二甲双胍抑制高糖培养大鼠肾小球系膜细胞内P38MAPK信号通路与氧化应激  

Metformin inhibits p38MAPK signaling pathway and oxidativestress in rat glomerular mesangial cells cultured with high glucose

作　　者：姚新明[1] 孔祥[1] 赵咏莉[1] 华强[1] 何春玲[1] 余丹[2] 李业琼 戴晓康 叶山东[2] YAO Xinming;KONG Xiang;ZHAO Yongli;HUA Qiang;HE Chunling;YU Dan;LI Yeqiong;DAI Xiaokang;YE Shandong(Department of Endocrinology,Yijishan Hospital Affiliated to Wannan Medical College,Wuhu 241001,Anhui,China;Department of Endocrinology,Anhui Provincial Hospital,Hefei 230001,Anhui,China)

机构地区：[1]皖南医学院弋矶山医院内分泌科 [2]安徽省立医院内分泌科

出　　处：《中国临床药理学与治疗学》2018年第6期614-620,共7页Chinese Journal of Clinical Pharmacology and Therapeutics

基　　金：安徽省自然科学基金(1608085QH191);安徽省自然科学基金(1508085SMH227);安徽省高校优秀青年人才支持计划项目(gxyq2017035);皖南医学院弋矶山医院人才引进基金(YR201507)

摘　　要：目的:观察高浓度葡萄糖培养大鼠肾小球系膜细胞内p38丝裂原活化蛋白激酶(p38 mitogenactivated protein kinase,p38MAPK)信号通路与氧化应激的关系及二甲双胍的调控作用。方法:大鼠肾小球系膜细胞(HBZY-1)在完全培养基中传代培养,并分成以下5组:(1)正常对照组(NC组);(2)高浓度葡萄糖培养组(HG组);(3)二甲双胍治疗组(MET组);(4)SB203580治疗组(SB组);(5)N-乙酰半胱氨酸治疗组(NAC)组。流式细胞术检测大鼠肾小球系膜细胞内活性氧(ROS)水平。用比色法和ELISA法分别检测细胞上清液中超氧化物歧化酶(SOD)活性与丙二醛(MDA)含量。实时定量PCR法检测肾小球系膜细胞内P22phox mRNA表达。Western blot法检测肾小球系膜细胞内P22phox蛋白和磷酸化p38MAPK(p-p38MAPK)蛋白表达。结果:与正常对照组相比,高浓度葡萄糖培养组细胞上清液中SOD活性降低,相反细胞上清液中MDA含量、肾小球系膜细胞内ROS水平、P22phox mRNA和蛋白及磷酸化p38MAPK蛋白表达均上升(P<0.05)。当二甲双胍干预后,细胞上清液中SOD活性上升,同时细胞上清液中MDA含量、肾小球系膜细胞内ROS水平、P22phox mRNA和蛋白及磷酸化p38MAPK蛋白表达均下降(P<0.05)。相似的结果出现在SB203580或N-乙酰半胱氨酸干预后(P<0.05)。结论:二甲双胍可以减轻高浓度葡萄糖培养大鼠肾小球系膜细胞氧化应激和磷酸化p38MAPK蛋白表达,进而起到了保护肾脏的作用。AIM： To observe relationship between p38 MAPK signaling pathway and oxidative stress in rat glomerular mesangial cells（ MCs） cultured in high glucose medium and the regulation of metformin. METHODS： Rat glomerular MCs were cultured in a complete medium,and were divided into five groups： normal control group（ group NC）,high glucose group（ group HG）,metformin-treated group（ group MET）, SB203580-treated group（ group SB）,N-acetylcysteine-treated group（ group NAC）. Intracellular reactive oxygen species（ ROS）content of rat glomerular MCs was tested by flow cytometry. Superoxide dismutase（ SOD） activity and malondialdehyde（ MDA） content in supernatant fluid were detected by colorimetry and ELISA,respectively. P22 phox mRNA of rat glomerular MCs was determined by real-time quantitative PCR. P22 phox protein and phosphorylation of p38 MAPK（ pp38 MAPK） protein of rat glomerular MCs were detected by Western blot. RESULTS： Compared with group NC, the activity of SOD was decreased,whereas the level of MDA,intracellular ROS content, P22 phox mRNA and protein, p-p38 MAPK protein expression were increased in group HG（ P〈0. 05）. When metformin was added to the high glucose medium,the activity of SOD was increased,meanwhile,the level of MDA, intracellular ROS content,P22 phox mRNA and protein,p-p38 MAPK protein expression were decreased（ P〈0. 05）,and the results were similar with SB203580 or N-acetylcysteine was added to the high glucose medium（ P〈0. 05）. CONCLUSION： Metformin can alleviate high glucose induced oxidative stress and pp38 MAPK protein expression of rat glomerular MCs,which maybe contribute to its reno-protection.

关 键 词：二甲双胍 氧化应激 p38MAPK 肾小球系膜细胞 

分 类 号：R965.2[医药卫生—药理学]