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Mesenchymal Stem Cells Derived from Rat Epicardial Versus Epididymal Adipose Tissue  [PDF]
Mohamadreza Baghaban Eslaminejad,Soura Mardpour,Marzieh Ebrahimi
Iranian Journal of Basic Medical Sciences , 2011,
Abstract: Objective(s)Some investigation has indicated that adipose-derived stem cells possess different surface epitopes and differentiation potential according to the localization of fat pad from which the cells were derived. In the present study proliferation capacity and aging of such cells were explored.Materials and MethodsAdherent cells were isolated from the collagenase digests of adipose tissues excised from rat epicardial and epididymal regions and propagated with several subcultures. The cells were then investigated whether or not they were able to differentiate into bone, cartilage and adipose cell lineages. Studied cells from two adipose tissues were also compared with respect to their in vitro proliferation capacity. The presence of senescent cells in the culture was determined and compared using senescence-associated (SA) -galactosidase staining method. ResultsSuccessful differentiations of the cells were indicative of their mesenchymal stem cells (MSCs) identity. Epicardial adipose-derived cells tended to have a short population doubling time (45±9.6 hr) than the epididymal adipose-derived stem cells (69±16 hr, P< 0.05). Colonogenic activity and the growth curve characteristics were all better in the culture of stem cells derived from epicardial compared to epididymal adipose tissue. Comparatively more percentage of senescent cells was present at the cultures derived from epididymal adipose tissue (P< 0.05).ConclusionOur data emphasize on the differences existed between the stem cells derived from adipose depots of different anatomical sites in terms of their proliferative capacity and in vitro aging. Such data can help understand varying results reported by different laboratories involved in adipose stem cell investigations.
Use of adipose tissue as a source of mesenchymal stem cells  [PDF]
Katarzyna Jezierska-Wo?niak,Dorota Nosarzewska,Anna Tutas,Anita Miko?ajczyk
Post?py Higieny i Medycyny Do?wiadczalnej , 2010,
Abstract: Enormous expectations are associated with stem cells with regard to cell therapy and tissue engineering. Stem cells have unlimited potential for self-renewal and develop into various cell types. For the mesodermal tissue engineering such a source of cells is the bone marrow stroma. However, isolation of the bone marrow requires general or spinal anesthesia and yields low number of mesodermal stem cells (MSCs) upon processing (1 MSC per 105 adherent stromal cells). An alternative source of autologous stem cells seems to be, apart from bone marrow: periosteum, muscular tissue or synovial membrane and adipose tissue. The adipose tissue is derived from the embryonic mesenchyme, contains a large number of stromal stem cells and is relatively easy to obtain in large quantities. It covers a widespread area of human body, and can be classified as white and brown adipose tissue in terms of location and function. Specimens of the adipose tissue are usually obtained from elective, laparoscopic or liposuction surgeries. Stromal stem cells, isolated from this tissue, exhibit characteristics common to mesodermal tissues, including: adherence to plastic, formation of fibroblastic- like colonies, extensive proliferative capacity, ability to differentiate into several mesodermal lineages (including bone, cartilage, muscle and fat), and expression of several common cell surface antigens. Recent evidence suggest that these cells can also form non-mesodermal tissues – neuron-like cells. The aim of this publication is to describe the application of the adipose tissue as a source of mesenchymal stem cells based on current literature data.
