氯化锂抑制β-淀粉样蛋白诱导细胞Tau蛋白磷酸化

目的探讨GSK-3β抑制剂氯化锂(LiCl)在β-淀粉样蛋白25～35片断(Aβ25-35)诱导的细胞Tau蛋白磷酸化中的作用及机制。方法MTT法观察不同浓度的LiCl(1、5、10、20mmol.L-1)单独作用24h,对SH-SY5Y细胞存活率的影响;20μmol.L-1的凝聚态Aβ25-35作用于SH-SY5Y细胞不同时间点,蛋白免疫印迹法研究Tau蛋白磷酸化位点(Ser199、Ser396及Tau1)水平的变化;LiCl(20mmol.L-1)预处理细胞1h后,观察Aβ的作用有无变化及可能的作用机制。结果不同浓度的LiCl作用24h后,MTT法示细胞存活率无明显变化(P>0.05);20μmol.L-1Aβ25-35作用不同时间点后,Tau蛋白在Ser396、Ser199位点的磷酸化水平在3h逐渐增加,6h达到最高峰,12h后又逐渐下降,在这3个时间点的增加量均具有统计学意义(P<0.05),而对非磷酸化Tau1没有影响(P>0.05);LiCl预处理可抑制Aβ25-35的作用(P<0.05),并可诱导失活形式的GSK-3β在Ser9位点的磷酸化(p-GSK-3βSer9)表达增高。结论GSK-3β参与了Aβ25-35诱导细胞Tau蛋白磷酸化的作用,LiCl可通过诱导失活形式的p-GSK-3βSer9表达增高而抑制GSK-3β的活性,进而抑制Aβ诱导的细胞Tau蛋白磷酸化。该实验为研究AD的发病机制及探索有效治疗药物提供了重要的理论基础。

The effects of LiCl on Tau phosphorylation induced by β-amyloid peptide

Aim To investigate the effects of LiCl,an inhibitor of glycogen synthase kinase 3β(GSK-3β)on tau hyperphosphorylation induced by β-amyloid peptide in SH-SY5Y cells and its possible mechanism.Methods The viability of cells and the level of tau phosphorylation at Ser396,Ser199 and Tau1 were detected by MTT and Western blot,respectively.The effects and possible mechanism of LiCl(20 mmol·L-1)on Aβ-induced cytotoxicity were studied.Results The viability of SH-SY5Y cells was unchanged after the exposure to different doses of LiCl.Western blot showed that Aβ25-35 induced the increased phosphorylation of two sites(Ser396,Ser199)tested,began to increase at 3h,reached its perk at 6 h,and tended to decline at 12 h(P<0.05).But totle tau recognized by Tau 1 was unchanged(P>0.05).Pretreatment of LiCl was able to suppress Aβ25-35-induced tau phosphorylation and also increase the expression of p-GSK-3βSer9.Conclusions The GSK-3β was involved in the Aβ25-35-induced tau phosphorylation.The block of Aβ25-35-induced-tau phosphorylation by LiCl is dependent on suppressing the GSK-3β activity through increasing the expression of p-GSK-3βSer9.