, Yoshiaki; Ishizuka, Erdal Tan; Murata, Miyuki; Shinmei, Yasuhiro; Ohguchi, Takeshi; Kanda, Atsuhiro; Noda, Kousuke; Ishida, Susumu

2016-08-01

Vascular endothelial growth factor C (VEGF-C) plays an important role in the development of a pterygium through lymphangiogenesis. We examined the association between VEGF-C and tumor necrosis factor-α (TNF-α) in the pathogenesis of pterygia. Cultured conjunctival epithelial cells were treated with TNF-α, and the gene expression levels of VEGFC were evaluated by quantitative polymerase chain reaction (qPCR) and VEGF-C protein expression levels were measured using an enzyme-linked immunosorbent assay (ELISA). In addition, using ELISA, we evaluated the VEGF-C protein expression in the supernatants of cultured conjunctival epithelial cells, in which we neutralized TNF-α using anti‑TNF-α antibody. The gene expression of tumor necrosis factor receptor superfamily, member 1A (TNFRSF1A), known as TNF receptor 1 (TNFR1), was confirmed using reverse transcription PCR in cultured conjunctival epithelial cells. Immunofluorescence microscopy was used to examine the localization of VEGF-C and TNFR1 in pterygium tissues and TNFR1 expression in cultured conjunctival epithelial cells. Immunohistochemistry was used to examine the localization of TNFR1 in pterygia and normal conjunctival tissues. VEGFC gene expression increased in cultured conjunctival epithelial cells 24 h after the addition of TNF-α. The secretion of VEGF-C protein was significantly increased 48 h after the stimulation of cultured conjunctival epithelial cells with TNF-α. Increased VEGF-C protein secretion stimulated by TNF-α was significantly reduced by anti-TNF-α neutralizing antibody treatment. In cultured conjunctival epithelial cells, TNFRSF1A and TNFR1 were expressed. TNFR1 was immunolocalized in normal conjunctival tissues and in human pterygium tissues as well as in VEGF‑C‑positive epithelial cells from human pterygia. Our data demonstrate that TNF-α mediates VEGF-C expression, which plays a critical role in the pathogenesis of pterygia.

Upregulated epidermal growth factor receptor expression following near-infrared irradiation simulating solar radiation in a three-dimensional reconstructed human corneal epithelial tissue culture model

PubMed Central

Tanaka, Yohei; Nakayama, Jun

2016-01-01

Background and objective Humans are increasingly exposed to near-infrared (NIR) radiation from both natural (eg, solar) and artificial (eg, electrical appliances) sources. Although the biological effects of sun and ultraviolet (UV) exposure have been extensively investigated, the biological effect of NIR radiation is still unclear. We previously reported that NIR as well as UV induces photoaging and standard UV-blocking materials, such as sunglasses, do not sufficiently block NIR. The objective of this study was to investigate changes in gene expression in three-dimensional reconstructed corneal epithelial tissue culture exposed to broad-spectrum NIR irradiation to simulate solar NIR radiation that reaches human tissues. Materials and methods DNA microarray and quantitative real-time polymerase chain reaction analysis were used to assess gene expression levels in a three-dimensional reconstructed corneal epithelial model composed of normal human corneal epithelial cells exposed to water-filtered broad-spectrum NIR irradiation with a contact cooling (20°C). The water-filter allowed 1,000–1,800 nm wavelengths and excluded 1,400–1,500 nm wavelengths. Results A DNA microarray with >62,000 different probes showed 25 and 150 genes that were up- or downregulated by at least fourfold and twofold, respectively, after NIR irradiation. In particular, epidermal growth factor receptor (EGFR) was upregulated by 19.4-fold relative to control cells. Quantitative real-time polymerase chain reaction analysis revealed that two variants of EGFR in human corneal epithelial tissue were also significantly upregulated after five rounds of 10 J/cm2 irradiation (P<0.05). Conclusion We found that NIR irradiation induced the upregulated expression of EGFR in human corneal cells. Since over half of the solar energy reaching the Earth is in the NIR region, which cannot be adequately blocked by eyewear and thus can induce eye damage with intensive or long-term exposure, protection from both UV and NIR radiation may prevent changes in gene expression and in turn eye damage. PMID:27536083

Study of surface phenomena in biomaterials: The influence of physical factors

NASA Astrophysics Data System (ADS)

Sachelarie, Liliana; Vasiliu, Mihaela Papusa; Ciobanu, Catalina

2015-10-01

This study's purpose is pointing out the phenomenon that occurs at time of interaction between the tissue with implant. The materials used are Ti and its alloys. The oral tissue must be compatible with the materials used in surgical implant to human body. The bio-materials surface behavior is influenced by physical characteristics. The methods we use show a number of bio-compatibility aspects. The success of an implant in a hard tissue depends not only on the initial attachment and the osteogenic cells consecutive proliferation, but also on their capacity to create a new bone.

Within-Host Evolution of Human Influenza Virus.

PubMed

Xue, Katherine S; Moncla, Louise H; Bedford, Trevor; Bloom, Jesse D

2018-03-10

The rapid global evolution of influenza virus begins with mutations that arise de novo in individual infections, but little is known about how evolution occurs within hosts. We review recent progress in understanding how and why influenza viruses evolve within human hosts. Advances in deep sequencing make it possible to measure within-host genetic diversity in both acute and chronic influenza infections. Factors like antigenic selection, antiviral treatment, tissue specificity, spatial structure, and multiplicity of infection may affect how influenza viruses evolve within human hosts. Studies of within-host evolution can contribute to our understanding of the evolutionary and epidemiological factors that shape influenza virus's global evolution. Copyright © 2018 Elsevier Ltd. All rights reserved.

A mathematical model for calculation of 90Sr absorbed dose in dental tissues: elaboration and comparison to EPR measurements.

PubMed

Shishkina, E A; Lyubashevskii, N M; Tolstykh, E I; Ignatiev, E A; Betenekova, T A; Nikiforov, S V

2001-09-01

A mathematical model for calculation of the 90Sr absorbed doses in dental tissues is presented. The results of the Monte-Carlo calculations are compared to the data obtained by EPR measurements of dental tissues. Radiometric measurements of the 90Sr concentrations. TLD and EPR dosimetry investigations were performed in animal (dog) study. The importance of the irregular 90Sr distribution in the dentine for absorbed dose formation has been shown. The dominant dose formation factors (main source-tissues) were identified for the crown dentine and enamel. The model has shown agreement with experimental data which allows to determine further directions of the human tooth model development.

Human cartilage tissue fabrication using three-dimensional inkjet printing technology.

PubMed

Cui, Xiaofeng; Gao, Guifang; Yonezawa, Tomo; Dai, Guohao

2014-06-10

Bioprinting, which is based on thermal inkjet printing, is one of the most attractive enabling technologies in the field of tissue engineering and regenerative medicine. With digital control cells, scaffolds, and growth factors can be precisely deposited to the desired two-dimensional (2D) and three-dimensional (3D) locations rapidly. Therefore, this technology is an ideal approach to fabricate tissues mimicking their native anatomic structures. In order to engineer cartilage with native zonal organization, extracellular matrix composition (ECM), and mechanical properties, we developed a bioprinting platform using a commercial inkjet printer with simultaneous photopolymerization capable for 3D cartilage tissue engineering. Human chondrocytes suspended in poly(ethylene glycol) diacrylate (PEGDA) were printed for 3D neocartilage construction via layer-by-layer assembly. The printed cells were fixed at their original deposited positions, supported by the surrounding scaffold in simultaneous photopolymerization. The mechanical properties of the printed tissue were similar to the native cartilage. Compared to conventional tissue fabrication, which requires longer UV exposure, the viability of the printed cells with simultaneous photopolymerization was significantly higher. Printed neocartilage demonstrated excellent glycosaminoglycan (GAG) and collagen type II production, which was consistent with gene expression. Therefore, this platform is ideal for accurate cell distribution and arrangement for anatomic tissue engineering.

Macromolecular crowding meets oxygen tension in human mesenchymal stem cell culture - A step closer to physiologically relevant in vitro organogenesis

NASA Astrophysics Data System (ADS)

Cigognini, Daniela; Gaspar, Diana; Kumar, Pramod; Satyam, Abhigyan; Alagesan, Senthilkumar; Sanz-Nogués, Clara; Griffin, Matthew; O'Brien, Timothy; Pandit, Abhay; Zeugolis, Dimitrios I.

2016-08-01

Modular tissue engineering is based on the cells’ innate ability to create bottom-up supramolecular assemblies with efficiency and efficacy still unmatched by man-made devices. Although the regenerative potential of such tissue substitutes has been documented in preclinical and clinical setting, the prolonged culture time required to develop an implantable device is associated with phenotypic drift and/or cell senescence. Herein, we demonstrate that macromolecular crowding significantly enhances extracellular matrix deposition in human bone marrow mesenchymal stem cell culture at both 20% and 2% oxygen tension. Although hypoxia inducible factor - 1α was activated at 2% oxygen tension, increased extracellular matrix synthesis was not observed. The expression of surface markers and transcription factors was not affected as a function of oxygen tension and macromolecular crowding. The multilineage potential was also maintained, albeit adipogenic differentiation was significantly reduced in low oxygen tension cultures, chondrogenic differentiation was significantly increased in macromolecularly crowded cultures and osteogenic differentiation was not affected as a function of oxygen tension and macromolecular crowding. Collectively, these data pave the way for the development of bottom-up tissue equivalents based on physiologically relevant developmental processes.

Macromolecular crowding meets oxygen tension in human mesenchymal stem cell culture - A step closer to physiologically relevant in vitro organogenesis

PubMed Central

Cigognini, Daniela; Gaspar, Diana; Kumar, Pramod; Satyam, Abhigyan; Alagesan, Senthilkumar; Sanz-Nogués, Clara; Griffin, Matthew; O’Brien, Timothy; Pandit, Abhay; Zeugolis, Dimitrios I.

2016-01-01

Modular tissue engineering is based on the cells’ innate ability to create bottom-up supramolecular assemblies with efficiency and efficacy still unmatched by man-made devices. Although the regenerative potential of such tissue substitutes has been documented in preclinical and clinical setting, the prolonged culture time required to develop an implantable device is associated with phenotypic drift and/or cell senescence. Herein, we demonstrate that macromolecular crowding significantly enhances extracellular matrix deposition in human bone marrow mesenchymal stem cell culture at both 20% and 2% oxygen tension. Although hypoxia inducible factor - 1α was activated at 2% oxygen tension, increased extracellular matrix synthesis was not observed. The expression of surface markers and transcription factors was not affected as a function of oxygen tension and macromolecular crowding. The multilineage potential was also maintained, albeit adipogenic differentiation was significantly reduced in low oxygen tension cultures, chondrogenic differentiation was significantly increased in macromolecularly crowded cultures and osteogenic differentiation was not affected as a function of oxygen tension and macromolecular crowding. Collectively, these data pave the way for the development of bottom-up tissue equivalents based on physiologically relevant developmental processes. PMID:27478033

The urea decomposition product cyanate promotes endothelial dysfunction

PubMed Central

El-Gamal, Dalia; Rao, Shailaja Prabhakar; Holzer, Michael; Hallström, Seth; Haybaeck, Johannes; Gauster, Martin; Wadsack, Christian; Kozina, Andrijana; Frank, Saša; Schicho, Rudolf; Schuligoi, Rufina; Heinemann, Akos; Marsche, Gunther

2014-01-01

The dramatic cardiovascular mortality of chronic kidney disease patients is attributable in a significant proportion to endothelial dysfunction. Cyanate, a reactive species in equilibrium with urea, is formed in excess in chronic kidney disease. Cyanate is thought to have a causal role in promoting cardiovascular disease, but the underlying mechanisms remain unclear. Immunohistochemical analysis performed in the present study revealed that carbamylated epitopes associate mainly with endothelial cells in human atherosclerotic lesions. Cyanate treatment of human coronary artery endothelial cells reduced expression of endothelial nitric oxide synthase and increased tissue factor and plasminogen activator inhibitor-1 expression. In mice, administration of cyanate - promoting protein carbamylation at levels observed in uremic patients - attenuated arterial vasorelaxation of aortic rings in response to acetylcholine, without affecting sodium nitroprusside-induced relaxation. Total endothelial nitric oxide synthase and nitric oxide production were significantly reduced in aortic tissue of cyanate-treated mice. This coincided with a marked increase of tissue factor and plasminogen activator inhibitor-1 protein levels in aortas of cyanate-treated mice. These data provide evidence that cyanate compromises endothelial functionality in vitro and in vivo and may contribute to the dramatic cardiovascular risk of patients suffering from chronic kidney disease. PMID:24940796

Integrative Radiation Biology

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

Barcellos-Hoff, Mary Helen

We plan to study tissue-level mechanisms important to human breast radiation carcinogenesis. We propose that the cell biology of irradiated tissues reveals a coordinated multicellular damage response program in which individual cell contributions are primarily directed towards suppression of carcinogenesis and reestablishment of homeostasis. We identified transforming growth factor β1 (TGFβ) as a pivotal signal. Notably, we have discovered that TGFβ suppresses genomic instability by controlling the intrinsic DNA damage response and centrosome integrity. However, TGFβ also mediates disruption of microenvironment interactions, which drive epithelial to mesenchymal transition in irradiated human mammary epithelial cells. This apparent paradox of positive andmore » negative controls by TGFβ is the topic of the present proposal. First, we postulate that these phenotypes manifest differentially following fractionated or chronic exposures; second, that the interactions of multiple cell types in tissues modify the responses evident in this single cell type culture models. The goals are to: 1) study the effect of low dose rate and fractionated radiation exposure in combination with TGFβ on the irradiated phenotype and genomic instability of non-malignant human epithelial cells; and 2) determine whether stromal-epithelial interactions suppress the irradiated phenotype in cell culture and the humanized mammary mouse model. These data will be used to 3) develop a systems biology model that integrates radiation effects across multiple levels of tissue organization and time. Modeling multicellular radiation responses coordinated via extracellular signaling could have a significant impact on the extrapolation of human health risks from high dose to low dose/rate radiation exposure.« less

ATZ11 recognizes not only Z-α1-antitrypsin-polymers and complexed forms of non-Z-α1-antitrypsin but also the von Willebrand factor.

PubMed

Goltz, Diane; Hittetiya, Kanishka; Yadegari, Hamideh; Driesen, Julia; Kirfel, Jutta; Neuhaus, Thomas; Steiner, Susanne; Esch, Christiane; Bedorf, Jörg; Hertfelder, Hans-Jörg; Fischer, Hans-Peter

2014-01-01

The ATZ11 antibody has been well established for the identification of α1-anti-trypsin (AAT) molecule type PiZ (Z-AAT) in blood samples and liver tissue. In this study, we systematically analyzed the antibody for additional binding sites in human tissue. Ultrastructural ATZ11 binding was investigated immunoelectron microscopically in human umbilical vein endothelial cells (HUVECs) and in platelets of a healthy individual. Human embryonic kidney (HEK293) cells were transiently transfected with Von Willebrand factor (VWF) and analyzed immunocytochemically using confocal microscopy and SDS-PAGE electrophoresis followed by western blotting (WB). Platelets and serum samples of VWF-competent and VWF-deficient patients were investigated using native PAGE and SDS-PAGE electrophoresis followed by WB. The specificity of the ATZ11 reaction was tested immunohistochemically by extensive antibody-mediated blocking of AAT- and VWF-antigens. ATZ11-positive epitopes could be detected in Weibel-Palade bodies (WPBs) of HUVECs and α-granules of platelets. ATZ11 stains pseudo-WBP containing recombinant wild-type VWF (rVWF-WT) in HEK293 cells. In SDS-PAGE electrophoresis followed by WB, anti-VWF and ATZ11 both identified rVWF-WT. However, neither rVWF-WT-multimers, human VWF-multimers, nor serum proteins of VWF-deficient patients were detected using ATZ11 by WB, whereas anti-VWF antibody (anti-VWF) detected rVWF-WT-multimers as well as human VWF-multimers. In human tissue specimens, AAT-antigen blockade using anti-AAT antibody abolished ATZ11 staining of Z-AAT in a heterozygous AAT-deficient patient, whereas VWF-antigen blockade using anti-VWF abolished ATZ11 staining of endothelial cells and megakaryocytes. ATZ11 reacts with cellular bound and denatured rVWF-WT and human VWF as shown using immunocytochemistry and subsequent confocal imaging, immunoelectron microscopy, SDS-PAGE and WB, and immunohistology. These immunoreactions are independent of the binding of Z-AAT-molecules and non-Z-AAT complexes.

Quantification of various growth factors in different demineralized bone matrix preparations.

PubMed

Wildemann, B; Kadow-Romacker, A; Haas, N P; Schmidmaier, G

2007-05-01

Besides autografts, allografts, and synthetic materials, demineralized bone matrix (DBM) is used for bone defect filling and treatment of non-unions. Different DBM formulations are introduced in clinic since years. However, little is known about the presents and quantities of growth factors in DBM. Aim of the present study was the quantification of eight growth factors important for bone healing in three different "off the shelf" DBM formulations, which are already in human use: DBX putty, Grafton DBM putty, and AlloMatrix putty. All three DBM formulations are produced from human donor tissue but they differ in the substitutes added. From each of the three products 10 different lots were analyzed. Protein was extracted from the samples with Guanidine HCL/EDTA method and human ELISA kits were used for growth factor quantification. Differences between the three different products were seen in total protein contend and the absolute growth factor values but also a large variability between the different lots was found. The order of the growth factors, however, is almost comparable between the materials. In the three investigated materials FGF basic and BMP-4 were not detectable in any analyzed sample. BMP-2 revealed the highest concentration extractable from the samples with approximately 3.6 microg/g tissue without a significant difference between the three DBM formulations. In DBX putty significantly more TGF-beta1 and FGFa were measurable compared to the two other DBMs. IGF-I revealed the significantly highest value in the AlloMatrix and PDGF in Grafton. No differences were accessed for VEGF. Due to the differences in the growth factor concentration between the individual samples, independently from the product formulation, further analyzes are required to optimize the clinical outcome of the used demineralized bone matrix. Copyright 2006 Wiley Periodicals, Inc.

Erythropoietin Improves the Survival of Fat Tissue after Its Transplantation in Nude Mice

PubMed Central

Hamed, Saher; Egozi, Dana; Kruchevsky, Danny; Teot, Luc; Gilhar, Amos; Ullmann, Yehuda

2010-01-01

Background Autologous transplanted fat has a high resorption rate, providing a clinical challenge for the means to reduce it. Erythropoietin (EPO) has non-hematopoietic targets, and we hypothesized that EPO may improve long-term fat graft survival because it has both pro-angiogenic and anti-apoptotic properties. We aimed to determine the effect of EPO on the survival of human fat tissue after its transplantation in nude mice. Methodology/Principal Findings Human fat tissue was injected subcutaneously into immunologically-compromised nude mice, and the grafts were then treated with either 20 IU or 100 IU EPO. At the end of the 15-week study period, the extent of angiogenesis, apoptosis, and histology were assessed in the fat grafts. The results were compared to vascular endothelial growth factor (VEGF)-treated and phosphate-buffered saline (PBS)-treated fat grafts. The weight and volume of the EPO-treated grafts were higher than those of the PBS-treated grafts, whose weights and volumes were not different from those of the VEGF-treated grafts. EPO treatment also increased the expression of angiogenic factors and microvascular density, and reduced inflammation and apoptosis in a dose-dependent manner in the fat grafts. Conclusions/Significance Our data suggest that stimulation of angiogenesis by a cluster of angiogenic factors and decreased fat cell apoptosis account for potential mechanisms that underlie the improved long-term survival of fat transplants following EPO treatment. PMID:21085572

Erythropoietin improves the survival of fat tissue after its transplantation in nude mice.