Adipose-Derived Mesenchymal Stromal/Stem Cells: Tissue Localization, Characterization, and Heterogeneity  [PDF]
Patrick C. Baer,Helmut Geiger
Stem Cells International , 2012, DOI: 10.1155/2012/812693
Abstract: Adipose tissue as a stem cell source is ubiquitously available and has several advantages compared to other sources. It is easily accessible in large quantities with minimal invasive harvesting procedure, and isolation of adipose-derived mesenchymal stromal/stem cells (ASCs) yields a high amount of stem cells, which is essential for stem-cell-based therapies and tissue engineering. Several studies have provided evidence that ASCs in situ reside in a perivascular niche, whereas the exact localization of ASCs in native adipose tissue is still under debate. ASCs are isolated by their capacity to adhere to plastic. Nevertheless, recent isolation and culture techniques lack standardization. Cultured cells are characterized by their expression of characteristic markers and their capacity to differentiate into cells from meso-, ecto-, and entodermal lineages. ASCs possess a high plasticity and differentiate into various cell types, including adipocytes, osteoblasts, chondrocytes, myocytes, hepatocytes, neural cells, and endothelial and epithelial cells. Nevertheless, recent studies suggest that ASCs are a heterogeneous mixture of cells containing subpopulations of stem and more committed progenitor cells. This paper summarizes and discusses the current knowledge of the tissue localization of ASCs in situ, their characterization and heterogeneity in vitro, and the lack of standardization in isolation and culture methods. 1. Introduction: Mesenchymal Stromal/Stem Cells The pathologist Cohnheim first observed the presence of nonhematopoietic stem cells in the bone marrow in 1867 [1]. He hypothesized that cells with a fibroblast-like morphology migrate to the sites of injury and help to regenerate damaged tissue. The pioneering work of Friedenstein and coworkers in the 1960s [2, 3] on the isolation, culture, and osteogenic differentiation of bone-marrow-derived cells opened a new field of stem cell research. Nearly 20 years later, Owen [4] and Caplan [5] introduced the terms stromal stem cells and mesenchymal stem cells (MSCs) to the scientific community. Whereas, in these initial works, MSCs were isolated from adult bone marrow, in the next decades, MSCs were also found in nearly all adult tissues (e.g., adipose tissue, synovium, dermis, periosteum, deciduous teeth), in peripheral blood, menstrual blood, and in solid organs (e.g., liver, spleen, lung) [6–8]. MSCs are a rare and quiescent population in their niche within fully specialized tissues. At present, there is a strong amount of data indicating that MSCs represent independent population(s) of stem cells
Murine mesenchymal progenitor cells from different tissues differentiated via mesenchymal microspheres into the mesodermal direction
Florian B?hrnsen, Ulrich Lindner, Markus Meier, Abdelalim Gadallah, Peter Schlenke, Hendrik Lehnert, Jürgen Rohwedel, Jan Kramer
BMC Cell Biology , 2009, DOI: 10.1186/1471-2121-10-92
Abstract: We introduced the mesenchymal microsphere method as a feasible time- and cell saving screening method to analyse multilineage differentiation properties of adult progenitor cells in a three-dimensional system. For this purpose we isolated, characterized and analyzed new sources of adult murine mesenchymal progenitor cells from perirenal adipose tissue and mediastinal stromal tissue in comparison to bone marrow progenitor cells. The proliferation capacity of the cells was demonstrated by determination of the daily doubling index. Although the flow cytometry analysis of undifferentiated cells revealed differences in the expression of CD marker molecules, all isolates have the capacity for multilineage differentiation following the mesenchymal microsphere protocol as well as the classical "micro mass body" protocol for chondrogenic and the monolayer cultivation protocol for osteogenic and adipogenic differentiation. Differentiation was characterized using histochemical and immunhistochemical staining as well as RT-PCR.We were able to show that the mesenchymal microsphere method is an efficient test system for chondro-, osteo- and adipogenic differentiation of adult progenitor cells. The advantage of this system in comparison to classical protocols is that approximately 7 times lower cell numbers are necessary. Since classical culture procedures are time intensive because high cell numbers have to be obtained, the new differentiation method may also save cells and time in future clinical applications using human mesenchymal stromal cells.Many tissues exhibit the capacity for renewal after trauma, disease or aging because of dormant stem cell reservoirs. Different types of stem cells have been described within the adult bone marrow including haematopoietic [1] and colony forming units-fibroblast-like-cells [2], later termed bone marrow stromal cells [3] which have been found to differentiate along multiple mesenchymal lineages [4-7]. Other sources of adult stem cells hav
Method for Obtaining Committed Adult Mesenchymal Precursors from Skin and Lung Tissue  [PDF]
Aurora Bernal, María Fernández, Laura M. Pérez, Nuria San Martín, Beatriz G. Gálvez
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0053215
Abstract: Aims The present study reports an easy and efficient method for obtaining adult mesenchymal precursors from different adult mouse tissues. Materials and Methods We describe the isolation and expansion of mesenchymal precursors from skin and lung by a non-enzymatic method. Skin and lung mesenchymal precursors isolated by a modified explant technique were characterized in vitro by defined morphology and by a specific gene expression profile and surface markers. Results and Conclusions Our results show that these precursors express stem cell and mesenchymal surface markers as well as epithelial markers. However, they are negative for markers of endothelium, cardiac and skeletal muscle or adipose tissue, indicating that they have initiated commitment to the tissues from which were isolated. These precursors can migrate without any stimulus and in response to stimuli as SDF1, MCP1 and TNFα and can be differentiated into epithelial lineages. Based on the properties of these precursors from adult tissues, we propose their use as tools for regenerative biomedicine.
