Objective:To investigate whether bone marrow multipotent adult progenitor cells(MAPCs)undergo cardiac specification and its dependablity with octamer-binding transcription factor-4(Oct-4)expression during their cardiomyocyte differentiation in vitro.Methods:MAPCs were isolated from rat bone marrow

treated with 5-azacytidine(5-aza

1 μmol/L)for 24 h

and cultured in a serum-free medium for cardiac differentiation for up to 35 days.Results:The cells started to express early cardiac-specific genes Nkx2.5 and GATA-4 with a significant increase in their mRNA level after 24h with 5-aza treatment.Western blot analysis and immunofluorescence staining revealed that the cardiac-specific proteins connexin-43 and troponin Ⅰ were expressed in the cells 7 days after 5-aza treatment.Flow cytometry analysis demonstrated that over 37% of the cells were positive for troponin Ⅰ by 35 days of differentiation

though the cells did not display spontaneous contraction.Meanwhile

the undifferentiated MAPCs expressed a significant level of the stem-cell-specific marker Oct-4 that was dramatically decreased in the cells shortly after the initiation of cardiomyocyte differentiation as evaluated using real-time(RT)-polymerase chain reaction

Western blot

immunofiuorescence staining

and flow cytometry.Conclusion:Downregulation of Oct-4 expression effectively accelerate cardiomyocyte differentiation of rat MAPCs after 5-aza induction in in vitro culture.