S1P/S1PR1信号通路通过调控ROS/NLRP3对高糖诱导的大鼠肾小管上皮细胞上皮间充质转化的影响  

Effects of S1P/S1PR1 pathway on high glucose induced epithelial-mesenchymal transition of rat renal tubular epithelial cells by regulating ROS/NLRP3

作　　者：黄太平 王菁[1] 徐焕雨 杨佳[1] 薛媛[1] 张婷婷[1] 田继华[1] HUANG Tai-ping;WANG Jing;XU Huan-yu;YANG Jia;XUE Yuan;ZHANG Ting-ting;TIAN Ji-hua(Dept of Microbiology and Immunology,Shanxi Medical University,Taiyuan 030001,China)

机构地区：[1]山西医科大学微生物学与免疫学教研室,山西太原030001

出　　处：《中国药理学通报》2023年第6期1143-1148,共6页Chinese Pharmacological Bulletin

基　　金：山西省自然科学基金资助项目(No 201901D111188);山西省回国留学人员科研资助项目(No 2021-079)。

摘　　要：目的探究S1P/S1PR1信号通路对高糖诱导的大鼠肾小管上皮细胞上皮间充质转化(epithelial-mesenchymal transition,EMT)的影响及其可能的作用机制。方法采用不同浓度葡萄糖处理细胞,ELISA法检测细胞内S1P表达,Western blot检测S1PR1蛋白表达;将细胞分为正常对照组、HG组及HG+siS1PR1组,RT-PCR检测细胞E-cadherin、Vimentin、Fibronectin及Twist mRNA的表达,Western blot检测E-cadherin、α-SMA、Vimentin、NLRP3、ASC及NF-κB蛋白表达,流式细胞术检测活性氧(reactive oxygen species,ROS)水平;将细胞分为正常对照组、S1P组及S1P+siS1PR1组,Western blot检测Vimentin、Snail、α-SMA、NLRP3、ASC及NF-κB蛋白表达,荧光显微镜测定ROS水平。结果ELISA结果显示,高糖刺激下细胞内S1P含量明显升高。Western blot结果显示,30 mmol·L^(-1)浓度时S1PR1蛋白表达量与对照组相比明显升高;阻断S1PR1后可以抑制高糖诱导的Fibronectin、Vimentin及Twist mRNA表达,上调E-cadherin mRNA的表达,Western blot结果显示,E-cadherin蛋白表达升高,Vimentin,α-SMA蛋白表达下降,同时可抑制细胞内ROS的产生和NLRP3,ASC,NF-κB蛋白水平;加入S1P激活剂后细胞内ROS水平,Vimentin、α-SMA、Snail、NLRP3、ASC及NF-κB蛋白水平表达均升高,阻断S1PR1后表达均降低。结论高糖能诱导肾小管上皮细胞EMT,其机制可能与S1P/S1PR1信号通路作用于ROS/NLRP3轴有关。Aim To explore the effect of S1P/S1PR1 signaling pathway on high glucose(HG)-induced epithelial-mesenchymal transition of rat renal tubular epithelial cells and its possible mechanism.Methods Cells were treated with different concentrations of glucose,and intracellular S1P expression was detected by ELISA and S1PR1 protein expression was detected by Western blot.The cells were divided into normal control group,HG group and HG+siS1PR1 group.The expression of E-cadherin,Vimentin,Fibronectin and Twist mRNA were detected by RT-qPCR and E-cadherin,α-SMA,Vimentin,NLRP3,ASC and NF-κB protein expression were detected by Western blot,and the levels of reactive oxygen species(ROS)were detected by flow cytometry.The cells were divided into normal control group,S1P group and S1P+siS1PR1 group.Vimentin,Snail,α-SMA,NLRP3,ASC and NF-κB protein expressions were detected by Western blot,and ROS levels were measured by fluorescence microscopy.Results ELISA results showed that the content of S1P in cells increased significantly under high glucose stimulation.Western blot results showed that S1PR1 protein expression was significantly higher at 30 mmol·L^(-1) compared with the control group.Blocking S1PR1 inhibited high glucose-induced Fibronectin,Vimentin and Twist mRNA expression,and up-regulated E-cadherin mRNA expression.Western blot showed that E-cadherin protein expression increased,Vimentin andα-SMA protein expression decreased,while intracellular ROS production and NLRP3,ASC and NF-κB protein levels were inhibited.The expression of intracellular ROS,Vimentin,α-SMA,Snail,NLRP3,ASC and NF-κB protein levels increased after adding S1P activator,and decreased after blocking S1PR1.Conclusions High glucose induces EMT in renal tubular epithelial cells,and the mechanism may be related to the S1P/S1PR1 signaling pathway acting on the ROS/NLRP3 axis.

关 键 词：S1P/S1PR1信号通路 ROS NF-κB NLRP3 上皮间充质转化 肾小管上皮细胞 

分 类 号：R-332[医药卫生] R322.61R392.11R394.2R692.39