Effect Of GPER On Estrogen-induced Cerebral Ischemia-reperfusion Injury

Objective: To investigate whether estrogen can regulate the expression of endoplasmic reticulum stress-related proteins and mediate cerebral ischemia-reperfusion injury(Gerebral ischemia-reperfusion injury)by mediating its specific protein receptor GPER(G protein-coupled estrogen receptor).CIRI).This further clarifies the possible mechanism by which estrogen-related receptors play a role in improving cerebral ischemia-reperfusion injury.Methods: The normal female SD rats selected in this study were required to establish an Ovariectomy(OVX)model,rats were divided into OVX group,OVX+I/R group,OVX+I/R+vehicle(DMSO),OVX+I/R+E2,OVX+I/R+G1(GPER agonist),OVX+I/R+E2+G15(GPER blocker).In this study,the animal model of cerebral ischemia-reperfusion injury was prepared by middle cerebral artery occlusion(MCAO).After 90 minutes of ischemia and 24 hours of reperfusion,Longa score was used to detect neurobehavioral changes,brainstem wet weight ratio was used to detect brain edema,2,3,5-triphenyltetrazolium chloride(TTC)The cerebral infarction volume was detected by staining.The morphological changes of neurons were observed by Nissl staining.The deoxynucleotidyl transferase-mediated deoxynucleotide triphosphate nick end labeling(TUNEL staining)was used to detect the apoptosis of neurons in each group.Immunofluorescence was used to detect the distribution and expression of Caspase-12 in the endoplasmic reticulum.The distribution and expression of caspase12 were detected by immunofluorescence.The endoplasmic reticulum emergency related proteins GRP78 and Caspase were detected by PCR and Western-blot.12 and CHOP mRNA and protein levels.Result:(1)Compared with the OVX+I/R group,the neurobehavioral scores of the OVX+I/R+E2 group and the OVX+I/R+G1 group were significantly lower(p < 0.05),and OVX+I/ The neurobehavioral scores of the R+E2 group were lower than those of the OVX+I/R+G1 group(p < 0.05).Compared with the OVX+I/R+E2 group,the neurobehavioral score of the OVX+I/R+E2+G15 group increased(p < 0.05).(2)Compared with the OVX+I/R group,OVX+I The brain edema degree was significantly decreased in the /R+E2 group and the OVX+I/R+G1 group(p < 0.05),and the cerebral edema degree in the OVX+I/R+E2 group was lower than OVX+I/R+G1.Group(p < 0.05).Compared with the OVX+I/R+E2 group,the degree of cerebral edema in the OVX+I/R+E2+G15 group increased(p < 0.05).(3)Compared with the OVX+I/R group,OVX+I/ The cerebral infarction volume of R+E2 group and OVX+I/R+G1 group was significantly decreased(p < 0.05),and the degree of cerebral infarction in OVX+I/R+E2 group was lower than OVX+I/R+G1.Group(p < 0.05).Compared with the OVX+I/R+E2 group,the degree of cerebral infarction increased in the OVX+I/R+E2+G15 group(p < 0.05).(4)Nissl staining showed that compared with the OVX+I/R group The degree of neuronal damage in hippocampus of OVX+I/R+E2 group and OVX+I/R+G1 group was significantly decreased,and the degree of neuronal damage in OVX+I/R+E2 group was lower than OVX+I/R+G1.group.Compared with the OVX+I/R+E2 group,the degree of neuronal damage in the OVX+I/R+E2+G15 group increased.(5)TUNEL staining showed that compared with OVX+I/R group,neuronal apoptosis in hippocampus of OVX+I/R+E2 group and OVX+I/R+G1 group was significantly decreased(p < 0.05).),and the neurons in the OVX+I/R+E2 group were lower than the OVX+I/R+G1 group(p < 0.05).Compared with OVX+I/R+E2 group,neuronal apoptosis in OVX+I/R+E2+G15 group increased(p < 0.05).(6)Immunofluorescence detection of Caspase-12 expression in hippocampus of rats The results showed that the expression of Caspase-12 was decreased in the hippocampus of OVX+I/R+E2 group and OVX+I/R+G1 group compared with OVX+I/R group(p < 0.05),and Caspase-12 of OVX+I/R+E2 group.The expression was lower than the OVX+I/R+G1 group(p < 0.05).Compared with the OVX+I/R+E2 group,the expression of Caspase-12 was increased in the OVX+I/R+E2+G15 group(p < 0.05).(7)The protein levels of GRP78,Caspase-12 and CHOP in rat hippocampal CA1 region were detected by Western-blot technique.The results showed that OVX+I/R+E2 group and OVX+I/R compared with OVX+I/R group.The expression of GRP78,Caspase-12 and CHOP protein was decreased in the +G1 group(p < 0.05),and the protein expression in the OVX+I/R+E2 group was lower than that in the OVX+I/R+G1 group(p < 0.05).Compared with the OVX+I/R+E2 group,the expression of GRP78,Caspase-12 and CHOP protein in the OVX+I/R+E2+G15 group increased(p < 0.05).(8)The mRNA levels of GRP78,Caspase-12 and CHOP in rat hippocampus were detected by PCR.The results showed that OVX+I/R+E2 group and OVX+I/R+G1 compared with OVX+I/R group.The mRNA expression of GRP78,Caspase-12 and CHOP was decreased(p < 0.05),and the expression of OVX+I/R+E2 group was lower than that of OVX+I/R+G1 group(p < 0.05).Compared with the OVX+I/R+E2 group,the mRNA expression of GRP78,Caspase-12 and CHOP in the OVX+I/R+E2+G15 group increased(p < 0.05).Conclusion: GPER is involved in the process of estrogen inhibiting the expression of endoplasmic reticulum stress-related proteins,alleviating neuronal apoptosis and improving dysfunction caused by cerebral ischemia-reperfusion injury,suggesting that estrogen inhibits endoplasmic reticulum stress and thus protects the brain.In the middle,there is GPER participation.