α-突触核蛋白对N-甲基-D-天门冬氨酸所致细胞毒性作用的影响

Effect of Alpha-synuclein on N-methyl-D-aspartic Acid-induced Cytotoxicity

目的研究α-突触核蛋白(α-Syn)对N-甲基-D-天门冬氨酸(NMDA)引起的多巴胺能神经细胞毒性作用的影响。方法在MES23.5多巴胺能神经细胞建立NMDA细胞毒模型,MTS法检测细胞活力,AO/PI荧光标记以及免疫印记测定caspase-3表达检测细胞凋亡,细胞外添加α-Syn蛋白,观察α-Syn对NMDA所致细胞毒性作用的影响。结果 NMDA(5mmol/L)引起明显的细胞内Ca2+升高以及细胞毒性作用,表现为细胞活力下降、凋亡细胞增加以及caspase-3表达升高。NMDA的细胞毒性作用可被NMDA受体特异性阻断剂MK801阻断。预先用α-Syn(2.5μmol/L,5μmol/L,10μmol/L)处理细胞明显抑制NMDA引起的细胞内Ca2+升高以及细胞毒性作用,其作用随α-Syn浓度增高而增强。结论α-Syn对NMDA所致多巴胺能神经元的细胞毒性作用具有抑制作用,其机制可能与其抑制与NMDA引起的Ca2+内流以及caspase-3激活有关。

Objective To investigate the effect of α-Synuclein（α-Syn） on N-methyl-D-aspartic acid（NMDA）-induced cytotoxicity.Methods NMDA cytotoxicity was produced on MES 23.5 dopaminergic cells.MTS cell viability assay,AO/PI fluorescent labeling of apoptotic cells,and western blot analysis of caspase-3 expression were used for assessment of the NMDA cytotoxicity.Confocal microscopy with Ca2＋ probe was applied for monitoring the change of intracellular Ca2＋ concentration.Results NMDA of 5 mmol/L triggered a significant increase of the cell death rate as revealed by decreased cell viability and increased apoptotic cells,which was blocked by a specific NMDA receptor antagonist MK801.In addition,NMDA induced significant increase in caspase-3 expression and intracellular Ca2＋ concentration.Pretreatment of the cells with α-Syn attenuated the NMDA-induced cell death,[Ca2＋]i up-regulation and caspase-3 increase.Conclusion α-Syn reduced NMDA-induced cytotoxicity through a mechanism of preventing caspase-3 activation and NMDA-induced Ca2＋ influx.