PubMed

Hamed, Saher; Egozi, Dana; Kruchevsky, Danny; Teot, Luc; Gilhar, Amos; Ullmann, Yehuda

2010-11-15

Autologous transplanted fat has a high resorption rate, providing a clinical challenge for the means to reduce it. Erythropoietin (EPO) has non-hematopoietic targets, and we hypothesized that EPO may improve long-term fat graft survival because it has both pro-angiogenic and anti-apoptotic properties. We aimed to determine the effect of EPO on the survival of human fat tissue after its transplantation in nude mice. Human fat tissue was injected subcutaneously into immunologically-compromised nude mice, and the grafts were then treated with either 20 IU or 100 IU EPO. At the end of the 15-week study period, the extent of angiogenesis, apoptosis, and histology were assessed in the fat grafts. The results were compared to vascular endothelial growth factor (VEGF)-treated and phosphate-buffered saline (PBS)-treated fat grafts. The weight and volume of the EPO-treated grafts were higher than those of the PBS-treated grafts, whose weights and volumes were not different from those of the VEGF-treated grafts. EPO treatment also increased the expression of angiogenic factors and microvascular density, and reduced inflammation and apoptosis in a dose-dependent manner in the fat grafts. Our data suggest that stimulation of angiogenesis by a cluster of angiogenic factors and decreased fat cell apoptosis account for potential mechanisms that underlie the improved long-term survival of fat transplants following EPO treatment.

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

NASA Astrophysics Data System (ADS)

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

2014-02-01

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

Correlations between quantitative fat–water magnetic resonance imaging and computed tomography in human subcutaneous white adipose tissue

PubMed Central

Gifford, Aliya; Walker, Ronald C.; Towse, Theodore F.; Brian Welch, E.

2015-01-01

Abstract. Beyond estimation of depot volumes, quantitative analysis of adipose tissue properties could improve understanding of how adipose tissue correlates with metabolic risk factors. We investigated whether the fat signal fraction (FSF) derived from quantitative fat–water magnetic resonance imaging (MRI) scans at 3.0 T correlates to CT Hounsfield units (HU) of the same tissue. These measures were acquired in the subcutaneous white adipose tissue (WAT) at the umbilical level of 21 healthy adult subjects. A moderate correlation exists between MRI- and CT-derived WAT values for all subjects, R2=0.54, p<0.0001, with a slope of −2.6, (95% CI [−3.3,−1.8]), indicating that a decrease of 1 HU equals a mean increase of 0.38% FSF. We demonstrate that FSF estimates obtained using quantitative fat–water MRI techniques correlate with CT HU values in subcutaneous WAT, and therefore, MRI-based FSF could be used as an alternative to CT HU for assessing metabolic risk factors. PMID:26702407

Allelic Expression of Deleterious Protein-Coding Variants across Human Tissues

PubMed Central

Kukurba, Kimberly R.; Zhang, Rui; Li, Xin; Smith, Kevin S.; Knowles, David A.; How Tan, Meng; Piskol, Robert; Lek, Monkol; Snyder, Michael; MacArthur, Daniel G.; Li, Jin Billy; Montgomery, Stephen B.

2014-01-01

Personal exome and genome sequencing provides access to loss-of-function and rare deleterious alleles whose interpretation is expected to provide insight into individual disease burden. However, for each allele, accurate interpretation of its effect will depend on both its penetrance and the trait's expressivity. In this regard, an important factor that can modify the effect of a pathogenic coding allele is its level of expression; a factor which itself characteristically changes across tissues. To better inform the degree to which pathogenic alleles can be modified by expression level across multiple tissues, we have conducted exome, RNA and deep, targeted allele-specific expression (ASE) sequencing in ten tissues obtained from a single individual. By combining such data, we report the impact of rare and common loss-of-function variants on allelic expression exposing stronger allelic bias for rare stop-gain variants and informing the extent to which rare deleterious coding alleles are consistently expressed across tissues. This study demonstrates the potential importance of transcriptome data to the interpretation of pathogenic protein-coding variants. PMID:24786518

Depth distributions of light action spectra for skin chromophores

NASA Astrophysics Data System (ADS)

Barun, V. V.; Ivanov, A. P.

2010-03-01

Light action spectra over wavelengths of 300-1000 nm are calculated for components of the human cutaneous covering: melanin, basal (bloodless) tissue, and blood oxy- and deoxyhemoglobin. The transformation of the spectra with depth in biological tissue results from two factors. The first is the wavelength dependence of the absorption coefficient corresponding to a particular skin chromophore and the second is the spectral selectivity of the radiation flux in biological tissue. This factor is related to the optical properties of all chromophores. A significant change is found to take place in the spectral distribution of absorbed radiant power with increasing depth. The action spectrum of light for the molecular oxygen contained in all components of biological tissue is also studied in the 625-645 nm range. The spectra are found to change with both the volume fraction of blood vessels and the degree of oxygenation of the blood. These results are useful for analyzing processes associated with optical absorption that are possible mechanisms for the interaction of light with biological tissues: photodissociation of oxyhemoglobin and the light-oxygen effect.

Engineering anisotropic biomimetic fibrocartilage microenvironment by bioprinting mesenchymal stem cells in nanoliter gel droplets.

PubMed

Gurkan, Umut A; El Assal, Rami; Yildiz, Simin E; Sung, Yuree; Trachtenberg, Alexander J; Kuo, Winston P; Demirci, Utkan

2014-07-07

Over the past decade, bioprinting has emerged as a promising patterning strategy to organize cells and extracellular components both in two and three dimensions (2D and 3D) to engineer functional tissue mimicking constructs. So far, tissue printing has neither been used for 3D patterning of mesenchymal stem cells (MSCs) in multiphase growth factor embedded 3D hydrogels nor been investigated phenotypically in terms of simultaneous differentiation into different cell types within the same micropatterned 3D tissue constructs. Accordingly, we demonstrated a biochemical gradient by bioprinting nanoliter droplets encapsulating human MSCs, bone morphogenetic protein 2 (BMP-2), and transforming growth factor β1 (TGF- β1), engineering an anisotropic biomimetic fibrocartilage microenvironment. Assessment of the model tissue construct displayed multiphasic anisotropy of the incorporated biochemical factors after patterning. Quantitative real time polymerase chain reaction (qRT-PCR) results suggested genomic expression patterns leading to simultaneous differentiation of MSC populations into osteogenic and chondrogenic phenotype within the multiphasic construct, evidenced by upregulation of osteogenesis and condrogenesis related genes during in vitro culture. Comprehensive phenotypic network and pathway analysis results, which were based on genomic expression data, indicated activation of differentiation related mechanisms, via signaling pathways, including TGF, BMP, and vascular endothelial growth factor.

Engineering Anisotropic Biomimetic Fibrocartilage Microenvironment by Bioprinting Mesenchymal Stem Cells in Nanoliter Gel Droplets

PubMed Central

2015-01-01

Over the past decade, bioprinting has emerged as a promising patterning strategy to organize cells and extracellular components both in two and three dimensions (2D and 3D) to engineer functional tissue mimicking constructs. So far, tissue printing has neither been used for 3D patterning of mesenchymal stem cells (MSCs) in multiphase growth factor embedded 3D hydrogels nor been investigated phenotypically in terms of simultaneous differentiation into different cell types within the same micropatterned 3D tissue constructs. Accordingly, we demonstrated a biochemical gradient by bioprinting nanoliter droplets encapsulating human MSCs, bone morphogenetic protein 2 (BMP-2), and transforming growth factor β1 (TGF- β1), engineering an anisotropic biomimetic fibrocartilage microenvironment. Assessment of the model tissue construct displayed multiphasic anisotropy of the incorporated biochemical factors after patterning. Quantitative real time polymerase chain reaction (qRT-PCR) results suggested genomic expression patterns leading to simultaneous differentiation of MSC populations into osteogenic and chondrogenic phenotype within the multiphasic construct, evidenced by upregulation of osteogenesis and condrogenesis related genes during in vitro culture. Comprehensive phenotypic network and pathway analysis results, which were based on genomic expression data, indicated activation of differentiation related mechanisms, via signaling pathways, including TGF, BMP, and vascular endothelial growth factor. PMID:24495169

Pattern-Recognition Receptor Signaling Regulator mRNA Expression in Humans and Mice, and in Transient Inflammation or Progressive Fibrosis

PubMed Central

Günthner, Roman; Kumar, Vankayala Ramaiah Santhosh; Lorenz, Georg; Anders, Hans-Joachim; Lech, Maciej

2013-01-01

The cell type-, organ-, and species-specific expression of the pattern-recognition receptors (PRRs) are well described but little is known about the respective expression profiles of their negative regulators. We therefore determined the mRNA expression levels of A20, CYLD, DUBA, ST2, CD180, SIGIRR, TANK, SOCS1, SOCS3, SHIP, IRAK-M, DOK1, DOK2, SHP1, SHP2, TOLLIP, IRF4, SIKE, NLRX1, ERBIN, CENTB1, and Clec4a2 in human and mouse solid organs. Humans and mice displayed significant differences between their respective mRNA expression patterns of these factors. Additionally, we characterized their expression profiles in mononuclear blood cells upon bacterial endotoxin, which showed a consistent induction of A20, SOCS3, IRAK-M, and Clec4a2 in human and murine cells. Furthermore, we studied the expression pattern in transient kidney ischemia-reperfusion injury versus post-ischemic atrophy and fibrosis in mice. A20, CD180, ST2, SOCS1, SOCS3, SHIP, IRAK-M, DOK1, DOK2, IRF4, CENTB1, and Clec4a2 were all induced, albeit at different times of injury and repair. Progressive fibrosis was associated with a persistent induction of these factors. Thus, the organ- and species-specific expression patterns need to be considered in the design and interpretation of studies related to PRR-mediated innate immunity, which seems to be involved in tissue injury, tissue regeneration and in progressive tissue scarring. PMID:24009023

Basis for the Induction of Tissue-Level Phase-2 Reentry as a Repolarization Disorder in the Brugada Syndrome

PubMed Central

Bueno-Orovio, Alfonso; Cherry, Elizabeth M.; Evans, Steven J.; Fenton, Flavio H.

2015-01-01

Aims. Human action potentials in the Brugada syndrome have been characterized by delayed or even complete loss of dome formation, especially in the right ventricular epicardial layers. Such a repolarization pattern is believed to trigger phase-2 reentry (P2R); however, little is known about the conditions necessary for its initiation. This study aims to determine the specific mechanisms that facilitate P2R induction in Brugada-affected cardiac tissue in humans. Methods. Ionic models for Brugada syndrome in human epicardial cells were developed and used to study the induction of P2R in cables, sheets, and a three-dimensional model of the right ventricular free wall. Results. In one-dimensional cables, P2R can be induced by adjoining lost-dome and delayed-dome regions, as mediated by tissue excitability and transmembrane voltage profiles, and reduced coupling facilitates its induction. In two and three dimensions, sustained reentry can arise when three regions (delayed-dome, lost-dome, and normal epicardium) are present. Conclusions. Not only does P2R induction by Brugada syndrome require regions of action potential with delayed-dome and lost-dome, but in order to generate a sustained reentry from a triggered waveback multiple factors are necessary, including heterogeneity in action potential distribution, tissue coupling, direction of stimulation, the shape of the late plateau, the duration of lost-dome action potentials, and recovery of tissue excitability, which is predominantly modulated by tissue coupling. PMID:26583094

The Use of Human Wharton's Jelly Cells for Cochlear Tissue Engineering.

PubMed

Mellott, Adam J; Detamore, Michael S; Staecker, Hinrich

2016-01-01

Tissue engineering focuses on three primary components: stem cells, biomaterials, and growth factors. Together, the combination of these components is used to regrow and repair damaged tissues that normally do not regenerate easily on their own. Much attention has been focused on the use of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), due to their broad differentiation potential. However, ESCs and iPSCs require very detailed protocols to differentiate into target tissues, which are not always successful. Furthermore, procurement of ESCs is considered ethically controversial in some regions and procurement of iPSCs requires laborious transformation of adult tissues and characterization. However, mesenchymal stem cells are an adult stem cell population that are not ethically controversial and are readily available for procurement. Furthermore, mesenchymal stem cells exhibit the ability to differentiate into a variety of cell types arising from the mesoderm. In particular, human Wharton's jelly cells (hWJCs) are mesenchymal-type stem cells found in umbilical cords that possess remarkable differentiation potential. hWJCs are a highly desirable stem cell population due to their abundance in supply, high proliferation rates, and ability to differentiate into multiple cell types arising from all three germ layers. hWJCs are used to generate several neurological phenotypes arising from the ectoderm and are considered for engineering mechanosensory hair cells found in the auditory complex. Here, we report the methods for isolating hWJCs from human umbilical cords and non-virally transfected for use in cochlear tissue engineering studies.

MOLD-SHAPED, NANOFIBER SCAFFOLD-BASED CARTILAGE ENGINEERING USING HUMAN MESENCHYMAL STEM CELLS AND BIOREACTOR

PubMed Central

Janjanin, Sasa; Li, Wan-Ju; Morgan, Meredith T.; Shanti, Rabie M.; Tuan, Rocky S.

2008-01-01

Background Mesenchymal stem cell (MSC)-based tissue engineering is a promising future alternative to autologous cartilage grafting. This study evaluates the potential of using MSCs, seeded into electrospun, biodegradable polymeric nanofibrous scaffolds, to engineer cartilage with defined dimensions and shape, similar to grafts used for subcutaneous implantation in plastic and reconstructive surgery. Materials and methods Human bone marrow derived MSCs seeded onto nanofibrous scaffolds and placed in custom-designed molds were cultured for up to 42 days in bioreactors. Chondrogenesis was induced with either transforming growth factor-β1 (TGF-β1) alone or in combination with insulin-like growth factor-I (IGF-I). Results Constructs exhibited hyaline cartilage histology with desired thickness and shape as well as favorable tissue integrity and shape retention, suggesting the presence of elastic tissue. Time-dependent increase in cartilage matrix gene expression was seen in both types of culture; at Day 42, TGF-β1/IGF-I treated cultures showed higher collagen type II and aggrecan expression. Both culture conditions showed significant time-dependent increase in sulfated glycosaminoglycan and hydroxyproline contents. TGF-β1/IGF-I treated samples were significantly stiffer; with equilibrium compressive Young’s modulus values reaching 17 kPa by Day 42. Conclusions The successful ex vivo development of geometrically defined cartilaginous construct using customized molding suggests the potential of cell-based cartilage tissue for reconstructive surgery. PMID:18316094

Detection and a possible link between parvovirus B19 and thyroid cancer.

PubMed

Etemadi, Ashkan; Mostafaei, Shayan; Yari, Kheirollah; Ghasemi, Amir; Minaei Chenar, Hamzeh; Moghoofei, Mohsen

2017-06-01

Human parvovirus B19 (B19) is a small, non-enveloped virus and belongs to Parvoviridae family. B19 persists in many tissues such as thyroid tissue and even thyroid cancer. The main aim of this study was to determine the presence of B19, its association with increased inflammation in thyroid tissue, and thus its possible role in thyroid cancer progression. Studies have shown that virus replication in non-permissive tissue leads to overexpression of non-structural protein and results in upregulation of proinflammatory cytokines such as interleukin 6 and tumor necrosis factor alpha. A total of 36 paraffin-embedded thyroid specimens and serum were collected from patients and 12 samples were used as control. Various methods were employed, including polymerase chain reaction, real-time polymerase chain reaction, and enzyme-linked immunosorbent assay. The results have shown the presence of B19 DNA in 31 of 36 samples (86.11%). Almost in all samples, the levels of non-structural protein 1, nuclear factor kappa B, tumor necrosis factor alpha, and interleukin 6 were simultaneously high. The presence of parvovirus B19 has a significant positive correlation with nuclear factor kappa B, tumor necrosis factor alpha, and interleukin 6 levels. This study suggests that B19 infection may play an important role in tumorigenesis and thyroid cancer development via the inflammatory mechanisms.

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.

21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

Code of Federal Regulations, 2012 CFR

2012-04-01

... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Tissue culture media for human ex vivo tissue and... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell culture...

21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

Code of Federal Regulations, 2013 CFR

2013-04-01

... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Tissue culture media for human ex vivo tissue and... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell culture...

21 CFR 876.5885 - Tissue culture media for human ex vivo tissue and cell culture processing applications.

Code of Federal Regulations, 2014 CFR

2014-04-01

... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Tissue culture media for human ex vivo tissue and... DEVICES Therapeutic Devices § 876.5885 Tissue culture media for human ex vivo tissue and cell culture processing applications. (a) Identification. Tissue culture media for human ex vivo tissue and cell culture...

Clinical EPR: Unique Opportunities and Some Challenges

PubMed Central

Swartz, Harold M.; Williams, Benjamin B.; Zaki, Bassem I.; Hartford, Alan C.; Jarvis, Lesley A.; Chen, Eunice; Comi, Richard J.; Ernstoff, Marc S.; Hou, Huagang; Khan, Nadeem; Swarts, Steven G.; Flood, Ann B.; Kuppusamy, Periannan

2014-01-01

Electron paramagnetic resonance (EPR) spectroscopy has been well established as a viable technique for measurement of free radicals and oxygen in biological systems, from in vitro cellular systems to in vivo small animal models of disease. However, the use of EPR in human subjects in the clinical setting, although attractive for a variety of important applications such as oxygen measurement, is challenged with several factors including the need for instrumentation customized for human subjects, probe and regulatory constraints. This paper describes the rationale and development of the first clinical EPR systems for two important clinical applications, namely, measurement of tissue oxygen (oximetry), and radiation dose (dosimetry) in humans. The clinical spectrometers operate at 1.2 GHz frequency and use surface loop resonators capable of providing topical measurements up to 1 cm depth in tissues. Tissue pO2 measurements can be carried out noninvasively and repeatedly after placement of an oxygen-sensitive paramagnetic material (currently India ink) at the site of interest. Our EPR dosimetry system is capable of measuring radiation-induced free radicals in the tooth of irradiated human subjects to determine the exposure dose. These developments offer potential opportunities for clinical dosimetry and oximetry, which include guiding therapy for individual patients with tumors or vascular disease, by monitoring of tissue oxygenation. Further work is in progress to translate this unique technology to routine clinical practice. PMID:24439333

Dirofilaria repens: emergence of autochthonous human infections in the Czech Republic (case reports).

PubMed

Matějů, Jana; Chanová, Marta; Modrý, David; Mitková, Barbora; Hrazdilová, Kristýna; Žampachová, Víta; Kolářová, Libuše

2016-04-19

Human dirofilariasis is a zoonotic infection that continues to spread to previously unaffected areas of Europe. In the South Moravian Region of the Czech Republic (CR), imported as well as autochthonous canine infections were recorded in the last decade, and parasite DNA was detected in mosquitoes of Aedes vexans. In the present paper, human Dirofilaria infections are reported from the country for the first time. The samples from five patients with suspected tissue helminthiases were investigated. In particular cases, nematodes were isolated from various tissues including skin of lower leg, soft tissues of finger, subcutaneous tissue of hypogastrium, lymph node and peritoneum. The diagnosis was based on light microscopic morphology and/or DNA analysis of the worms. In addition, ELISA examination of patients' sera for anti-filaria IgG antibodies was performed. In the CR, five cases of human dirofilariasis caused by Dirofilaria repens were recorded during 2010-2014 (species determination for three of them was confirmed besides morphological also by DNA analysis). At least, three of the cases were of autochthonous origin (the patients are Czech citizens residing in South Moravian Region who have never travelled abroad). The findings confirm the natural setting of D. repens in South Moravian Region of the CR. Dirofilariasis should be therefore considered as endemic in this area where it may represent a significant risk factor for public health.

