Zeste基因增强子同源物2抑制剂减轻氧糖剥夺/再灌注诱导的神经元氧化应激和铁死亡  

Enhancer of Zeste homolog 2 inhibitor mitigates oxygen glucose deprivation/recovery induced oxidative stress and ferroptosis in neurons

作　　者：柳亚珍 孙海伟[1] 汪继 朱建良[1] 马丽梅 肖盐[1] 刘励军[1] Liu Ya-zhen;Sun Hai-wei;Wang Ji;Zhu Jian-liang;Ma Li-mei;Xiao Yan;Liu Li-jun(Department of Emergency and Intensive Care,the Second Affiliated Hospital of Soochow University,Suzhou 215000,China)

机构地区：[1]苏州大学附属第二医院急诊科,江苏苏州215000 [2]广州医科大学附属第二医院神经外科,广东广州510260

出　　处：《中国急救医学》2023年第6期472-480,共9页Chinese Journal of Critical Care Medicine

基　　金：苏州市临床重点病种诊疗专项项目(LCZX201404);苏州市学科建设托举工程项目(XKTJ-XK202008)。

摘　　要：目的探讨Zeste基因增强子同源物2(EZH2)抑制剂GSK126在神经元缺血-再灌注损伤中的作用及机制。方法采用海马神经元HT22细胞制备氧糖剥夺/再灌注(OGD/R)模型。细胞分为正常对照组(Control组)、OGD/R模型组(OGD/R组)和OGD/R模型+低、中、高浓度EZH2抑制剂组(OGD/R+GSK1260.5、1、5μmol/L组)。采用CCK-8法、流式细胞术检测各组HT22细胞损伤,相应试剂盒检测细胞氧化应激指标活性氧(ROS)水平、丙二醛(MDA)含量和谷胱甘肽/氧化型谷胱甘肽(GSH/GSSG)比值。通过荧光探针C11 BODIPY 581/591和FerroOrange检测细胞内铁死亡指标脂质过氧化物(Lipid ROS)和Fe^(2+)含量。实时荧光定量逆转录聚合酶链反应(RT-qPCR)和蛋白免疫印记法(Western blot)实验检测细胞EZH2、NRF2、HO-1、SLC7A11和GPX4的mRNA和蛋白表达。结果与Control组比较,OGD/R组HT22细胞活力降低,细胞死亡率升高,细胞内ROS水平升高,MDA含量增加,GSH/GSSG比值减少,Lipid ROS、Fe^(2+)含量增加,EZH2的mRNA和蛋白表达升高,NRF2、HO-1、SLC7A11和GPX4的mRNA和蛋白表达均降低(P值均<0.05)。与OGD/R组比较,OGD/R+GSK1260.5、1、5μmol/L组HT22细胞活力均升高,细胞死亡率均下降,细胞内ROS水平均下降,MDA含量均减少,GSH/GSSG比值均增加,Lipid ROS、Fe^(2+)含量均减少,EZH2的mRNA和蛋白表达均降低,NRF2、HO-1、SLC7A11和GPX4的mRNA和蛋白表达均升高(P值均<0.05)。结论EZH2抑制剂GSK126可减轻OGD/R诱导的神经元细胞损伤,其机制可能与NRF2/HO-1通路抑制氧化应激和SLC7A11/GPX4通路抑制铁死亡有关。Objective To investigate the role and mechanism of enhancer of Zeste homolog 2(EZH2)inhibitor(GSK126)in neuronal ischemia-reperfusion injury.Methods An model of oxygen glucose deprivation/recovery(OGD/R)was established by hippocampal neurons HT22 cells.The cells were divided into the following groups:normal control group(Control group),OGD/R model group(OGD/R group),and OGD/R model+low,medium and high concentration EZH2 inhibitor group(OGD/R+GSK1260.5,1,5μmol/L group).Cell injury of HT22 cell was measured by CCK-8 method and flow cytometry in different groups.Intracellular oxidative stress indicator,such as the levels of reactive oxygen species(ROS),the contents of malondialdehyde(MDA)and the ratio of glutathione to oxidized glutathione(GSH/GSSG),were tested by corresponding assay kits in cells.Levels of intracellular ferroptosis indicators like lipid peroxides(Lipid ROS)and Fe^(2+)were tested by fluorescent probes C11 BODIPY 581/591 and FerroOrange in cells.The mRNA and protein expression of EZH2,NRF2,HO-1,SLC7A11 and GPX4 were measured by RT-qPCR and Western blot in cells.Results Compared with the Control group,cell viability in the OGD/R group was decreased and cell mortality was increased.The levels of intracellular ROS and MDA were increased,but the GSH/GSSG were decreased.The contents of Lipid ROS and Fe^(2+)were increased.The mRNA and protein expression of EZH2 were increased,but the mRNA and protein expression of NRF2,HO-1,SLC7A11 and GPX4 were all decreased(all P<0.05).In comparison with OGD/R group,the OGD/R+GSK1260.5,1,5μmol/L groups’cell viability was all increased,and cell mortality was all decreased.The levels of intracellular ROS and the contents of MDA were all decreased,but the GSH/GSSG were increased.The contents of Lipid ROS and Fe 2+were all decreased.The mRNA and protein expression of EZH2 were all decreased,but the mRNA and protein expression of NRF2,HO-1,SLC7A11 and GPX4 were all increased(all P<0.05).Conclusions GSK126 can mitigate neuronal ischemia-reperfusion injury induced by OGD/R,

关 键 词：Zeste基因增强子同源物2(EZH2) GSK126(EZH2抑制剂) 氧糖剥夺/再灌注(OGD/R) 铁死亡 

分 类 号：R743.3[医药卫生—神经病学与精神病学]