Role of 5-azacytidine in the differentiation of rat bone marrow mesenchymal stem cells into cardiomyogenic cells

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Abstract

Introduction

Increasing attention is being paid to the use of mesenchymal stem cells (MSCs) for treatment of human diseases such as myocardial infarction.

Aim

To study the differentiation of the bone marrow mesenchymal stem cells (BM-MSCs) into cardiomyogenic cells using 5-azacytidine.

Materials and Methods

Forty adult male albino rats were used in this study. BM-MSCs were isolated and cultured in a complete Dulbecco's modified Eagle's medium containing 1% antibiotics and 10% fetal bovine serum (the control group). Second passaged cells were treated with 10μmol/l 5-azacytidine for 72h. Then, the medium was removed and kept in a 5-azacytidine-free medium for 4 weeks (the 5-azacytidine-treated group). The adherent cells of both groups were examined using a phase-contrast microscope and a transmission electron microscope. Expressions of cytoskeleton protein desmin and cardiac muscle-specific cardiac troponin T were assessed by immunohistochemistry.

Results

BM-MSCs of the control group were spindle and star shaped with multiple processes and vesicular nuclei. After adding 5-azacytidine for 1 week, the cells showed multinucleation. On the second week, the cells formed stick-like structures. The cells showed extensive cytoplasmic striations in the third week. Finally, in the fourth week, the cells formed myotube-like structures. Immunohistochemical staining of cells of the 5-azacytidine-treated group revealed a positive immune reaction for desmin and cardiac troponin-T. Ultrastructural examination of the 5-azacytidine-treated group revealed that the cells were elongated with central oval large nuclei. The mitochondria were elongated with well developed cristae. There were abundant free ribosomes and extensive dilated rough endoplasmic reticulum. Myofibrils started to appear in the peripheral part of the cytoplasm and T-tubules appeared.

Conclusion

MSCs can be differentiated in vitro by 5-azacytidine into cardiomyogenic cells, which are important for repairing infracted myocardium.

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