氯化锂对冈田酸所致SK-N-SH神经元突触萎缩的保护作用

摘要

目的：考察氯化锂( LiCl)对蛋白磷酸酶抑制剂冈田酸( OA)诱导的SK-N-SH细胞分化神经元损伤的保护作用和tau蛋白Ser-262位点磷酸化水平的影响。方法利用全反式维甲酸( ATRA)诱导SK-N-SH细胞分化为成熟的神经元细胞；采用OA诱导成熟神经元细胞建立AD模型；采用磺酰罗丹明B( SRB)比色法考察LiCl对成熟的神经元细胞增殖的抑制作用；Giemsa染色观察SK-N-SH细胞形态学变化；并采用Image-Proplus软件测定神经元细胞的突触长度；采用Western blot检测synaptophysin蛋白和tau蛋白Ser-262位点磷酸化水平。结果10μmol · L-1 ATRA连续处理7 d,可诱导SK-N-SH细胞突触生长和synaptophysin蛋白表达等典型分化神经元的特征。20~100 nmol · L-1 OA作用于分化神经元,可浓度和时间依赖性抑制细胞增殖,同时致分化神经元突触萎缩, tau蛋白 Ser-262位点磷酸化水平也明显升高。10 mmol·L-1 LiCl预处理可维持synaptophysin蛋白高表达,抑制tau蛋白Ser-262位点磷酸化水平( P<0．01)。结论 LiCl能够改善 OA 所致分化神经元的突触损伤,并伴随着synaptophysin表达的升高、tau蛋白Ser-262位点异常磷酸化水平的降低。%Aim To explore the protective effects of lithium chloride ( LiCl ) on neurous injuries and phos-phorylation of tau protein at serine262 induced by okada-ic acid( OA) . Methods The neuroblastoma SK-N-SH cells were differentiated by all-trans-retinoic acid ( AT-RA) . The differentiated SK-N-SH cells were treated with OA to establish the Alzheimer′s disease cellular model. SK-N-SH cells′ viability and proliferation were measured by SRB test. Giemsa staining was used to observe cell morphology. The neurite length of SK-N-SH cells was measured by Image-Proplus software. Syn-aptophysin and phosphorylated tau protein at serine262 expression levels were tested by Western blot. Results The SK-N-SH cells which were treated with 10 μmol ·L-1 ATRA for 7 days displayed mature neuronal fea-tures. The synaptic length of SK-N-SH cells became longer. And the levels of serine262 phospho-tau was sig-nificantly elevated. 20~100 nmol·L-1 OA effectively inhibited the viability of differentiated SK-N-SH cells in a concentration-dependent manner and in a time-de-pendent manner. The OA treatment induced obvious synaptic atrophy in differentiated SK-N-SH cells. And the phosphorylation level of tau protein serine262 also greatly increased. The pretreatment with 10 mmol · L-1 LiCl significantly ameliorated the synaptic atrophy, the decrease of synaptophysin expression and the in-crease of tau phosphorylation at serine262 induced by OA in differentiated SK-N-SH cells. Conclusion LiCl could effectively inhibit OA-induced synaptic atro-phy in differentiated SK-N-SH cells, and it could also result in the increase of synaptophysin expression and the decrease of the phosphorylation of tau protein at serine262 .