摘要
目的 探讨红景天苷调节核因子-E2 相关因子 2(Nrf2)/抗氧化反应元件(ARE)信号通路对氧糖剥夺再灌注(OGD/R)诱导的神经元铁死亡的影响.方法 将小鼠神经元细胞HT22 随机分为对照组、模型组、红景天苷低剂量(20 μmol/L)组、红景天苷高剂量(40 μmol/L)组、红景天苷(40 μmol/L)+ML385(Nrf2 抑制剂,1 μmol/L)组,除对照组外其余组细胞均进行OGD/R诱导.通过MTT和CCK-8法检测各组细胞活性和增殖能力;铁试剂盒检测细胞内铁浓度;商品化试剂盒检测各组细胞GPX4、SOD、MDA、ROS活性;透射电镜观察大鼠神经元超微结构的变化;Western Blot检测GPX4、COX2、ACSL4 及NRF2/ARE信号通路相关蛋白表达.结果 与对照组比较,模型组神经元细胞超微结构被破坏,线粒体缩小,嵴消失,外膜破裂,细胞活力和增殖率、GPX4、SOD活性、Nrf2、HO-1、GPX4 蛋白表达显著降低,Fe2+浓度、MDA、ROS活性、COX2、ACSL4 蛋白表达显著升高(P<0.05).与模型组比较,红景天苷低、高剂量组细胞超微结构明显改善,细胞活力和增殖率、GPX4、SOD活性、Nrf2、HO-1、GPX4 蛋白表达显著升高,Fe2+浓度、MDA、ROS活性、COX2、ACSL4 蛋白表达显著降低(P<0.05).与红景天苷高剂量组比较,红景天苷+ML385 组显著逆转了上述指标变化.结论 红景天苷能够激活Nrf2/ARE信号通路抑制OGD/R诱导的神经元铁死亡.
Abstract
Objective To investigate the regulatory effects of salidroside on neuronal iron death induced by oxygen glucose deprivation/reperfusion(OGD/R)by regulating the nuclear factor E2 related factor 2(Nrf2)/antioxidant response element(ARE)signaling pathway.Methods Mouse neuronal cells HT22 were treated with blank control,induced with OGD/R,OGD/R+low-dose salidroside(20μmol/L),OGD/R+high-dose salidroside(40μmol/L),OGD/R+ high-dose salidroside(40μmol/L)+ ML385(Nrf2 inhibitor,1μmol/L).Cell viability and proliferation were detected by MTT assay and CCK-8 assay,respectively.Intracellular iron concentration was measured using an iron assay kit.Activities of glutathione peroxidase 4(GPX4),superoxide dismutase(SOD),malondialdehyde(MDA),and reactive oxygen species(ROS)were measured using commercial kits.Changes in the ultrastructure of mouse neurons were observed by transmission electron microscopy.Western Blot was applied to detect the expressions of GPX4,COX2,ACSL4,and associated proteins in the Nrf2/ARE signaling pathway.Results Compared with those treated with blank control,mouse neurons induced with OGD/R presented damaged ultrastructure of neuronal cells,mitochondrial shrinkage,disappearance of the mitochondrial cristae,ruptured outer membrane,significantly lower cell viability,proliferation rate,activities of GPX4 and SOD,and protein expressions of Nrf2,HO-1,and GPX4,but significantly higher Fe2+ concentration,activities of MDA and ROS,and protein expressions of COX2 and ACSL4(P<0.05).Compared with those induced with OGD/R,cells induced with OGD/R+low-dose salidroside(20μmol/L)and OGD/R+high-dose salidroside(40μmol/L)presented significantly improved ultrastructure,significantly increased cell viability,proliferation rate,activities of GPX4 and SOD,and protein expressions of Nrf2,HO-1,and GPX4,and significantly reduced Fe2+ concentration,activities of MDA and ROS,and protein expressions of COX2 and ACSL4(P<0.05).Compared with those induced with OGD/R + high-dose salidroside(40μmol/L),the above indexes were significantly reversed in cells induced with OGD/R+ high-dose salidroside(40μmol/L)+ML385(Nrf2 inhibitor,1μmol/L).Conclusion Salidroside can inhibit OGD/R-induced neuronal ferroptosis by activating the Nrf2/ARE signaling pathway.
维普
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