过氧化氢诱导人视网膜色素上皮细胞衰老及其机制探讨  

Investigation of human retinal pigment epithelial cells senescence induced by hydrogen peroxide and its mechanisms

作　　者：赵颖[1] 牛膺筠[2] 周占宇[2] 

机构地区：[1]青岛大学医学院附属市立医院眼科,山东省青岛市266011 [2]青岛大学医学院附属医院眼科,山东省青岛市266003

出　　处：《眼科新进展》2011年第9期804-806,共3页Recent Advances in Ophthalmology

基　　金：国家自然科学基金资助(编号:30572010)~~

摘　　要：目的研究过氧化氢诱导人视网膜色素上皮(retinal pigment epithelium,RPE)细胞氧化损伤中的细胞衰老现象并探讨其机制。方法培养的人RPE细胞随机分为正常对照组和过氧化氢处理组,过氧化氢处理组根据用600μmol·L-1过氧化氢处理的不同时段分为1h、6h、12h、24h及72h组。流式细胞术分析各组细胞周期,计算G0/G1期细胞所占比率;流式细胞术检测各组细胞线粒体膜电位的变化;免疫组织化学法检测各组细胞Caspase-9的变化。结果与正常对照组相比,过氧化氢作用6h组G0/G1期细胞所占比率增高(P<0.05),且随作用时间的延长而增加,72h组最高(P<0.01),达到(89.12±0.21)%;过氧化氢损伤导致RPE细胞膜电位降低,且随时间延长逐渐降低,72h组最低(62.48±0.59);Caspase-9蛋白在过氧化氢作用1h时表达开始增高,且随时间延长逐渐增加,在24h时达到高峰(36·2±0.4),72h时略有回落(26.4±0.8),但仍较正常组(3.2±0.2)高。结论过氧化氢作用下,人RPE细胞生长停滞于G/G期,呈现衰老现象。线粒体膜电位的降低和线粒体凋亡途径上游因子Caspase-9的活化可能是细胞衰老的机制之一。Objective To investigate the human retinal pigment epithelial（RPE） cells senescence in oxidative injury induced by hydrogen peroxide（H2O2）,and explore its mechanisms.Methods Cultured human RPE cells were randomly divided into normal group and H2O2 treatment group.Based on the time of treating with 600 μmol·L-1 H2O2,the H2O2 treatment group was subdivided into 1 hour,6 hours,12 hours,24 hours and 72 hours groups.Cell cycles were analyzed by flow cytometry to calculate the percentage of G0/G1 phase cells.Flow cytometry was used to detect the mitochondrial membrane potential changes.The expression of Caspase-9 protein was assessed by immunohistochemistry.Results Compared with normal group,the percentage of G0/G1 phase cells increased in 6 hours group（P0.05）,reached a maximum in 72 hours group（P0.01）,reached（89.12±0.21）%.H2O2 caused a time-dependent drop of mitochondrial membrane potential in RPE cells,the 72 hours group was the least（62.48±0.59）.The expression of Caspase-9 protein was increased in 1 hour group,reached a maximum in 24 hours group（36.2±0.4）,then decreased in 72 hours group（26.4±0.8）,but still higher than the normal group（3.2±0.2）.Conclusion Treated with H2O2,human RPE cells growth is arrested in G0/G1 phase.The drop of mitochondrial membrane potential and Caspase-9 protein expression may be one of the mechanisms of RPE cells senescence.

关 键 词：视网膜色素上皮细胞 过氧化氢 细胞衰老 线粒体膜电位 CASPASE-9 

分 类 号：R777.2[医药卫生—眼科]