淫羊藿苷抑制内质网应激介导HT22细胞抗谷氨酸诱导凋亡

目的探讨淫羊藿苷(ICA)通过调控内质网应激(ERS)通路保护小鼠海马神经元HT22细胞抗谷氨酸(Glu)损伤的机制。方法筛选适当的ICA预处理浓度,以Glu(5 mmol/L)处理18 h构建细胞损伤模型。将细胞分为对照组、Glu损伤组、低剂量ICA+Glu(L-ICA+Glu)组和高剂量ICA+Glu(H-ICA+Glu)组。CCK-8法检测细胞活性,TUNEL染色显示凋亡细胞,比色法检测细胞乳酸脱氢酶(LDH)释放量,JC-1染色检测线粒体膜电位(MMP)变化,DCFH-DA检测细胞活性氧(ROS)水平,Western blot检测ERS通路和凋亡相关蛋白含量。此外,以上各组经CHOP siRNA预处理后,分为Glu+ControlsiRNA组、Glu+CHOPsiRNA组、H-ICA+Glu+ControlsiRNA组、H-ICA+Glu+CHOPsiRNA组,检测细胞活性、凋亡率、ROS含量、MMP水平及凋亡相关蛋白表达。结果ICA在浓度低于10μmol/L时无明显毒性作用。ICA能够显著提高HT22细胞活性,降低LDH释放量,抑制细胞凋亡,降低ROS水平,恢复MMP,且呈剂量依赖性(P<0.05)。此外,ICA下调损伤后p-eIF2α和CHOP表达,同时增加Bcl2,抑制Bax(P<0.05)。CHOP siRNA预处理后,Glu的损伤作用明显减弱;同时CHOP siRNA预处理能够明显增强ICA对HT22细胞的保护作用。结论ICA可能通过部分抑制ERS诱导的氧化应激及凋亡,保护HT22细胞抗Glu损伤。

Icariin protects HT22 cells against glutamate induced apoptosis by inhibition of endoplasmic reticulum stress

Objective To investigate the mechanism of icariin(ICA)protecting mouse hippocampal neuron HT22 cells against glutamate(Glu)injury by regulating the endoplasmic reticulum stress(ERS)pathway.Methods The appropriate pretreatment concentration of ICA were screened,and the cell injury model was constructed by Glu(5 mmol/L)treatment for 18 hours.The cells were divided into the control group,the Glu injury group,the low-dose ICA+Glu(L-ICA+Glu)group and the high-dose ICA+Glu(H-ICA+Glu)group.The cell viability was detected by CCK-8 assay,the apoptotic cell was assessed by TUNEL staining,the release of cell lactate dehydrogenase(LDH)was detected by colorimetric method,the change of mitochondrial membrane potential(MMP)was detected by JC-1 staining,the cellular reactive oxygen species(ROS)was detected by DCFH-DA,and ERS pathway and apoptosis-related protein content were detected by Western blot.In addition,after pretreated with CHOP siRNA in the above groups,they were divided into the Glu+ControlsiRNA group,Glu+CHOPsiRNA group,H-ICA+Glu+ControlsiRNA group,H-ICA+Glu+CHOPsiRNA group.Cell viability,apoptosis rate,ROS content,MMP level and apoptosis-related protein expression were detected.Results ICA had no significant toxic effect at concentrations below 10μmol/L.ICA could significantly increase the viability of HT22 cells,decrease LDH release,inhibit cell apoptosis,reduce ROS levels,and restore MMP in a dose-dependent manner(P<0.05).In addition,ICA down-regulated the expression of p-eIF2αand CHOP after injury,while increased Bcl2 level and inhibited Bax level(P<0.05).After CHOP siRNA pretreatment,the injury effect of Glu was significantly weakened.At the same time,CHOP siRNA pretreatment could significantly enhance the protective effect of ICA on HT22 cells.ConclusionICA may protect HT22 cells against Glu injury by partially inhibiting ERS-induced oxidative stress and apoptosis.