聚集素蛋白在聚集体形成中的参与机制  

The participation of clusterin in the formation of aggresome

作　　者：马建芳[1] 陈生弟[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院神经科,上海交通大学医学院神经病学研究所,上海200025

出　　处：《中华神经医学杂志》2007年第8期757-761,共5页Chinese Journal of Neuromedicine

基　　金：国家重点基础研究规划(2006cb500706);国家自然科学基金(30471918);上海市医学领军人才项目(06033)

摘　　要：目的探讨聚集素蛋白在聚集体形成中的参与机制。方法本实验分为处理组、对照组和未处理组,处理组用100nmol/L鱼藤酮,对照组用0.01%的二甲基亚砜处理多巴胺能SK-N-SH细胞。然后用Trypan Blue法检测细胞存活率,Western blot检测细胞内聚集素蛋白表达的变化。在处理组和对照组中,用HE法检测聚集体的形成,免疫荧光双标检测聚集素蛋白与细胞内聚集体的关系,免疫共沉淀检测α-突触核蛋白与聚集素蛋白的相互作用情况。结果鱼藤酮损伤多巴胺能SK-N-SH细胞,并形成类似Lewy体的聚集体结构,其中聚集素蛋白和α-突触核蛋白、波形蛋白和泛素共存在聚集体中。100nmol/L鱼藤酮在6h后,引起聚集素蛋白的表达升高,与α-突触核蛋白的相互作用无变化;24h时,聚集素蛋白的表达降低,与α-突触核蛋白的相互作用也减弱。结论聚集素蛋白确实存在于聚集体中,可能是通过与α-突触核蛋白蛋白相互作用而沉积于聚集体中。Objective To investigate the participation of clusterin in the formation of aggresome. Methods Dopaminergic SK-N-SH cells were divided into three groups： the treatment group, control group and non-treatment group. Rotenone （100 nmol/L） and 0.01% DMSO were used respectively on dopaminergic SK-N-SH cells in the treatment and control groups. Then the trypan blue and Western blot assays were applied respectively to detect cell viability and the changes of intracellular clusterin expression. In the treatment and control groups, HE staining was employed to detect the formation of aggresome, the immunofluorescence double-labeling assay was employed to detect the relationship between clusterin and intracellular aggresome, and the co-immunoprecipitation assay was employed to detect interaction between α-synuclein and clusterin. Results Rotenone injured the dopaminergic SK-N-SH cells and induced the formation of Lewy body-like aggresome, in which clusterin coexisted with α-synuclein, vimentin and ubiquitin. Rotenone （100 nmol/L） increased the expression of clusterin at 6 h since its application, but no impact on its interaction with α-synuclein; the expression of clusterin began to decrease at 24 h and its interaction with α-synuclein weakened. Conclusion Clusterin does exist in aggresome and it may probably interact with α-synuclein to deposit in aggresome.

关 键 词：聚集素蛋白 聚集体 Α-突触核蛋白 鱼藤酮 多巴胺 

分 类 号：R33[医药卫生—人体生理学]