Effects Of STC2 In METH-Induced Nerve Cell Apoptosis

Objective: To investigate the effect of STC2 on METH-induced nerve cell apoptosis and to explore the possible mechanism on the molecular level.Methods: In vitro experiments,PC12 cells were treated with concentration gradient METH(0,0.1,0.5,1,1.5,2,2.5,3 m M)for 24 h,CCK-8 was used to detect cell viability,to preliminarily determine the cytotoxic effect of METH;PC12 cells were treated with concentration gradient METH(0,1,1.5,2,2.5 m M)for 24 h,m RNA and protein expression levels of STC2 and ERS marker molecules(GRP78,CHOP)were detected by q RT-PCR and Western blot,then PC12 cells were treated with 2.5m M METH according to time gradient(0,3,6,12,24 h),m RNA and protein expression levels of STC2,GRP78,and CHOP were detected by q RT-PCR and Western blot;According to the experimental results,the follow-up in vitro experiment was finally carried out with 2.5 m M METH for 24 h;The ERS specific inhibitor 4-PBA was used to pretreat PC12 cells for 30 min before METH intervention to verify the role of ERS in the apoptosis induced by METH;The expression of STC2 was silenced by the si RNA sequential transfection method,the PC12 cells were divided into the si RNA-NC group,the si RNA-NC+METH group,the si RNA-STC2 group,and the si RNA-STC2+METH group,the expression levels of STC2 and ERS related apoptotic proteins(GRP78,CHOP,Bcl-2,Bax,cleaved caspase-3,cleaved caspase-12)were detected by Western blot,cell viability was measured by CCK-8,and cell apoptosis rate was measured by TUNEL and flow cytometry.In vivo experiment,12 SD rats were randomly divided into the Control group and the METH group,adopting METH acute poisoning model.The damage of striatal neurons was detected by HE staining and nissl staining,and the expression levels of STC2 and ERS related apoptotic proteins were detected by Western blot,to determine the toxic effect of METH on striatal neurons;The expression of STC2 was silenced in the striatum by si RNA in vivo,24 male SD rats were randomly divided into the si RNA-NC group,the si RNA-NC+METH group,the si RNA-STC2 group,and the si RNA-STC2+METH group,Western blot was used to detect the expression levels of STC2 and ERS related apoptotic proteins,TUNEL staining was used to detect the apoptosis,the damage of striatal neurons was detected by HE staining and nissl staining.Results: METH treatment significantly inhibited the cell viability,and the cell viability decreased to 75% after 2.5m M METH treatment(P<0.01).m RNA and protein expression levels of STC2,GRP78,and CHOP in PC12 cells were significantly increased after METH treatment,and in a concentration and time dependence(P<0.01).Compared with the si RNA-NC group,the protein level of STC2 in si RNA-STC2 group was significantly decreased(P<0.01).Compared with the si RNA-NC+METH group,the si RNA-STC2+METH group had higher expression levels of GRP78,CHOP,Bax,cleaved caspase-3,and cleaved caspase-12(P<0.01),cell viability was further decreased(P<0.01)and apoptosis rate was further increased(P<0.01).After acute exposure to METH,the expression levels of STC2 and ERS related apoptotic proteins in the rat striatum were significantly increased(P<0.01).The results of HE staining and nissl staining showed that,compared with the control group,the structure of neurons in the striatum of METH group was not clear,neuron shrinkage and nissl bodies decreased.After STC2 silenced in the striatum,compared with the si RNA-NC+METH group,the damage of striatal neurons in the si RNA-STC2+METH group was more serious(P<0.01),the apoptosis rate was further increased(P<0.01),and the expression levels of ERS related apoptotic proteins was further up-regulated(P<0.01).Conclusion: STC2 promotes METH-induced nerve cell apoptosis through negative regulation of ERS.