| In recent years, the understanding that regeneration progresses exist at the level of the myocardium has placed stem cell research at the center stage in cardiology. Several groups have reported that putative mesenchymal stem cells (MSCs) have shown great promise in regenerating and repopulating the damaged myocardium, restoring its function. However, the use of this approach is limited by the fact that the majority of the transplanted MSCs are readily lost, potentially triggered by the ischemic environment in vivo. The precise events leading to MSC loss is however unclear and very little is known about the cellular mechanisms that may mediate this process. We were therefore interested in unrave2ling why transplanted MSCs fail to survive in ischemic tissue and identifying the factors and examining the mechanisms that may trigger this response. In the present study, we have therefore investigated the effects of hypoxia and serum deprivation (SD), two components that may mimic conditions under ischemia, to delineate the signaling pathway of ischemia—induced apoptosis in MSCs.1. The method for the isolation and culture of bone marrow derived MSCs from patients with coronary artery disease and Sprague-Dawley rats was established. Cells are negative for the pan leukocyte marker CD45 and the endothelial/hematopoietic progenitor marker CD34; but express marker CD29 and CD44. The cells can differentiate into osteoblasts and adipose cells. These data indicateds that the cells we isolated belong to mesenchymal stem cells.2. The cellular hypoxia and SD system in vitro to mimic conditions under ischemia was established, It was found that hypoxia/SD induced MSCs apoptosis based the results of Hoechst 33342 staining , cell size and fluorescent dye Annexin-Ⅴ FITC flow cytometric analysis. The combination of hypoxia and SD induced a time-depended activity of caspase-3, 8, 9 monitored with the substrate-pNA, and cleavage of pro-caspase-3 detected by Western blotting. General caspase...
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