PI3K/Akt/GSK-3β通路在体外培养神经元缺氧缺血损伤中的作用

The effect of PI3K/Akt/GSK-3β signaling pathway on the injury of neurons after hypoxia-ischemia in vitro

目的探讨神经元缺氧缺血损伤时,PI3K/Akt/GSK-3β信号通路各分子间的调控机制及干预该信号通路对神经突损伤修复的作用。方法培养新生SD大鼠皮质神经元,建立体外神经元氧糖剥夺(OGD)模型。实验分为4组:正常对照组,OGD组,LY294002干预组,SB415286干预组。Western blot方法检测神经元丝氨酶/苏氨酸蛋白激酶(Akt)、磷酸化Akt(p-Akt)、糖原合成激酶-3β(GSK-3β)、p-GSK-3β的蛋白表达变化。应用神经元微管蛋白(TUJ1)免疫荧光染色观察神经突受损情况。结果与正常对照组比较,体外培养神经元OGD后,p-Akt、p-GSK-3β表达降低(P<0.05)。与OGD组比较,LY294002干预后p-Akt和p-GSK-3β表达减少(P<0.05),神经突损伤明显加重。与OGD组比较,SB415286干预后p-GSK-3β表达增加(P<0.05),神经突损伤减轻。结论 PI3K/Akt/GSK-3β信号通路调节OGD后神经突损伤。

Objective To investigate the regulation mechanisms among PI3 K/Akt/GSK-3βsignaling pathway in cultured cortical neurons after hypoxia-ischemia(HI),and explore the neurites injury reparation after intervening this signaling pathway.Methods The cerebral cortical neurons from newborn Sprague-Dawley(SD)rats were cultured in vitro and established oxygen-glucose deprivation(OGD)model which imitated HI environment.Cells were divided randomly into 4 groups:the control group,the OGD group,the LY294002 intervention group and the SB415286 intervention group.Western blot was used to detect the protein expression of serine/threonine protein kinase(Akt),phospho-serine/threonine protein kinase(p-Akt),glycogen synthase kinase-3β(GSK-3β),phospho glycogen synthase kinase-3β(p-GSK-3β)in neurons.Cells were immunostained with anti-neuron specific classⅢβ-tubulin(TUJ1)to observe the injury of neurites.Results As compared with the control group,the expression of p-Akt and p-GSK-3βwas significantly decreased after OGD(P<0.05).As compared with the OGD group,the expression of p-Akt and p-GSK-3β was significantly decreased in the LY294002 intervention group(P<0.05),and the neurites injury was obviously aggravated.As compared with the OGD group,the p-GSK-3β expression was significantly increased in the SB415286 intervention group(P<0.05),and the immunofluorescence also showed a significant reduction in neurites injury.Conclusion PI3K/Akt/GSK-3β signaling pathway is involved in the regulation of neurites injury after OGD.