Objective To investigate the potential of adult mesenchymal stem cells(MSCs)derived from human bonemarrow to undergo cardiomyogenic differentiation after exposure to 5-azacytidine(5-aza)in vitro.Methods Asmall bone marrow aspirate was taken from the iliac crest of human volunteers,and hMSCs were isolated by 1.073g/mL Percoll and propagated in the right cell culturing medium as previously described.The phenotypes of hMSCswere characterized with the use of flow cytometry.The hMSCs were cultured in cell culture medium(as control)andmedium mixed with 5-aza for cellular differentiation.We examined by immunohistochemistry at 21 days theinducement of desmin,cardiac-specific cardiac troponin I(cTnI),GATA 4 and connexin-43 respectively.ResultsThe hMSCs are fibroblast-like morphology and express CD44~+ CD29~+ CD90~+/CD34~-CD45~-CD31~-CD11_a~-.After5-aza treatment,20-30% hMSCs connected with adjoining cells and coalesced into myotube structures after 14days.Twenty-one days after 5-aza treatment,immunofluorescence showed that some cells expressed desmin,GATA4,cTnI and connexin-43 in 5,10 μmol/L 5-aza groups,but no cardiac specific protein was found in neither 3μmol/L 5-aza group nor in the control group.The ratio of cTnI positively stained cells in 10 μmol/L group was higherthan that in 5 μmol/L group(65.3±4.7% vs 48.2±5.4%,P<0.05).Electron microscopy revealed thatmyofilaments were formed.The induced cells expressed cardiac-myosin heavy chain(MyHC)gene by reversetranscription-polymerase chain reaction(RT-PCR).Conclusions Theses findings suggest that hMSCs from adultbone marrow can be differentiated into cardiac-like muscle cells with 5-aza inducement in vitro and the differentiationis in line with the 5-aza concentration.(J Geriatr Cardiol 2004;1(2):101-107.) Objective To investigate the potential of adult mesenchymal stem cells (MSCs) derived from human bone marrow to undergo cardiomyogenic differentiation after exposure to 5-azacytidine (5-aza) in vitro. Methods Asmall bone marrow aspirate was taken from the iliac crest of human volunteers, and hMSCs were isolated by 1.073 g / mL Percoll and propagated in the right cell culturing medium as previously described. The phenotypes of hMSCs were characterized with the use of flow cytometry. hMSCs were cultured in cell culture medium (as control) and medium mixed with 5 -aza for cellular differentiation. We examined by immunohistochemistry at 21 days the induction of desmin, cardiac-specific cardiac troponin I (cTnI), GATA 4 and connexin-43 respectively. Results The hMSCs are fibroblast-like morphology and express CD44 ~ + CD29 ~ 20-90% hMSCs connected with adjoining cells and coalesced into myotube structures after 14 days. Twenty-one days after 5-aza treatment, i mmunofluorescence showed that some cells expressed desmin, GATA4, cTnI and connexin-43 in 5, 10 μmol / L 5-aza groups, but no cardiac specific protein was found in 3 μmol / L 5-aza group nor in the control group. ratio of cTnI positively stained cells in 10 μmol / L group was higherthan that in 5 μmol / L group (65.3 ± 4.7% vs 48.2 ± 5.4%, P <0.05). Electron microscopy revealed that myofilaments were the induced cells expressed cardiac- myosin heavy chain (MyHC) gene by reversetranscription-polymerase chain reaction (RT-PCR) .Conclusions Theses findings suggest that hMSCs from adultbone marrow can be differentiated into cardiac-like muscle cells with 5-aza inducement in vitro and the differentiation in in line with the 5-aza concentration. (J Geriatr Cardiol 2004; 1 (2): 101-107.)