Effects of cyclic compressive loading on chondrogenesis of rabbit bone-marrow derived mesenchymal stem cells.
(41/4797)
The objective of this study was to examine the effects of cyclic compressive loading on chondrogenic differentiation of rabbit bone-marrow mesenchymal stem cells (BM-MSCs) in agarose cultures. Rabbit BM-MSCs were obtained from the tibias and femurs of New Zealand white rabbits. After the chondrogenic potential of BM-MSCs was verified by pellet cultures, cell-agarose constructs were made by suspending BM-MSCs in 2% agarose (10(7) cells/ml) for a cyclic, unconfined compression test performed in a custom-made bioreactor. Specimens were divided into four groups: control; transforming growth factor (TGF-beta) (with TGF-beta1 treatment); loading (with stimulation of cyclic, unconfined compressive loading); and TGF-beta loading (with TGF-beta1 treatment and loading stimulation) groups. In the loading experiment, specimens were subjected to sinusoidal loading with a 10% strain magnitude at a frequency of 1 Hz for 4 hours a day. Experiments were conducted for 3, 7, and 14 consecutive days. While the experimental groups (TGF-beta, loading, and TGF-beta loading) exhibited significantly higher levels of expressions of chondrogenic markers (collagen II and aggrecan) at three time periods, there were no differences among the experimental groups after an extra 5-day culture. This suggests that compressive loading alone induces chondrogenic differentiation of rabbit BM-MSCs as effectively as TGF-beta or TGF-beta plus loading treatment. Moreover, both the compressive loading and the TGF-beta1 treatment were found to promote the TGF-beta1 gene expression of rabbit BM-MSCs. These findings suggest that cyclic compressive loading can promote the chondrogenesis of rabbit BM-MSCs by inducing the synthesis of TGF-beta1, which can stimulate the BM-MSCs to differentiate into chondrocytes. (+info)
Transfer and stable transgene expression of a mammalian artificial chromosome into bone marrow-derived human mesenchymal stem cells.
(42/4797)
Mammalian artificial chromosomes (ACEs) transferred to autologous adult stem cells (SCs) provide a novel strategy for the ex vivo gene therapy of a variety of clinical indications. Unlike retroviral vectors, ACEs are stably maintained, autonomous, and nonintegrating. In this report we assessed the delivery efficiency of ACEs and evaluated the subsequent differentiation potential of ACE-transfected bone marrow-derived human mesenchymal stem cells (hMSCs). For this, an ACE carrying multiple copies of the red fluorescent protein (RFP) reporter gene was transferred under optimized conditions into hMSCs using standard cationic transfection reagents. RFP expression was detectable in 11% of the cells 4-5 days post-transfection. The RFP-expressing hMSCs were enriched by high-speed flow cytometry and maintained their potential to differentiate along adipogenic or osteogenic lineages. Fluorescent in situ hybridization and fluorescent microscopy demonstrated that the ACEs were stably maintained as single chromosomes and expressed the RFP transgenes in both differentiated cultures. These findings demonstrate the potential utility of ACEs for human adult SC ex vivo gene therapy. (+info)
Mesenchymal stem cells can be differentiated into endothelial cells in vitro.
