Differentiation of mesenchymal stem cells into cardiomyocyte-like cells induced by H9C2 cell culture medium / 中国组织工程研究

ABSTRACT

BACKGROUND:Mesenchymal stem cels can differentiate into cardiomyocytesin vitro induced by different methods, such as chemical drug induction, autologous transplantationin vivo, andin vitro simulation of cardiac-like microenvironment, but these inducible methods show low induction rate, complex operation, and toxic side effects. OBJECTIVE:To investigate the role of H9C2 cellculture medium in the differentiation of mesenchymal stem cels into cardiomyocyte-like cels. METHODS: Mesenchymal stem cels were obtained by the whole bone marrow adherent culture and H9C2 cellculture medium was prepared as a culture medium. Then mesenchymal stem cels were co-cultured with H9C2 cellculture medium for 1, 3, 5, 7 days. H9C2 cels cultured in 10% Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 served as positive control groups, while mesenchymal stem cels cultured in 10%Dulbecco’s modified Eagle’s medium/Ham’s nutrient mixture F-12 as negative control group. Immunofluorescence and western blot assay were used to detect expression of myocardial celljunction protein (desmin) and troponin T. Real-time quantitative PCR was applied to detect mRNA expression of myocardial celltrait genes, α-cardiac myosin heavy chain and β-myosin heavy chain. RESULTS AND CONCLUSION:After co-culture of H9C2 cellculture medium and mesenchymal stem cels for 7 days, the troponin T positive cels were up to (16±7)%, which was significantly higher than that of non-induced mesenchymal stem cels. Compared with the negative control group, the expression of troponin T protein and desmin after induction were significantly increased (P < 0.05) by western-blot detection; real-time PCR showed that the mRNA expression ofα-cardiac myosin heavy chain and β-myosin heavy chain in differentiated cels were both up-regulated (P < 0.05). These findings suggest that H9C2 cellculture medium may induce the differentiation of mesenchymal stem cels into cardiomyocyte-like cels.