microRNA-21通过促进成纤维细胞的增殖和分化调节心肌梗死后的心脏重塑

MicroRNA-21 Regulates Cardiac Remodeling by Promoting Proliferation and Differentiation of Fibroblast after Myocardial Infarction

目的探讨小鼠心肌梗死模型中microRNA-21(miR-21)对心脏成纤维细胞增殖和分化的影响。方法利用左前降支结扎法构建小鼠心肌梗死模型(心肌梗死组),以心电图及病理改变作为建模成功的观察指标。采用荧光定量PCR检测各组心肌组织miR-21的表达。利用miR-21的模拟物(miR-21 mimic)转染小鼠成纤维细胞(miR-21 mimic组),使其在细胞中呈过表达状态,然后标记5-乙炔基-2′脱氧尿嘧啶核苷(EdU),通过荧光显微镜观察成纤维细胞的增殖情况,蛋白质印迹法检测成纤维细胞α平滑肌肌动蛋白(α-SMA)和小鼠Smad同源物7(Smad7)的表达,并与随机对照组及空白对照组比较。结果与假手术组比较,心肌梗死组梗死交界区的miR-21的表达量明显上调[(6.043±0.231)×10-4vs(1.620±0.451)×10-4,P<0.01],miR-21 mimic组成纤维细胞miR-21基因表达较随机对照组和空白对照组显著上调[(4.839±0.705)×10-4vs(1.143±0.064)×10-4vs(1.017±0.201)×10-4,P<0.01],miR-21mimic组成纤维细胞EdU染色阳性率较随机对照组和空白对照组显著增加[(27.892±1.645)%vs(12.553±1.227)%vs(13.946±1.550)%,P<0.01];同时,miR-21 mimic组成纤维细胞Smad7蛋白的表达明显下调,而α-SMA的表达显著上调。结论心肌梗死后,上调的miR-21可以通过调节转化生长因子(TGF)-β信号通路中Smad7的表达促进成纤维细胞的增殖和分化,进而调节心梗后的心脏重塑。

Objective To investigate the role of microRNA-21（miR-21） on cardiac fibroblast proliferation and dif-ferentiation in the mouse model of myocardial infarction. Methods The mouse model of myocardial infarction （MI） was es-tablished by ligation of the left coronary artery in male C57BL/6 mice（MI group）. The echocardiographic assessment and his-tological evaluation were performed after ligation. The expression levels of miR-21 were measured by quantitative real-time PCR in the various myocardial tissues. The cardiac fibroblasts transfected with miR-21 mimic were over-expressed miR-21. The proliferation was assessed by immunostaining for 5-ethynyl-2’-deoxyuridine （EdU）. Western blot assay was used to detect the expression ofα-SMA and Smad7 in the cardiac fibroblasts,and compared with control group and blank group. Results The expression of miR-21 was significantly increased in border area in MI group than that of sham group [（6.043 ±177; 0.231）±215;10-4 vs（1.620±177;0.451）±215;10-4,P〈0.01]. There was a higher expression of miR-21 in miR-21 mimic group than that of control group and blank group [（4.839±177;0.705）±215;10-4 vs（1.143±177;0.064）±215;10-4 vs（1.017±177;0.201）±215;10-4,P〈0.01]. The EdU positive rate was significantly higher in miR-21 mimic group than that of control group and blank group[（27.892±177;1.645）%vs（12.553 ±177; 1.227）% vs（13.946 ±177; 1.550）%,P〈0.01]. The expression of α-SMA was significantly increased in miR-21 mimic group, while the expression of Smad7, a target gene of miR-21, was significantly decreased. Conclusion The over-expression of miR-21 in cardiac fibroblasts disrupts TGF-βsignaling pathway by reducing the expression of Smad7, which promotes the proliferation and differentiation of cardiac fibroblast, and finally regulates cardiac remodeling after myocardial infarction.