富硒黄芪皂苷抑制Aβ25-35诱导的阿尔茨海默病细胞模型细胞凋亡的作用机制

Mechanism of selenium-enriched astragalus saponins in inhibiting apoptosis of Alzheimer's disease cell model induced by Aβ25-35

目的:观察富硒黄芪皂苷对Aβ25-35诱导的阿尔茨海默病细胞模型细胞增殖、β-微管蛋白III(β-tubulin III)蛋白表达及磷脂酰肌醇-3-激酶/蛋白激酶B(PI3K/Akt)信号通路的影响。方法:利用40μmol·L^(-1)Aβ25-35处理大鼠肾上腺髓质嗜铬瘤分化细胞株PC12建立AD细胞模型。实验分为AD模型组、富硒黄芪皂苷干预组和对照组,富硒黄芪皂苷干预组加入1、2.5、5、10、20、50、100、200μmol·L^(-1)富硒黄芪皂苷干预,MTT法检测各组细胞的增殖活性,倒置显微镜观察富硒黄芪皂苷对细胞形态学的影响。免疫荧光法检测对照组、AD模型组和100μmol·L^(-1)富硒黄芪皂苷干预组β-tubulin III的表达,Western-blot法检测对照组、AD模型组和100μmol·L^(-1)富硒黄芪皂苷干预组PI3K、p-Akt、Bcl-2、Bax和Caspase-3的蛋白表达。结果:富硒黄芪皂苷50、100、200μmol·L^(-1)组贴壁细胞数目均明显多,细胞分布密度明显增加,以100μmol·L^(-1)组差异最显著。富硒黄芪皂苷100μmol·L^(-1)组干预24 h后能显著减轻Aβ25-35的毒性损伤,能明显提升AD细胞的β-tubulin III、PI3K、p-Akt和Bcl-2的蛋白水平(P<0.05),能明显降低Bax和Caspase-3的蛋白水平(P<0.05)。结论:富硒黄芪皂苷能显著改善AD细胞模型细胞形态以及提高细胞活力,其机制可能是通过激活PI3K/Akt信号通路抑制其细胞凋亡实现的。

Objective:To observe the effects of selenium-enriched astragalosides on cell proliferation,β-tubulin III(Iβ-tubulin III)protein expression and phosphatidylinositol-3-kinase/protein kinase B(PI3K/Akt)signal pathway induced by Aβ25-35.Methods:The AD cell model was established by treating rat adrenal medullary pheochromocytoma cell line PC12 with 40μmol·L^(-1)Aβ25-35.The experiment was divided into AD model group,selenium-rich astragaloside intervention group and control group.Selenium-rich astragaloside intervention group was treated with 1,2.5,5,10,20,50,100,200μmol·L^(-1)selenium-rich astragaloside.The proliferation activity of cells in each group was detected by MTT method,and the effect of selenium-rich astragaloside on cell morphology was observed under inverted microscope.The expression ofβ-tubulin III in control group,AD model group and 100μmol·L^(-1)selenium-enriched astragalus saponins intervention group was detected by immunofluorescence.The protein expressions of PI3K,p-Akt,Bcl-2,Bax and Caspase-3 in control group,AD model group and 100μmol·L^(-1)selenium-rich astragalus saponins intervention group were detected by Western-blot.Results:The number of adherent cells and the distribution density of adherent cells were significantly increased in selenium-rich astragalus saponins 50,100,200μmol·L^(-1)groups,and the difference was the most significant in 100μmol·L^(-1)group.After 24 hours of intervention,selenium-rich astragaloside 100μmol·L^(-1)could significantly reduce the toxic injury of Aβ25-35,increase the protein levels ofβ-tubulin III,PI3K,p-Akt and Bcl-2 of AD cells(P<0.05),and decrease the protein levels of Bax and Caspase-3(P<0.05).Conclusion:Selenium-enriched astragalosides can significantly improve the morphology and viability of AD cells,which may be achieved by activating PI3K/Akt signal pathway to inhibit cell apoptosis.