砷暴露诱导人肝细胞氧化应激过程中NADPH氧化酶作用机制  

The Role of NADPH Oxidase in the Oxidative Stress of Human Liver Cells Induced by Arsenic Exposure

作　　者：李影娜[1] 章琳[1] 李秀丽[1] 郑华东[1] 于俊霞[1] 秦绪军[2] 杨渭临[1] 

机构地区：[1]西安交通大学第二附属医院老年病科,陕西西安710004 [2]第四军医大学预防医学院毒理学教研室 陕西省自由基生物学与医学重点实验室 特殊作业环境危害评估与防治教育部重点实验室,陕西西安710032

出　　处：《现代生物医学进展》2013年第27期5212-5216,共5页Progress in Modern Biomedicine

基　　金：国家自然科学基金项目(31070766);陕西省科技新星项目(2010KJXX-06)

摘　　要：目的:探讨砷暴露诱导细胞氧化应激的分子机制。方法:采用人正常肝细胞进行亚砷酸钠和砷酸钠的暴露处理,并设相应对照组,采用SOD模拟物MnTMPyP和还原型谷胱甘肽(reduced glutathione,GSH)预处理,检测细胞超氧阴离子(O2·-)和细胞整体ROS的水平。Western Blot方法检测细胞氧化/抗氧化重要酶微粒体谷胱甘肽硫转移酶(microsomal glutathione S-transferase-1,Mgst-1)、半胱氨酸双加氧酶1(cysteine dioxygenase 1,Cdo1)和NADPH氧化酶的催化亚基NOX4的表达。针对NADPH氧化酶,采用特异性抑制剂(diphenyleneiodonium chloride,DPI)进行预处理,观察对砷暴露引起的细胞ROS水平及细胞凋亡的影响。结果:砷暴露能够显著诱导细胞超氧阴离子的产生,提高细胞整体ROS水平,其中三价砷(亚砷酸钠,As3+)诱导氧化应激作用显著强于五价砷(砷酸钠,As5+)。亚砷酸钠能够显著提高NOX4的表达。针对NADPH氧化酶的抑制剂DPI能够显著抑制砷暴露引起的细胞ROS水平升高以及细胞凋亡的增加。结论:NADPH氧化酶是砷暴露诱导人肝细胞的作用靶点,砷能够通过NADPH氧化酶产生大量超氧阴离子,提高ROS水平,造成氧化应激,诱导人正常肝细胞凋亡。Objective： To investigate the molecular mechanism of oxidative stress of human liver cells induced by arsenic expo- sure. Methods： Human normal liver cell line HL-7702 was treated with sodium arsenite （10 μM） or sodium arsenate （10μM） for 12 h. MnTMPyP （5 μM）, reduced glutathione （GSH） （5 mM） and NADPH oxidase inhibitor （diphenyleneiodonium chloride, DPI） （10 μM） was used in the experiments indicated in the methods. The superoxide anion radical （Oi） and reactive oxygen species （ROS） produc- tion and cell apoptosis were determined by flow cytometry. The expressions of Mgst-1, Cdol and NOX4 were determined by Western Blot. Results： Sodium arsenite instead of sodium arsenate significantly induced O2- production, NOX4 expression and cell apoptosis. NADPH oxidase inhibitor DPI pre-treatment efficiently inhibited the arsenite-induced ROS production and cell apoptosis. Conclusion： NADPH oxidase was the molecular target of arsenite in the arsenite-induced oxidative stress to the human liver cells. Arsenite stimulated the ROS production by activating NADPH oxidase, and leading to the oxidative stress and the cell apoptosis.

关 键 词：砷 氧化应激 活性氧 NADPH氧化酶 凋亡 

分 类 号：R595.2[医药卫生—内科学] R995[医药卫生—临床医学]