[Cardiomyocyte-like differentiation of human bone marrow mesenchymal stem cells after exposure of 5-azacytidine in vitro].
To investigate the potential of adult mesenchymal stem cells (hMSCs) derived from human bone marrow to undergo cardiomyogenic differentiation after exposure of 5-azacytidine in vitro. A small bone marrow aspirate was taken from the iliac crest of human volunteers, and hMSCs were isolated by 1.073 g/mL Percoll and cultured in the right cell culturing medium as previously described. The phonotypes of hMSCs were identified by flow cytometry. The stem cells were cultured in cell culture medium (as control) and medium mixed with 5-azacytidine (5-aza, 3, 5, 10 micromol/L) (n=5, respectively) for cellular differentiation. We examined respectively with immunohischemistry at 21 days of inducement on desmin, cardiac-specific cardiac troponin I (cTnI), GATA4 & connexin43. The ultrastructures of induced cells were examined by transmission electron microscope. The results indicated that the hMSCs showed a fibroblast-like morphology with vortex distribution in their peak propagation, and express high level of CD44 but negative for CD34 and CD45. 20%-30% cells grown after 5, 10 microl/L 5-aza treatment connected with adjoining cells and coalesced into myotube structures after 14 days. After 21 days of culturing, immunohistochemistry revealed expression of desmin, GATA4, cTnI and connexin43 in 5, 10 micromol/L showed positive, but no cardiac specific protein were found in neither 3 micromol/L nor in control group. The ratio of cTnI positive stained cells in 10 micromol/L group were higher than that in 5 micromol/L group (65.3+/-4.7% vs 48.2+/-5.4%, p<0.05). Electron microscopy revealed myofilaments were formed. The results indicated that purified hMSCs from adult bone marrow can be differentiated into cardiac-like muscle cells with 5-aza inducement in vitro and the differentiation is in line with the 5-aza concentration.
Cao
Feng
F
Beijing Institute of Transfusion medicine, Beijing 100850, China.
Niu
Li Li
LL
Meng
Ling
L
Zhao
Lian Xu
LX
Zheng
Ming
M
Yue
Wen
W
Bai
Ci Xian
CX
Jia
Guo Liang
GL
Pei
Xue Tao
XT
D000328:Adult; D001374:Azacitidine / Q000494:pharmacology*; D001854:Bone Marrow Cells / Q000166:cytology*; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D032383:Myocytes, Cardiac / Q000378:metabolism* / Q000648:ultrastructure
D001374:Azacitidine