微小RNA-29b在肾小管上皮细胞的表达及调节上皮间质转分化的作用机制  

Expression of microRNA-29b in renal tubular epithelial cells and its effects on epithelial mesenchymal transition

作　　者：胡洪涛[1] 曾芳[1] 高月[1] 廖星[1] 水华[1] 

机构地区：[1]武汉大学中南医院肾内科,武汉430071

出　　处：《临床肾脏病杂志》2016年第7期427-431,共5页Journal Of Clinical Nephrology

基　　金：湖北省自然科学基金(NO.2013CFB227)

摘　　要：目的研究血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)对大鼠肾小管上皮细胞(NRK-52E)中微小核糖核酸(miRNA)-29b(miR-29b)表达的影响,以及miR-29b在细胞间质转分化中的作用,为研究肾间质纤维化机制提供新思路。方法体外培养NRK-52E,将不加入AngⅡ的细胞作为对照组,加入AngⅡ的细胞为实验组,按加入不同浓度的AngⅡ(10^(-9)mol/L、10^(-7)mol/L)及AngⅡ刺激时间(12 h、24 h、48 h)的长短又分为实验亚组,同时设立miR-29b mimic转染组及阴性对照组。显微镜下观察对照组与实验组细胞的形态学改变,运用实时荧光定量聚合酶链反应检测rniR-29b表达,蛋白质免疫印迹试验检测Snail、E钙黏附素(E-cadherin)、α平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)蛋白表达。两两比较用t检验。结果实验组中AngⅡ刺激12 h后,细胞形态呈梭形改变;随着AngⅡ刺激浓度的增加,α-SMA、Snail蛋白表达显著升高,E-cadherin表达量显著下降,与对照组相比,差异均有统计学意义(P<0.05)。实验组中随着AngⅡ刺激时间的延长,细胞α-SMA、Snail和E-cadherin的表达呈时间依赖性,α-SMA、Snail的表达显著增加,E-cadherin表达量显著降低,与对照组相比,P<0.05。同时AngⅡ呈浓度和时间依赖性下调NRK-52E细胞中miR-29b表达,与对照组相比,P<0.05。此外,miR-29b mimic转染组和阴性对照组相比,α-SMA、Snail蛋白表达显著降低(P<0.05),E-cadherin表达量显著升高(P<0.05)。结论 AngⅡ可诱导NRK-52E细胞发生间质转分化,且miR-29b可抑制AngⅡ诱导的NRK-52E细胞发生间质转分化。Objective To observe the effects of angiotensin H （Ang Ⅱ ） on the expression of microRNA-29b （miR-29b） in renal tubular epithelial cells （NRK-52E cells）, and to investigate wheth- er miR-29b regulated the fibrogenic epithelial mesenchymal transition （EMT） in NRK-52E cells and explore the underlying mechanisms. Methods The cells in the absence of Ang Ⅱ served as control group, and those treated with Ang Ⅱ as experimental group. According to the different concentra- tions of Ang Ⅱ （10-9, and 10-Tmol/L） and the different time of Ang Ⅱ （12 h, 24 h, and 48 h）, the cells were divided into some subgroups, miR-29b mimic group and negative control group were also conducted. With 10-7 mol/L Ang Ⅱ stimulating NRK-52E cells, we observed the changes in cellular morphology. Real-time PCR was used to detect the expression of miR-29b after Ang Ⅱ induction. Western blotting was used to detect the changes of Snail, E-cadherin and α-SMA proteins. T-test was used to evaluate the differences in means among the groups. Results After stimulation with Ang Ⅱ , NRK-52E cells showed morphological changes in spindle. The expression of α-SMA and Snail protein was markedly increased, and that of E-cadherin protein markedly decreased in a dose-dependent man- ner after stimulation with Ang Ⅱ as compared with control groups, P〈0. 05. Ang Ⅱ stimulation markedly increased the expression of a-SMA and Snail protein, and decreased the expression of E-cad- herin protein in a time-dependent manner as compared with control groups, P〈0. 05. Treatment of NRK-52E cells with Ang Ⅱ caused a remark decrease in the expression of miR-29b in a dose- and time-dependent manner, as compared with control groups, P〈0. 05. Besides, miR-29b mimic group displayed a decrease in the expression of a-SMA and Snail protein, and an increase in the expression of E-cadherin protein compared to negative control group （P〈0. 05）. Conclusions The results suggest that inducing miR-29b expression may be one of the mechanisms

关 键 词：miR-29b 上皮间质转化 血管紧张素Ⅱ 肾小管上皮细胞 

分 类 号：R692[医药卫生—泌尿科学]