Arborvitae (Thuja plicata) essential oil significantly inhibited critical inflammation- and tissue remodeling-related proteins and genes in human dermal fibroblasts.

PubMed

Han, Xuesheng; Parker, Tory L

2017-06-01

Arborvitae ( Thuja plicata ) essential oil (AEO) is becoming increasingly popular in skincare, although its biological activity in human skin cells has not been investigated. Therefore, we sought to study AEO's effect on 17 important protein biomarkers that are closely related to inflammation and tissue remodeling by using a pre-inflamed human dermal fibroblast culture model. AEO significantly inhibited the expression of vascular cell adhesion molecule 1 (VCAM-1), intracellular cell adhesion molecule 1 (ICAM-1), interferon gamma-induced protein 10 (IP-10), interferon-inducible T-cell chemoattractant (I-TAC), monokine induced by interferon gamma (MIG), and macrophage colony-stimulating factor (M-CSF). It also showed significant antiproliferative activity and robustly inhibited collagen-I, collagen-III, plasminogen activator inhibitor-1 (PAI-1), and tissue inhibitor of metalloproteinase 1 and 2 (TIMP-1 and TIMP-2). The inhibitory effect of AEO on increased production of these protein biomarkers suggests it has anti-inflammatory property. We then studied the effect of AEO on the genome-wide expression of 21,224 genes in the same cell culture. AEO significantly and diversely modulated global gene expression. Ingenuity pathway analysis (IPA) showed that AEO robustly affected numerous critical genes and signaling pathways closely involved in inflammatory and tissue remodeling processes. The findings of this study provide the first evidence of the biological activity and beneficial action of AEO in human skin cells.

Role of Insulin-like growth factors in initiation of follicle growth in normal and polycystic human ovaries.

PubMed

Stubbs, Sharron A; Webber, Lisa J; Stark, Jaroslav; Rice, Suman; Margara, Raul; Lavery, Stuart; Trew, Geoffrey H; Hardy, Kate; Franks, Stephen

2013-08-01

Polycystic ovary syndrome (PCOS), the commonest cause of anovulatory infertility, is characterized by disordered follicle development including increased activation and accelerated growth of preantral follicles. Data from experimental animals and preliminary results from studies of human ovarian tissue suggest that IGFs affect preantral follicle development. Our objectives were to investigate the expression of the type-1 IGF receptor (IGFR-1) in the human ovary and to determine whether IGFs are involved in stimulating the transition of follicles from primordial to primary stage in normal and polycystic ovaries. We used archived ovarian tissue for protein expression studies and small cortical biopsies for follicle isolation and for tissue culture. This was a laboratory-based study, using clinical tissue samples. A total of 54 women, 33 with normal ovaries and 21 with polycystic ovaries, were classified by reference to menstrual cycle history and ultrasonography. We evaluated expression of IGFR-1 mRNA in isolated preantral follicles and of IGFR-1 protein in archived ovarian tissue samples from normal and polycystic ovaries and effects of exogenous IGF-1 on preantral follicle development and survival in cultured fragments of normal and polycystic ovaries. IGFR-1 mRNA and protein was expressed in preantral follicles at all stages of development and enhanced expression was noted in PCOS follicles during early preantral development. IGF-1 stimulated initiation of follicle growth in normal tissue but had little effect on preantral follicle growth in polycystic ovaries in which, characteristically, there was a higher proportion of follicles that had entered the growing phase even before culture. IGFs are plausible candidates in regulation of initiation of human follicle growth, and accelerated preantral follicle growth in PCOS may be due to increased activity of endogenous IGFs.

Topically applied connective tissue growth factor/CCN2 improves diabetic preclinical cutaneous wound healing: potential role for CTGF in human diabetic foot ulcer healing.

PubMed

Henshaw, F R; Boughton, P; Lo, L; McLennan, S V; Twigg, S M

2015-01-01

Topical application of CTGF/CCN2 to rodent diabetic and control wounds was examined. In parallel research, correlation of CTGF wound fluid levels with healing rate in human diabetic foot ulcers was undertaken. Full thickness cutaneous wounds in diabetic and nondiabetic control rats were treated topically with 1 μg rhCTGF or vehicle alone, on 2 consecutive days. Wound healing rate was observed on day 14 and wound sites were examined for breaking strength and granulation tissue. In the human study across 32 subjects, serial CTGF regulation was analyzed longitudinally in postdebridement diabetic wound fluid. CTGF treated diabetic wounds had an accelerated closure rate compared with vehicle treated diabetic wounds. Healed skin withstood more strain before breaking in CTGF treated rat wounds. Granulation tissue from CTGF treatment in diabetic wounds showed collagen IV accumulation compared with nondiabetic animals. Wound α-smooth muscle actin was increased in CTGF treated diabetic wounds compared with untreated diabetic wounds, as was macrophage infiltration. Endogenous wound fluid CTGF protein rate of increase in human diabetic foot ulcers correlated positively with foot ulcer healing rate (r = 0.406; P < 0.001). These data collectively increasingly substantiate a functional role for CTGF in human diabetic foot ulcers.

Nanoparticle-mediated transcriptional modification enhances neuronal differentiation of human neural stem cells following transplantation in rat brain.

PubMed

Li, Xiaowei; Tzeng, Stephany Y; Liu, Xiaoyan; Tammia, Markus; Cheng, Yu-Hao; Rolfe, Andrew; Sun, Dong; Zhang, Ning; Green, Jordan J; Wen, Xuejun; Mao, Hai-Quan

2016-04-01

Strategies to enhance survival and direct the differentiation of stem cells in vivo following transplantation in tissue repair site are critical to realizing the potential of stem cell-based therapies. Here we demonstrated an effective approach to promote neuronal differentiation and maturation of human fetal tissue-derived neural stem cells (hNSCs) in a brain lesion site of a rat traumatic brain injury model using biodegradable nanoparticle-mediated transfection method to deliver key transcriptional factor neurogenin-2 to hNSCs when transplanted with a tailored hyaluronic acid (HA) hydrogel, generating larger number of more mature neurons engrafted to the host brain tissue than non-transfected cells. The nanoparticle-mediated transcription activation method together with an HA hydrogel delivery matrix provides a translatable approach for stem cell-based regenerative therapy. Copyright © 2016 Elsevier Ltd. All rights reserved.

Isolation of Human Innate Lymphoid Cells.

PubMed

Krabbendam, Lisette; Nagasawa, Maho; Spits, Hergen; Bal, Suzanne M

2018-06-29

Innate lymphoid cells (ILCs) are innate immune cells of lymphoid origin that have important effector and regulatory functions in the first line of defense against pathogens, but also regulate tissue homeostasis, remodeling, and repair. Their function mirrors T helper cells and cytotoxic CD8 + T lymphocytes, but they lack expression of rearranged antigen-specific receptors. Distinct ILC subsets are classified in group 1 ILCs (ILC1s), group 2 ILCs (ILC2s), and group 3 ILCs (ILC3s and lymphoid tissue-inducer cells), based on the expression of transcription factors and the cytokines they produce. As the frequency of ILCs is low, their isolation requires extensive depletion of other cell types. The lack of unique cell surface antigens further complicates the identification of these cells. Here, methods for ILC isolation and characterization from human peripheral blood and different tissues are described. © 2018 by John Wiley & Sons, Inc. © 2018 John Wiley & Sons, Inc.

Fracture healing: mechanisms and interventions

PubMed Central

Einhorn, Thomas A.; Gerstenfeld, Louis C.

2015-01-01

Fractures are the most common large-organ, traumatic injuries to humans. The repair of bone fractures is a postnatal regenerative process that recapitulates many of the ontological events of embryonic skeletal development. Although fracture repair usually restores the damaged skeletal organ to its pre-injury cellular composition, structure and biomechanical function, about 10% of fractures will not heal normally. This article reviews the developmental progression of fracture healing at the tissue, cellular and molecular levels. Innate and adaptive immune processes are discussed as a component of the injury response, as are environmental factors, such as the extent of injury to the bone and surrounding tissue, fixation and the contribution of vascular tissues. We also present strategies for fracture treatment that have been tested in animal models and in clinical trials or case series. The biophysical and biological basis of the molecular actions of various therapeutic approaches, including recombinant human bone morphogenetic proteins and parathyroid hormone therapy, are also discussed. PMID:25266456

Engineering a humanized bone organ model in mice to study bone metastases.

PubMed

Martine, Laure C; Holzapfel, Boris M; McGovern, Jacqui A; Wagner, Ferdinand; Quent, Verena M; Hesami, Parisa; Wunner, Felix M; Vaquette, Cedryck; De-Juan-Pardo, Elena M; Brown, Toby D; Nowlan, Bianca; Wu, Dan Jing; Hutmacher, Cosmo Orlando; Moi, Davide; Oussenko, Tatiana; Piccinini, Elia; Zandstra, Peter W; Mazzieri, Roberta; Lévesque, Jean-Pierre; Dalton, Paul D; Taubenberger, Anna V; Hutmacher, Dietmar W

2017-04-01

Current in vivo models for investigating human primary bone tumors and cancer metastasis to the bone rely on the injection of human cancer cells into the mouse skeleton. This approach does not mimic species-specific mechanisms occurring in human diseases and may preclude successful clinical translation. We have developed a protocol to engineer humanized bone within immunodeficient hosts, which can be adapted to study the interactions between human cancer cells and a humanized bone microenvironment in vivo. A researcher trained in the principles of tissue engineering will be able to execute the protocol and yield study results within 4-6 months. Additive biomanufactured scaffolds seeded and cultured with human bone-forming cells are implanted ectopically in combination with osteogenic factors into mice to generate a physiological bone 'organ', which is partially humanized. The model comprises human bone cells and secreted extracellular matrix (ECM); however, other components of the engineered tissue, such as the vasculature, are of murine origin. The model can be further humanized through the engraftment of human hematopoietic stem cells (HSCs) that can lead to human hematopoiesis within the murine host. The humanized organ bone model has been well characterized and validated and allows dissection of some of the mechanisms of the bone metastatic processes in prostate and breast cancer.

Enhanced HIV-1 replication in ex vivo ectocervical tissues from post-menopausal women correlates with increased inflammatory responses.

PubMed

Rollenhagen, C; Asin, S N

2011-11-01

Knowledge about early innate immune responses at the mucosal surfaces of the female genital tract is important in understanding the pathogenesis of heterosexual transmission of human immunodeficiency virus type-1 (HIV-1). As estradiol decreases inflammatory responses, we postulated that an estradiol-deficient state such as post-menopause could enhance expression of inflammatory factors that stimulate HIV-1 replication. We compare HIV-1 integration, transcription, and viral p24 release levels among ectocervical tissues obtained from pre- and post-menopausal donors. We detected enhanced HIV-1 p24 release levels in post- compared with pre-menopausal tissues (P<0.0001), but saw no difference in HIV-1 integration. Overall, 100% of post-menopausal tissues exhibited levels of HIV-1 transcription above background compared with only 60% of pre-menopausal tissues. Increased HIV-1 transcription was associated with enhanced interleukin (IL)-1β, IL-6, monocyte chemotactic protein-1, growth-regulated oncogene-α, and interferon-γ-inducible protein-10 expression. Neutralization and nuclear factor-κB-targeting small-interfering RNA experiments both decreased HIV-1 transcription, suggesting that the early inflammatory response may facilitate HIV-1 replication in ex vivo ectocervical tissues from post-menopausal women.

A nanoparticulate injectable hydrogel as a tissue engineering scaffold for multiple growth factor delivery for bone regeneration.

PubMed

Dyondi, Deepti; Webster, Thomas J; Banerjee, Rinti

2013-01-01

Gellan xanthan gels have been shown to be excellent carriers for growth factors and as matrices for several tissue engineering applications. Gellan xanthan gels along with chitosan nanoparticles of 297 ± 61 nm diameter, basic fibroblast growth factor (bFGF), and bone morphogenetic protein 7 (BMP7) were employed in a dual growth factor delivery system to promote the differentiation of human fetal osteoblasts. An injectable system with ionic and temperature gelation was optimized and characterized. The nanoparticle loaded gels showed significantly improved cell proliferation and differentiation due to the sustained release of growth factors. A differentiation marker study was conducted, analyzed, and compared to understand the effect of single vs dual growth factors and free vs encapsulated growth factors. Dual growth factor loaded gels showed a higher alkaline phosphatase and calcium deposition compared to single growth factor loaded gels. The results suggest that encapsulation and stabilization of growth factors within nanoparticles and gels are promising for bone regeneration. Gellan xanthan gels also showed antibacterial effects against Pseudomonas aeruginosa, Staphylococcus aureus, and Staphylococcus epidermidis, the common pathogens in implant failure.

The Placenta: Applications in Orthopaedic Sports Medicine.

PubMed

McIntyre, James Alexander; Jones, Ian A; Danilkovich, Alla; Vangsness, C Thomas

2018-01-01

Placenta has a long history of use for treating burns and wounds. It is a rich source of collagen and other extracellular matrix proteins, tissue reparative growth factors, and stem cells, including mesenchymal stem cells (MSCs). Recent data show its therapeutic potential for orthopaedic sports medicine indications. To provide orthopaedic surgeons with an anatomic description of the placenta, to characterize its cellular composition, and to review the literature reporting the use of placenta-derived cells and placental tissue allografts for orthopaedic sports medicine indications in animal models and in humans. Systematic review. Using a total of 63 keyword combinations, the PubMed and MEDLINE databases were searched for published articles describing the use of placental cells and/or tissue for orthopaedic sports medicine indications. Information was collected on placental tissue type, indications, animal model, study design, treatment regimen, safety, and efficacy outcomes. Results were categorized by indication and subcategorized by animal model. Outcomes for 29 animal studies and 6 human studies reporting the use of placenta-derived therapeutics were generally positive; however, the placental tissue source, clinical indication, and administration route were highly variable across these studies. Fourteen animal studies described the use of placental tissue for tendon injuries, 13 studies for osteoarthritis or articular cartilage injuries, 3 for ligament injuries, and 1 for synovitis. Both placenta-derived culture-expanded cells (epithelial cells or MSCs) and placental tissue allografts were used in animal studies. In all human studies, commercial placental allografts were used. Five of 6 human studies examined the treatment of foot and ankle pathological conditions, and 1 studied the treatment of knee osteoarthritis. A review of the small number of reported studies revealed a high degree of variability in placental cell types, placental tissue preparation, routes of administration, and treatment regimens, which prohibits making any definitive conclusions. Currently, the clinical use of placenta is limited to only commercial placental tissue allografts, as there are no placenta-derived biological drugs approved for the treatment of orthopaedic sports medicine conditions in the United States. However, this review shows that the application of placental cells or tissue allografts appears to be safe and has potential to improve outcomes for orthopaedic sports medicine indications.

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

UPREGULATION OF TISSUE FACTOR IN HUMAN ENDOTHELIAL CELLS FOLLOWING ULTRAFINE PARTICLE EXPOSURE

EPA Science Inventory

Epidemiology studies have linked the exposure to air pollutant particles with increased cardiovascular mortality and morbidity, but the mechanisms remain unknown. In our laboratory we have tested the hypothesis that the ultrafine fraction of ambient pollutant particles would cau...

Possible Muscle Repair in the Human Cardiovascular System.

PubMed

Sommese, Linda; Zullo, Alberto; Schiano, Concetta; Mancini, Francesco P; Napoli, Claudio

2017-04-01

The regenerative potential of tissues and organs could promote survival, extended lifespan and healthy life in multicellular organisms. Niches of adult stemness are widely distributed and lead to the anatomical and functional regeneration of the damaged organ. Conversely, muscular regeneration in mammals, and humans in particular, is very limited and not a single piece of muscle can fully regrow after a severe injury. Therefore, muscle repair after myocardial infarction is still a chimera. Recently, it has been recognized that epigenetics could play a role in tissue regrowth since it guarantees the maintenance of cellular identity in differentiated cells and, therefore, the stability of organs and tissues. The removal of these locks can shift a specific cell identity back to the stem-like one. Given the gradual loss of tissue renewal potential in the course of evolution, in the last few years many different attempts to retrieve such potential by means of cell therapy approaches have been performed in experimental models. Here we review pathways and mechanisms involved in the in vivo repair of cardiovascular muscle tissues in humans. Moreover, we address the ongoing research on mammalian cardiac muscle repair based on adult stem cell transplantation and pro-regenerative factor delivery. This latter issue, involving genetic manipulations of adult cells, paves the way for developing possible therapeutic strategies in the field of cardiovascular muscle repair.

Computational Analysis of Intersubject Variability and Thrombin Generation in Dilutional Coagulopathy

DTIC Science & Technology

2012-11-01

proteins: Factor (F)II, FV, FVII , FVIII, F IX, and FX, as well as the anticoagulants antithrombin (AT) and TF pathway inhibi- tor (TFPI). The results...coagulation factors FII, FV, FVII , FVIIa, FVIII, F IX and FX, as well as the anticoagulants TFPI and AT and the throm- bin generation inducer TF. The model...scenario and tissue factor concentration. CONCLUSION: Dilutional effects on thrombin genera- tion in a human population can be predicted from trends

Tissue repair

PubMed Central

2010-01-01

As living beings that encounter every kind of traumatic event from paper cut to myocardial infarction, we must possess ways to heal damaged tissues. While some animals are able to regrow complete body parts following injury (such as the earthworm who grows a new head following bisection), humans are sadly incapable of such feats. Our means of recovery following tissue damage consists largely of repair rather than pure regeneration. Thousands of times in our lives, a meticulously scripted but unseen wound healing drama plays, with cells serving as actors, extracellular matrix as the setting and growth factors as the means of communication. This article briefly reviews the cells involved in tissue repair, their signaling and proliferation mechanisms and the function of the extracellular matrix, then presents the actors and script for the three acts of the tissue repair drama. PMID:21220961

Pulp regeneration in a full-length human tooth root using a hierarchical nanofibrous microsphere system.

PubMed

Li, Xiangwei; Ma, Chi; Xie, Xiaohua; Sun, Hongchen; Liu, Xiaohua

2016-04-15

While pulp regeneration using tissue engineering strategy has been explored for over a decade, successful regeneration of pulp tissues in a full-length human root with a one-end seal that truly simulates clinical endodontic treatment has not been achieved. To address this challenge, we designed and synthesized a unique hierarchical growth factor-loaded nanofibrous microsphere scaffolding system. In this system, vascular endothelial growth factor (VEGF) binds with heparin and is encapsulated in heparin-conjugated gelatin nanospheres, which are further immobilized in the nanofibers of an injectable poly(l-lactic acid) (PLLA) microsphere. This hierarchical microsphere system not only protects the VEGF from denaturation and degradation, but also provides excellent control of its sustained release. In addition, the nanofibrous PLLA microsphere integrates the extracellular matrix-mimicking architecture with a highly porous injectable form, efficiently accommodating dental pulp stem cells (DPSCs) and supporting their proliferation and pulp tissue formation. Our in vivo study showed the successful regeneration of pulp-like tissues that fulfilled the entire apical and middle thirds and reached the coronal third of the full-length root canal. In addition, a large number of blood vessels were regenerated throughout the canal. For the first time, our work demonstrates the success of pulp tissue regeneration in a full-length root canal, making it a significant step toward regenerative endodontics. The regeneration of pulp tissues in a full-length tooth root canal has been one of the greatest challenges in the field of regenerative endodontics, and one of the biggest barriers for its clinical application. In this study, we developed a unique approach to tackle this challenge, and for the first time, we successfully regenerated living pulp tissues in a full-length root canal, making it a significant step toward regenerative endodontics. This study will make positive scientific impact and interest the broad and multidisciplinary readership in the dental biomaterials and craniofacial tissue engineering community. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

CFTR expression and organ damage in cystic fibrosis

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

Tizzano, E.; Chitayat, D.; Buchwald, M.

