白藜芦醇对HSC-T6细胞NLRP3炎性体活化的影响  

Effects of resveratrol on activation of NLRP3 inflammasome in HSC-T6 cells

作　　者：朱毅 阙任烨 李勇[3] Yi Zhu;Ren-Ye Que;Yong Li(First Department of Internal Medicine, Jiading Hospital of Traditional Chinese Medicine, Shanghai 201800, China;Department of Gastroenterology, Shanghai Hospital of Integrated Traditional Chinese and Western Medicine,Shanghai University of Traditional Chinese Medicine, Shanghai 200082, China;Department of Gastroenterology, Shanghai Hospital of Traditional Chinese Medicine, Shanghai University of Traditional Chinese Medicine, Shanghai 200071, Chin)

机构地区：[1]上海市嘉定区中医医院内一科,上海市201800 [2]上海中医药大学附属上海市中西医结合医院脾胃病科,上海市200082 [3]上海中医药大学附属上海市中医医院脾胃病科,上海市200071

出　　处：《世界华人消化杂志》2018年第8期479-487,共9页World Chinese Journal of Digestology

基　　金：国家自然科学基金;No.81573775;上海市卫生系统百人计划基金;No.XBR2013120~~

摘　　要：目的研究白藜芦醇(resveratrol,Res)对肝星状细胞(hepatic stellate cell,HSC)-T6细胞内Nod样受体蛋白3(nodlike receptor protein 3,NLRP3)炎性体活化的影响,探讨Res抗肝纤维化的作用机制,为Res的临床应用提供理论依据.方法体外培养大鼠HSC系HSC-T6,细胞用含10%胎牛血清的DMEM高糖培养基培养种板,细胞贴壁后加入Res(4、8、16μmol/L)或乙酰半胱氨酸(5 mmol/L)孵育24 h.随后加入过氧化氢(0.2 mmol/L)孵育4 h制作氧化应激模型.采用MTT法检测细胞增殖水平.ELISA法检测细胞培养上清液中的I型胶原(collagen type 1,COL-1)、转化生长因子(transforming growth factorβ1,TGF-β1)、白介素-1β(interleukin-1β,IL-1β)、IL-18、丙二醛(malondialdehyde,MDA)和超氧化物歧化酶(superoxide dismutase,SOD)的含量.荧光酶标仪检测细胞内活性氧(reactive oxygen species,ROS)含量.Western blot法检测细胞α-平滑肌肌动蛋白(alpha smooth muscle actin,α-SMA)、NLRP3、凋亡相关斑点样蛋白(apoptosis-associated speck-like protein,ASC)、caspase1的表达.结果与对照组比较,Res在浓度范围4-64μmol/L对HSC-T6细胞均有显著的抑制效应(P<0.01).与对照组比较,氧化应激模型组细胞增殖率,细胞上清液中COL-1、TGF-β1、MDA、IL-1β、IL-18含量,细胞内ROS产量及α-SMA、NLRP3、caspase1-p10蛋白表达均呈显著增高趋势(P<0.01);细胞上清液中SOD含量呈明显下降趋势(P<0.01).与模型组比较,Res低、中、高剂量及阳性对照药NAC均可显著抑制HSC-T6细胞增殖率,减少细胞上清液中COL-1、TGF-β1、MDA、IL-1β、I L-18含量,细胞内ROS产量及α-SMA、NLRP3、ASC、caspase1-p10蛋白表达(P<0.01);显著提高细胞上清液中SOD的含量(P<0.01).结论R e s能够通过调节ROS-NLRP3炎性体通路抑制HSC-T6细胞的增殖与活化.AIM To investigate the effect of resveratrol（Res）on the activation of nod-like receptor protein 3（NLRP3）inflammasome in hepatic stellate cell（HSC）-T6 cells and to explore the anti-fibrotic mechanism of Res.METHODS Rat hepatic stellate cell（HSC）line HSC-T6 was used.HSC-T6 cells were seeded into cell culture plates with high glucose DMEM medium containing 10% fetal bovine serum for 24 h.Then,the cells were incubated with Res（4,8,and 16 μmol/L）or acetylcysteine（NAC;5 mrnol/L）for 24 h.Oxidative stress（OS）was induced by exposure to hydrogen peroxide（H202;0.2 mmol/L）for 4 h.MTT method was used to observe the effect of Res on HSC-T6 cell proliferation.ELISA was used to detect the contents of type I collagen（COL-I）,transforming growth factor 131（TGF-I31）,interleukin（IL）-1β,IL-18,malondialdehyde（MDA）,and superoxide dismutase（SOD）in cell culture supernatant.Reactive oxygen species（ROS）production was measured with a fluorescence microplate reader following staining with DCFH-DA probe.Western blot analysis was used to detect the expression of alpha-smooth muscle actin（α-SMA）,NLRP3,apoptosis-associated specklike protein（ASC）,and cysteinyl aspartate specific proteinase I（caspase 1）in HSC-T6 cells.RESULTS Compared with control cells,Res at concentrations from 4 μmol/L to 64 μmol/L significantly suppressed the proliferation of HSC-T6 cells.Compared with control cells,OS induction significantly increased the proliferation of HSC-T6 cells,the contents of COL-1,TGF-β1,MDA,IL-1β,and IL-18 in cell culture supernatant,intracellular ROS production,and the protein expression of α-SMA,NLRP3,ASC,and caspase 1-p10（P 〈 0.01）,but decreased the content of SOD in cell culture supernatant（P 〈 0.01）.Compared with the OS group,treatment with low-,medium-,or high-dose Res or positive control NAC significantly decreased the proliferation of HSC-T6 cells,the contents of COL-1,TGF-β1,MDA,IL-1β,and IL-18 in cell culture supernatant,intracellcular ROS production,a

关 键 词：白藜芦醇 肝纤维化 肝星状细胞 NLRP3炎性体 

分 类 号：R285.5[医药卫生—中药学]