Background: Due to their immunomodulatory and trophic support functions, mesenchymal stem cells (MSCs) are promising in the field of cell-based regenerative medicine. However, MSC survival post-transplantation is challenged by various microenvironment stress factors. Here, we investigated the role of vitronectin (VTN) in the survival strategy of MSCs under serum deprivation stress condition. Methods: Proliferation kinetics and cell adhesion of MSCs under serum deprivation were determined from population doublings and cell-matrix de-adhesion studies, respectively. mRNA and protein expression levels of VTN were confirmed by qRT-PCR and Western blotting, respectively. Immunofluorescence technique revealed distribution of VTN under serum deprivation stress. siRNA and inhibitor-based studies were performed to confirm the role and regulation of VTN. Apoptosis and cell cycle status of MSCs were assessed using flow cytometric analysis. Results: Subjecting MSCs to serum deprivation led to significant increase in cell spread area and cell-matrix adhesion. An upregulation of VTN expression was noted with an arrest in G0/G1 phase of cell cycle and no appreciable apoptotic change. Pro-survival PI3kinase pathway inhibition led to further increase in VTN expression with no apoptotic change. siRNA-mediated inhibition of VTN resulted in reversal in G0/G1 cell cycle arrest and a marked increase in apoptosis, suggesting a role of VTN in preventing serum deprivation-induced apoptotic cell death. In addition, p65 knockdown resulted in downregulation of VTN establishing an association between NF-κβ pathway and VTN. Conclusions: VTN was identified as a survival factor in providing protection from serum deprivation-induced apoptosis in MSCs.
The efficacy of mesenchymal stem cell (MSC) therapy is currently limited by low retention and poor survival of transplanted cells as demonstrated by clinical studies. This is mainly due to the harsh microenvironment created by oxygen and nutrient deprivation and inflammation at the injured sites. The choice of MSC source could be critical in determining fate and cellular function of MSCs under stress. Our objective here was to investigate the influence of ischemia-like stress on Wharton's jelly MSCs (WJ-MSCs) from human umbilical cord to assess their therapeutic relevance in ischemic diseases. We simulated conditions of ischemia in vitro by culturing WJ-MSCs in 2% oxygen in serum deprived and low glucose medium. Under these conditions, WJ-MSCs retained viable population of greater than 80%. They expressed the characteristic MSC surface antigens at levels comparable to the control WJ-MSCs and were negative for the expression of costimulatory molecules. An upregulation of many ECM and adhesion molecules and growth and angiogenic factors contributing to wound healing and regeneration was noted in the ischemic WJ-MSC population by a PCR array. Their migration ability, however, got impaired. Our findings provide evidence that WJ-MSCs might be therapeutically beneficial and potent in healing wounds under ischemic conditions.
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