Sigma-1受体在喷他佐辛减轻SH-SY5Y细胞氧糖剥夺-复氧复糖损伤中的作用:与内质网应激的关系

Role of Sigma-1 receptor in pentazoxine-induced reduction of oxygen-glucose deprivation and restoration injury in SH-SY5Y cells:relationship with endoplasmic reticulum stress

目的:评价Sigma-1受体(Sigma-1R)在喷他佐辛减轻SH-SY5Y细胞氧糖剥夺-复氧复糖(OGD/R)损伤中的作用及其与内质网应激的关系。方法:将生长良好的SH-SY5Y细胞采用随机数字表法分为4组( n=6):对照组(C组)、OGD/R组(O组)、OGD/R+喷他佐辛组(OP组)和OGD/R+喷他佐辛+BD1047组(OPB组)。C组细胞正常培养;O组、OP组和OPB组将培养基更换为EBSS培养基,置于培养箱中培养4 h,然后更换为含10%胎牛血清的DMEM/F12培养基复氧复糖18 h,OP组复氧复糖时加入喷他佐辛(终浓度10 μmol/L),OPB组复氧复糖时加入喷他佐辛(终浓度10 μmol/L)和Sigma-1R阻断剂BD1047(终浓度20 μmol/L)。复氧复糖18 h时采用流式细胞仪检测细胞凋亡率,采用Western blot法检测Sigma-1R、C/EBP同源蛋白(CHOP)、磷酸化需肌醇酶1(p-IRE1)、剪接型X-框结合蛋白1(XBP1s)和活化的半胱氨酸天冬氨酸蛋白酶3(c-cas-3)表达。 结果:与C组比较,O组、OP组和OPB组细胞凋亡率升高,CHOP和p-IRE1表达上调,O组和OPB组XBP1s和c-cas-3表达上调( P<0.05),OP组Sigma-1R、XBP1s和c-cas-3表达差异无统计学意义( P>0.05);与O组比较,OP组细胞凋亡率降低,Sigma-1R表达上调,CHOP、p-IRE1、XBP1s和c-cas-3表达下调( P<0.05);与OP组比较,OPB组细胞凋亡率升高,Sigma-1R表达下调,CHOP、p-IRE1、XBP1s和c-cas-3表达上调( P<0.05)。 结论:Sigma-1R参与了喷他佐辛减轻SH-SY5Y细胞氧糖剥夺-复氧复糖损伤的过程,机制可能与抑制内质网应激有关。

Objective To evaluate the role of Sigma-1 receptor(Sigma-1R)in pentazoxine-induced reduction of oxygen-glucose deprivation and restoration(OGD/R)injury in SH-SY5Y cells and the relationship with endoplasmic reticulum stress(ERS).Methods The well-growing SH-SY5Y cells were divided into 4 groups(n=6 each)using a random number table method:control group(group C),OGD/R group(group O),OGD/R+pentazoxin group(group OP)and OGD/R+pintazoxin+BD1047 group(group OPB).The cells in group C were normally cultured.In O group,OP group and OPB group,the culture medium was replaced with EBSS medium,and then the cells were cultured in an incubator of 5%CO2-95%N2 at 37℃for 4 h,then replaced with DMEM/F12 medium containing 10%fetal bovine serum for restoration of O2-glucose supply for 18 h,and in addition pentazoxin(final concentration 10μmmol/L)was added during restoration in OP group,and pentazoxin(final concentration 10μmmol/L)and Sigma-1R blocker BD1047(final concentration 20μmol/L)were added during restoration in OPB group.The apoptosis rate was detected by flow cytometry at the end of restoration,and the expression of Sigma-1R,C/EBP homologous protein(CHOP),phosphorylated inositol-requiring enzyme 1(p-IRE1),spliced X-box binding protein 1(XBP1s),and activated caspase-3(c-cas-3)was detected by Western blot.Results Compared with group C,the apoptosis rate was significantly increased,and the expression of CHOP and p-IRE1 was up-regulated in O group,OP group and OPB group,the expression of XBP1s and c-cas-3 was significantly up-regulated in O group and OPB group(P<0.05),and no significant change was found in the expression of Sigma-1R,XBP1s and c-cas-3 in OP group(P>0.05).Compared with O group,the apoptosis rate was significantly decreased,the expression of Sigma-1R was up-regulated,and the expression of CHOP,p-IRE1,XBP1s and c-cas-3 was down-regulated in OP group(P<0.05).Compared with OP group,the apoptosis rate was significantly increased,the expression of Sigma-1R was down-regulated,and the expression of CHOP,p-IRE1,XBP1s and c-cas-3 was up-regulated in OPB group(P<0.05).Conclusions Sigma-1R is involved in pentazoxine-induced reduction of OGD/R injury in SH-SY5Y cells,and the mechanism may be related to inhibition of endoplasmic reticulum stress.