Icariin On Bone Marrow Mesenchymal Stem Cell Proliferation And Differentiation To Cardiac Cells

Objective:To study the influnce on proliferation and differentiation of cardiomyocytes from marrow mesenchymal stem cells (MSCs) in vitro by icarilin (ICA) which is the active component of epimedium. Epimedium has the function of warming yang and nourishing kidney.Methods: MSCs were isolated and cultured from human bone marrow. MSCs derived from the third or the forth were cultured in vitro for experiment. The effect of icarilin (four concentration groups :20ug/l, 40ug/l,80ug/l,160ug/l)on proliferation of MSCs was tested by MTT method. During the inducing process,ICA, 5-azacytidine and ICA+5-aza were used as inducing factors. The MSCs differentiation was observed for four weeks under phase-contrast microscope. The Immunofluorescence technique was used for the expression of TroponinI (cTnI), Connexin43 (CX43), and using flow cytometry to test the cardiomyocytes different-tiation rate(percentage of the cells with expression of cTnI, CX43). The expression of GATA-4 and Nkx2-5 was tested by semi-quanti- tative PCR and Western-blotting.Results: The OD of ICA group was higher than the control group(P< 0.01), and the ICA concentration of 80ug/l was best. Fluorescence result showed that, both 5-aza group and ICA+5-aza group had expression of cTnI and CX43. The expression of cTnI and CX43 was weak for ICA group, and it was no expression for control group. Flow cytometry showed that the percentage of cTnI, CX43 expression was 47.22% and 44.43% for ICA+5-aza group, 29. 10% and 27. 15% for 5-aza group. The rate of positive cells with ICA+5-aza induction was higher than the induction of 5-aza(P< 0. 01). The ICA group had lower expression(7. 02%,7. 69%) and the control group had almost no expression. Semi-quantitative PCR and Western-blotting showed that the expression levels of GATA-4 and Nkx2-5 with ICA+5-aza and 5-aza induction were significantly higher than the induction of simplex ICA and no induction. The expression levels of GATA-4 and Nkx2-5 with induction of ICA+5-aza were higher than that of 5-aza. Conclusion: ICA can accelerate the proliferation of MSCs. ICA can accelerate the differentiation of cardiomyocytes from MSCs combining with 5-aza.The effect of 5-aza plus ICA is the better factor in inducing MSCs to differentiate into cardiomyocytes. ICA might has the trend of inducing MSCs to differentiate into cardiomyocytes, which needs further study.