Aβ_(25-35)诱导SH-SY5Y细胞Tau蛋白的过度磷酸化

β-Amyloid Peptide 25-35 Induce Tau Hyperphosporylation in SH-SY5Y Cells

目的 采用凝聚态 Aβ2 5- 35建立 Tau蛋白过度磷酸化的阿尔茨海默病样细胞模型。 方法 用不同浓度凝聚态 Aβ2 5- 35作用于 SH - SY5 Y细胞 2 4 ,4 8,72 h,通过四唑盐比色法及乳酸脱氢酶释放实验观察 Aβ2 5- 35对 SH-SY5 Y细胞存活的影响 ;选择对细胞存活影响较小的适宜浓度和作用时间处理细胞 ,以蛋白免疫印迹法研究 Tau蛋白磷酸化水平的变化。 结果 Aβ2 5- 35对细胞存活的影响随着作用时间的延长或作用浓度的增加而增强。 1 0μm ol/ L Aβ2 5- 35作用于细胞 72 h或 2 0μm ol/ L Aβ2 5- 35作用于细胞≥ 4 8h时 ,MTT代谢率均明显下降 (P<0 .0 5 )。Aβ2 5- 35作用浓度 >2 0μmol/ L作用 72 h后 ,细胞 L DH释放率明显增加 (P<0 .0 5 )。选用细胞毒性小的 1 0 ,2 0μmol/L Aβ2 5- 35作用于 SH- SY5 Y细胞 0 ,3,6 ,1 2 ,2 4 ,4 8h,蛋白免疫印迹示 Tau蛋白在 Ser396、Ser1 99/ 2 0 2位点的磷酸化水平增高 ,于 6 h达到最高峰。 结论 凝聚态 Aβ2 5- 35可诱导建立阿尔茨海默病样

Objective To establish a cell model of Alzheimer like tau hyperphosphorylation in SH SY5Y cells induced by aggregated β amyloid peptide 25 35 (Aβ 25 35 ). Methods SH SY5Y cells were treated with different concentrations of aggregated Aβ 25 35 for 24, 48, 72 h, respectively. 3 [4,5 Dimethylthiazol 2 yl] 2, 5 diphenyltetrazolium bromide(MTT) and lactate dehydrogenase(LDH) assay were used to measure the viability of SH SY5Y cells after Aβ 25 35 exposure, in order to choose an optimal concentration and exposure time of Aβ 25 35 . Western blotting was used to detect the level of tau phosphorylation in SH SY5Y cells. Results Treatment of SH SY5Y cells with 10 μmol/L Aβ 25 35 for 72 h and 20 μmol/L Aβ 25 35 for 48 h caused a decrease in the metabolic rate of MTT. Treatment of SH SY5Y cells with 20 μmol/L Aβ 25 35 for 72 h caused a increase in the rate of LDH release. 10 and 20 μmol/L Aβ 25 35 induced tau hyperphosphorylation in SH SY5Y cells, and the immunoreactivity of anti phosphorylated tau at Ser 396 and Ser 199/202 reached a maximum level within 6 h after Aβ 25 35 exposure. Conclusion Aggregated Aβ 25 35 can induce tau hyperphosphorylation in SH SY5Y cells.