Effects of 5-Azacytidine on Differentiation of Ovine Mesenchymal Stem Cells

Abstract

Background: Mesenchymal stem cells (MSCs) have potential of self-renewal and differentiation into many cell types and can be used for cells therapy. Addition of epigenetic drugs can also convert these cells into cardiomyocytes. Objective: The aim of this study was to evaluate the effect of 5-azacytidine on differentiation of sheep fetal bone marrow MSCs into cardiomyocytes. Materials and Methods: Isolated bone sheep fetal marrow MSCs was cultured in DMEM: F12 medium. Passaged-3 cells were examined for their differentiation capacity into osteocytes and adipose cells. Surface antigens of isolated cells were analyzed using flow cytometry. Cells after third passage treated with 5 μM 5-azacytidine for 48h followed by medium without 5-azacytidine for twenty-one days. Expression of cardiac alpha-actinin and Troponin I (cTnI) were analyzed by immunohistochemistry. Results: MSCs expressed CD44, CD166 and weak expression of CD34 and CD45, apart from this expression a very weak expression of CD105 was also observed. Oil red for adipose cells and alizarin red staining for osteocytes indicated that the cells from all studied groups maintained the differentiation potential into adipose and osteocytes cells. Expression of Gata4, connexin 43, Atrial Natriuretic Peptide (ANP) was also detected by reverse transcriptase polymerase chain reaction and weak expression of troponinI. Sarcomeric alpha-actinin and cTnI were also partially detected by immunohistochemistry on these cells. Conclusion: This result indicated that addition of 5-azacytidine into culture medium could be effective on MSCs differentiation into cardiomyocytes