Evaluation of the differentiation process of human adipose tissue-derived mesenchymal stem cells to skeletal muscle-like cells: An in vitro study

Background This study was carried out to evaluate the development of skeletal muscle-like cells derived from human adipose tissue mesenchymal stem cells induced by 5-azacytidine [5-aza] under in vitro condition. Human adipose tissue mesenchymal stem cells [ADSCs] were purified. These cells were cultured in osteogenic or adipogenic induction medium to induce osteogenic and adipogenic differentiation. On the other hand، the skeletal myogenic differentiation potential of these cells was investigated using 3µmol 5-azacytidine [5-aza] treatment for 24 hours. Then the culture was washed out by PBS and were put in the culture medium free of 5-aza for another 4 weeks until sampling. RT-PCR assay was performed to detect the expression of specific skeletal muscle genes including Myogenin، Myh، alpha-actin، tropomyosin and myosin at 1-4 weeks after the first induction. The adherent adipose tissue-derived mesenchymal stem cells exhibited a significant proliferative capacity and also showed osteogenic and adipogenic differentiation as seen in previous studies. The expression of four skeletal muscle cells genes was detected 1st to 4th week after induction in a time-dependent manner. There was a continuous increase in expression of Myogenin gene from the 1st to 4th week، whereas that of the Myh was higher after two weeks and lower after 4 weeks in comparison to the other weeks. The levels of mRNA of alpha-actin and Myosin expression were at the highest level in the fourth week. Expression of tropomyosin gene was not significant in the samples. The current study indicated that stimulating human adipose tissue-derived mesenchymal stem cells by 5-aza with the concentration of 3µmol under in vitro condition can lead to differentiating skeletal muscle-like cells. Furthermore prolonged culture duration may lead to more differentiated cells.