Establishment and Molecular Characterization of Mesenchymal Stem Cell Lines Derived From Human Visceral & Subcutaneous Adipose Tissues  [cached]
Pravin Dattatray Potdar,Jyoti Prakash Sutar
Journal of Stem Cells and Regenerative Medicine , 2010,
Abstract: Mesenchymal stem cells (MSCs), are multipotent stem cells that can differentiate into osteoblasts, chondrocytes, myocytes and adipocytes. We utilized adipose tissue as our primary source, since it is a rich source of MSCs as well as it can be harvested using a minimally invasive surgical procedure. Both visceral and subcutaneous adipose tissue (VSAT, SCAT respectively) samples were cultured using growth medium without using any substratum for their attachment. We observed growth of mesenchymal like cells within 15 days of culturing. In spite of the absence of any substratum, the cells adhered to the bottom of the petri dish, and spread out within 2 hours. Presently VSAT cells have reached at passage 10 whereas; SCAT cells have reached at passage 14. Morphologically MSCs obtained from visceral adipose tissue were larger in shape than subcutaneous adipose tissue. We checked these cells for presence or absence of specific stem cell molecular markers. We found that VSAT and SCAT cells confirmed their MSC phenotype by expression of specific MSC markers CD 105 and CD13 and absence of CD34 and CD 45 markers which are specific for haematopoietic stem cells. These cells also expressed SOX2 gene confirming their ability of self-renewal as well as expressed OCT4, LIF and NANOG for their properties for pluripotency & plasticity. Overall, it was shown that adipose tissue is a good source of mesenchymal stem cells. It was also shown that MSCs, isolated from adipose tissue are multipotent stem cells that can differentiate into osteoblasts, chondrocytes, cardiomyocytes, adipocytes and liver cells which may open a new era for cell based regenerative therapies for bone, cardiac and liver disorders.
Synergistic Actions of Hematopoietic and Mesenchymal Stem/Progenitor Cells in Vascularizing Bioengineered Tissues  [PDF]
Eduardo K. Moioli, Paul A. Clark, Mo Chen, James E. Dennis, Helaman P. Erickson, Stanton L. Gerson, Jeremy J. Mao
PLOS ONE , 2008, DOI: 10.1371/journal.pone.0003922
Abstract: Poor angiogenesis is a major road block for tissue repair. The regeneration of virtually all tissues is limited by angiogenesis, given the diffusion of nutrients, oxygen, and waste products is limited to a few hundred micrometers. We postulated that co-transplantation of hematopoietic and mesenchymal stem/progenitor cells improves angiogenesis of tissue repair and hence the outcome of regeneration. In this study, we tested this hypothesis by using bone as a model whose regeneration is impaired unless it is vascularized. Hematopoietic stem/progenitor cells (HSCs) and mesenchymal stem/progenitor cells (MSCs) were isolated from each of three healthy human bone marrow samples and reconstituted in a porous scaffold. MSCs were seeded in micropores of 3D calcium phosphate (CP) scaffolds, followed by infusion of gel-suspended CD34+ hematopoietic cells. Co-transplantation of CD34+ HSCs and CD34? MSCs in microporous CP scaffolds subcutaneously in the dorsum of immunocompromized mice yielded vascularized tissue. The average vascular number of co-transplanted CD34+ and MSC scaffolds was substantially greater than MSC transplantation alone. Human osteocalcin was expressed in the micropores of CP scaffolds and was significantly increased upon co-transplantation of MSCs and CD34+ cells. Human nuclear staining revealed the engraftment of transplanted human cells in vascular endothelium upon co-transplantation of MSCs and CD34+ cells. Based on additional in vitro results of endothelial differentiation of CD34+ cells by vascular endothelial growth factor (VEGF), we adsorbed VEGF with co-transplanted CD34+ and MSCs in the microporous CP scaffolds in vivo, and discovered that vascular number and diameter further increased, likely owing to the promotion of endothelial differentiation of CD34+ cells by VEGF. Together, co-transplantation of hematopoietic and mesenchymal stem/progenitor cells may improve the regeneration of vascular dependent tissues such as bone, adipose, muscle and dermal grafts, and may have implications in the regeneration of internal organs.
Mitochondrial Respiration Regulates Adipogenic Differentiation of Human Mesenchymal Stem Cells  [PDF]
Yanmin Zhang, Glenn Marsboom, Peter T. Toth, Jalees Rehman
PLOS ONE , 2013, DOI: 10.1371/journal.pone.0077077
Abstract: Human mesenchymal stem cells (MSCs) are adult multipotent stem cells which can be isolated from bone marrow, adipose tissue as well as other tissues and have the capacity to differentiate into a variety of mesenchymal cell types such as adipocytes, osteoblasts and chondrocytes. Differentiation of stem cells into mature cell types is guided by growth factors and hormones, but recent studies suggest that metabolic shifts occur during differentiation and can modulate the differentiation process. We therefore investigated mitochondrial biogenesis, mitochondrial respiration and the mitochondrial membrane potential during adipogenic differentiation of human MSCs. In addition, we inhibited mitochondrial function to assess its effects on adipogenic differentiation. Our data show that mitochondrial biogenesis and oxygen consumption increase markedly during adipogenic differentiation, and that reducing mitochondrial respiration by hypoxia or by inhibition of the mitochondrial electron transport chain significantly suppresses adipogenic differentiation. Furthermore, we used a novel approach to suppress mitochondrial activity using a specific siRNA-based knockdown of the mitochondrial transcription factor A (TFAM), which also resulted in an inhibition of adipogenic differentiation. Taken together, our data demonstrates that increased mitochondrial activity is a prerequisite for MSC differentiation into adipocytes. These findings suggest that metabolic modulation of adult stem cells can maintain stem cell pluripotency or direct adult stem cell differentiation.
