胰岛素对APPsw细胞内tau蛋白磷酸化的影响及其机制研究

Effect of insulin on tau protein hyperphosphorylation in APPsw cell and its mechanism exploration

目的 采用过度表达APPsw基因的SH-SY5Y细胞（APPsw细胞）,体外观察不同浓度的胰岛素对APPsw细胞内tau蛋白磷酸化的影响.方法 应用Western blot技术检测APPsw细胞内tau蛋白磷酸化的表达;应用免疫荧光双标与共聚焦激光扫描显微镜检测APPsw细胞内p-GSK-3β和p-tau的共表达.结果 Western blot结果显示,与对照组相比,1 000 nM的胰岛素使APPsw细胞内Thr231和Ser396位点上的tau蛋白磷酸化表达水平分别降低到（68.91±13.55）和（45.53±22.16） （P＜ 0.05）;共聚焦显微镜检测结果显示p-tau和p-GSK-3β主要分布于细胞质中,而且在APPsw细胞中二者的表达有一定关系.结论 胰岛素抑制APPsw细胞内tau蛋白的过度磷酸化,其机制可能与GSK-3β蛋白活性减小有关.

Objective To observe the effects of insulin at different concentrations on tau protein phosphorylation in vitro using an SH-SY5Y cell line overexpressing APPsw gene （APPsw cell）.Methods Western blot was used to detect the expression of tau phosphorylation.The coexpression of between p-GSK-3β and p-tau in APPsw cells was detected by double immunofluorescence and confocal laser scanning microscopy.Results Western blot results showed that 1 000 nM insulin treatment decreased the phosphorylation of tau protein at Thr231 and Ser 396 by（68.91 ± 13.55） and （45.53±22.16）,respectively,compared with the normal control （P＜ 0.05） in APPsw cells.The confocal microscopic analysis showed that p-tau and p-GSK-3β colocalized predominantly in cell cytoplasm,and there is some interaction between intracellular expression of p-GSK-3β and p-tau.Conclusion Insulin can suppress the tau protein hyperphosphorylation by potentially inhibiting the activity of GSK-3β.