miR-219与TGFBR2的调控关系在肾脏纤维化中的作用  

Role of miR-219 and TGFBR2 in pathogenesis of renal fibrosis

作　　者：李霜青[1] 应俊[1] 许旭春[1] LI Shuang-qing;YING Jun;XU Xu-chun(Jinhua Municipal Central Hospital, Jinhua 321000 , China)

机构地区：[1]金华市中心医院,浙江金华321000

出　　处：《中国病理生理杂志》2019年第4期703-709,共7页Chinese Journal of Pathophysiology

基　　金：金华市中心医院课题(No.JY2016-02-14)

摘　　要：目的:研究微小RNA-219(miR-219)通过靶向调控转化生长因子βⅡ型受体(TGFBR2)在肾脏纤维化中发挥的作用。方法:收集2017年9月～2018年3月于我院就诊的70例肾脏纤维化患者,选取同期来本院体检的20例健康人设为对照组,RT-qPCR检测肾脏纤维化患者及对照组血清中miR-219的表达水平,并检测血管紧张素Ⅱ(AngⅡ)刺激大鼠肾成纤维细胞NRK49F后miR-219的表达水平。Western blot检测转染miR-219模拟物(miR-219 mimics)的NRK49F细胞在AngⅡ刺激后α-平滑肌肌动蛋白(α-SMA)的蛋白表达情况。筛选出miR-219的潜在靶基因TGFBR2,并通过萤光素酶报告基因法进行验证。RT-qPCR和Western blot检测miR-219 mimics对TGFBR2的mRNA及蛋白表达的影响。RT-qPCR检测miR-219 mimics对α-SMA、结缔组织生长因子(CTGF)、Ⅰ型胶原α1链(COL1A1)和COL3A1的mRNA表达水平的影响。构建单侧输尿管闭塞(UUO)小鼠模型并检测其肾脏组织中miR-219的表达水平,对UUO小鼠注射miR-219后观察肾脏纤维化的变化情况,并检测COL1A1和COL3A1的mRNA表达水平。结果:肾脏纤维化患者血清中miR-219的表达水平明显低于对照组,UUO小鼠肾脏组织中miR-219的表达显著下降(P<0.01);AngⅡ刺激NRK49F细胞后miR-219的表达水平明显降低,且miR-219 mimics可抑制α-SMA蛋白的表达(P<0.01);miR-219 mimics对TGFBR2具有靶向调控作用,可抑制TGFBR2的mRNA及蛋白表达;miR-219 mimics可抑制α-SMA、CTGF、COL1A1和COL3A1的mRNA表达水平;miR-219可下调UUO小鼠中COL1A1和COL3A1的mRNA表达水平并抑制其肾脏纤维化进程。结论:miR-219可通过抑制TGFBR2的表达从而抑制肾脏纤维化的发展,可能成为肾脏纤维化诊断及治疗的新靶点。AIM: To study the role of microRNA-219(miR-219) in regulation of transforming growth factor-β receptor type 2(TGFBR2) in renal fibrosis. METHODS: The renal fibrosis patients(n=70) were selected in this stu-dy, and 20 cases of healthy people were selected as control group. RT-qPCR was used to detect the expression of miR-219 in the serum of the patients with renal fibrosis and control group, and the expression of miR-219 in NRK49 F cells after stimulation with angiotensin Ⅱ(AngⅡ) was detected. The protein expression of α-smooth muscle actin(α-SMA) in the NRK49 F cells transfected with miR-219 mimics after stimulation with AngⅡ was determined by Western blot. The potential target gene TGFBR2 of miR-219 was screened and verified by the method of luciferase reporter gene. RT-qPCR and Western blot were used to detected the effect of miR-219 mimics on the expression of TGFBR2 at mRNA and protein levels, and the mRNA expression of α-SMA, connective tissue growth factor(CTGF), type Ⅰ collagen α1(COL1A1) and COL3A1 in the NRK49 F cells was also detected, respectively. The unilateral ureteral occlusion(UUO) mouse model was established and the expression of miR-219 in the renal tissue was monitored. The morphological change of renal fibrosis was observed in the UUO mice after injection of miR-219, and the mRNA expression levels of COL1A1 and COL3A1 were detected. RESULTS: The expression level of miR-219 in the patients with renal fibrosis was significantly lower than that in control group, and the expression of miR-219 in the UUO mice was decreased significantly(P<0.01). The expression level of miR-219 was significantly decreased in the NRK49 F cells after AngⅡ stimulation, and miR-219 mimics inhibited the protein expression of α-SMA(P<0.01). miR-219 mimics had a targeted regulatory effect on TGFBR2 gene, which inhibited the mRNA and protein expression of TGFBR2. miR-219 mimics inhibited the mRNA expression of α-SMA, CTGF, COL1A1 and COL3A1. miR-219 also down-regulated the mRNA expression of COL1A1 and COL3

关 键 词：肾脏纤维化 微小RNA-219 转化生长因子βⅡ型受体 单侧输尿管闭塞 NRK49F细胞 

分 类 号：R692[医药卫生—泌尿科学] R363[医药卫生—外科学]