1994-09-01

To assist our understanding of the origin of organ damage caused by cystic fibrosis (CF) disease, we have analyzed the pattern of expression of the CF gene (CFTR). mRNA in situ hybridization analysis was carried out in human fetal, newborn, infant and adult tissues and the abundance of the mRNA was correlated with the known pathology at the various stages of human development. Analysis of the pattern of expression indicates a constitutive level of mRNA in gastrointestinal tissues starting during early development and maintained throughout life. Prenatal respiratory tissues show qualitative and quantitative major differences in comparison to postnatal lungmore » samples. Male reproductive tissues show high levels of expression in the head of the epididymis compared with the rest of the male ducts. Female reproductive tissues show a variable pattern of expression at different stages during fetal development and during puberty probably due to changes in hormonal levels. Gastrointestinal and male reproductive tissues have a consistent pathology at birth, whereas no lung abnormalities have been described in newborns affected by CF. Our results show that there is no exact correlations between organ damage present at birth and the degree of CFTR expression. To explain these observations, we hypothesize that the pathogenesis of organ damage in CF depend on at least three factors: the rate of CFTR-mediated fluid secretion, differences in genotype and environmental factors, such as the amount of macromolecules in the lumen of the ducts. This last element predicts that damage will occur in tissues with high protein loads and low flow rates (e.g. pancreas, epididymis), where the absence of CFTR function leads to obstruction and pathology. Organs that express CFTR but with no significant damage (e.g. prenatal lung, female reproductive tissues), will have a low protein load and a high flow rates.« less

Synthesis of embryonic tendon-like tissue by human marrow stromal/mesenchymal stem cells requires a three-dimensional environment and transforming growth factor β3.

PubMed

Kapacee, Zoher; Yeung, Ching-Yan Chloé; Lu, Yinhui; Crabtree, David; Holmes, David F; Kadler, Karl E

2010-10-01

Tendon-like tissue generated from stem cells in vitro has the potential to replace tendons and ligaments lost through injury and disease. However, thus far, no information has been available on the mechanism of tendon formation in vitro and how to accelerate the process. We show here that human mesenchymal stem cells (MSCs) and bone marrow-derived mononuclear cells (BM-MNCs) can generate tendon-like tissue in 7days mediated by transforming growth factor (TGF) β3. MSCs cultured in fixed-length fibrin gels spontaneously synthesized narrow-diameter collagen fibrils and exhibited fibripositors (actin-rich, collagen fibril-containing plasma membrane protrusions) identical to those that occur in embryonic tendon. In contrast, BM-MNCs did not synthesize tendon-like tissue under these conditions. We performed real-time PCR analysis of MSCs and BM-MNCs. MSCs upregulated genes encoding type I collagen, TGFβ3, and Smad2 at the time of maximum contraction of the tendon-like tissue (7days). Western blot analysis showed phosphorylation of Smad2 at maximum contraction. The TGFβ inhibitor SB-431542, blocked the phosphorylation of Smad2 and stopped the formation of tendon-like tissue. Quantitative PCR showed that BM-MNCs expressed very low levels of TGFβ3 compared to MSCs. Therefore we added exogenous TGFβ3 protein to BM-MNCs in fibrin gels, which resulted in phosphorylation of Smad2, synthesis of collagen fibrils, the appearance of fibripositors at the plasma membrane, and the formation of tendon-like tissue. In conclusion, MSCs that self-generate TGFβ signaling or the addition of TGFβ3 protein to BM-MNCs in fixed-length fibrin gels spontaneously make embryonic tendon-like tissue in vitro within 7days. Copyright © 2010 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.

Hemoglobin enhances tissue factor expression on human malignant cells.

PubMed

Siddiqui, F A; Amirkhosravi, A; Amaya, M; Meyer, T; Biggerstaff, J; Desai, H; Francis, J L

2001-04-01

Tissue Factor (TF) is a transmembrane glycoprotein that complexes with factor VII/activated factor VII to initiate blood coagulation. TF may be expressed on the surface of various cells including monocytes and endothelial cells. Over-expression of TF in human tumor cell lines promotes metastasis. We recently showed that hemoglobin (Hb) forms a specific complex with TF purified from human malignant melanoma cells and enhances its procoagulant activity (PCA). To further study this interaction, we examined the effect of Hb on the expression of TF on human malignant (TF+) cells and KG1 myeloid leukemia (TF-) cells. Human melanoma A375 and J82 bladder carcinoma cells, which express TF at moderate and relatively high levels, respectively, were incubated with varying concentrations (0-1.5 mg/ml) of Hb. After washing, cells were analyzed for Hb binding and TF expression using flow cytometry and confocal microscopy. Hb bound to the cells in a concentration-dependent manner, and increased both TF expression and PCA. The human A375 malignant melanoma cells incubated with Hb (1 mg/ml) expressed up to six times more TF antigen than cells without Hb. This increase in TF expression and PCA of intact cells incubated with Hb was significantly inhibited by cycloheximide at a concentration of 10 microg/ml (P < 0.01). An increase in total cellular TF antigen content was demonstrated by specific immunoassay. In contrast, Hb (5 mg/ml) did not induce TF expression and PCA on KG1 cells as determined by flow cytometry and TF (FXAA) activity. We conclude that Hb specifically binds to TF-bearing malignant cells and increases their PCA. This effect seems to be at least partly due to de novo synthesis of TF and increased surface expression. However, the exact mechanism by which Hb binds and upregulates TF expression remains to be determined.

Protein-releasing polymeric scaffolds induce fibrochondrocytic differentiation of endogenous cells for knee meniscus regeneration in sheep.

PubMed

Lee, Chang H; Rodeo, Scott A; Fortier, Lisa Ann; Lu, Chuanyong; Erisken, Cevat; Mao, Jeremy J

2014-12-10

Regeneration of complex tissues, such as kidney, liver, and cartilage, continues to be a scientific and translational challenge. Survival of ex vivo cultured, transplanted cells in tissue grafts is among one of the key barriers. Meniscus is a complex tissue consisting of collagen fibers and proteoglycans with gradient phenotypes of fibrocartilage and functions to provide congruence of the knee joint, without which the patient is likely to develop arthritis. Endogenous stem/progenitor cells regenerated the knee meniscus upon spatially released human connective tissue growth factor (CTGF) and transforming growth factor-β3 (TGFβ3) from a three-dimensional (3D)-printed biomaterial, enabling functional knee recovery. Sequentially applied CTGF and TGFβ3 were necessary and sufficient to propel mesenchymal stem/progenitor cells, as a heterogeneous population or as single-cell progenies, into fibrochondrocytes that concurrently synthesized procollagens I and IIα. When released from microchannels of 3D-printed, human meniscus scaffolds, CTGF and TGFβ3 induced endogenous stem/progenitor cells to differentiate and synthesize zone-specific type I and II collagens. We then replaced sheep meniscus with anatomically correct, 3D-printed scaffolds that incorporated spatially delivered CTGF and TGFβ3. Endogenous cells regenerated the meniscus with zone-specific matrix phenotypes: primarily type I collagen in the outer zone, and type II collagen in the inner zone, reminiscent of the native meniscus. Spatiotemporally delivered CTGF and TGFβ3 also restored inhomogeneous mechanical properties in the regenerated sheep meniscus. Survival and directed differentiation of endogenous cells in a tissue defect may have implications in the regeneration of complex (heterogeneous) tissues and organs. Copyright © 2014, American Association for the Advancement of Science.

A case study of technology transfer: Rehabilitative engineering at Rancho Los Amigos Hospital. [prosthetic devices engineering

NASA Technical Reports Server (NTRS)

Hildred, W.

1973-01-01

The transfer of NASA technolgy to rehabilitative applications of artificial limbs is studied. Human factors engineering activities range from orthotic manipulators to tiny dc motors and transducers to detect and transmit voluntary control signals. It is found that bicarbon implant devices are suitable for medical equipment and artificial limbs because of their biological compatibility with human body fluids and tissues.

Local application of periodontal ligament stromal cells promotes soft tissue regeneration.

PubMed

Baik, H S; Park, J; Lee, K J; Chung, C

2014-09-01

To test the potential stimulatory effect of local application of periodontal ligament (PDL) stromal cells on soft tissue regeneration. Fluorescently labeled PDL cells outgrown from extracted human premolars or phosphate-buffered saline were locally injected to the cutaneous wounds created on mice. Soft tissue regeneration was evaluated for 14 days using photographs and histomorphometry. PDL cell engraftment was tracked with confocal microscopy. To detect the paracrine effect of the PDL cells on soft tissue regeneration, PDL cell-conditioned medium (CM) was evaluated for the concentration of secretory factors, transforming growth factor-beta 1 (TGFβ1). The effect of PDL CM on the proliferation and migration of dermal fibroblast and keratinocyte was tested using MTT assay and migration assay. The application of PDL cells significantly promoted soft tissue regeneration compared with the application of PBS. Self-replicating PDL cells were engrafted into the hair follicles of the host tissue. Dermal fibroblast proliferation and keratinocyte migration were significantly enhanced by the treatment with PDL CM. Physiologically significant amount of TGFβ1 was secreted from PDL cells into the CM. Local injection of PDL cells promoted soft tissue regeneration in part by the enhancement of fibroblast proliferation and keratinocyte migration through a paracrine mechanism. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

DMSO‐ and Serum‐Free Cryopreservation of Wharton's Jelly Tissue Isolated From Human Umbilical Cord

PubMed Central

Shivakumar, Sharath Belame; Bharti, Dinesh; Subbarao, Raghavendra Baregundi; Jang, Si‐Jung; Park, Ji‐Sung; Ullah, Imran; Park, Ji‐Kwon; Byun, June‐Ho

2016-01-01

ABSTRACT The facile nature of mesenchymal stem cell (MSC) acquisition in relatively large numbers has made Wharton's jelly (WJ) tissue an alternative source of MSCs for regenerative medicine. However, freezing of such tissue using dimethyl sulfoxide (DMSO) for future use impedes its clinical utility. In this study, we compared the effect of two different cryoprotectants (DMSO and cocktail solution) on post‐thaw cell behavior upon freezing of WJ tissue following two different freezing protocols (Conventional [−1°C/min] and programmed). The programmed method showed higher cell survival rate compared to conventional method of freezing. Further, cocktail solution showed better cryoprotection than DMSO. Post‐thaw growth characteristics and stem cell behavior of Wharton's jelly mesenchymal stem cells (WJMSCs) from WJ tissue cryopreserved with a cocktail solution in conjunction with programmed method (Prog‐Cock) were comparable with WJMSCs from fresh WJ tissue. They preserved their expression of surface markers, pluripotent factors, and successfully differentiated in vitro into osteocytes, adipocytes, chondrocytes, and hepatocytes. They also produced lesser annexin‐V‐positive cells compared to cells from WJ tissue stored using cocktail solution in conjunction with the conventional method (Conv‐Cock). Real‐time PCR and Western blot analysis of post‐thaw WJMSCs from Conv‐Cock group showed significantly increased expression of pro‐apoptotic factors (BAX, p53, and p21) and reduced expression of anti‐apoptotic factor (BCL2) compared to WJMSCs from the fresh and Prog‐Cock group. Therefore, we conclude that freezing of fresh WJ tissue using cocktail solution in conjunction with programmed freezing method allows for an efficient WJ tissue banking for future MSC‐based regenerative therapies. J. Cell. Biochem. 117: 2397–2412, 2016. © 2016 The Authors. Journal of Cellular Biochemistry published by Wiley Periodicals, Inc. PMID:27038129

Radiation Fibrosis of the Vocal Fold: From Man to Mouse

PubMed Central

Johns, Michael M.; Kolachala, Vasantha; Berg, Eric; Muller, Susan; Creighton, Frances X.; Branski, Ryan C.

2013-01-01

Objectives To characterize fundamental late tissue effects in the human vocal fold following radiation therapy. To develop a murine model of radiation fibrosis to ultimately develop both treatment and prevention paradigms. Design Translational study using archived human and fresh murine irradiated vocal fold tissue. Methods 1) Irradiated vocal fold tissue from patients undergoing laryngectomy for loss of function from radiation fibrosis were identified from pathology archives. Histomorphometry, immunohistochemistry, and whole-genome microarray as well as real-time transcriptional analyses was performed. 2) Focused radiation to the head and neck was delivered to mice in a survival fashion. One month following radiation, vocal fold tissue was analyzed with histomorphometry, immunohistochemistry, and real-time PCR transcriptional analysis for selected markers of fibrosis. Results Human irradiated vocal folds demonstrated increased collagen transcription with increased deposition and disorganization of collagen in both the thyroarytenoid muscle and the superficial lamina propria. Fibronectin were increased in the superficial lamina propria. Laminin decreased in the thyroarytenoid muscle. Whole genome microarray analysis demonstrated increased transcription of markers for fibrosis, oxidative stress, inflammation, glycosaminoglycan production and apoptosis. Irradiated murine vocal folds demonstrated increases in collagen and fibronectin transcription and deposition in the lamina propria. Transforming growth factor (TGF)-β increased in the lamina propria. Conclusion Human irradiated vocal folds demonstrate molecular changes leading to fibrosis that underlie loss of vocal fold pliability that occurs in patients following laryngeal irradiation. Irradiated murine tissue demonstrates similar findings, and this mouse model may have utility in creating prevention and treatment strategies for vocal fold radiation fibrosis. PMID:23242839

Are clinical findings of systemic titanium dispersion following implantation explained by available in vitro evidence? An evidence-based analysis.

PubMed

Curtin, Justin Paul; Wang, Minji

2017-08-01

Although the presence of titanium wear particles released into tissues is known to induce local inflammation following the therapeutic implantation of titanium devices into humans, the role that titanium ions play in adverse tissue responses has received little attention. Support that ongoing titanium ion release occurs is evidenced by the presence of ionic titanium bound to transferrin in blood, and ongoing excretion in the urine of patients with titanium devices. However, as reports documenting the presence of titanium within tissues do not distinguish between particulate and ionic forms due to technical challenges, the degree to which ionic titanium is released into tissues is unknown. To determine the potential for titanium ion release into tissues, this study evaluates available in vitro evidence relating to the release of ionic titanium under physiological conditions. This is a systematic literature review of studies reporting titanium ion release into solutions from titanium devices under conditions replicating the interstitial pH and constituents. Inclusion and exclusion criteria were defined. Of 452 articles identified, titanium ions were reported in nine media relevant to human biology in seventeen studies. Only one study, using human serum replicated both physiological pH and the concentration of constituents while reporting the presence of titanium ions. While there is insufficient information to explain the factors that contribute to the presence of titanium ions in serum of humans implanted with titanium devices, currently available information suggests that areas of future inquiry include the role of transferrin and organic acids.

Plasma NOV/CCN3 Levels Are Closely Associated with Obesity in Patients with Metabolic Disorders

PubMed Central

Pakradouni, Jihane; Le Goff, Wilfried; Calmel, Claire; Antoine, Bénédicte; Villard, Elise; Frisdal, Eric; Abifadel, Marianne; Tordjman, Joan; Poitou, Christine; Bonnefont-Rousselot, Dominique; Bittar, Randa; Bruckert, Eric; Clément, Karine; Fève, Bruno; Martinerie, Cécile; Guérin, Maryse

2013-01-01

Objective Evidence points to a founder of the multifunctional CCN family, NOV/CCN3, as a circulating molecule involved in cardiac development, vascular homeostasis and inflammation. No data are available on the relationship between plasma NOV/CCN3 levels and cardiovascular risk factors in humans. This study investigated the possible relationship between plasma NOV levels and cardiovascular risk factors in humans. Methods NOV levels were measured in the plasma from 594 adults with a hyperlipidemia history and/or with lipid-lowering therapy and/or a body mass index (BMI) >30 kg/m2. Correlations were measured between NOV plasma levels and various parameters, including BMI, fat mass, and plasma triglycerides, cholesterol, glucose, and C-reactive protein. NOV expression was also evaluated in adipose tissue from obese patients and rodents and in primary cultures of adipocytes and macrophages. Results After full multivariate adjustment, we detected a strong positive correlation between plasma NOV and BMI (r = 0.36 p<0.0001) and fat mass (r = 0.33 p<0.0005). According to quintiles, this relationship appeared to be linear. NOV levels were also positively correlated with C-reactive protein but not with total cholesterol, LDL-C or blood glucose. In patients with drastic weight loss induced by Roux-en-Y bariatric surgery, circulating NOV levels decreased by 28% (p<0.02) and 48% (p<0.0001) after 3 and 6 months, respectively, following surgery. In adipose tissue from obese patients, and in human primary cultures NOV protein was detected in adipocytes and macrophages. In mice fed a high fat diet NOV plasma levels and its expression in adipose tissue were also significantly increased compared to controls fed a standard diet. Conclusion Our results strongly suggest that in obese humans and mice plasma NOV levels positively correlated with NOV expression in adipose tissue, and support a possible contribution of NOV to obesity-related inflammation. PMID:23785511

MAFB Determines Human Macrophage Anti-Inflammatory Polarization: Relevance for the Pathogenic Mechanisms Operating in Multicentric Carpotarsal Osteolysis.

PubMed

Cuevas, Víctor D; Anta, Laura; Samaniego, Rafael; Orta-Zavalza, Emmanuel; Vladimir de la Rosa, Juan; Baujat, Geneviève; Domínguez-Soto, Ángeles; Sánchez-Mateos, Paloma; Escribese, María M; Castrillo, Antonio; Cormier-Daire, Valérie; Vega, Miguel A; Corbí, Ángel L

2017-03-01

Macrophage phenotypic and functional heterogeneity derives from tissue-specific transcriptional signatures shaped by the local microenvironment. Most studies addressing the molecular basis for macrophage heterogeneity have focused on murine cells, whereas the factors controlling the functional specialization of human macrophages are less known. M-CSF drives the generation of human monocyte-derived macrophages with a potent anti-inflammatory activity upon stimulation. We now report that knockdown of MAFB impairs the acquisition of the anti-inflammatory profile of human macrophages, identify the MAFB-dependent gene signature in human macrophages and illustrate the coexpression of MAFB and MAFB-target genes in CD163 + tissue-resident and tumor-associated macrophages. The contribution of MAFB to the homeostatic/anti-inflammatory macrophage profile is further supported by the skewed polarization of monocyte-derived macrophages from multicentric carpotarsal osteolysis (Online Mendelian Inheritance in Man #166300), a pathology caused by mutations in the MAFB gene. Our results demonstrate that MAFB critically determines the acquisition of the anti-inflammatory transcriptional and functional profiles of human macrophages. Copyright © 2017 by The American Association of Immunologists, Inc.

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.

Connective tissue growth factor stimulates the proliferation, migration and differentiation of lung fibroblasts during paraquat-induced pulmonary fibrosis.

PubMed

Yang, Zhizhou; Sun, Zhaorui; Liu, Hongmei; Ren, Yi; Shao, Danbing; Zhang, Wei; Lin, Jinfeng; Wolfram, Joy; Wang, Feng; Nie, Shinan

2015-07-01

It is well established that paraquat (PQ) poisoning can cause severe lung injury during the early stages of exposure, finally leading to irreversible pulmonary fibrosis. Connective tissue growth factor (CTGF) is an essential growth factor that is involved in tissue repair and pulmonary fibrogenesis. In the present study, the role of CTGF was examined in a rat model of pulmonary fibrosis induced by PQ poisoning. Histological examination revealed interstitial edema and extensive cellular thickening of interalveolar septa at the early stages of poisoning. At 2 weeks after PQ administration, lung tissue sections exhibited a marked thickening of the alveolar walls with an accumulation of interstitial cells with a fibroblastic appearance. Masson's trichrome staining revealed a patchy distribution of collagen deposition, indicating pulmonary fibrogenesis. Western blot analysis and immunohistochemical staining of tissue samples demonstrated that CTGF expression was significantly upregulated in the PQ-treated group. Similarly, PQ treatment of MRC-5 human lung fibroblast cells caused an increase in CTGF in a dose-dependent manner. Furthermore, the addition of CTGF to MRC-5 cells triggered cellular proliferation and migration. In addition, CTGF induced the differentiation of fibroblasts to myofibroblasts, as was evident from increased expression of α-smooth muscle actin (α-SMA) and collagen. These findings demonstrate that PQ causes increased CTGF expression, which triggers proliferation, migration and differentiation of lung fibroblasts. Therefore, CTGF may be important in PQ-induced pulmonary fibrogenesis, rendering this growth factor a potential pharmacological target for reducing lung injury.

