NADPH氧化酶在人真皮成纤维细胞氧化应激损伤中的作用  

Role of NADPH oxidase in oxidative stress injury of human dermal fibroblasts

作　　者：陈颖[1] 黄昊[1] 汤红峰[1] 郑秀芬[1] 胡勇[1] 王瑞华[1] 

机构地区：[1]南方医科大学附属顺德第一人民医院皮肤科,广东佛山528300

出　　处：《南方医科大学学报》2016年第3期391-395,共5页Journal of Southern Medical University

基　　金：佛山市科学技术局医学类科技攻关项目(2014AB001923)

摘　　要：目的探讨NADPH氧化酶在人真皮成纤维细胞氧化应激损伤中的作用。方法 H_2O_2构建氧化应激模型,将实验分为正常组、氧化损伤组和NADPH氧化酶抑制剂(DPI)组,MTT检测细胞活力,DCFH-DA荧光探针检测细胞内活性氧(ROS)改变,Western blot分析NADPH氧化酶胞膜亚基gp91phox表达变化。结果 H_2O_2对成纤维细胞的氧化损伤呈时间和浓度依赖,H_2O_2700μmol/L处理24 h后细胞活力下降约40%(P<0.05),ROS升高2倍(P<0.05)。而抑制剂组细胞活力较氧化损伤组增加20%(P<0.05),ROS下降至正常水平(P<0.05)。Western blotting结果显示氧化损伤组gp91phox表达随H_2O_2浓度逐渐升高,而抑制剂组表达接近正常水平。结论 H_2O_2可通过影响NADPH氧化酶特别是胞膜亚基gp91phox引发成纤维细胞氧化损伤。Objective To investigate the role of NADPH oxidase（Nox） in the oxidative stress injury of human dermal fibroblasts（HFbs）. Methods An oxidative stress injury model was established in HFbs by exposure to H_2O_2. Normal HFbs and HFbs exposed to H_2O_2 with and without pretreatment with NADPH oxidase inhibitor were tested for cell viability using MTT assay,and the intracellular reactive oxygen species（ROS） were determined with a DCFH-DA fluorescent probe. Western blotting was used to measure the protein expressions of membrane- bound subunit gp91 phox of NADPH oxidase in the cells. Result H_2O_2time- and concentration-dependently induced oxidative stress injury in the fibroblasts, causing a reduction of the cell viability to40% after a 24-h exposure at 700 μmol/L（P0.05） and an increase of ROS by 2 folds after a 2-h exposure at 700 μmol/L（P0.05）.Compared with the cells with oxidative stress injury, the cells with NADPH oxidase inhibitor pretreatment showed a 20% higher cell viability（P0.05） and normal ROS level（P0.05） following H_2O_2 exposure. Western blotting demonstrated increased expression of gp91 phox in the cells exposed to increasing H_2O_2 concentrations, but gp91 phox expression remained normal in cells pretreated with NADPH oxidase inhibitor. Conclusion H_2O_2 can induce oxidative stress injury in the fibroblasts by affecting NADPH oxidase, especially its membrane-bound subunit gp91 phox.

关 键 词：人真皮成纤维细胞 活性氧 氧化应激 NADPH氧化酶 NADPH氧化酶抑制剂 

分 类 号：R36[医药卫生—病理学]