MicroRNA-29c通过TGF-β1通路对扩张性心肌病心肌纤维化调控的实验研究

Experimental study on the regulation of myocardial fibrosis in dilated cardiomyopathy by microRNA-29c via the TGF-β1 pathway

目的研究microRNA-29c(miR-29c)在扩张性心肌病(DCM)心肌纤维化过程中的作用。方法收集因DCM行心脏移植患者的心肌组织标本3例,作为DCM组,意外创伤性脑死亡而无心脏疾病患者的心肌组织3例,作为对照组;通过Masson染色观察心肌纤维化状况,阿霉素处理心肌成纤维细胞,上调miR-29c表达,观察对心肌纤维化的影响;应用RT-PCR、Western Bolt法检测转化生长因子-β1(TGF-β1)、ColⅠ、ColⅢmRNA和蛋白表达水平;CCK8检测心肌成纤维细胞的活力。结果DCM组心肌组织中心肌纤维化严重,miR-29c表达下降,TGF-β1mRNA和蛋白表达增加(P<0.05);阿霉素处理后的心肌成纤维细胞活动度增加(P<0.05);转染miR-29c mimic后的细胞TGF-β1、Co.lⅠ、ColⅢmRNA表达显著降低(P<0.05)。结论DCM心肌组织心肌纤维化严重,miR-29c能减弱TGF-β1对心肌纤维化的诱导作用,抑制细胞病理性增生,可能通过TGF-β1通路调控DCM心肌纤维化。

Objective To investigate the role of microRNA-29c(miR-29c)in myocardial fibrosis associated with dilated cardiomyopathy(DCM).Methods Myocardial tissue samples were collected from three patients undergoing heart transplantation due to DCM(DCM group)and from three patients who suffered accidental traumatic brain death without heart disease(control group).Myocardial fibrosis was assessed using Masson's trichrome staining.Cardiac fibroblasts were cultured and treated with doxorubicin.miR-29c mimic was transfected into cardiac fibroblasts to upregulate miR-29c expression,and its effects on myocardial fibrosis were observed.RT-PCR and Western blot analyses were used to detect the expression levels of transforming growth factor-β1(TCF-β1),collagen I(Col I),and collagen II(Col II)mRNA and proteins.The CCK8 assay was employed to assess the viability of cardiac fibroblasts.Results Severe myocardial fibrosis was observed in the myocardial tissues of the DCM group,with a significant downregulation of miR-29c expression and a significant increase in TGF-β1 mRNA and protein expression(P<0.05).CCK8 assays indicated increased activity in cardiac fibroblasts treated with doxorubicin(P<0.05).Furthermore,after transfection with the miR-29c mimic,there was a significant decrease in TCF-β1,Col I,and Col II mRNA expression in cardiac fibroblasts(P<0.05).Conclusion Severe myocardial fibrosis is present in the myocardial tissues of DCM patients.miR-29c can attenuate the induction of myocardial fibrosis by TGF-β1 and inhibit the pathological proliferation of cardiac fibroblasts.miR-29c may regulate DCM myocardial fibrosis through the TGF-β1 pathway.