Study Of Bone Marrow Mesenchymal Stem Cells Differentiation Intervened By Traditional Chinese Medicine

Bone marrow mesenchymal stem cells (BMMSCs),which can be isolatedfrom bone marrow and expanded in vitro without any apparent modification inphenotype, are a group of cells with the capability of self-renewal and potentialof multilineage differentiation. Under certain conditions, BMMSCs candifferentiate into the osteogenic, chondrogenic, myogenic, endothelial andadipogenic lineages. As BMMSCs could be easily obtained, they are verysuitable for self-transplantation and have tremendous potential for cell therapyand tissue engineering. Accordingly, the investigation of the differentiation ofBMMSCs into cardiomyocytes is of great theoretical and clinical significationfor treating myocardial infarction by cell transplantation.In this study, we established the models of isolating and purifyingmesenchymal stem cells from marrow and inducing rat-BMMSCs todifferentiate into cardiomyocytes by 5-azacytidine in vitro, and identified thecells both before and after treated with 5-azacytidine using the ways of flowcytometry analysis, immunohistochemical method and RT-PCR. The resultsshowed that the original cells, which were a fibroblast-like morphology,attached to the culture dishes tightly, and proliferated quickly in the culturemedium, uniformly expressed the cell surface markers CD29, CD44, CD105 andCD166 but not CD14, CD34 and CD45, and also expressed the genes of stem cellmarkers nestin, nucleostemin (NST) and Oct-4. When cultured BMMSCs weretreated with 10 μmol/L 5-azacytidine for 24 hours on day 3 of a 4-week culture, thecells were lengthened in configuration and slowered in proliferation gradually, andexpressed the cardiac muscle cell markers cardiac-specific troponin I, β-myosinheavy chain and myosin light chain-2v after 4 weeks. All of above, we demonstratedthat plastic adherent cultures elaborated from rat bone marrow were mesenchymalstem cells, which could differentiate into cardiomyocytes by 5-azacytidinetreatment in vitro.On the basis of primary study, we investigated the effect of medicated sera ofTraditional Chinese Medicine (TCM) on the proliferation and differentiation ofswine-BMMSCs by MTT assay and proteomic approach, respectively. The resultsshowed that when the concentration of medicated sera was below 10% theproliferation of BMMSCs could be improved. However, when the concentration wasabove 10%, they could be suppressed. By studying the differentiation of BMMSCsinduced by 5-Azacytidine and intervened by Shuanglong medicated sera using theproteomic approach, we found the expressions of 41 proteins were remarkably alteredcompared with the control, and 18 proteins were remarkably altered during the courseof intervention. Of the 13 proteins which were subsequently identified, PROX2,Hsp27, stathmin-1 and BMP-7 may be involved in the cell proliferation anddifferentiation. Our study indicated that under certain condition, medicated sera ofTCM could promote the proliferation and 5-azacytidine-induced differentiation ofBMMSCs in vitro.