Abstract: AIM: To observe the effect of 5-azacytidine (5-Aza) on bone marrow mesenchymal stem cells (MSCs) and to determine the cardiomyocyte-like cells that were differentiated from MSCs. METHODS: Bone marrow mononuclear cells (BM-MNC) were isolated from bone marrow by the method of density gradient centrifugation and were cultured in low glucose DMEM containing 200 ml/L fetal bovine serum. The cells were purified by the different abilities of adhesion. Cell surface antigens were tested by flow cytometry. MSCs were induced by 5-Aza at the third passage for 24 h. After being treated by 5-Aza for 4 weeks, the cells were harvested. Muscle tissue antigen such as α-actin and the cardiac muscle heterologous protein, e.g., cTnT, were detected by immunocytochemical staining. Ultramicrostructure of the cells was observed under transmission electron microscope. RESULTS: After being induced by 5-Aza, α-actin and cTnT were expressed in MSCs. However, expression in MSC without 5-Aza treatment was negative. Ultramicrostructure of the cells such as myofilament was observed under transmission electron microscope. CONCLUSION: Human bone marrow MSCs can differentiate into cardiomyocyte-like cells under 5-Aza induction in vitro.