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基于miR-21/Smad7探讨芍药苷在抗血管紧张素Ⅱ诱导心肌肥大中的作用
作者:张会超1  徐里2  芮浩淼1  曹程浩1  杨凤鸣1  袁彬3 
单位:1. 河南省中医院 心病科, 河南 郑州 450000;
2. 河南中医药大学 基础医学院, 河南 郑州 450000;
3. 河南中医药大学 研究生院, 河南 郑州 450000
关键词:心肌肥厚 芍药苷 microRNA-21 Smad7 
分类号:R-33;R541
出版年·卷·期(页码):2018·37·第二期(191-197)
摘要:

目的:探讨芍药苷(Pae)在对抗血管紧张素Ⅱ(Ang-Ⅱ)诱导心肌肥大中的作用及其机制。方法:Ang-Ⅱ诱导建立心肌细胞肥大模型,分为正常对照组、Ang-Ⅱ组、Pae组及Ang-Ⅱ+Pae组,脂质体转染法将微小RNA-21模拟物(miR-21 mimic)、信号转导分子7(Smad7) siRNA及其对照转入模型细胞。图像分析系统测算心肌细胞相对表面积,实时PCR检测miR-21及心房利尿钠肽(ANP)、脑利尿钠肽(BNP) mRNA表达,双荧光素酶实验检测miR-21对Smad7的靶向作用。Western blotting观察Smad7、ANP及BNP蛋白表达。结果:Ang-Ⅱ诱导显著增加心肌细胞表面积,ANP、BNP mRNA及蛋白表达,上调miR-21,Pae能部分逆转Ang-Ⅱ诱导的miR-21表达增加(P<0.05),降低心肌细胞表面积,ANP、BNP mRNA及蛋白相对表达量(均P<0.05)。MiR-21靶向Smad7 3'-UTR,发挥对Smad7转录后翻译的抑制作用,Pae能降低miR-21对Smad7的抑制(P<0.05),Smad7敲除后削弱了Pae对心肌细胞作用的相对表面积以及对ANP、BNP mRNA及蛋白表达的抑制作用(均P<0.05)。结论:Pae能通过负性调节miR-21降低靶蛋白Smad7表达,发挥抵抗心肌细胞肥大的作用。

Objective:To identify the role of paeoniflorin(Pae) on attenuating Ang-Ⅱ induced cardiomyocyte hypertrophy. Methods:A model of cardiac hypertrophy induced by Ang-Ⅱ in vitro was developed. Cardiomyocytes were separated and divided into four groups:control group, Ang-Ⅱ group, Pae group and Ang-Ⅱ plus Pae group. Cells were transfected with miR-21 mimic, Smad7 siRNA or their negative control group. Myocardial cell image analysis system was used to measure the relative surface area of cardiomyocytes; Real-time PCR was used to assay the mRNA expression of miR-21, ANP and BNP. A dual luciferase reporter assay was used to detect the relationship between Smad7 activity and miR-21 expression.Western blotting was used to evaluate the protein level of Smad7, ANP and BNP. Results:Ang Ⅱ pretreatment significantly increased the myocardial cell surface area, as well as up-regulated the miR-21, ANP and BNP expression, while paeoniflorin can partly reverse these effects brought by Ang Ⅱ (all P<0.05). Results of dual luciferase reporter assay showed that miR-21 directly targeted the 3'-UTR of Smad7, resulting in the post-transcriptional translation inhibition of Smad7 (P<0.05). Pae can partly reduce the inhibition effect of miR-21 on Smad7 (P<0.05), Smad7 knockdown remarkably attenuated the inhibition effect on cell relative surface area, ANP and BNP expression induced by Pae (P<0.05). Conclusion:These results indicate that Pae may resistance to Ang-Ⅱ induced cardiac hypertrophy via miR-21/Smad7 pathway.

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