前列腺素E2受体1亚型在转化生长因子β1诱导小鼠系膜细胞损伤中的作用机制  

Role of prostaglandin E2 receptor subtype 1 on TGF-β1-induced mouse mesangial cell

作　　者：马雯[1] 徐玉音[1] 仇芝[1] 范亚平[1] 陈晓岚[1] 

机构地区：[1]江苏南通大学附属医院肾内科,南通226001

出　　处：《肾脏病与透析肾移植杂志》2015年第1期44-49,共6页Chinese Journal of Nephrology,Dialysis & Transplantation

基　　金：国家自然科学基金(81170656);南通市科技项目(HS2011021);南通大学研究生科技创新计划项目(YKC13078)

摘　　要：目的:探讨前列腺素E2受体1亚型(EP1)对转化生长因子β1(TGF-β1)诱导的小鼠肾小球系膜细胞(MCs)增殖、细胞外基质的影响及可能机制。方法:体外原代培养MCs。采用脂质体Lipofectamine TM 2000化学合成法将siRNA转染至MCs中沉默EP1受体,筛选干扰效率最大的EP1-siRNA片段。实验分组:(1)正常对照组;(2)TGF-β1(10 ng/ml)组;(3)NC-siRNA+TGF-β1(10 ng/ml)组;(4)EP1-siRNA+TGF-β1(10 ng/ml)组;(5)EP1-siRNA组。ELISA法检测各组细胞上清中前列腺素E2(PGE2)的表达,实时荧光定量PCR方法检测各组细胞纤维连接蛋白(FN)、结缔组织生长因子(CTGF)、环氧化酶2(COX2)、膜结合型前列腺素E2合酶1(m PGES-1)mRNA的表达。Western blot法检测FN、CTGF、COX2、m PGES-1蛋白及p38MAPK、ERK活性的变化。结果:与正常对照组相比,TGF-β1组FN、CTGF、COX2、m PGES-1的mRNA和蛋白的表达上调(P<0.05);与TGF-β1组相比,EP1-siRNA+TGF-β1组FN、CTGF、COX2、m PGES-1的mRNA和蛋白的表达下调(P<0.05);TGF-β1刺激后,p38MAPK、ERK磷酸化水平明显增强(P<0.05),EP1-siRNA+TGF-β1组,p38MAPK、ERK磷酸化水平较对照组明显下调(P<0.05)。结论:EP1-siRNA可能通过抑制ERK和p38MAPK磷酸化减轻TGF-β1诱导的小鼠系膜细胞损伤。Objective：To investigate the effect and mechanisms of prostaglandin E2 receptor subtype 1 （EP1） on transforming growth factor-β1 （TGF-β1）-induced mouse mesangial cells （MCs）. Methodology：The primary MCs were generated. Three siRNAs were designed and synthesized. One of them that was capable of silencing EP1 most effectively in MCs. They were divided into five cell groups： （1）control group; （2） TGF-β1 （10 ng/ml） group; （3） NC-siRNA＋TGF- β1 （ 10 ng/ml） group; （4） EPI-siRNA＋TGF-β1 （ 10 ng/ml） group; （5） EPI-siRNA group. The expression of PGE2 in cell supernatant of each group was detected by using ELISA, the mRNA expression of fibronectin （FN）, connective tissue growth factor （CTGF） , cyclooxygenase-2 （COX2） , and mPGES-1 of each group was determined by using Real-Time PCR, and the proteins of FN, CTGF, COX2, mPGES-1 and the variation of phosphorylation proteins relating to MAPKs signaling pathway, such as p38, ERK, were examined by using Western blot. Results： Compared with normal control group, mRNA and protein expressions of FN, CTGF, COX-2 and mPGES-1 were up-regulated in group 2 （TGF-β1 group） （P〈0. 05）. Compared with group 2 （TGF-β1 group）, the expressions of FN, CTGF, COX-2 and mPGES-1 mRNA and proteins were down-regulated in group 4 （EPI-siRNA＋TGF-β1 group）（ P〈0. 05）. The silencing EP1 also declined TGF- β1-induced p38 and ERK phosphorylation （P〈0.05）. Conclusion：EPl-siRNA may mitigate the damage of TGF-β1 induced MCs by inhabiting p38 and ERK1/2 phosphorylation.

关 键 词：前列腺素E2受体1亚型 SIRNA 转化生长因子Β1 系膜细胞 信号通路 

分 类 号：R692[医药卫生—泌尿科学]