(43/4797)
Human bone marrow-derived mesenchymal stem cells (MSCs) have the potential to differentiate into mesenchymal tissues like osteocytes, chondrocytes, and adipocytes in vivo and in vitro. The aim of this study was to investigate the in vitro differentiation of MSCs into cells of the endothelial lineage. MSCs were generated out of mononuclear bone marrow cells from healthy donors separated by density gradient centrifugation. Cells were characterized by flow cytometry using a panel of monoclonal antibodies and were tested for their potential to differentiate along different mesenchymal lineages. Isolated MSCs were positive for the markers CD105, CD73, CD166, CD90, and CD44 and negative for typical hematopoietic and endothelial markers. They were able to differentiate into adipocytes and osteocytes after cultivation in respective media. Differentiation into endothelial-like cells was induced by cultivation of confluent cells in the presence of 2% fetal calf serum and 50 ng/ml vascular endothelial growth factor. Laser scanning cytometry analysis of the confluent cells in situ showed a strong increase of expression of endothelial-specific markers like KDR and FLT-1, and immunofluorescence analysis showed typical expression of the von Willebrand factor. The functional behavior of the differentiated cells was tested with an in vitro angiogenesis test kit where cells formed characteristic capillary-like structures. We could show the differentiation of expanded adult human MSCs into cells with phenotypic and functional features of endothelial cells. These predifferentiated cells provide new options for engineering of artificial tissues based on autologous MSCs and vascularized engineered tissues. (+info)
Functional expression of HGF and HGF receptor/c-met in adult human mesenchymal stem cells suggests a role in cell mobilization, tissue repair, and wound healing.
(44/4797)
Human mesenchymal stem cells (hMSC) are adult stem cells with multipotent capacities. The ability of mesenchymal stem cells to differentiate into many cell types, as well as their high ex vivo expansion potential, makes these cells an attractive therapeutic tool for cell transplantation and tissue engineering. hMSC are thought to contribute to tissue regeneration, but the signals governing their mobilization, diapedesis into the bloodstream, and migration into the target tissue are largely unknown. Here we report that hepatocyte growth factor (HGF) and the cognate receptor HGFR/c-met are expressed in hMSC, on both the RNA and the protein levels. The expression of HGF was downregulated by transforming growth factor beta. HGF stimulated chemotactic migration but not proliferation of hMSC. Therefore the HGF/c-met signaling system may have an important role in hMSC recruitment sites of tissue regeneration. The controlled regulation of HGF/c-met expression may be beneficial in tissue engineering and cell therapy employing hMSC. (+info)
Interactions of chemokines and chemokine receptors mediate the migration of mesenchymal stem cells to the impaired site in the brain after hypoglossal nerve injury.
(45/4797)
Mesenchymal stem cells (MSCs), cultured ex vivo, recently were shown to be able to migrate into sites of brain injuries when transplanted systemically or locally, suggesting that MSCs possess migratory capacity. However, the mechanisms underlying the migration of these cells remain unclear. In this study, we examined the role of some chemokines and their receptors in the trafficking of rat MSCs (rMSCs) in a rat model of left hypoglossal nerve injury. rMSCs transplanted into the lateral ventricles of the rat brain migrated to the avulsed hypoglossal nucleus, where the expression of chemokines, stromal-cell-derived factor 1 (SDF-1), and fractalkine was observed to be increased. This increase temporally paralleled the migration of rMSCs into the avulsed nucleus at 1 and 2 weeks after operation. It has been found that rMSCs express CXCR4 and CX3CR1, the respective receptors for SDF-1 and fractalkine, and other chemokine receptors, CCR2 and CCR5. Furthermore, in vitro analysis revealed that recombinant human SDF-1 alpha (rhSDF-1alpha) and recombinant rat fractalkine (rrfractalkine) induced the migration of rMSCs in a G-protein-dependent manner. Intracerebral injection of rhSDF-1alpha has also been shown to stimulate the homing of transplanted rMSCs to the site of injection in the brain. These data suggest that the interactions of fractalkine-CX3CR1 and SDF-1-CXCR4 could partially mediate the trafficking of transplanted rMSCs. This study provides an important insight into the understanding of the mechanisms governing the trafficking of transplanted rMSCs and also significantly expands the potential role of MSCs in cell therapy for brain injuries and diseases. (+info)
Decreased number and impaired angiogenic function of endothelial progenitor cells in patients with chronic renal failure.
