敲低微小RNA-377表达可抑制高糖诱导的人肾小球系膜细胞增殖及炎性反应  

MicroRNA- 377 knockdown suppresses high glucose- induced proliferation and inflammation in human mesangial cells

作　　者：杨光[1] 黎小燕[2] 吴乙偲 钟树妹[2] Yang Guang;Li Xiaoya.n;Wu Yicai;Zhong Shumei(Department of Nephrology,Ganzhou People＇s Hospital,Ganzhou 341000,China)

机构地区：[1]赣州市人民医院肾内科,341000 [2]赣州市人民医院内分泌科,341000

出　　处：《中华肾脏病杂志》2018年第6期446-452,共7页Chinese Journal of Nephrology

基　　金：赣州市科技计划项目(GZ2016ZSF148)

摘　　要：目的探讨微小RNA（miRNA）-377对高糖诱导的人肾小球系膜细胞（HMC）增殖和炎性反应的影响，并初步分析其可能的作用机制。方法细胞分组为正常对照组（5．5mmol／L葡萄糖）、高糖组（30．0mmol／L葡萄糖）、阴性miRNA抑制物转染＋高糖组、阴性miRNA类似物转染＋高糖组、miRNA-377抑制物转染＋高糖组（miR-377i＋高糖组）、miRNA-377类似物转染＋高糖组（miR-377m＋高糖组）。实时定量PCR检测miRNA-377的表达。BrdU染色及流式细胞术分别检测HMC的增殖及细胞周期。ELISA检测促炎因子肿瘤坏死因子α（TNF-α）、白细胞介素（IL）-18、IL-6及单核细胞趋化蛋白1（MCP-1）的释放。Western印迹评估核因子KB（NF-κB）信号通路的活性，评估指标包括磷酸化（P）-IκBα、p-P65和核内P65。结果与正常对照组比较，高糖组HMC的细胞活力、miRNA-377表达、细胞增殖率均增加（均P〈0．05），细胞周期中S期、G2期／M期细胞比例增加（均P〈0．05），炎性因子TNF-α、IL-18、IL-6、MCP-1的表达均增加（均P〈0．05），胞内p-IκBα/IκBα、p-P65／P65的表达及核内P65水平均升高（均P〈0．05）。与高糖组比较．miR-377i＋高糖组细胞增殖率降低（P〈0．05），细胞周期中S期、G2期／M期细胞比例均降低（均P〈0．05），TNF-α、IL-18、IL-6、MCP-1的表达均减少（均P〈0．05），细胞内p-IκBα/IκBα、p-P65／P65的表达及核内P65水平均降低（均P〈0．05）。miR-377m＋高糖组与miR。377i＋高糖组结果相反（均P〈0．05）。结论miRNA-377表达下调可部分逆转高糖诱导的HMC增殖和周期转换，并可抑制炎性因子的释放。其作用机制可能与抑制NF—KB的活化有关。Objective To investigate the effect and potential mechanism of microRNA （miRNA）- 377 on high glucose- induced proliferation and inflammation in human mesangial cells. Methods Cells were randomly divided into six groups： control group （5.5 mmol/L glucose）, high glucose group （30.0 mmol/L glucose）, negtive miRNA inhibitor transfection＋high glucose group, negtive miRNA mimic transfection＋high glucose group, miRNA-377 inhibitor transfection＋high glucose group （miR-377i＋high glucose group）, miRNA-377 mimic transfection＋high glucose group （miR-377m＋high glucose group）, milRNA-377 expression was detected by real-time PCR. Cell proliferation and cell cycle were detected by BrdU assay and flow cytometry, respectively. The release of tumor necrosis factor- （TNF- α）, interleukin （IL）- 18, IL-6 and macrophages ehemotaxis protein-1 （MCP-l） were evaluated by ELISA. The activations of NF-κB pathway, including the expressions of phosporylated （p）-IκBα, p-P65 and nuclear P65, were measured by Western blotting. Results Compared with those in control group, in high glucose group cell viability, miRNA-377 expression and cell proliferation rate increased （all P 〈 0.05）, proportions of S phase cell and G2/M phase cell in cell cycle increased （all P 〈 0.05）, the levels of TNF-α, IL-18, IL-6 and MCP-1 were higher （all P 〈 0.05）, as well as the expressions of p-IκBα/IκBα, p-P65/P65 and nuclear P65 were increased （all P 〈 0.05）. Compared with high glucose group, cell proliferation rate was restrained （P 〈 0.05）, proportions of S phase cell and G2/M phase cell in cell cycle was descreased （all P 〈 0.05）, the levels of TNF-α, IL-18, IL-6 and MCP-1 were lower （all P 〈 0.05）, as well as the expressions of p-IκBα/IκBα, p-P65/P65 and nuclear P65 were reduced （all P 〈 0.05） in miR- 377i ＋high glucose group. However, miR- 377m＋high glucose group presented opposite results （all P 〈 0.05）. Conclusions miRNA- 377 knockdown can p

关 键 词：微RNAS 肾小球系膜细胞 细胞增殖 炎症 信号传导 NF-κB 

分 类 号：R587.2[医药卫生—内分泌] R692.9[医药卫生—内科学]