热休克蛋白70对氧化应激所致核仁素裂解的影响  

Effect of Heat Shock Protein 70 on Cleavage of Nucleolin Induced by Oxidative Stress

作　　者：王慷慨[1] 蒋磊[1] 刘可[1] 刘梅冬[1] 王浩[1] 易宇欣[1] 袁灿[1] 鄂顺梅[1] 石永忠[1] 肖献忠[1] 

机构地区：[1]中南大学湘雅医学院病理生理学教研室,湖南省长沙市410078

出　　处：《中国动脉硬化杂志》2004年第4期373-377,共5页Chinese Journal of Arteriosclerosis

基　　金：国家自然科学基金 ( 3 0 3 0 0 177;3 0 2 70 5 3 3 );国家 973重点项目 (G2 0 0 0 0 5 690 8);教育部博士点专项基金 ( 2 0 0 2 0 5 3 3 0 3 2 )资助

摘　　要：为探讨热休克蛋白 70对氧化应激 (0 .5mmol L过氧化氢 )所致核仁素裂解的影响及其机制 ,采用免疫印迹技术检测氧化应激诱导核仁素的裂解 ;通过热休克反应和构建热休克蛋白 70转基因细胞观察热休克反应与热休克蛋白 70对核仁素裂解的影响 ,同时采用免疫共沉淀技术检测热休克蛋白 70与核仁素之间的相互作用。结果发现 ,0 .5mmol L过氧化氢可导致多种细胞的核仁素发生裂解 (有 80kDa裂解片断出现 ) ,而且最早出现裂解片段的时间都在过氧化氢处理 30min到 1h左右 ;热休克预处理及转热休克蛋白 70后均引起热休克蛋白 70的表达明显增加 ,同时显著抑制抑制氧化应激所致核仁素裂解片段的出现 ,免疫共沉淀结果显示在氧化应激时热休克蛋白70与核仁素直接结合 ,形成复合物。以上结果提示 ,热休克反应与热休克蛋白 70可以显著抑制氧化应激所致核仁素的裂解 ,其机理与氧化应激时热休克蛋白 70与核仁素直接结合有关。Aim To observe the cleavage of nucleolin (also named C23) during apoptosis induced by oxidative stress and clarify the effect of heat shock protein 70 (HSP70) on cleavage of nucleolin and its possible molecular mechanism. Methods 0.5 mmol/L hydrogen peroxide (H 2O 2) was added into cultured cells to mimic oxidative stress. Cleavage of C23 were detected by using immunoblotting; Heat shock response (HSR) and HSP70 transgenic cell lines were used to observe the effect of HSR and HSP70 on cleavage of C23 induced by oxidative stress, and the relationship between HSP70 and C23 was evaluated by using co-immunoprecipitation. Results Activity of caspase-3 increased significantly after 2 h of 0.5 mmol/L H 2O 2 treatment, and reached peak at 12 h. The cleavage of C23 appeared 30 min to 1 h after treatment of H 2O 2 as indicated by a cleaved fragmentation of 80 kDa, which was significantly inhibited by HSR and HSP70. Furthermore co-immunoprecipitation study indicated that HSP70 could bind directly to C23 during H 2O 2 treatment. Conclusions Oxidative stress could induce the activation of caspase-3, cleavage of C23 and apoptosis; and HSP70 could inhibit significantly the cleavage of C23 induced by oxidative stress and its mechanism was related to the interaction between HSP70 and C23 during oxidative stress.

关 键 词：病理学与病理生理学 热休克蛋白70对氧化应激所致核仁素裂解的影响 免疫印迹法 热休克蛋白70 核仁素 氧化应激 过氧化氢 热休克反应 

分 类 号：R363[医药卫生—病理学]