miR-99a对高胰岛素诱导小鼠主动脉血管平滑肌细胞增殖的影响及机制研究  

Effect of miR-99a on insulin-stimulated proliferation of vascular smooth muscle cells in mice and its mechanism

作　　者：张紫微[1,2] 杨丽霞[1] 郭瑞威[1] 吕晋琳[1,2] 陆霓虹 王先梅[1] 石燕昆[1] ZHANG Ziwei YANG Lixia GUO Ruiwei LYU Jinlin LU Nihong WANG Xianmei SHI Yankun(Department of Cardiology, Kunming General Hospital of Chengdu Military Area, Kunming 650032, Yunnan Province, China Postgraduate Department, Kunming Medical University, Kunming 650500, Yunnan Province, China)

机构地区：[1]成都军区昆明总医院心内科,云南昆明650032 [2]昆明医科大学昆明总医院临床学院,云南昆明650500

出　　处：《解放军医学院学报》2016年第11期1164-1168,共5页Academic Journal of Chinese PLA Medical School

基　　金：国家自然科学基金面上项目(81170250)~~

摘　　要：目的观察mi R-99a对高浓度胰岛素诱导小鼠主动脉血管平滑肌细胞(vascular smooth muscle cell,VSMC)增殖的影响及可能机制。方法原代培养小鼠主动脉血管平滑肌细胞,用超生理浓度剂量胰岛素(100 nmol/L)作用于血管平滑肌细胞,用荧光实时定量PCR(quantitative real-time PCR,q RT-PCR)检测mi R-99a表达变化情况。Brdu法检测转染mi R-99a对高胰岛素促细胞增殖的影响。q RT-PCR及Western blot检测转染mi R-99a组及阴性对照组哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,m TOR)m RNA及蛋白变化情况,并检测转染mi R-99a对高胰岛素促进m TOR表达及对其下游通路激活的影响。Western blot检测转染mi R-99a组及阴性对照组细胞周期素D1(Cyclin D1)的变化。结果高浓度胰岛素显著降低mi R-99a的表达。转染mi R-99a mimic后可减少基础的及胰岛素促进的VSMC增殖作用。mi R-99a负向调节m TOR m RNA及蛋白,转染mi R-99a mimic可显著削弱高胰岛素促进的m TOR/P70S6K1通路激活作用,而转染mi R-99a inhibitor可增加高胰岛素促进m TOR/p70s6k1通路激活作用。转染mi R-99a mimic后可减少基础的及胰岛素促进的Cyclin D1表达。结论 mi R-99a可通过负向调节m TOR抑制高胰岛素诱导的VSMC细胞增殖作用。Objective To explore the effect of mi R-99 a on proliferation of insulin-stimulated vascular smooth muscle cell（VSMC） in mice and investigate its detailed mechanism. Methods The vascular smooth muscle cells derived from C57 mice were cultured by adherent method, and the cells were treated with 100 nmol/L of insulin for 48 h. Quantitative real-time PCR（q RT-PCR） was adopted to investigate the changes of mi R-99 a expression. Brdu assay was used to evaluate the effect of transfected mi R-99 a on insulininduced proliferation of VSMCs. q RT-PCR and western blot analysis were used to detect m RNA and protein expression changes of Mammalian target of rapamycin（m TOR） after mi R-99 a transfection in experimental group and negative control and explore the effect of mi R-99 a on insulin-enforced m TOR/P70S6 K signaling pathway. And western blot analysis was used to detect the expression of Cyclin D1 after mi R-99 a transfection in experimental group and negative control. Results High dose of insulin significantly decreased the expression of mi R-99 a. Transfection of mi R-99 a mimics remarkably attenuated basal and insulin-stimulated VSMCs proliferation, and downregulation insulin-activated m TOR/P70S6 K signaling pathway and insulin-enforced expression of Cyclin D1. In contrast, transfection of mi R-99 a inhibitors further enhanced insulin-activated m TOR/P70S6 K signaling pathway. ConclusionMi R-99 a can inhibit insulin-stimulated VSMC proliferation by downregulation of m TOR.

关 键 词：微小RNA 99a 胰岛素 血管平滑肌细胞 

分 类 号：R587.2[医药卫生—内分泌]