维生素K3诱导氧化应激经ERK信号途径介导HeLa细胞发生自噬  

Autophagy of HeLa cells mediated by oxidative stress induced by vitamin K3 through ERK signal pathway

作　　者：于春艳[1] 刘希[2] 于春荣 张钰[4] 苏静[4] 刘玉和[1] 李洪岩[4] 

机构地区：[1]北华大学基础医学院病理学教研室,吉林吉林132013 [2]北华大学化学与生物学院实验中心,吉林吉林132013 [3]北京昭衍新药研究中心有限公司毒理部,北京100176 [4]吉林大学基础医学院病理生理学教研室,吉林长春130021

出　　处：《吉林大学学报（医学版）》2014年第2期229-232,共4页Journal of Jilin University：Medicine Edition

基　　金：国家自然科学基金资助课题(81202552;81272876);吉林省教育厅科研基金资助课题(吉教科合字2011第117号;吉教科合字2012第394号);吉林省科技厅自然科学基金资助课题(201215103)

摘　　要：目的:利用维生素K3(VK3)复制HeLa细胞氧化应激模型,探讨细胞外信号调节激酶(ERK)途径在氧化应激诱导自噬过程中的作用。方法:将人宫颈癌HeLa细胞随机分为对照组、VK3组(30μmol·L-1)、U0126组(10μmol·L-1)和VK3(30μmol·L-1)+U0126(10μmol·L-1)组。MTT法检测各组细胞生存率;Western blotting法检测HeLa细胞中ERK1/2、磷酸化ERK1/2(Phospho-ERK1/2)、自噬相关蛋白LC3-Ⅱ、凋亡相关蛋白caspase-3及其活化片段cleaved caspase-3的表达水平;Hoechest 33342染色激光共聚焦观察凋亡细胞核形态和细胞凋亡率。结果:与VK3组比较,VK3+U0126组HeLa细胞生存率降低(P<0.05)。VK3组作用2、4和8hHeLa细胞中Phospho-ERK1/2表达水平增加(P<0.05)。与对照组比较,VK3组HeLa细胞中LC3-Ⅱ蛋白表达水平增加(P<0.05);与VK3组比较,VK3+U0126组HeLa细胞中LC3-Ⅱ蛋白表达水平降低(P<0.05),cleaved caspase-3蛋白表达水平增加(P<0.05)。与VK3组比较,VK3+U0126组HeLa细胞呈现凋亡细胞核形态,细胞凋亡率升高(P<0.05)。结论:氧化应激通过ERK信号途径诱导HeLa细胞发生自噬,被激活的细胞自噬能抑制VK3诱导HeLa细胞凋亡。Objective To establish oxidative stress model of HeLa cells copied by vitamin K3 （VK3）, and to investigate the role of external signal-regulated kinase （ERK） pathway in autophagy induced by oxidative stress in HeLa cells. Methods HeLa cells were divided into control group, VK3 group （30 μmol·L^-1）, U0126 group （10 μmol·L^-1）, and VK3 （30μmol·L^-1）＋ U0126 group （10 μmol·L^-1）. The survival rate of HeLa cells in each group was measured by MTT assay; the expression levels of external signal-regulated kinase 1/2 （ERK1/2）, phosphorylated ERK 1/2 （Phospho-ERK1/2）, microtubule-associated protein light chain 3-Ⅱ （LC3-Ⅱ）, and cysteine aspartic acid specific protease-3 （caspase-3）, and its activated fragment cleaved caspase-3 were determined by Western blotting method; the morphological changes of nuclear and apoptotic rate of HeLa cells in each group were detected by Hoechst staining. Results Compared with VK3 group, the survival rate of HeLa cells in VK3＋ U0126 group was decreased （P〈0.05）. The expressions of Phospho-ERK1/2 in HeLa cells were increased after treated with VK3 for 2, 4, and 8 h （P〈0.05）. Compared with control group, the expression of LC3-Ⅱ protein in VK3 group was increased（P〈0. 05）. Compared with VK3 group, the expression of LC3-Ⅱ in HeLa cells in VK3q-U0126 group was decreased（P〈0.05）, and the expression of cleaved caspase-3 protein was increased （P〈0.05）. Compared with VK3 group, the morphological changes of nuclear appeared and the apoptotic rate of HeLa cells in VK3q-U0126 group was increased（P〈0.05）. Gonclusion Oxidative stress activates autophagy of HeLa cells induced by ERK signal pathway, and the autophagy can inhibit the apoptosis of HeLa cells induced by VK3.

关 键 词：氧化应激 细胞外信号调节激酶 自噬 HELA细胞 

分 类 号：R363.2[医药卫生—病理学]