冈田酸诱导人神经母细胞瘤细胞系SH-SY5Y损伤

Okadaic acid induces damage of human neuroblastoma cell line SH-SY5Y

目的探索冈田酸(OA)诱导神经元微管蛋白相关蛋白(Tau)异常磷酸化对人神经母细胞瘤细胞系(SH-SY5Y)损伤的影响。方法将OA按照不同浓度(20和40 nmol/L)和/或时间(0、12、24、48和72 h)处理SH-SY5Y细胞;光学显微镜下观察细胞形态学;透射电子显微镜观察细胞超微结构;TUNEL法检测细胞凋亡;Western blot检测p-Tau和caspase3蛋白的表达;RT-qPCR检测细胞凋亡抑制家族蛋白(IAPs)的mRNA表达。结果与对照组相比,OA显著增加SH-SY5Y细胞Tau蛋白磷酸化(P<0.001),促进细胞损伤凋亡(P<0.001),损伤线粒体结构,同时也可显著上调IAPs表达(P<0.05)。结论OA诱导的SH-SY5Y细胞的Tau蛋白异常磷酸化可导致细胞损伤凋亡,抑制内源凋亡通路或上调IAPs可能作为阿尔茨海默病(AD)防治的重要靶标。

Objective To investigate the effect of okadaic acid(OA)-induced phosphorylation of neuron micro-tubule associated protein tau(Tau)on the damage of human neuroblastoma SH-SY5Y cells.Methods SH-SY5Y cells were incubated with different concentrations(20 and 40 nmol/L)of OA for 0,12,24,48 and 72 hours followed by microscopy for cell morphology.Cell ultrastructure was observed by transmission electron microscope and cell apoptosis was detected by TUNEL assay.The protein expression of p-Tau and caspase3 was analyzed by Western blot and mRNA level of inhibitor of apoptosis family proteins(IAPs)was determined by RT-qPCR.Results OA induced phosphorylation of Tau protein in SH-SY5Y cells as compared to the result from control goup(P<0.001),promoted cell damage and apoptosis(P<0.001).OA also injured mitochondrial structure in a concentration and time dependent manner.It can also up-regulate IAPs expression significantly(P<0.05).Conclusions The aberrant phosphorylation of Tau protein in SH-SY5Y cells induced by OA may lead to cell injury and apoptosis.Inhibition of endogenous apoptotic pathways or up-regulation of IAPs family proteins provides potential targets for AD prevention and treatment.