晚期氧化蛋白产物通过内质网应激调节人肾小管上皮细胞自噬的研究  

Advanced oxidation protein products regulate autophagy in human renal tubular epithelial cells via endoplasmic reticulum stress

作　　者：李敏晖 汤珣[2] 姜婷婷[2] 张雯英 谢唯 章俊[2] LI Minhui;TANG Xun;JIANG Tingting;ZHANG Wenying;XIE Wei;ZHANG Jun(Blood Purification Center,Nanhai Hospital of Southern Medical University, Foshan,Guangdong 528244,China;Department of Nephrology,Zhujiang Hospital of Southern Medical University,Guangzhou,Guangdong 510280,China)

机构地区：[1]南方医科大学南海医院血液净化中心,广东佛山528244 [2]南方医科大学珠江医院肾内科,广州510280

出　　处：《重庆医学》2018年第29期3735-3738,3742,共5页Chongqing medicine

基　　金：2017佛山市卫生和计划生育局科研课题立项项目(20170214)

摘　　要：目的探讨内质网应激(ERS)介导的晚期氧化蛋白产物(AOPPs)诱导肾小管上皮细胞(HK-2)的自噬抑制及其信号通路。方法 (1)AOPPs刺激HK-2细胞,检测信号通路腺苷酸活化蛋白激酶/乳动物雷帕霉素靶蛋白(AMPK/mTOR)的表达;(2)AMPK抑制剂Compoun C和激活剂AICAR分别预处理细胞,并予AOPPs刺激,检测自噬标志物LC3Ⅱ/LC3Ⅰ、Beclin1、P62及AMPK/mTOR蛋白的表达;(3)ERS激活剂Thapsigargin与抑制剂Salubrinal分别预处理细胞,并予AOPPs刺激,检测ERS标志蛋白葡萄糖调节蛋白78(GRP78)、自噬及AMPK/mTOR蛋白的表达。结果 AOPPs可抑制AMPK的磷酸化和诱导mTOR的磷酸化(P<0.05)。Compoun C使LC3Ⅱ/LC3Ⅰ、Beclin1表达下降,p62表达上升,同时抑制AMPK的磷酸化和诱导mTOR的磷酸化(P<0.05),而AICAR表现出相反的效果。Thapsigargin能抑制AMPK的磷酸化和诱导mTOR的磷酸化,LC3Ⅱ/LC3Ⅰ、Beclin1表达下降,p62表达上升,而Salubrinal表现出相反的效果。结论ERS可能通过激活AMPK/mTOR信号通路介导AOPPs诱导的自噬抑制的发生。Objective To investigate the effects of endoplasmic reticulum stress(ERS)on advanced oxidation protein products(AOPPs)-inhibited autophagy in human renal tubular epithelial cells(HK-2)and its possible signaling pathway.Methods (1)HK-2 cells were stimulated by AOPPs,and the expression of AMP-activated protein kinase and mammalian target of rapamycin(AMPK/mTOR)was detected by Western blot.(2)HK-2 cells were treated with AMPK inhibitor(Compoun C)or AMPK activator(AICAR)respectively,and then stimulated by AOPPs.Western blot was used to determine the expression of autophagy related proteins,including LC3Ⅱ/LC3Ⅰ,Beclin1,P62 and AMPK/mTOR signaling pathway related proteins.(3)HK-2 cells were treated with ERS activator(Thapsigargin)or ERS inhibitor(Salubrinal),and then stimulated by AOPPs.The levels of ERS related protein,GRP78,and autopahgy related proteins as well as AMPK/mTOR signaling pathway related proteins were detected via Western blot.Results AOPPs inhibited the phosphorylation of AMPK and induced the phosphorylation of mTOR(P<0.05).Compoun C decreased the protein expression levels of LC3Ⅱ/LC3Ⅰand Beclin 1,increased the expression level of p62 protein,inhibited the phosphorylation of AMPK,and induced the phosphorylation of mTOR(P<0.05),while the AICAR showed the opposite effects.Thapsigargin inhibited the phosphorylation of AMPK and induced the phosphorylation of mTOR,decreased the protein expression levels of LC3Ⅱ/LC3Ⅰand Beclin 1,increased the p62 protein level(P<0.05),while the Salubrinal showed the opposite results.Conclusion ERS could mediate AOPPs-induced autophagy inhibition by activation of the AMPK/mTOR signaling pathway in HK-2 cells.

关 键 词：晚期氧化蛋白产物 内质网应激 丝氨酸苏氨酸蛋白激酶/哺乳动物雷帕霉素靶蛋白通路 自噬 

分 类 号：R692.5[医药卫生—泌尿科学]