Different populations and sources of human mesenchymal stem cells (MSC): A comparison of adult and neonatal tissue-derived MSC
Ralf Hass, Cornelia Kasper, Stefanie B?hm, Roland Jacobs
Cell Communication and Signaling , 2011, DOI: 10.1186/1478-811x-9-12
Abstract: These functions underlie the important physiological roles of MSC and underscore a significant potential for the clinical use of distinct populations from the various tissues. MSC derived from different adult (adipose tissue, peripheral blood, bone marrow) and neonatal tissues (particular parts of the placenta and umbilical cord) are therefore compared in this mini-review with respect to their cell biological properties, surface marker expression and proliferative capacities. In addition, several MSC functions including in vitro and in vivo differentiation capacities within a variety of lineages and immune-modulatory properties are highlighted. Differences in the extracellular milieu such as the presence of interacting neighbouring cell populations, exposure to proteases or a hypoxic microenvironment contribute to functional developments within MSC populations originating from different tissues, and intracellular conditions such as the expression levels of certain micro RNAs can additionally balance MSC function and fate.During the last few years isolations of adult mesenchymal stem cells from different sources have been reported. Bone marrow derived stem cells first described by Friedenstein et al. are still the most frequently investigated cell type and often designated as the gold standard [1]. Mesenchymal stem cells derived from adipose tissue [2], peripheral blood [3], the lung [4] or the heart [5] however have also shown promising potential for proliferation and differentiation into different cell types. In this section we focus on the comparison of adult mesenchymal stem cells derived from bone marrow (BM), adipose tissue (AT) and peripheral blood (PB) (Figure 1).BM-MSC are isolated from bone marrow aspirate. This invasive procedure is painful for the patient and is accompanied by a risk of infection. The commonly applied preparation method for the generation of MSC from bone marrow is density gradient centrifugation [6]. The collected fraction containing mon
Transcriptomics Comparison between Porcine Adipose and Bone Marrow Mesenchymal Stem Cells during In Vitro Osteogenic and Adipogenic Differentiation  [PDF]
Elisa Monaco, Massimo Bionaz, Sandra Rodriguez-Zas, Walter L. Hurley, Matthew B. Wheeler
PLOS ONE , 2012, DOI: 10.1371/journal.pone.0032481
Abstract: Bone-marrow mesenchymal stem cells (BMSC) are considered the gold standard for use in tissue regeneration among mesenchymal stem cells (MSC). The abundance and ease of harvest make the adipose-derived stem cells (ASC) an attractive alternative to BMSC. The aim of the present study was to compare the transcriptome of ASC and BMSC, respectively isolated from subcutaneous adipose tissue and femur of 3 adult pigs, during in vitro osteogenic and adipogenic differentiation for up to four weeks. At 0, 2, 7, and 21 days of differentiation RNA was extracted for microarray analysis. A False Discovery Rate ≤0.05 for overall interactions effect and P<0.001 between comparisons were used to determine differentially expressed genes (DEG). Ingenuity Pathway Analysis and DAVID performed the functional analysis of the DEG. Functional analysis of highest expressed genes in MSC and genes more expressed in MSC vs. fully differentiated tissues indicated low immunity and high angiogenic capacity. Only 64 genes were differentially expressed between ASC and BMSC before differentiation. The functional analysis uncovered a potential larger angiogenic, osteogenic, migration, and neurogenic capacity in BMSC and myogenic capacity in ASC. Less than 200 DEG were uncovered between ASC and BMSC during differentiation. Functional analysis also revealed an overall greater lipid metabolism in ASC, while BMSC had a greater cell growth and proliferation. The time course transcriptomic comparison between differentiation types uncovered <500 DEG necessary to determine cell fate. The functional analysis indicated that osteogenesis had a larger cell proliferation and cytoskeleton organization with a crucial role of G-proteins. Adipogenesis was driven by PPAR signaling and had greater angiogenesis, lipid metabolism, migration, and tumorigenesis capacity. Overall the data indicated that the transcriptome of the two MSC is relatively similar across the conditions studied. In addition, functional analysis data might indicate differences in therapeutic application.
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