Human papilloma virus and survival of oropharyngeal cancer patients treated with surgery and adjuvant radiotherapy.

PubMed

Broglie, Martina A; Soltermann, Alex; Haile, Sarah R; Huber, Gerhard F; Stoeckli, Sandro J

2015-07-01

Impact of p16 protein, a surrogate marker for human papilloma virus induced cancer, p53 and EGFR as well as clinical factors on survival in a patient cohort with oropharyngeal squamous cell carcinoma (OPSCC) treated by surgical resection and adjuvant radiotherapy (RT) ± concomitant chemotherapy (CT). This is a retrospective analysis of patient's charts and tumor tissue. 57 patients were consecutively included and their tumor tissue assembled on a tissue microarray following immunohistochemical analysis. Survival times were estimated by means of Kaplan-Meier analysis. The importance of clinical and immunohistochemical factors for outcome was estimated by cox proportional hazard models. With 88% 5-year overall survival, 91% 5-year disease-specific survival and 91% 5-year disease-free survival, respectively, we found excellent survival rates in this surgically treated patient cohort of mainly advanced OPSCC (93% AJCC stage III or IV). The only factors positively influencing survival were p16 overexpression as well as p53 negativity and even more pronounced the combination of those biomarkers. Survival analysis of patients classified into three risk categories according to an algorithm based on p16, smoking, T- and N-category revealed a low, intermediate and high-risk group with significant survival differences between the low and the high-risk group. Patients with OPSCC can be successfully treated by surgery and adjuvant RT ± CT with a clear survival benefit of p16 positive, p53 negative patients. We recommend considering a combination of immunohistochemical (p16, p53) and clinical factors (smoking, T- and N-category) for risk stratification.

Quantification of phase I / II metabolizing enzyme gene expression and polycyclic aromatic hydrocarbon-DNA adduct levels in human prostate

PubMed Central

John, Kaarthik; Ragavan, Narasimhan; Pratt, M. Margaret; Singh, Paras B.; Al-Buheissi, Salah; Matanhelia, Shyam S.; Phillips, David H.; Poirier, Miriam C.; Martin, Francis L.

2008-01-01

BACKGROUND Studies of migrant populations suggest that dietary and/or environmental factors play a crucial role in the aetiology of prostatic adenocarcinoma (CaP). The human prostate consists of the peripheral zone (PZ), transition zone (TZ) and central zone (CZ); CaP occurs most often in the PZ. METHODS To investigate the notion that an underlying differential expression of phase I/II genes, and/or the presence of polycyclic aromatic hydrocarbon (PAH)-DNA adducts might explain the elevated PZ susceptibility, we examined prostate tissues (matched tissue sets consisting of PZ and TZ) from men undergoing radical retropubic prostatectomy for CaP (n=26) or cystoprostatectomy (n=1). Quantitative gene expression analysis was employed for cytochrome P450 (CYP) isoforms CYP1A1, CYP1B1 and CYP1A2, as well as N-acetyltransferase 1 and 2 (NAT1 and NAT2) and catechol-O-methyl transferase (COMT). RESULTS CYP1B1, NAT1 and COMT were expressed in all tissue sets; levels of CYP1B1 and NAT1 were consistently higher in the PZ compared to TZ. Immunohistochemistry confirmed the presence of CYP1B1 (nuclear-associated and primarily in basal epithelial cells) and NAT1. Tissue sections from 23 of these aforementioned 27 matched tissue sets were analyzed for PAH-DNA adduct levels using antiserum elicited against DNA modified with r7, t8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydro-benzo[a]pyrene (BPDE). PAH-DNA adduct levels were highest in glandular epithelial cells, but a comparison of PZ and TZ showed no significant differences. CONCLUSION Although expression of activating and/or detoxifying enzymes may be higher in the PZ, PAH-DNA adduct levels appear to be similar in both zones. Therefore, factors other than PAH-DNA adducts may be responsible for promotion of tumour formation in the human prostate. PMID:19143007

Quantitative Ultrasound Imaging Using Acoustic Backscatter Coefficients.

NASA Astrophysics Data System (ADS)

Boote, Evan Jeffery

Current clinical ultrasound scanners render images which have brightness levels related to the degree of backscattered energy from the tissue being imaged. These images offer the interpreter a qualitative impression of the scattering characteristics of the tissue being examined, but due to the complex factors which affect the amplitude and character of the echoed acoustic energy, it is difficult to make quantitative assessments of scattering nature of the tissue, and thus, difficult to make precise diagnosis when subtle disease effects are present. In this dissertation, a method of data reduction for determining acoustic backscatter coefficients is adapted for use in forming quantitative ultrasound images of this parameter. In these images, the brightness level of an individual pixel corresponds to the backscatter coefficient determined for the spatial position represented by that pixel. The data reduction method utilized rigorously accounts for extraneous factors which affect the scattered echo waveform and has been demonstrated to accurately determine backscatter coefficients under a wide range of conditions. The algorithms and procedures used to form backscatter coefficient images are described. These were tested using tissue-mimicking phantoms which have regions of varying scattering levels. Another phantom has a fat-mimicking layer for testing these techniques under more clinically relevant conditions. Backscatter coefficient images were also formed of in vitro human liver tissue. A clinical ultrasound scanner has been adapted for use as a backscatter coefficient imaging platform. The digital interface between the scanner and the computer used for data reduction are described. Initial tests, using phantoms are presented. A study of backscatter coefficient imaging of in vivo liver was performed using several normal, healthy human subjects.

Harnessing the therapeutic potential of mesenchymal stem cells in multiple sclerosis

PubMed Central

Darlington, Peter J; Boivin, Marie-Noëlle; Bar-Or, Amit

2011-01-01

Phase I clinical trials exploring the use of autologous mesenchymal stem cell (MSC) therapy for the treatment of multiple sclerosis (MS) have begun in a number of centers across the world. MS is a complex and chronic immune-mediated and neurodegenerative disease influenced by genetic susceptibility and environmental risk factors. The ideal treatment for MS would involve both attenuation of detrimental inflammatory responses, and induction of a degree of tissue protection/regeneration within the CNS. Preclinical studies have demonstrated that both human-derived and murine-derived MSCs are able to improve outcomes in the animal model of MS, experimental autoimmune encephalomyelitis. How MSCs ameliorate experimental autoimmune encephalomyelitis is being intensely investigated. One of the major mechanisms of action of MSC therapy is to inhibit various components of the immune system that contribute to tissue destruction. Emerging evidence now supports the idea that MSCs can access the CNS where they can provide protection against tissue damage, and may facilitate tissue regeneration through the production of growth factors. The prospect of cell-based therapy using MSCs has several advantages, including the relative ease with which they can be extracted from autologous bone marrow or adipose tissue and expanded in vitro to reach the purity and numbers required for transplantation, and the fact that MSC therapy has already been used in other human disease settings, such as graft-versus-host and cardiac disease, with initial reports indicating a good safety profile. This article will focus on the theoretical and practical issues relevant to considerations of MSC therapy in the context of MS. PMID:21864075

Soluble tissue factor has unique angiogenic activities that selectively promote migration and differentiation but not proliferation of endothelial cells

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

He Yingbo; Chang Guodong; Zhan Shunli

2008-06-06

The level of circulating tissue factor (TF) is up-regulated in human angiogenesis-related malignancies. However, whether circulating TF has angiogenic activities has not been determined. Soluble TF (sTF) is the main domain of circulating TF. Here, using cell migration, wound healing, and tubule formation assays, human recombinant sTF was found to significantly promote the migration and differentiation of endothelial cells. The stress fiber formation and rearrangement induced by sTF observed through immunofluorescence microscope may be responsible for the stimulatory migration effect of sTF. Nevertheless, sTF had no effects on endothelial cell proliferation. Interestingly, sTF can be internalized by endothelial cells, whichmore » implies a novel mechanism for sTF in angiogenesis. These results suggest that sTF has unique angiogenic activities and may serve as a potential therapeutic target to treat diseases associated with angiogenesis such as cancer and rheumatoid arthritis.« less

Production of colony-stimulating factor in human dental pulp fibroblasts.

PubMed

Sawa, Y; Horie, Y; Yamaoka, Y; Ebata, N; Kim, T; Yoshida, S

2003-02-01

Class II major histocompatilibity complex (MHC)-expressing cells are usually distributed in dental pulp, and it was postulated that the colony-stimulating factor (CSF) derived from dental pulp fibroblasts contributes to the migration of class II MHC-expressing cells into pulp tissue. This study aimed to investigate the CSF production of human dental pulp fibroblasts. In pulp tissue sections, granulocyte (G)-CSF was detected from normal teeth, while G-CSF, macrophage (M)-CSF, and granulocyte-macrophage (GM)-CSF were detected from teeth with dentinal caries. In cultured dental pulp fibroblasts, G-CSF was detected by immunostaining, immunoprecipitation, and ELISA, and mRNAs of G-CSF, M-CSF, and GM-CSF were detected by RT-PCR. The dental pulp fibroblasts cultured with TNF-alpha were found to increase the G-CSF expression and to produce M-CSF and GM-CSF. These findings suggest that dental pulp fibroblasts usually produce G-CSF. In the presence of TNF-alpha, dental pulp fibroblast express M-CSF and GM-CSF.

FDTD chiral brain tissue model for specific absorption rate determination under radiation from mobile phones at 900 and 1800 MHz

NASA Astrophysics Data System (ADS)

Zamorano, M.; Torres-Silva, H.

2006-04-01

A new electrodynamics model formed by chiral bioplasma, which represents the human head inner structure and makes it possible to analyse its behaviour when it is irradiated by a microwave electromagnetic field from cellular phones, is presented. The finite-difference time-domain (FDTD) numeric technique is used, which allows simulation of the electromagnetic fields, deduced with Maxwell's equations, and allows us to simulate the specific absorption rate (SAR). The results show the SAR behaviour as a function of the input power and the chirality factor. In considering the chiral brain tissue in the proposed human head model, the two more important conclusions of our work are the following: (a) the absorption of the electromagnetic fields from cellular phones is stronger, so the SAR coefficient is higher than that using the classical model, when values of the chiral factor are of order of 1; (b) 'inverse skin effect' shows up at 1800 MHz, with respect to a 900 MHz source.

Detection and quantification of mast cell, vascular endothelial growth factor, and microvessel density in human inflammatory periapical cysts and granulomas.

PubMed

Fonseca-Silva, T; Santos, C C O; Alves, L R; Dias, L C; Brito, M; De Paula, A M B; Guimarães, A L S

2012-09-01

To identify and quantify mast cell (MC), vascular endothelial growth factor (VEGF) expression and microvessel density (MVD) in human periapical cysts and granulomas. Archived samples of cysts (n = 40) and granulomas (n = 28) were sectioned and stained with toluidine blue. MCs were identified and counted. Immunohistochemical reactions were employed to evaluate the tissue expression of VEGF and vessels. MVD was estimated by determining the areas of tissue labelled with CD31 antibody. The data were analysed using the Mann-Whitney test (P < 0.05). MCs were observed in the peripheral regions of both lesion types, whilst VEGF and MVD were distributed in the stroma. The presence of MCs was higher in cysts than in granulomas (P < 0.05). VEGF and MVD expression were similar in these lesions. The highest number of MCs was observed in cysts. Moreover, the identification of VEGF and MVD was consistent with the immune mechanisms involved in the lesions. © 2012 International Endodontic Journal.

A Role for the NF-kb/Rel Transcription Factors in Human Breast Cancer

DTIC Science & Technology

1998-07-01

binding proteins present in a series of nuclear extracts from cell lines and from breast tumor tissues as well as normal mammary epithelium. Finally, we...RelA is nuclear in several examples. Our recent data on nuclear extracts of breast tumors shows that there is a significant increase in NF-KB binding...Figure 2 in the appendix). Additionally, immunoblotting of nuclear extracts versus adjacent tissue controls showed that NF-KB p50, p52 and c-Rel were

Aldo-keto Reductase Family 1 B10 as a Novel Target for Breast Cancer Treatment

DTIC Science & Technology

2010-08-01

overexpressed in tested human breast cancer tissues and mediates acetyl-CoA carboxylase-α ( ACCA ) stability, affecting fatty acid de novo synthesis and...9703; Fax. 217-545-3227; E-mail: dcao@siumed.edu Running title: AKR1B10 as a new risk factor for breast cancer Abbreviations used: ACCA , acetyl...The effect of AKR1B10 expression in cancer tissue on patient survival was evaluated with Kaplan - Meier plots, and results showed that AKR1B10

Apparent diffusion coefficient of the normal human brain for various experimental conditions

NASA Astrophysics Data System (ADS)

Moraru, Luminita; Dimitrievici, Lucian

2017-01-01

Diffusion-Weighted Magnetic Resonance Imaging (DW-MRI) is being increasingly used to assess both brain tissues and cerebrospinal fluid integrity. In this paper we study inter-site reproducibility of the apparent diffusion coefficient values for the main cerebral tissues such as gray matter, white matter and into cerebrospinal fluid and for three different stacks of slices that were spaced at L = 79.8, 84.9 and 90 mm. We assessed the impact of the attenuation factor and diffusion gradient on the results reproducibility.

NADE (p75NTR-associated cell death executor) suppresses cellular growth in vivo.

PubMed

Tong, Xiangjun; Xie, Dong; Roth, Wilfried; Reed, John; Koeffler, H Phillip

2003-06-01

NADE, a p75NTR (low-affinity neurotrophin receptor p75) -associated cell death executor, was initially cloned from a human ovarian granulosa cell cDNA library, as an unknown protein with the name, pHGR74. It was reported to mediate nerve growth factor-induced apoptosis. We independently isolated human NADE (pHGR74) from breast cancer cell lines. Expression of NADE in various human cancer cell lines, and human and murine tissues was examined. NADE was highly expressed in human endocrine-related organs and embryotic murine tissues. Forced expression of NADE in CHO (Chinese hamster ovary) cells and MDA-MB-231 human breast cancer cells had little effect on the growth of the cells in vitro, while it dramatically suppressed cellular growth in vivo. We used the yeast two-hybrid system to search for NADE binding protein. Dynactin was identified as a candidate. The p75NTR was not found in this assay and did not co-immunoprecipitate with human NADE. Furthermore, the cells stably transfected with NADE did not respond to NGF or TNF. Thus, human and murine NADE appear to have different functions.

Porphyromonas gingivalis: An Overview of Periodontopathic Pathogen below the Gum Line

PubMed Central

How, Kah Yan; Song, Keang Peng; Chan, Kok Gan

2016-01-01

Periodontal disease represents a group of oral inflammatory infections initiated by oral pathogens which exist as a complex biofilms on the tooth surface and cause destruction to tooth supporting tissues. The severity of this disease ranges from mild and reversible inflammation of the gingiva (gingivitis) to chronic destruction of connective tissues, the formation of periodontal pocket and ultimately result in loss of teeth. While human subgingival plaque harbors more than 500 bacterial species, considerable research has shown that Porphyromonas gingivalis, a Gram-negative anaerobic bacterium, is the major etiologic agent which contributes to chronic periodontitis. This black-pigmented bacterium produces a myriad of virulence factors that cause destruction to periodontal tissues either directly or indirectly by modulating the host inflammatory response. Here, this review provides an overview of P. gingivalis and how its virulence factors contribute to the pathogenesis with other microbiome consortium in oral cavity. PMID:26903954

DNA methylation links genetics, fetal environment, and an unhealthy lifestyle to the development of type 2 diabetes.

PubMed

Nilsson, Emma; Ling, Charlotte

2017-01-01

Type 2 diabetes is a complex trait with both environmental and hereditary factors contributing to the overall pathogenesis. One link between genes, environment, and disease is epigenetics influencing gene transcription and, consequently, organ function. Genome-wide studies have shown altered DNA methylation in tissues important for glucose homeostasis including pancreas, liver, skeletal muscle, and adipose tissue from subjects with type 2 diabetes compared with nondiabetic controls. Factors predisposing for type 2 diabetes including an adverse intrauterine environment, increasing age, overweight, physical inactivity, a family history of the disease, and an unhealthy diet have all shown to affect the DNA methylation pattern in target tissues for insulin resistance in humans. Epigenetics including DNA methylation may therefore improve our understanding of the type 2 diabetes pathogenesis, contribute to development of novel treatments, and be a useful tool to identify individuals at risk for developing the disease.

Anti-inflammatory and anti-fibrinolytic effects of thrombomodulin alfa through carboxypeptidase B2 in the presence of thrombin.

PubMed

Tawara, Shunsuke; Sakai, Takumi; Matsuzaki, Osamu

2016-11-01

Thrombomodulin (TM) alfa, a recombinant human soluble TM, enhances activation of pro-carboxypeptidase B2 (pro-CPB2) by thrombin. Activated pro-CPB2 (CPB2) exerts anti-inflammatory and anti-fibrinolytic activities. Therefore, TM alfa may also have anti-inflammatory and anti-fibrinolytic effects through CPB2. However, these effects of TM alfa have not been elucidated. In the present study, we investigated the effects of TM alfa on inactivation of complement component C5a as an anti-inflammatory effect and prolongation of clot lysis time as an anti-fibrinolytic effect via CPB2 in vitro. CPB2 activity and tissue factor-induced thrombin generation was examined by a chromogenic assay. C5a inactivation was evaluated by C-terminal cleavage of C5a and inhibition of C5a-induced human neutrophil migration. Clot lysis time prolongation was examined by a tissue-type plasminogen activator-induced clot lysis assay. CPB2 activity in human plasma was increased by TM alfa and thrombin in a concentration-dependent manner. TM alfa inhibited tissue factor-induced thrombin generation and enhanced pro-CPB2 activation in human plasma simultaneously. The mass spectrum of C5a treated with TM alfa, thrombin, and pro-CPB2 was decreased at 156m/z, indicating that TM alfa enhanced the processing of C5a to C-terminal-cleaved C5a, an inactive form of C5a. C5a-induced human neutrophil migration was decreased after C5a treatment with TM alfa, thrombin, and pro-CPB2. TM alfa prolonged the clot lysis time in human plasma, and this effect was completely abolished by addition of a CPB2 inhibitor. TM alfa exerts anti-inflammatory and anti-fibrinolytic effects through CPB2 in the presence of thrombin in vitro. Copyright © 2016 Elsevier Ltd. All rights reserved.

Distribution of phthalocyanines and Raman reporters in human cancerous and noncancerous breast tissue as studied by Raman imaging.

PubMed

Brozek-Pluska, Beata; Jarota, Arkadiusz; Jablonska-Gajewicz, Joanna; Kordek, Radzislaw; Czajkowski, Wojciech; Abramczyk, Halina

2012-08-01

There is a considerable interest in the developing new diagnostic techniques allowing noninvasive tracking of the progress of therapies used to treat a cancer. Raman imaging of distribution of phthalocyanine photosensitizers may open new possibilities of Photodynamic Therapy (PDT) to treat a wide range of neoplastic lesions with improved effectiveness of treatment through precise identification of malignant areas. We have employed Raman imaging and Raman spectroscopy to analyze human breast cancer tissue that interacts with photosensitizers used in the photodynamic therapy of cancer. PCA (Principal Component Analysis) has been employed to analyze various areas of the noncancerous and cancerous breast tissues. The results show that the emission spectra combined with the Raman images are very sensitive indicators to specify the aggregation state and the distribution of phthalocyanines in the cancerous and noncancerous breast tissues. Our results provide experimental evidence on the role of aggregation of phthalocyanines as a factor of particular significance in differentiation of the normal and tumourous (cancerous or benign pathology) breast tissues. We conclude that the Raman imaging reported here has a potential to be a novel and effective photodynamic therapeutic method with improved selectivity for the treatment of breast cancer.

