Bone Morphogenetic Protein 9 and 13 Induce C3H10T1/2 Cell Differentiation to Cardiomyocyte-Like Cells In Vitro

The present study aimed to evaluate the effect of bone morphogenetic protein 9 (BMP9) and BMP13 on cardiac differentiation of C3H10T1/2 cells in vitro and to characterize the differentiated cells on their ultrastructure and transmembrane electrophysiological features. C3H10T1/2 cells were transfected with the vectors for BMP9 or BMP13 and differentiated into cardiomyocytes in vitro for up to 28 days. The expression of cardiac-specific genes Gata4 and Mef2c and proteins troponin T (cTnT) and connexin 43 (Cx43) was significantly increased in the cells transfected with BMP9 or BMP13 after differentiation over the controls as evaluated using quantitative RT-PCR, Western blotting, and immunofluorescence staining. Transmission electron microscopy and Masson trichrome staining showed that the specific myocardial leap dish and myofilament-like structure were present in the cells overexpressing BMP9 or BMP13, not in the control cells. Whole-cell patch-clamping study demonstrated the presence of delayed rectifier potassium current, inward rectifier potassium current, and T-type calcium current in the cells overexpressing BMP9 or BMP13. Sodium current was detected in a small number of cells overexpressing BMP9, not in the BMP13-transfected cells or the control cells. The expression of Mef2c gene and Cx43 and cTnT proteins was also significantly higher in the cells overexpressing BMP9 than those overexpressing BMP13. Our data indicate that BMP9 and BMP13 (BMP9 might be more effective) promoted the differentiation of C3H10T1/2 cells into cardiomyocyte-like cells with cellular ultrastructures and ion channel currents similar to mature cardiomyocytes in vitro.