Experimental study on MyoD gene induced differentiation of bone marrow mesen-chymal stem cells into myoblasts in vitro

摘要

Objective: To explore the possibility of the transfection of MyoD gene induced bone marrow mesenchymal stem cells ( MSCs) to differentiate into myoblasts in vitro. Methods: The eukaryotic expression plasmid vector pIRES2-EGFP-MyoD was transfected into MSCs with lipotransfection method, and the positive cells were selected by G418; The expression of MyoD was detected in the transfected MSCs with RT-PCR and the amplified, purified product was identified by sequencing; The reporter gene enhanced green fluorescence protein ( EFGP) was observed in the transfected cells under a fluorescent and a laser confocal microscopes; Immunohistochemical methods was used to examine the expressions of MyoD, myogenin, myosin, myoglobin and desmin in the differentiated cells. The ultrastructure changes of the cells before and after transfection were observed with electron microscopy. Results: The expression of MyoD was detected in the transfected MSCs with RT-PCR and the amplified, purified product was as same in sequence as that from Genbank; Green fluorescence was observed in the transfected cells under a fluorescent and a laser confocal microscopes; Immunohistochemical methods indicated that MyoD, myogenin, myosin, myoglobin and desmin were expressed in the transfected cells; The transfected cells showed the morphological characteristics of mature cells with filaments in their cytoplasm. Conclusion: MyoD gene can induce cultured MSCs to successfully differentiate into myoblasts , probably providing an experimental foundation for trauma repair.