Bladder Smooth Muscle Cells Differentiation from Dental Pulp Stem Cells: Future Potential for Bladder Tissue Engineering

PubMed Central

Song, Bing; Jiang, Wenkai; Alraies, Amr; Liu, Qian; Gudla, Vijay; Oni, Julia; Wei, Xiaoqing; Sloan, Alastair; Ni, Longxing; Agarwal, Meena

2016-01-01

Dental pulp stem cells (DPSCs) are multipotent cells capable of differentiating into multiple cell lines, thus providing an alternative source of cell for tissue engineering. Smooth muscle cell (SMC) regeneration is a crucial step in tissue engineering of the urinary bladder. It is known that DPSCs have the potential to differentiate into a smooth muscle phenotype in vitro with differentiation agents. However, most of these studies are focused on the vascular SMCs. The optimal approaches to induce human DPSCs to differentiate into bladder SMCs are still under investigation. We demonstrate in this study the ability of human DPSCs to differentiate into bladder SMCs in a growth environment containing bladder SMCs-conditioned medium with the addition of the transforming growth factor beta 1 (TGF-β1). After 14 days of exposure to this medium, the gene and protein expression of SMC-specific marker (α-SMA, desmin, and calponin) increased over time. In particular, myosin was present in differentiated cells after 11 days of induction, which indicated that the cells differentiated into the mature SMCs. These data suggested that human DPSCs could be used as an alternative and less invasive source of stem cells for smooth muscle regeneration, a technology that has applications for bladder tissue engineering. PMID:26880982

Biomimetic 3D tissue printing for soft tissue regeneration.

PubMed

Pati, Falguni; Ha, Dong-Heon; Jang, Jinah; Han, Hyun Ho; Rhie, Jong-Won; Cho, Dong-Woo

2015-09-01

Engineered adipose tissue constructs that are capable of reconstructing soft tissue with adequate volume would be worthwhile in plastic and reconstructive surgery. Tissue printing offers the possibility of fabricating anatomically relevant tissue constructs by delivering suitable matrix materials and living cells. Here, we devise a biomimetic approach for printing adipose tissue constructs employing decellularized adipose tissue (DAT) matrix bioink encapsulating human adipose tissue-derived mesenchymal stem cells (hASCs). We designed and printed precisely-defined and flexible dome-shaped structures with engineered porosity using DAT bioink that facilitated high cell viability over 2 weeks and induced expression of standard adipogenic genes without any supplemented adipogenic factors. The printed DAT constructs expressed adipogenic genes more intensely than did non-printed DAT gel. To evaluate the efficacy of our printed tissue constructs for adipose tissue regeneration, we implanted them subcutaneously in mice. The constructs did not induce chronic inflammation or cytotoxicity postimplantation, but supported positive tissue infiltration, constructive tissue remodeling, and adipose tissue formation. This study demonstrates that direct printing of spatially on-demand customized tissue analogs is a promising approach to soft tissue regeneration. Copyright © 2015 Elsevier Ltd. All rights reserved.

Molecular biology and genetics of embryonic eyelid development.

PubMed

Rubinstein, Tal J; Weber, Adam C; Traboulsi, Elias I

2016-09-01

The embryology of the eyelid is a complex process that includes interactions between the surface ectoderm and mesenchymal tissues. In the mouse and human, the eyelids form and fuse before birth; they open prenatally in the human and postnatally in the mouse. In the mouse, cell migration is stimulated by different growth factors such as FGF10, TGF-α, Activin B, and HB-EGF. These growth factors modulate downstream BMP4 signaling, the ERK cascade, and JNK/c-JUN. Several mechanisms, such as the Wnt/β-catenin signaling pathway, may inhibit and regulate eyelid fusion. Eyelid opening, on the other hand, is driven by the BMP/Smad signaling system. Several human genetic disorders result from dysregulation of the above molecular pathways.

Concise Review: Translating Regenerative Biology into Clinically Relevant Therapies: Are We on the Right Path?

PubMed Central

2017-01-01

Abstract Despite approaches in regenerative medicine using stem cells, bio‐engineered scaffolds, and targeted drug delivery to enhance human tissue repair, clinicians remain unable to regenerate large‐scale, multi‐tissue defects in situ. The study of regenerative biology using mammalian models of complex tissue regeneration offers an opportunity to discover key factors that stimulate a regenerative rather than fibrotic response to injury. For example, although primates and rodents can regenerate their distal digit tips, they heal more proximal amputations with scar tissue. Rabbits and African spiny mice re‐grow tissue to fill large musculoskeletal defects through their ear pinna, while other mammals fail to regenerate identical defects and instead heal ear holes through fibrotic repair. This Review explores the utility of these comparative healing models using the spiny mouse ear pinna and the mouse digit tip to consider how mechanistic insight into reparative regeneration might serve to advance regenerative medicine. Specifically, we consider how inflammation and immunity, extracellular matrix composition, and controlled cell proliferation intersect to establish a pro‐regenerative microenvironment in response to injuries. Understanding how some mammals naturally regenerate complex tissue can provide a blueprint for how we might manipulate the injury microenvironment to enhance regenerative abilities in humans. Stem Cells Translational Medicine 2018;7:220–231 PMID:29271610

Human adipose-derived stem cell spheroid treated with photobiomodulation irradiation accelerates tissue regeneration in mouse model of skin flap ischemia.

PubMed

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

2017-11-01

Skin flap grafting is a form of transplantation widely used in plastic surgery. However, ischemia/reperfusion injury is the main factor which reduces the survival rate of flaps following grafting. We investigated whether photobiomodulation (PBM) precondition prior to human adipose-derived stromal cell (hASC) spheroid (PBM-spheroid) transplantation improved skin tissue functional recovery by the stimulation of angiogenesis and tissue regeneration in skin flap of mice. The LED had an emission wavelength peaked at 660 ± 20 nm (6 J/cm 2 , 10 mW/cm 2 ). The expression of angiogenic growth factors in PBM-spheroid hASCs was much greater than that of not-PBM-treated spheroid or monolayer-cultured hASCs. From immunochemical staining analysis, the hASCs of PBM-spheroid were CD31 + , KDR + , and CD34 + , whereas monolayer-cultured hASCs were negative for these markers. To evaluate the therapeutic effect of hASC PBM-spheroid in vivo, PBS, monolayer-cultured hASCs, and not-PBM-spheroid were transplanted into a skin flap model. The animals were observed for 14 days. The PBM-spheroid hASCs transplanted into the skin flap ischemia differentiated into endothelial cells and remained differentiated. Transplantation of PBM-spheroid hASCs into the skin flap ischemia significantly elevated the density of vascular formations through angiogenic factors released by the skin flap ischemia and enhanced tissue regeneration at the lesion site. Consistent with these results, the transplantation of PBM-spheroid hASCs significantly improved functional recovery compared with PBS, monolayer-cultured hASCs, and not-PBM-spheroid treatment. These findings suggest that transplantation of PBM-spheroid hASCs may be an effective stem cell therapy for the treatment of skin flap ischemia.

Obesity does not promote tumorigenesis of localized patient-derived prostate cancer xenografts

PubMed Central

Ascui, Natasha; Frydenberg, Mark; Risbridger, Gail P.; Taylor, Renea A.; Watt, Matthew J.

2016-01-01

There are established epidemiological links between obesity and the severity of prostate cancer. We directly tested this relationship by assessing tumorigenicity of patient-derived xenografts (PDXs) of moderate-grade localized prostate cancer in lean and obese severe combined immunodeficiency (SCID) mice. Mice were rendered obese and insulin resistant by high-fat feeding for 6 weeks prior to transplantation, and PDXs were assessed 10 weeks thereafter. Histological analysis of PDX grafts showed no differences in tumor pathology, prostate-specific antigen, androgen receptor and homeobox protein Nkx-3.1 expression, or proliferation index in lean versus obese mice. Whilst systemic obesity per se did not promote prostate tumorigenicity, we next asked whether the peri-prostatic adipose tissue (PPAT), which covers the prostate anteriorly, plays a role in prostate tumorigenesis. In vitro studies in a cellularized co-culture model of stromal and epithelial cells demonstrated that factors secreted from human PPAT are pro-tumorigenic. Accordingly, we recapitulated the prostate-PPAT spatial relationship by co-grafting human PPAT with prostate cancer in PDX grafts. PDX tissues were harvested 10 weeks after grafting, and histological analysis revealed no evidence of enhanced tumorigenesis with PPAT compared to prostate cancer grafts alone. Altogether, these data demonstrate that prostate cancer tumorigenicity is not accelerated in the setting of diet-induced obesity or in the presence of human PPAT, prompting the need for further work to define the at-risk populations of obesity-driven tumorigenesis and the biological factors linking obesity, adipose tissue and prostate cancer pathogenesis. PMID:27351281

MicroRNA-300 targets hypoxia inducible factor-3 alpha to inhibit tumorigenesis of human non-small cell lung cancer.

PubMed

Zhang, Y; Guo, Y; Yang, C; Zhang, S; Zhu, X; Cao, L; Nie, W; Yu, H

2017-01-01

Non-small cell lung cancer (NSCLC) is one of the most deadly human cancers. MicroRNA-300 acts as both tumor promoter and suppressor in different types of cancer. Here, we try to identify the function of microRNA-300 in human NSCLC. We compared MicroRNA-300 levels between tumor tissues versus paired adjacent non-tumor lung tissues from NSCLC patients, and in NSCLC versus normal lung cell lines. Effects of microRNA-300 on cell proliferation, invasion and migration were examined in vitro, and on tumor growth in vivo using a xenograft mouse model. Potential mRNA targets of microRNA-300 were predicted and underlying mechanism was explored. MicroRNA-300 expression was lower in both NSCLC tissues and cell lines. Overexpression of microRNA-300 inhibited proliferation, invasion and migration of NSCLC cells in vitro, and tumor growth in vivo. MicroRNA-300 could directly bind to the 3'-UTR of hypoxia inducible factor-3 alpha (HIF3α) mRNA, and inhibit both its mRNA and protein expressions. Restoring HIF3α expression could rescue the inhibitory effects of microRNA-300 on tumorigenesis of NSCLC both in vitro and in vivo. MicroRNA-300 is a tumor suppressor microRNA in NSCLC by downregulating HIF3α expression. Both microRNA-300 and HIF3α may serve as potential therapeutic targets in NSCLC treatment.

Feasibility study of the Fab fragment of a monoclonal antibody against tissue factor as a diagnostic tool.

PubMed

Tsumura, Ryo; Sato, Ryuta; Furuya, Fumiaki; Koga, Yoshikatsu; Yamamoto, Yoshiyuki; Fujiwara, Yuki; Yasunaga, Masahiro; Matsumura, Yasuhiro

2015-12-01

Tissue factor (TF) is expressed strongly in various types of cancer, especially cancers that are often refractory to treatment, such as pancreatic cancer. In this study, we compared the differences in the biophysical and pharmacological properties of whole IgG and the Fab fragment of anti-human TF monoclonal antibody (1849 antibodies), in order to determine their suitability for application in the diagnosis and treatment of cancers. In the biophysical examination, we investigated the characteristics of 1849-whole IgG and 1849-Fab by SPR sensing and confocal fluorescence microscopy analysis using recombinant human TF antigen and TF-overexpressing human pancreatic cancer cell line, BxPC3, respectively. After conjugation with Alexa-Flour-647, in vivo imaging was conducted in mice bearing BxPC3 xenograft tumors. Furthermore, the distribution of the conjugates in tumors and major organs was evaluated by ex vivo study. The in vitro experiments showed that 1849 antibodies had high affinity against TF antigen. In addition, 1849-Fab showed a faster dissociation rate from the antigen than 1849-whole IgG. In mice, 1849-Fab-Alexa-Flour-647 showed rapid renal clearance and faster tumor accumulation, achieving a high contrast signal over nearby normal tissues in the early phase and enhanced tumor penetration after administration. On the other hand, 1849-whole IgG-Alexa-Flour-647 showed slow clearance from the blood and sustained high tumor accumulation. These results suggest that 1849-Fab may be a useful tool for pancreatic cancer diagnosis.

Compound parabolic concentrator probe for efficient light collection in spectroscopy of biological tissue

NASA Astrophysics Data System (ADS)

Tanaka, Kazunori; Pacheco, Marcos T. T.; Brennan, James F., III; Itzkan, Irving; Berger, Andrew J.; Dasari, Ramachandra R.; Feld, Michael S.

1996-02-01

We describe a compound parabolic concentrator (CPC)-based probe for enhanced signal collection in the spectroscopy of biological tissues. Theoretical considerations governing signal enhancement compared with conventional collection methods are given. A ray-tracing program was used to analyze the throughput of CPC's with shape deviations and surface imperfections. A modified CPC shape with 99% throughput was discovered. A 4.4-mm-long CPC was manufactured and incorporated into an optical fiber-based near-infrared Raman spectrometer system. For human tissue samples, light collection was enhanced by a factor of 7 compared with collection with 0.29-NA optical fibers.

Tissue-engineered microenvironment systems for modeling human vasculature.

PubMed

Tourovskaia, Anna; Fauver, Mark; Kramer, Gregory; Simonson, Sara; Neumann, Thomas

2014-09-01

The high attrition rate of drug candidates late in the development process has led to an increasing demand for test assays that predict clinical outcome better than conventional 2D cell culture systems and animal models. Government agencies, the military, and the pharmaceutical industry have started initiatives for the development of novel in-vitro systems that recapitulate functional units of human tissues and organs. There is growing evidence that 3D cell arrangement, co-culture of different cell types, and physico-chemical cues lead to improved predictive power. A key element of all tissue microenvironments is the vasculature. Beyond transporting blood the microvasculature assumes important organ-specific functions. It is also involved in pathologic conditions, such as inflammation, tumor growth, metastasis, and degenerative diseases. To provide a tool for modeling this important feature of human tissue microenvironments, we developed a microfluidic chip for creating tissue-engineered microenvironment systems (TEMS) composed of tubular cell structures. Our chip design encompasses a small chamber that is filled with an extracellular matrix (ECM) surrounding one or more tubular channels. Endothelial cells (ECs) seeded into the channels adhere to the ECM walls and grow into perfusable tubular tissue structures that are fluidically connected to upstream and downstream fluid channels in the chip. Using these chips we created models of angiogenesis, the blood-brain barrier (BBB), and tumor-cell extravasation. Our angiogenesis model recapitulates true angiogenesis, in which sprouting occurs from a "parent" vessel in response to a gradient of growth factors. Our BBB model is composed of a microvessel generated from brain-specific ECs within an ECM populated with astrocytes and pericytes. Our tumor-cell extravasation model can be utilized to visualize and measure tumor-cell migration through vessel walls into the surrounding matrix. The described technology can be used to create TEMS that recapitulate structural, functional, and physico-chemical elements of vascularized human tissue microenvironments in vitro. © 2014 by the Society for Experimental Biology and Medicine.

Optimization of culture conditions for stem cells derived from human anterior cruciate ligament and bone marrow.

PubMed

Cheng, Ming-Te; Liu, Chien-Lin; Chen, Tain-Hsiung; Lee, Oscar K

2014-01-01

Tissue engineering with stem cells is a fascinating approach for treating anterior cruciate ligament (ACL) injuries. In our previous study, stem cells isolated from the human anterior cruciate ligament were shown to possess extensive proliferation and differentiation capabilities when treated with specific growth factors. However, optimal culture conditions and the usefulness of fetal bovine serum (FBS) as a growth factor in in vitro culture systems are yet to be determined. In this study, we compared the effects of different culture media containing combinations of various concentrations of FBS and the growth factors basic fibroblastic growth factor (bFGF) and transforming growth factor-β1 (TGF-β1) on the proliferation and differentiation of ligament-derived stem cells (LSCs) and bone marrow mesenchymal stem cells (BMSCs). We found that α-MEM plus 10% FBS and bFGF was able to maintain both LSCs and BMSCs in a relatively undifferentiated state but with lower major extracellular matrix (ECM) component gene expression and protein production, which is beneficial for stem cell expansion. However, the differentiation and proliferation potentials of LSCs and BMSCs were increased when cultured in MesenPRO, a commercially available stem cell medium containing 2% FBS. MesenPRO in conjunction with TGF-β1 had the greatest ability to induce the differentiation of BMSCs and LSCs to ligament fibroblasts, which was evidenced by the highest ligamentous ECM gene expression and protein production. These results indicate that culture media and growth factors play a very important role in the success of tissue engineering. With α-MEM plus 10% FBS and bFGF, rapid proliferation of stem cells can be achieved. In this study, MesenPRO was able to promote differentiation of both LSCs and BMSCs to ligament fibroblasts. Differentiation was further increased by TGF-β1. With increasing understanding of the effects of different culture media and growth factors, manipulation of stem cells in the desired direction for ligament tissue engineering can be achieved.

Immunohistochemical profile of cytokines and growth factors expressed in vestibular schwannoma and in normal vestibular nerve tissue.

PubMed

Taurone, Samanta; Bianchi, Enrica; Attanasio, Giuseppe; Di Gioia, Cira; Ierinó, Rocco; Carubbi, Cecilia; Galli, Daniela; Pastore, Francesco Saverio; Giangaspero, Felice; Filipo, Roberto; Zanza, Christian; Artico, Marco

2015-07-01

Vestibular schwannomas, also known as acoustic neuromas, are benign tumors, which originate from myelin-forming Schwann cells. They develop in the vestibular branch of the eighth cranial nerve in the internal auditory canal or cerebellopontine angle. The clinical progression of the condition involves slow and progressive growth, eventually resulting in brainstem compression. The objective of the present study was to investigate the expression level and the localization of the pro-inflammatory cytokines, transforming growth factor-β1 (TGF-β1) interleukin (IL)-1β, IL-6 and tumor necrosis factor-α (TNF-α), as well as the adhesion molecules, intracellular adhesion molecule-1 and vascular endothelial growth factor (VEGF), in order to determine whether these factors are involved in the transformation and development of human vestibular schwannoma. The present study investigated whether changes in inflammation are involved in tumor growth and if so, the mechanisms underlying this process. The results of the current study demonstrated that pro-inflammatory cytokines, including TGF-β1, IL-1β and IL-6 exhibited increased expression in human vestibular schwannoma tissue compared with normal vestibular nerve samples. TNF-α was weakly expressed in Schwann cells, confirming that a lower level of this cytokine is involved in the proliferation of Schwann cells. Neoplastic Schwann cells produce pro-inflammatory cytokines that may act in an autocrine manner, stimulating cellular proliferation. In addition, the increased expression of VEGF in vestibular schwannoma compared with that in normal vestibular nerve tissue, suggests that this factor may induce neoplastic growth via the promotion of angiogenesis. The present findings suggest that inflammation may promote angiogenesis and consequently contribute to tumor progression. In conclusion, the results of the present study indicated that VEGF and pro-inflammatory cytokines may be potential therapeutic targets in vestibular schwannoma. Further studies are necessary to confirm the involvement of these factors in the growth of neoplasms and to develop inhibitors of pro-inflammatory cytokines as a potential treatment option in the future.

