高迁移率族蛋白1在鱼藤酮诱导的多巴胺能神经细胞损伤中的作用

The Role of High Mobility Group Box 1 in Dopaminergic Neuronal Damage Induced by Rotenone

目的:观察高迁移率族蛋白1(HMGB1)在鱼藤酮诱导的帕金森病(PD)细胞模型中的作用,探讨HMGB1在PD发病机制中的作用。方法:使用鱼藤酮作用于SH-SY5Y细胞建立PD细胞模型;免疫荧光染色观察细胞内HMGB1、α-synuclein及微管相关蛋白轻链3(LC3)的表达及定位;免疫印迹法检测不同浓度鱼藤酮作用下及siRNA-HMGB1干扰HMGB1后HMGB1、LC3Ⅱ/Ⅰ、自噬底物P62及Toll样受体4(TLR4)的表达水平;通过雷帕霉素(Rap)和3-甲基腺嘌呤(3-MA)分别激活或抑制自噬,实时荧光定量PCR检测HMGB1的mRNA转录水平。结果:在PD细胞模型中,HMGB1出现胞浆转位,并与胞浆中α-synuclein及LC3颗粒样聚集物共定位;随着鱼藤酮浓度增加,HMGB1蛋白表达上调,伴有LC3II/I比例增高、P62及TLR4蛋白水平降低;Rap和3-MA分别激活或抑制自噬可引起HMGB1 mRNA的转录水平相应增加或降低;干扰HMGB1表达可抑制自噬的激活。结论:胞浆转位的HMGB1通过作用于自噬溶酶体途径参与PD的发病,可能是PD治疗的新靶点。

Objective： To identify the role of high mobility group box 1 （HMGB1） in dopaminergic neuronal damage induced by rotenone. Methods： SH-SY5Y cells were treated with rotenone with different concentration to make cell model of Parkinson＇ s disease （PD）. The expression and location of HMGB1, a-synuclein and LC3 were analyzed by immnofluorescence staining. The protein expression of HMGB1, LC3II/I, P62 and TLR4 were assessed by western blot. Activated on inhibited autophagy by rapamycin or 3-MA, the transcription level of HMGB1 mRNA was detected by real-time fluorescent quantitative PCR. Results： HMGB1 translocated in cytoplasm and colocalized with a-synuclein and LC3 in PD cells, accompanied by enhanced autophagy. Increased autophagosomes of autophagy directly increased the expression of HMGB 1. Knocked down HMGB 1 expression using RNA interferenc inhibited the intracellular activation of autophagy pathway. Conclusion： HMGB 1 cytoplasmic translocation is involved in the activation of autophagy in PD, which may be a novel therapeutic target for PD.