O-糖基化在酪氨酸羟化酶功能调节中的作用  

Role of O-glycosylation in Regulation of Tyrosine Hydroxylase

作　　者：丁伟[1] 谢腾[1] 张捷[1] 付锴[1] 赵时雨[1] 黄鑫[1] 想象 杨振宇[1] 

机构地区：[1]武汉大学中南医院神经外科,湖北武汉430071

出　　处：《武汉大学学报（医学版）》2014年第4期519-524,共6页Medical Journal of Wuhan University

基　　金：湖北省自然科学基金资助项目(编号:2011CDB511)

摘　　要：目的:观察酪氨酸羟化酶(TH)O-糖基化和磷酸化水平在6-羟多巴胺(6-OHDA)诱导的SH-SY5Y细胞(人神经母细胞瘤细胞)帕金森病细胞模型中的变化,探讨O-糖基化在酪氨酸羟化酶功能调节中的作用。方法:用6-OHDA处理SH-SY5Y细胞作为模型组;用相同终浓度的6-OHDA加上四氧嘧啶(ALX,O-糖基化转移酶抑制剂)处理SH-SY5Y细胞作为模型治疗组;用ALX处理SH-SY5Y细胞作为治疗对照组。用Western印迹检测SHSY5Y细胞TH的表达以及其O-糖基化和磷酸化水平的变化,采用ELISA法检测各组细胞多巴胺的浓度。结果:6-OHDA浓度依赖性地引起SH-5YSY细胞存活率下降,12.5-50.0μmol/L 6-OHDA处理细胞24h,细胞存活率为对照组的96.9%-72.3%;100μmol/L 6-OHDA处理细胞24h后存活率低于50%。50μmol/L 6-OHDA处理SH-SY5Y细胞24h引起细胞TH表达的下降、糖基化水平升高和磷酸化水平下降;ALX处理SH-SY5Y细胞24h后,TH O-糖基化水平下降、磷酸化水平升高,但TH的表达无改变,总的多巴胺的浓度升高了11.5%±3.4%(P<0.01);用四氧嘧啶+50μmol/L 6-OHDA处理SH-SY5Y细胞24h后,与模型组比较,TH表达无明显改变,O-糖基化水平下降,磷酸化水平上升,总多巴胺浓度升高8.7%±2.0%(P<0.01)。结论:在6-羟多巴胺(6-OHDA)诱导的SH-SY5Y细胞帕金森病细胞模型中,TH的O-糖基化水平升高,磷酸化水平的下降;通过抑制酪氨酸羟化酶的O-糖基化水平可以上调其活性,引起多巴胺分泌增多,这一作用可能同O-糖基化与磷酸化的竞争性抑制有关。Objective： To explore the role of O-glycosylation of tyrosine hydroxylase （TH） in Parkinson＇s disease （PD） cellular model treated with 6-hydroxydopamine （6-OHDA）. Methods： The changes of O-glycosylation and phosphorylation of TH and the levels of TH were detected by Western blotting in a PD cellular model of SH-SY5Y cell （a human neuroblastoma cell line） trea- ted with 6-OHDA. The dopamine content was measured with ELISA assay. The effect of Alloxan （an inhibitor of O-linked N-acetylglucosamine transferase） on O-glycosylation and phosphoryl- ation of TH, the levels of TH and dopamine content were investigated. Results： Western blotting showed that 24 h after the treatment of 50μmol/L 6-OHDA, the expression of TH decreased obviously, and the levels of O-glycosylation of TH increased, with the decrease of phosphorylation of TH. Application of alloxan leaded to the decrease of O-glycosylation and increase of phospho- rylation of TH, but there was no change in the expression of TH. Compared with control, thetotal dopamine concentration increased by 11.5 % ±3.4 % （P〈0.01） in treatment control group and increased by 8.7%±2.0%（P〈0.05） in the model group with the treatment of Alloxan. Conclusion： In SH-SYSY cell model of Parkinson＇s disease treated with 6-OHDA, O-glycosyla- tion levels increased and the phosphorylation levels decreased in TH. Inhibiting the O-glycosylation of TH can activate the phosphorylation of TH without change the expression of TH, finally increase the secretion of dopamine.

关 键 词：帕金森病 酪氨酸羟化酶 O-糖基化 磷酸化 

分 类 号：R651.11[医药卫生—外科学]