Microfluidic extraction and microarray detection of biomarkers from cancer tissue slides

NASA Astrophysics Data System (ADS)

Nguyen, H. T.; Dupont, L. N.; Jean, A. M.; Géhin, T.; Chevolot, Y.; Laurenceau, E.; Gijs, M. A. M.

2018-03-01

We report here a new microfluidic method allowing for the quantification of human epidermal growth factor receptor 2 (HER2) expression levels from formalin-fixed breast cancer tissues. After partial extraction of proteins from the tissue slide, the extract is routed to an antibody (Ab) microarray for HER2 titration by fluorescence. Then the HER2-expressing cell area is evaluated by immunofluorescence (IF) staining of the tissue slide and used to normalize the fluorescent HER2 signal measured from the Ab microarray. The number of HER2 gene copies measured by fluorescence in situ hybridization (FISH) on an adjacent tissue slide is concordant with the normalized HER2 expression signal. This work is the first study implementing biomarker extraction and detection from cancer tissue slides using microfluidics in combination with a microarray system, paving the way for further developments towards multiplex and precise quantification of cancer biomarkers.

TISSUE-Tregs

PubMed Central

Panduro, Marisella; Benoist, Christophe; Mathis, Diane

2016-01-01

The immune system is responsible for defending an organism against the myriad of microbial invaders it constantly confronts. It has become increasingly clear that the immune system has a second major function: the maintenance of organismal homeostasis. Foxp3+CD4+ regulatory T cells (Tregs) are important contributors to both of these critical activities, defense being the primary purview of Tregs circulating through lymphoid organs, and homeostasis ensured mainly by their counterparts residing in parenchymal tissues. This review focuses on so-called tissue Tregs. We first survey existing information on the phenotype, function, sustaining factors, and human equivalents of the three best-characterized tissue-Treg populations—those operating in visceral adipose tissue, skeletal muscle, and the colonic lamina propria. We then attempt to distill general principles from this body of work—as concerns the provenance, local adaptation, molecular sustenance, and targets of action of tissue Tregs, in particular. PMID:27168246

Comparison Between Human and Porcine Thromboelastograph Parameters in Response to Ex-Vivo Changes to Platelets, Plasma, and Red Blood Cells

DTIC Science & Technology

2013-01-01

diluted with lactated Ringer’s solution. We demonstrated that the major factor affecting the MA and angle was the platelet count. In fact, reducing...with an accelerant, either kaolin or tissue factor or both as in the case of ‘rapid’ TEG. The TEG tracing represents the cell-based theory of...There are claims in the trauma literature that the pro- longation of the R-time reflects clotting factor deficiency or dilution, prolongation of K

miRNA-205 affects infiltration and metastasis of breast cancer

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

Wang, Zhouquan; Department of Tumor, SenGong Hospital of Shaanxi, Xi’an 710300; Liao, Hehe

2013-11-08

Highlights: •We detected expression of miR-205 in breast cancer cell lines and tissue samples. •We suggest miR-205 is downregulated in human breast cancer tissues and MCF7 cells. •We suggest the lower expression of miR-205 play a role in breast cancer onset. •These data suggest that miR-205 directly targets HER3 in human breast cancer. -- Abstract: Background: An increasing number of studies have shown that miRNAs are commonly deregulated in human malignancies, but little is known about the function of miRNA-205 (miR-205) in human breast cancer. The present study investigated the influence of miR-205 on breast cancer malignancy. Methods: The expressionmore » level of miR-205 in the MCF7 breast cancer cell line was determined by quantitative (q)RT-PCR. We then analyzed the expression of miR-205 in breast cancer and paired non-tumor tissues. Finally, the roles of miR-205 in regulating tumor proliferation, apoptosis, migration, and target gene expression were studied by MTT assay, flow cytometry, qRT-PCR, Western blotting and luciferase assay. Results: miR-205 was downregulated in breast cancer cells or tissues compared with normal breast cell lines or non-tumor tissues. Overexpression of miR-205 reduced the growth and colony-formation capacity of MCF7 cells by inducing apoptosis. Overexpression of miR-205 inhibited MCF7 cell migration and invasiveness. By bioinformation analysis, miR-205 was predicted to bind to the 3′ untranslated regions of human epidermal growth factor receptor (HER)3 mRNA, and upregulation of miR-205 reduced HER3 protein expression. Conclusion: miR-205 is a tumor suppressor in human breast cancer by post-transcriptional inhibition of HER3 expression.« less

Electrical stimulation: a novel tool for tissue engineering.

PubMed

Balint, Richard; Cassidy, Nigel J; Cartmell, Sarah H

2013-02-01

New advances in tissue engineering are being made through the application of different types of electrical stimuli to influence cell proliferation and differentiation. Developments made in the last decade have allowed us to improve the structure and functionality of tissue-engineered products through the use of growth factors, hormones, drugs, physical stimuli, bioreactor use, and two-dimensional (2-D) and three-dimensional (3-D) artificial extracellular matrices (with various material properties and topography). Another potential type of stimulus is electricity, which is important in the physiology and development of the majority of all human tissues. Despite its great potential, its role in tissue regeneration and its ability to influence cell migration, orientation, proliferation, and differentiation has rarely been considered in tissue engineering. This review highlights the importance of endogenous electrical stimulation, gathering the current knowledge on its natural occurrence and role in vivo, discussing the novel methods of delivering this stimulus and examining its cellular and tissue level effects, while evaluating how the technique could benefit the tissue engineering discipline in the future.

Genome and transcriptome studies of the protozoan parasites Trypanosoma cruzi and Giardia intestinalis

NASA Astrophysics Data System (ADS)

Farran, Alexandra J. E.

Vocal fold (VF) diseases and disorders are difficult to treat surgically or therapeutically. Tissue engineering offers an alternative strategy for the restoration of functional VF. In this work, we have developed tissue engineering methodologies for the functional reconstruction of VF. As a first step, the structure, composition and mechanical properties of native VF tissues have been investigated. In pigs ranging from fetal to 2+ years old, the VF structure and viscoelastic properties were found to be age-dependent. Adult tissues were more organized, displaying a denser lamina propria, and mature elastin fibers compared to fetal tissues, resulting in higher storage moduli. Secondly, biomimetic scaffolds which recaptured the mechanical properties of the native VF were developed. Chemically-defined collagen-hyaluronic acid (HA) composite hydrogels, and elastin-mimetic hybrid polymers (EMHPs) were successfully used as conducive 3D matrices, and 2D elastic scaffolds respectively, to in vitro static culture of fibroblasts. While the collagen-HA hydrogels allowed for in situ cell encapsulation and supported cell attachment and proliferation in 3D, the integrin-binding domain RGDSP was needed for cell proliferation on EMHPs. To emulate in vitro the mechanical environment of the native VF tissue, a dynamic culture system capable of generating vibratory stimulations at human phonation frequencies was successfully created and characterized. Gene expression analysis of fibroblasts subjected to 1 hour vibrations in 2D revealed that the expression of ECM-related genes was altered in response to changes in vibratory frequency and amplitude. Finally, expanding on our previous studies, the dynamic culture system was modified to accommodate for the long-term dynamic culture of cell-laden hydrogels. Human mesenchymal stem cells (hMSCs) encapsulated in a collagen/HA-based hydrogel, cultured in presence of connective tissue growth factor (CTGF), and subjected to high frequency vibrations were shown to respond to all three type of external factors. In summary, microenvironments such as biomimetic scaffolds, soluble factors, and mechanical stimuli are important modulator of cellular function. The strategic combination of those microenvironments into a biomimicking VF tissue engineering 3D system did not only provide an in vitro platform for the investigation of VF diseases, but also have the potential to offer alternative treatments for VF disorders.

Redundant roles of the phosphatidate phosphatase family in triacylglycerol synthesis in human adipocytes.

PubMed

Temprano, Ana; Sembongi, Hiroshi; Han, Gil-Soo; Sebastián, David; Capellades, Jordi; Moreno, Cristóbal; Guardiola, Juan; Wabitsch, Martin; Richart, Cristóbal; Yanes, Oscar; Zorzano, Antonio; Carman, George M; Siniossoglou, Symeon; Miranda, Merce

2016-09-01

In mammals, the evolutionary conserved family of Mg(2+)-dependent phosphatidate phosphatases (PAP1), involved in phospholipid and triacylglycerol synthesis, consists of lipin-1, lipin-2 and lipin-3. While mutations in the murine Lpin1 gene cause lipodystrophy and its knockdown in mouse 3T3-L1 cells impairs adipogenesis, deleterious mutations of human LPIN1 do not affect adipose tissue distribution. However, reduced LPIN1 and PAP1 activity has been described in participants with type 2 diabetes. We aimed to characterise the roles of all lipin family members in human adipose tissue and adipogenesis. The expression of the lipin family was analysed in adipose tissue in a cross-sectional study. Moreover, the effects of lipin small interfering RNA (siRNA)-mediated depletion on in vitro human adipogenesis were assessed. Adipose tissue gene expression of the lipin family is altered in type 2 diabetes. Depletion of every lipin family member in a human Simpson-Golabi-Behmel syndrome (SGBS) pre-adipocyte cell line, alters expression levels of adipogenic transcription factors and lipid biosynthesis genes in early stages of differentiation. Lipin-1 knockdown alone causes a 95% depletion of PAP1 activity. Despite the reduced PAP1 activity and alterations in early adipogenesis, lipin-silenced cells differentiate and accumulate neutral lipids. Even combinatorial knockdown of lipins shows mild effects on triacylglycerol accumulation in mature adipocytes. Overall, our data support the hypothesis of alternative pathways for triacylglycerol synthesis in human adipocytes under conditions of repressed lipin expression. We propose that induction of alternative lipid phosphate phosphatases, along with the inhibition of lipid hydrolysis, contributes to the maintenance of triacylglycerol content to near normal levels.

Preclinical development of a humanized neutralizing antibody targeting HGF.

PubMed

Kim, Hyori; Hong, Sung Hee; Kim, Jung Yong; Kim, In-Chull; Park, Young-Whan; Lee, Song-Jae; Song, Seong-Won; Kim, Jung Ju; Park, Gunwoo; Kim, Tae Min; Kim, Yun-Hee; Park, Jong Bae; Chung, Junho; Kim, In-Hoo

2017-03-24

Hepatocyte growth factor (HGF) and its receptor, cMET, play critical roles in cell proliferation, angiogenesis and invasion in a wide variety of cancers. We therefore examined the anti-tumor activity of the humanized monoclonal anti-HGF antibody, YYB-101, in nude mice bearing human glioblastoma xenografts as a single agent or in combination with temozolomide. HGF neutralization, The extracellular signal-related kinases 1 and 2 (ERK1/2) phosphorylation, and HGF-induced scattering were assessed in HGF-expressing cell lines treated with YYB-101. To support clinical development, we also evaluated the preclinical pharmacokinetics and toxicokinetics in cynomolgus monkeys, and human and cynomolgus monkey tissue was stained with YYB-101 to test tissue cross-reactivity. We found that YYB-101 inhibited cMET activation in vitro and suppressed tumor growth in the orthotopic mouse model of human glioblastoma. Combination treatment with YYB-101 and temozolomide decreased tumor growth and increased overall survival compared with the effects of either agent alone. Five cancer-related genes (TMEM119, FST, RSPO3, ROS1 and NBL1) were overexpressed in YYB-101-treated mice that showed tumor regrowth. In the tissue cross-reactivity assay, critical cross-reactivity was not observed. The terminal elimination half-life was 21.7 days. Taken together, the in vitro and in vivo data demonstrated the anti-tumor efficacy of YYB-101, which appeared to be mediated by blocking the HGF/cMET interaction. The preclinical pharmacokinetics, toxicokinetics and tissue cross-reactivity data support the clinical development of YYB-101 for advanced cancer.

Results from a horizon scan on risks associated with transplantation of human organs, tissues and cells: from donor to patient.

PubMed

Herberts, C A; Park, M V D Z; Pot, J W G A; de Vries, C G J C A

2015-03-01

The successful transplantation of human materials such as organs, tissues and cells into patients does not only depend on the benefits, but also on the mitigation of risks. To gain insight into recent publications on risks associated with the process of transferring human materials from donor to recipient we performed a horizon scan by reviewing scientific literature and news websites of 2011 on this subject. We found there is ample information on how extended donor criteria, such as donor age, affect the survival rates of organs or patients. Interestingly, gender mismatch does not appear to be a major risk factor in organ rejection. Data on risks of donor tumor transmission was very scarce; however, risk categories for various tumor types have been suggested. In order to avoid rejection, a lot of research is directed towards engineering tissues from a patient's own tissues and cells. Some but not all of these developments have reached the clinic. Developments in the field of stem cell therapy are rapid. However, many hurdles are yet to be overcome before these cells can be applied on a large scale in the clinic. The processes leading to genetic abnormalities in cells differentiated from stem cells need to be identified in order to avoid transplantation of aberrant cells. New insights have been obtained on storage and preservation of human materials, a critical step for success of their clinical use. Likewise, quality management systems have been shown to improve the quality and safety of human materials used for transplantation.

Pancreatic islet isolation variables in non-human primates (rhesus macaques).

PubMed

Andrades, P; Asiedu, C K; Gansuvd, B; Inusah, S; Goodwin, K J; Deckard, L A; Jargal, U; Thomas, J M

2008-07-01

Non-human primates (NHPs) are important preclinical models for pancreatic islet transplantation (PIT) because of their close phylogenetic and immunological relationship with humans. However, low availability of NHP tissue, long learning curves and prohibitive expenses constrain the consistency of isolated NHP islets for PIT studies. To advance preclinical studies, we attempted to identify key variables that consistently influence the quantity and quality of NHP islets. Seventy-two consecutive pancreatic islet isolations from rhesus macaques were reviewed retrospectively. A scaled down, semi-automated islet isolation method was used, and monkeys with streptozotocin-induced diabetes, weighing 3-7 kg, served as recipients for allotransplantation. We analysed the effects of 22 independent variables grouped as donor factors, surgical factors and isolation technique factors. Islet yields, success of isolation and transplantation results were used as quantitative and qualitative outcomes. In the multivariate analysis, variables that significantly affected islet yield were the type of monkey, pancreas preservation, enzyme lot and volume of enzyme delivered. The variables associated with successful isolation were the enzyme lot and volume delivered. The transplant result was correlated with pancreas preservation, enzyme lot, endotoxin levels and COBE collection method. Islet quantity and quality are highly variable between isolations. The data reviewed suggest that future NHP isolations should use bilayer preservation, infuse more than 80 ml of Liberase into the pancreas, collect non-fractioned tissue from the COBE, and strictly monitor for infection.

Cadmium, Zinc, and Selenium Levels in Carcinoma of the Human Prostate

DTIC Science & Technology

2007-04-01

tissue (4-6). Cadmium (Cd) possesses carcinogenic effect that is hormonally mediated (7, 8), and is recognized as a risk factor in development of...in prostatic cells [28], and that the carcinogenic effect of Cd can be hormonally mediated [13, 29]. Protective Factors - Selenium and Zinc Se...studies have shown that this generation of Pacific Islands people have traditional diets, eating more taro, shellfish and fresh vegetables, and

Chronic social isolation in the prairie vole induces endothelial dysfunction: implications for depression and cardiovascular disease

PubMed Central

Peuler, Jacob D.; Scotti, Melissa-Ann L.; Phelps, Laura E.; McNeal, Neal; Grippo, Angela J.

2012-01-01

Humans with depression show impaired endothelium-dependent vasodilation, one recent demonstration of which was in the form of a reduced acetylcholine (ACh)-induced relaxation of adrenergically-precontracted small arteries biopsied from older depressed patients. Results from such uses of ACh in general have been validated as the most predictive marker of endothelium-related cardiovascular diseases. Accordingly, we examined vascular reactivity to ACh in the socially isolated prairie vole, a new animal model relevant to human depression and cardiovascular disease. Thoracic aortas were carefully dissected from female prairie voles after one month of social isolation (versus pairing with a sibling). Only aortas that contracted to the adrenergic agent phenylephrine (PE) and then relaxed to ACh were evaluated. Among those, ACh-induced relaxations were significantly reduced by social isolation (p<0.05), with maximum relaxation reaching only 30% (of PE-induced precontraction) compared to 47% in aortas from paired (control) animals. Experimental removal of the endothelium from an additional set of aortic tissues abolished all ACh relaxations including that difference. In these same tissues, maximally-effective concentrations of the nitric oxide-donor nitroprusside still completely relaxed all PE-induced precontraction of the endothelial-free smooth muscle, and to the same degree in tissues from isolated versus paired animals. Finally, in the absence of PE-induced precontraction ACh did not relax but rather contracted aortic tissues, and to a significantly greater extent in tissues from socially isolated animals if the endothelium was intact (p<0.05). Thus, social isolation in the prairie vole may 1) impair normal release of protective anti-atherosclerotic factors like nitric oxide from the vascular endothelium (without altering the inherent responsiveness of the vascular smooth muscle to such factors) and 2) cause the endothelium to release contracting factors. To our knowledge this is the first demonstration of this phenomenon in an animal model of depression induced solely by social isolation. These findings have implications for understanding mechanisms involved in depression and cardiovascular disease. PMID:22469565

Keratin hydrogel carrier system for simultaneous delivery of exogenous growth factors and muscle progenitor cells1,2,3

PubMed Central

Tomblyn, Seth; Kneller, Elizabeth Pettit; Walker, Stephen J.; Ellenburg, Mary D.; Kowalczewski, Christine J.; Van Dyke, Mark; Burnett, Luke; Saul, Justin M.

2017-01-01

Ideal material characteristics for tissue engineering or regenerative medicine approaches to volumetric muscle loss (VML) include the ability to deliver cells, growth factors and molecules that support tissue formation from a system with a tunable degradation profile. Two different types of human hair-derived keratins were tested as options to fulfill these VML design requirements: (1) oxidatively extracted keratin (keratose) characterized by a lack of covalent crosslinking between cysteine residues, and (2) reductively extracted keratin (kerateine) characterized by disulfide crosslinks. Human skeletal muscle myoblasts cultured on coatings of both types of keratin had increased numbers of multinucleated cells compared to collagen or Matrigel™ and adhesion levels greater than collagen. Rheology showed elastic moduli from 102 – 105 Pa and viscous moduli from 101 – 104 Pa depending on gel concentration and keratin type. Kerateine and keratose showed differing rates of degradation due to the presence or absence of disulfide crosslinks, which likely contributed to observed differences in release profiles of several growth factors. In vivo testing in a subcutaneous mouse model showed that keratose hydrogels can be used to deliver mouse muscle progenitor cells and growth factors. Histological assessment showed minimal inflammatory responses and an increase in markers of muscle formation. PMID:25953729

A 3D thermal model to analyze the temperature changes of digits during cold stress and predict the danger of frostbite in human fingers.

PubMed

Fallahi, Amir; Reza Salimpour, Mohammad; Shirani, Ebrahim

2017-04-01

The existing computational models of frostbite injury are limited to one and two dimensional schemes. In this study, a coupled thermo-fluid model is applied to simulate a finger exposed to cold weather. The spatial variability of finger-tip temperature is compared to experimental ones to validate the model. A semi-realistic 3D model for tissue and blood vessels is used to analyze the transient heat transfer through the finger. The effect of heat conduction, metabolic heat generation, heat transport by blood perfusion, heat exchange between tissues and large vessels are considered in energy balance equations. The current model was then tested in different temperatures and air speeds to predict the danger of frostbite in humans for different gloves. Two prevalent gloves which are commonly used in cold climate are considered for investigation. The endurance time and the fraction of necrotic tissues are two main factors suggested for obtaining the response of digit tissues to different environmental conditions. Copyright © 2017 Elsevier Ltd. All rights reserved.

Prognostic significance of kynurenine 3-monooxygenase and effects on proliferation, migration, and invasion of human hepatocellular carcinoma.

