microRNA-1诱导大鼠骨髓间充质干细胞向心肌样细胞分化

miRNA-1 induces rat bone marrow mesenchymal stem cells differentiation into cardiomyocyte-like cells

目的研究microRNA-1(miRNA-1)能否诱导骨髓间充质干细胞(MSCs)向心肌样细胞分化。方法构建大鼠miRNA-1表达载体,分离扩增培养及鉴定大鼠MSCs。脂质体法转染大鼠第4代MSCs,实时定量RT-PCR(qRT-PCR)检测转染miRNA-1质粒后MSCs的miRNA-1表达水平。分别于转染2、4和6 d后用RT-PCR检测心肌重要转录因子GATA4、NKx2.5和MEF2C的mRNA表达,免疫荧光检测心肌特异蛋白I(cTnI)的表达。结果 90%以上的MSCs表达MSCs重要标志物CD29、CD44;未检测到造血前体细胞标志抗原CD34、白细胞标志抗原CD45的表达。转染miRNA-1质粒后,miRNA-1表达水平明显上调。转染miRNA-1质粒2、4和6 d后,GATA4、NKx2.5和MEF2C的mRNA表达逐渐增强。第4和6天后可见cTnI阳性表达细胞。结论 miRNA-1能诱导大鼠MSCs向心肌样细胞分化。

Objective To investigate may miRNA-1 induce rat bone marrow mesenchymal stem cells（rMSCs） differentiation into cardiomyocyte-like cells.Methods miRNA-1 expression vector was constructed,rMSCs were isolated,amplified and identified.The fourth passage of rMSCs were transfected with Lipofectamine2000 and miRNA-1 expression was then detected by quantitative real-time RT-PCR（qRT-PCR）.The mRNA expression of cardiac-specific transcription factor GATA4,NKx2.5 and MEF2C were detected by RT-PCR,cTnI expression was measured by immunofluorescence,respectively,2,4,6 days.Results Over 90% of the cultured rMSCs was positive for CD29 and CD44,as two significant markers of MSCs and negative for hematopoetic precursor cell markers CD34,leukocyte antigen CD45.After transfection of miRNA-1 in MSCs,the expression level of miRNA-1 was markedly enhanced.The mRNA expression of GATA4,NKx2.5 and MEF2C gradually increased from day 2 to day 6.The cTnI expression was detected at day 4,6.Conclusion miRNA-1 can induce MSCs differentiation into cardiomyocyte-like cells.