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OBJECTIVE: Increased risk of cardiovascular disease in patients with chronic renal failure (CRF) has been explained by accelerated atherosclerosis and impaired angiogenesis, in which endothelial progenitor cells (EPCs) may play key roles. We hypothesized that altered EPC biology may contribute to the pathophysiology of CRF. METHODS AND RESULTS: EPCs were isolated from CRF patients on maintenance hemodialysis (n=44) and from a normal control group (n=30). CRF patients showed markedly decreased numbers of EPC (44.6%) and colonies (75.3%) when compared with the controls (P<0.001). These findings were corroborated by 30.5% decrease in EPC migratory function in response to vascular endothelial growth factor (VEGF) (P=0.040) and 48.8% decrease in EPC incorporation into human umbilical vein endothelial cells (HUVEC) (P<0.001). In addition, Framingham's risk factor score of both CRF (r=-0.461, P=0.010) and normal group (r=-0.367, P=0.016) significantly correlated with the numbers of EPC. Indeed, the number of circulating EPC was significantly lower in CRF patients than in normal group under the same burden of risk factors (P<0.001). A significant correlation was also observed between dialysis dose (Kt/V) and EPC incorporation into HUVEC (r=0.427, P=0.004). CONCLUSIONS: EPC biology, which is critical for neovascularization and the maintenance of vascular function, is altered in CRF. Our data strongly suggest that dysfunction of circulating EPC has a role in the progression of cardiovascular disease in patients with CRF. (+info)
Msx2 and necdin combined activities are required for smooth muscle differentiation in mesoangioblast stem cells.
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Little is known about the molecular mechanism underlying specification and differentiation of smooth muscle (SM), and this is, at least in part, because of the few cellular systems available to study the acquisition of a SM phenotype in vitro. Mesoangioblasts are vessel-derived stem cells that can be induced to differentiate into different cell types of the mesoderm, including SM. We performed a DNA microarray analysis of a mesoangioblast clone that spontaneously expresses an immature SM phenotype and compared it with a sister clone mainly composed of undifferentiated progenitor cells. This study allowed us to define a gene expression profile for "stem" cells versus smooth muscle cells (SMCs) in the absence of differentiation inducers such as transforming growth factor beta. Two transcription factors, msx2 and necdin, are expressed at least 100 times more in SMCs than in stem cells, are coexpressed in all SMCs and tissues, are induced by transforming growth factor beta, and, when coexpressed, induce a number of SM markers in mesoangioblast, fibroblast, and endothelial cell lines. Conversely, their downregulation through RNA interference results in a decreased expression of SM markers. These data support the hypothesis that Msx2 and necdin act as master genes regulating SM differentiation in at least a subset of SMCs. (+info)
Circulating CD34-positive cells provide an index of cerebrovascular function.
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BACKGROUND: Increasing evidence points to a role for circulating endothelial progenitor cells, including populations of CD34- and CD133-positive cells present in peripheral blood, in maintenance of the vasculature and neovascularization. Immature populations, including CD34-positive cells, have been shown to contribute to vascular homeostasis, not only as a pool of endothelial progenitor cells but also as a source of growth/angiogenesis factors at ischemic loci. We hypothesized that diminished numbers of circulating immature cells might impair such physiological and reparative processes, potentially contributing to cerebrovascular dysfunction. METHODS AND RESULTS: The level of circulating immature cells, CD34-, CD133-, CD117-, and CD135-positive cells, in patients with a history of atherothrombotic cerebral ischemic events was analyzed to assess possible correlations with the degree of carotid atherosclerosis and number of cerebral infarctions. There was a strong inverse correlation between numbers of circulating CD34- and CD133-positive cells and cerebral infarction. In contrast, there was no correlation between the degree of atherosclerosis and populations of circulating immature cells. Analysis of patients with cerebral artery occlusion revealed a significant positive correlation between circulating CD34- and CD133-positive cells and regional blood flow in areas of chronic hypoperfusion. CONCLUSIONS: These results suggest a possible contribution of circulating CD34- and CD133-positive cells in maintenance of the cerebral circulation in settings of ischemic stress. Our data demonstrate the utility of a simple and precise method to quantify circulating CD34-positive cells, the latter providing a marker of cerebrovascular function. (+info)