PubMed

Jin, Haojie; Zhang, Yurong; You, Haiyan; Tao, Xuemei; Wang, Cun; Jin, Guangzhi; Wang, Ning; Ruan, Haoyu; Gu, Dishui; Huo, Xisong; Cong, Wenming; Qin, Wenxin

2015-06-23

Kynurenine 3-monooxygenase (KMO) is a pivotal enzyme in the kynurenine pathway of tryptophan degradation and plays a critical role in Huntington's and Alzheimer's diseases. This study aimed to examine the expression of KMO in human hepatocellular carcinoma (HCC) and investigate the relationship between its expression and prognosis of HCC patients. We first analyzed KMO expression in 120 paired HCC samples (HCC tissues vs matched adjacent non-cancerous liver tissues), and 205 clinical HCC specimens using immunohistochemistry (IHC). Kaplan-Meier survival and Cox regression analyses were executed to evaluate the prognosis of HCC. The results of IHC analysis showed that KMO expression was significantly higher in HCC tissues than that in normal liver tissues (all p < 0.05). Survival and recurrence analyses showed that KMO was an independent prognostic factor for overall survival (OS) and time to recurrence (TTR) (both p<0.01). And in vitro studies revealed that KMO positively regulated proliferation, migration, and invasion of HCC cells. These results suggest that KMO exhibits tumor-promoting effects towards HCC and it may serve as a novel prognostic marker in HCC.

Osteochondral Interface Tissue Engineering Using Macroscopic Gradients of Bioactive Signals

PubMed Central

Dormer, Nathan H.; Singh, Milind; Wang, Limin; Berkland, Cory J.; Detamore, Michael S.

2013-01-01

Continuous gradients exist at osteochondral interfaces, which may be engineered by applying spatially patterned gradients of biological cues. In the present study, a protein-loaded microsphere-based scaffold fabrication strategy was applied to achieve spatially and temporally controlled delivery of bioactive signals in three-dimensional (3D) tissue engineering scaffolds. Bone morphogenetic protein-2 and transforming growth factor-β1-loaded poly(d,llactic- co-glycolic acid) microspheres were utilized with a gradient scaffold fabrication technology to produce microsphere-based scaffolds containing opposing gradients of these signals. Constructs were then seeded with human bone marrow stromal cells (hBMSCs) or human umbilical cord mesenchymal stromal cells (hUCMSCs), and osteochondral tissue regeneration was assessed in gradient scaffolds and compared to multiple control groups. Following a 6-week cell culture, the gradient scaffolds produced regionalized extracellular matrix, and outperformed the blank control scaffolds in cell number, glycosaminoglycan production, collagen content, alkaline phosphatase activity, and in some instances, gene expression of major osteogenic and chondrogenic markers. These results suggest that engineered signal gradients may be beneficial for osteochondral tissue engineering. PMID:20379780

Induced pluripotent stem (iPS) cells from human fetal stem cells.

PubMed

Guillot, Pascale V

2016-02-01

Pluripotency defines the ability of stem cells to differentiate into all the lineages of the three germ layers and self-renew indefinitely. Somatic cells can regain the developmental potential of embryonic stem cells following ectopic expression of a set of transcription factors or, in certain circumstances, via modulation of culture conditions and supplementation with small molecule, that is, induced pluripotent stem (iPS) cells. Here, we discuss the use of fetal tissues for reprogramming, focusing in particular on stem cells derived from human amniotic fluid, and the development of chemical reprogramming. We next address the advantages and disadvantages of deriving pluripotent cells from fetal tissues and the potential clinical applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

Use of cryostat sections from snap-frozen nervous tissue for combining stereological estimates with histological, cellular, or molecular analyses on adjacent sections.

PubMed

Schmitz, C; Dafotakis, M; Heinsen, H; Mugrauer, K; Niesel, A; Popken, G J; Stephan, M; Van de Berg, W D; von Hörsten, S; Korr, H

2000-10-01

Adequate tissue preparation is essential for both modern stereological and immunohistochemical investigations. However, combining these methodologies in a single study presents a number of obstacles pertaining to optimal histological preparation. Tissue shrinkage and loss of nuclei/nucleoli from the unprotected section surfaces of unembedded tissue used for immunohistochemistry may be problematic with regard to adequate stereological design. In this study, frozen cryostat sections from hippocampal and cerebellar regions of two rat strains and cerebellar and cerebral regions from a human brain were analyzed to determine the potential impact of these factors on estimates of neuron number obtained using the optical disector. Neuronal nuclei and nucleoli were clearly present in thin sections of snap-frozen rat (3 microm) and human (6 microm) tissue, indicating that neuronal nuclei/nucleoli are not unavoidably lost from unprotected section surfaces of unembedded tissue. In order to quantify the potential impact of any nuclear loss, optical fractionator estimates of rat hippocampal pyramidal cells in areas CA1-3 and cerebellar granule and Purkinje cells were made using minimal (1 microm) upper guard zones. Estimates did not differ from data reported previously in the literature. This data indicates that cryostat sections of snap-frozen nervous tissue may successfully be used for estimating total neuronal numbers using optical disectors.

Human adipose tissue mesenchymal stromal cells and their extracellular vesicles act differentially on lung mechanics and inflammation in experimental allergic asthma.

PubMed

de Castro, Ligia Lins; Xisto, Debora Gonçalves; Kitoko, Jamil Zola; Cruz, Fernanda Ferreira; Olsen, Priscilla Christina; Redondo, Patricia Albuquerque Garcia; Ferreira, Tatiana Paula Teixeira; Weiss, Daniel Jay; Martins, Marco Aurélio; Morales, Marcelo Marcos; Rocco, Patricia Rieken Macedo

2017-06-24

Asthma is a chronic inflammatory disease that can be difficult to treat due to its complex pathophysiology. Most current drugs focus on controlling the inflammatory process, but are unable to revert the changes of tissue remodeling. Human mesenchymal stromal cells (MSCs) are effective at reducing inflammation and tissue remodeling; nevertheless, no study has evaluated the therapeutic effects of extracellular vesicles (EVs) obtained from human adipose tissue-derived MSCs (AD-MSC) on established airway remodeling in experimental allergic asthma. C57BL/6 female mice were sensitized and challenged with ovalbumin (OVA). Control (CTRL) animals received saline solution using the same protocol. One day after the last challenge, each group received saline, 10 5 human AD-MSCs, or EVs (released by 10 5  AD-MSCs). Seven days after treatment, animals were anesthetized for lung function assessment and subsequently euthanized. Bronchoalveolar lavage fluid (BALF), lungs, thymus, and mediastinal lymph nodes were harvested for analysis of inflammation. Collagen fiber content of airways and lung parenchyma were also evaluated. In OVA animals, AD-MSCs and EVs acted differently on static lung elastance and on BALF regulatory T cells, CD3 + CD4 + T cells, and pro-inflammatory mediators (interleukin [IL]-4, IL-5, IL-13, and eotaxin), but similarly reduced eosinophils in lung tissue, collagen fiber content in airways and lung parenchyma, levels of transforming growth factor-β in lung tissue, and CD3 + CD4 + T cell counts in the thymus. No significant changes were observed in total cell count or percentage of CD3 + CD4 + T cells in the mediastinal lymph nodes. In this immunocompetent mouse model of allergic asthma, human AD-MSCs and EVs effectively reduced eosinophil counts in lung tissue and BALF and modulated airway remodeling, but their effects on T cells differed in lung and thymus. EVs may hold promise for asthma; however, further studies are required to elucidate the different mechanisms of action of AD-MSCs versus their EVs.

The human interleukin-1 alpha gene is located on the long arm of chromosome 2 at band q13.

PubMed

Lafage, M; Maroc, N; Dubreuil, P; de Waal Malefijt, R; Pébusque, M J; Carcassonne, Y; Mannoni, P

1989-01-01

Interleukin-1 alpha (IL-1 alpha) and interleukin-1 beta (IL-1 beta) are two biochemically distinct, but distantly related, polypeptidic cytokines that play a key role in inflammation, immunologic reactions, and tissue repair. Recently, it has been shown that IL-1 alpha is identical to hematopoietin 1, which was described as a hematopoietic growth factor acting on early progenitor cells in synergy with other hematopoietic growth factors. In this report we discuss our use of in situ hybridization on human prometaphase cells with a human IL-1 alpha cDNA probe to localize the human IL-1 alpha gene on the proximal part of the long arm of chromosome 2 at band q13, in the same chromosomal region as the IL-1 beta gene.

Progeroid syndromes: models for stem cell aging?

PubMed

Bellantuono, I; Sanguinetti, G; Keith, W N

2012-02-01

Stem cells are responsible for tissue repair and maintenance and it is assumed that changes observed in the stem cell compartment with age underlie the concomitant decline in tissue function. Studies in murine models have highlighted the importance of intrinsic changes occurring in stem cells with age. They have also drawn the attention to other factors, such as changes in the local or systemic environment as the primary cause of stem cell dysfunction. Whilst knowledge in murine models has been advancing rapidly there has been little translation of these data to human aging. This is most likely due to the difficulties of testing the regenerative capacity of human stem cells in vivo and to substantial differences in the aging phenotype within humans. Here we summarize evidence to show how progeroid syndromes, integrated with other models, can be valuable tools in addressing questions about the role of stem cell aging in human degenerative diseases of older age and the molecular pathways involved.

[Tissue repositories for research at Sheba Medical Center(SMC].

PubMed

Cohen, Yehudit; Barshack, Iris; Onn, Amir

2013-06-01

Cancer is the number one cause of death in both genders. Breakthroughs in the understanding of cancer biology, the identification of prognostic factors, and the development of new treatments are increasingly dependent on access to human cancer tissues with linked clinicopathological data. Access to human tumor samples and a large investment in translational research are needed to advance this research. The SMC tissue repositories provide researchers with biological materials, which are essential tools for cancer research. SMC tissue repositories for research aim to collect, document and preserve human biospecimens from patients with cancerous diseases. This is in order to provide the highest quality and well annotated biological biospecimens, used as essential tools to achieve the growing demands of scientific research needs. Such repositories are partners in acceLerating biomedical research and medical product development through clinical resources, in order to apply best options to the patients. Following Institutional Review Board approval and signing an Informed Consent Form, the tumor and tumor-free specimens are coLLected by a designated pathologist at the operating room only when there is a sufficient amount of the tumor, in excess of the routine needs. Blood samples are collected prior to the procedure. Other types of specimens collected include ascites fluid, pleural effusion, tissues for Optimal Cutting Temperature [OCT] and primary culture etc. Demographic, clinical, pathologicaL, and follow-up data are collected in a designated database. SMC has already established several organ or disease-specific tissue repositories within different departments. The foundation of tissue repositories requires the concentrated effort of a multidisciplinary team composed of paramedical, medical and scientific professionals. Research projects using these specimens facilitate the development of 'targeted therapy', accelerate basic research aimed at clarifying molecular mechanisms involved in cancer, and support the development of novel diagnostic tools.

UP-REGULATION OF TISSUE FACTOR IN HUMAN PULMONARY ARTERY ENDOTHELIAL CELLS AFTER ULTRAFINE PARTICLE EXPOSURE

EPA Science Inventory

Background: Epidemiology studies have linked exposure to pollutant particles to

increased cardiovascular mortality and morbidity, but the mechanisms remain unknown.

Objectives: We tested the hypothesis that the ultrafine fraction of ambient pollutant

particle...

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.

Endogenous Generation of Singlet Oxygen and Ozone in Human and Animal Tissues: Mechanisms, Biological Significance, and Influence of Dietary Components.

PubMed

Onyango, Arnold N

2016-01-01

Recent studies have shown that exposing antibodies or amino acids to singlet oxygen results in the formation of ozone (or an ozone-like oxidant) and hydrogen peroxide and that human neutrophils produce both singlet oxygen and ozone during bacterial killing. There is also mounting evidence that endogenous singlet oxygen production may be a common occurrence in cells through various mechanisms. Thus, the ozone-producing combination of singlet oxygen and amino acids might be a common cellular occurrence. This paper reviews the potential pathways of formation of singlet oxygen and ozone in vivo and also proposes some new pathways for singlet oxygen formation. Physiological consequences of the endogenous formation of these oxidants in human tissues are discussed, as well as examples of how dietary factors may promote or inhibit their generation and activity.

Endogenous Generation of Singlet Oxygen and Ozone in Human and Animal Tissues: Mechanisms, Biological Significance, and Influence of Dietary Components

PubMed Central

2016-01-01

Recent studies have shown that exposing antibodies or amino acids to singlet oxygen results in the formation of ozone (or an ozone-like oxidant) and hydrogen peroxide and that human neutrophils produce both singlet oxygen and ozone during bacterial killing. There is also mounting evidence that endogenous singlet oxygen production may be a common occurrence in cells through various mechanisms. Thus, the ozone-producing combination of singlet oxygen and amino acids might be a common cellular occurrence. This paper reviews the potential pathways of formation of singlet oxygen and ozone in vivo and also proposes some new pathways for singlet oxygen formation. Physiological consequences of the endogenous formation of these oxidants in human tissues are discussed, as well as examples of how dietary factors may promote or inhibit their generation and activity. PMID:27042259

The effect of serum on monocyte tissue factor generation.

PubMed

Edwards, R L; Perla, D

1984-09-01

Human monocytes generate the procoagulant tissue factor (MTF) following exposure to a variety of immune stimuli in vitro. The generation of MTF is modified by T cells, lymphokines, and immunoregulatory lipoproteins, and recent studies have shown that MTF can be activated in an immune-specific manner following exposure to antigen. We have examined the role of serum factors in the regulation of MTF generation. Low concentrations (less than 1%) of heat-inactivated normal human serum greatly enhanced MTF generation in cultures of normal peripheral blood mononuclear cells. The stimulatory effect was observed in cultures of both unstimulated cells and cells exposed to bacterial lipopolysaccharide. Stimulation was not observed at high serum concentrations (greater than 10%) and could not be explained by endotoxin contamination or activation of the assay system. Stimulatory activity was present in plasma and BaSO4-adsorbed plasma as well as autologous and allogeneic serum, was not abolished by removal of serum lipoproteins, and did not require the presence of T cells for its expression. Sera from 28 different normal volunteers were screened for stimulatory activity and demonstrated a wide variation in potency. These results suggest that a potent factor is present in sera that enhances the expression of MTF activity in vitro. This factor is distinct from previously described lipoprotein regulators and may play a role in the initiation of coagulation in both normal hemostasis and pathologic states.

Muscle Tissue Engineering Using Gingival Mesenchymal Stem Cells Encapsulated in Alginate Hydrogels Containing Multiple Growth Factors.

PubMed

Ansari, Sahar; Chen, Chider; Xu, Xingtian; Annabi, Nasim; Zadeh, Homayoun H; Wu, Benjamin M; Khademhosseini, Ali; Shi, Songtao; Moshaverinia, Alireza

2016-06-01

Repair and regeneration of muscle tissue following traumatic injuries or muscle diseases often presents a challenging clinical situation. If a significant amount of tissue is lost the native regenerative potential of skeletal muscle will not be able to grow to fill the defect site completely. Dental-derived mesenchymal stem cells (MSCs) in combination with appropriate scaffold material, present an advantageous alternative therapeutic option for muscle tissue engineering in comparison to current treatment modalities available. To date, there has been no report on application of gingival mesenchymal stem cells (GMSCs) in three-dimensional scaffolds for muscle tissue engineering. The objectives of the current study were to develop an injectable 3D RGD-coupled alginate scaffold with multiple growth factor delivery capacity for encapsulating GMSCs, and to evaluate the capacity of encapsulated GMSCs to differentiate into myogenic tissue in vitro and in vivo where encapsulated GMSCs were transplanted subcutaneously into immunocompromised mice. The results demonstrate that after 4 weeks of differentiation in vitro, GMSCs as well as the positive control human bone marrow mesenchymal stem cells (hBMMSCs) exhibited muscle cell-like morphology with high levels of mRNA expression for gene markers related to muscle regeneration (MyoD, Myf5, and MyoG) via qPCR measurement. Our quantitative PCR analyzes revealed that the stiffness of the RGD-coupled alginate regulates the myogenic differentiation of encapsulated GMSCs. Histological and immunohistochemical/fluorescence staining for protein markers specific for myogenic tissue confirmed muscle regeneration in subcutaneous transplantation in our in vivo animal model. GMSCs showed significantly greater capacity for myogenic regeneration in comparison to hBMMSCs (p < 0.05). Altogether, our findings confirmed that GMSCs encapsulated in RGD-modified alginate hydrogel with multiple growth factor delivery capacity is a promising candidate for muscle tissue engineering.

The serum levels of connective tissue growth factor in patients with systemic lupus erythematosus and lupus nephritis.

PubMed

Wang, F-M; Yu, F; Tan, Y; Liu, G; Zhao, M-H

2014-06-01

The expression of connective tissue growth factor mRNA in human kidneys may serve as an early marker for lupus nephritis progression. Therefore, we speculated that connective tissue growth factor may be involved in the pathogenesis of systemic lupus erythematosus and lupus nephritis. In this study, we set out to investigate the associations between serum connective tissue growth factor levels and clinicopathological features of patients with systemic lupus erythematosus and lupus nephritis. Serum samples from patients with non-renal systemic lupus erythematosus, renal biopsy-proven lupus nephritis and healthy control subjects were detected by enzyme-linked immunosorbent assay for serum connective tissue growth factor levels. The associations between connective tissue growth factor levels and clinicopathological features of the patients were further analysed. The levels of serum connective tissue growth factor in patients with non-renal systemic lupus erythematosus and lupus nephritis were both significantly higher than those in the normal control group (34.14 ± 12.17 ng/ml vs. 22.8 ± 3.0 ng/ml, p<0.001; 44.1 ± 46.8 ng/ml vs. 22.8 ± 3.0 ng/ml, p = 0.035, respectively). There was no significant difference of the serum connective tissue growth factor levels between non-renal systemic lupus erythematosus and lupus nephritis group (34.14 ± 12.17 ng/ml vs. 44.1 ± 46.8 ng/ml, p = 0.183). Serum connective tissue growth factor levels were significantly higher in lupus nephritis patients with the following clinical manifestations, including anaemia (51.3 ± 51.4 ng/ml vs. 23.4 ± 9.7 ng/ml, p<0.001) and acute renal failure (85.5 ± 75.0 ng/ml vs. 31.2 ± 21.8 ng/ml, p = 0.002). Serum connective tissue growth factor levels in class IV were significantly higher than that in class II, III and V (57.6 ± 57.5 ng/ml vs. 18.7 ± 6.4 ng/ml, p = 0.019; 57.6 ± 57.5 ng/ml vs. 25.2 ± 14.9 ng/ml, p = 0.006; 57.6 ± 57.5 ng/ml vs. 30.5 ± 21.3 ng/ml, p = 0.017, respectively). Serum connective tissue growth factor levels were significantly higher in those with both active/chronic lesions than those in those with active lesions only in either class IV (84.9 ± 69.6 ng/ml vs. 40.0 ± 40.2 ng/ml, p = 0.001) or in combination of class III and IV lupus nephritis (63.3 ± 63.4 ng/ml vs. 38.3 ± 37.9 ng/ml, p = 0.035, respectively). Serum connective tissue growth factor levels were negatively associated with estimated glomerular filtration rate (r = -0.46, p<0.001) and positively associated with interstitial inflammation (r = 0.309, p = 0.002) and interstitial fibrosis (r = 0.287, p = 0.004). Serum connective tissue growth factor level was a risk factor for doubling of serum creatinine in lupus nephritis (p<0.001, hazard ratio = 1.015, 95% confidence intervals 1.008-1.022) in univariate analysis. Serum connective tissue growth factor levels were significantly higher in lupus and correlated with chronic renal interstitial injury and doubling of serum creatinine in patients with lupus nephritis. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.