The Role Of JAK2/STAT3 Pathway In Cerebral Ischemia-reperfusion Injury Induced By Intraoperative Hypotension Through Regulating HIF-1α

Organ ischemia and hypoxia caused by intraoperative hypotension(IOH)is associated with many negative postoperative outcomes.Brain is a sensitive organ to ischemia and hypoxia,although cerebral blood flow can be maintained within range of certain blood pressure through autonomic regulation.But when blood pressure drops to a certain level,the autonomic regulation mechanism of the brain will be shocked,and accompanies with ischemia-hypoxia and reperfusion injury.However,the extent and duration of hypotension that can cause brain damage is unknown.HIF-1α is one of the main regulatory elements of the cellular response to hypoxia,and there are currently different opinions about its role in cerebral ischemia-reperfusion injury.As one of the main upstream pathways of HIF-1α,the JAK2/STAT3 pathway has the functions of regulating cell proliferation,apoptosis,inflammatory response and angiogenesis.Therefore,this study will verify the role of HIF-1α in brain injury caused by intraoperative hypotension through in vivo and in vitro,and further explore its related molecular mechanisms.一.The role of HIF-1α in cerebral ischemia-reperfusion injury induced by perioperative hypotensionObjective:To explore the role of HIF-1α in cerebral ischemia-reperfusion injury caused by perioperative hypotension.Methods:In vitro,the appropriate drug dose was determined by the CCK-8 experiment,and the cells were divided into several groups,including Control group,YC-1 group,DMOG group,OGD/R group,OGD/R+YC-1 group,OGD/R+DMOG group,Afterwards,different groups were treated with drugs 24 hours before the start of the experiment,and the OGD4h and R24h experiments were performed to simulate cerebral ischemia-reperfusion injury.LDH assay and Western blotting were used to observed apoptotic protein associated with HIF-1α.In vivo,30 experimental healthy white rabbits were divided into 5 groups,namely the sham operation group,the solvent control group,the YC-1 group,the CH group,and the CH+YC-1 group.The marginal veins were treated with corresponding drugs,sodium nitroprusside was used to control blood pressure after the start of the experiment,the MAP was reduced to 50%of the basic value for 30 minutes,and the MAP was restored to the basic value after maintaining for 2 hours.Then the damage of hippocampal area of rabbits in different groups was judged,the apoptosis of hippocampal area was judged by TUNEL,and the ratio of BCL-2/BAX and HIF-la protein expression in different groups were detected by WB and Immunohistochemistry.Results:1.In vitro,compared with the control group,the ratio of BCL-2/BAX in the OGD/R group decreased,the expression of HIF-1α increased,and the LDH leakage rate increased(p<0.05);after stimulating the HIF-1α protein,the OGD/R+DMOG group Compared with the OGD/R group,the ratio of BCL-2/BAX was increased,the expression of HIF-1α was increased,and the leakage rate of LDH was decreased(p<0.05);after inhibiting HIF-1α,the ratio of BCL-2/BAX in the OGD/R+YC-1 group was higher than that in the OGD group The/R group decreased,the expression of HIF-1α decreased,and the LDH leakage rate increased(p<0.05).2.In vivo,after HE staining in the CH group and the CH+YC-1 group,the cells were loosely arranged,pyknosis,nuclear fragmentation,and nuclear edge aggregation were observed,among which the CH+YC-1 group was more obvious;The results of TUNEL and WB showed that there was no significant difference in the apoptosis and protein expression among the sham-operated group,the solvent control group,and the YC-1 group;Compared with the sham operation group,the CH group had increased apoptosis,and the expression of apoptotic proteins and HIF-1α were increased(p<0.05);Compared with the CH group,the CH+YC-1 group had increased apoptosis and the expression of apoptotic proteins were increased,the expression of HIF-1α protein were decreased(p<0.05).二.The role of JAK2/STAT3 in cerebral ischemia-reperfusion injury induced by intraoperative hypotension by regulating the expression of HIF-laObjective:To explore the effect of JAK2/STAT3 in cerebral ischemia-reperfusion injury caused by intraoperative hypotension by regulating the expression of HIF-1α.Methods:In vitro,the appropriate drug dose was determined by the CCK-8 experiment,and the cells were divided into several groups,includingControl group,OGD/R+AG-490 group,OGD/R group,OGD/R+AG-490 group,Afterwards,different groups were treated with drugs 24 hours before the start of the experiment,and the OGD4h and R24h experiments were performed to simulate cerebral ischemia-reperfusion injury.LDH assay and Western blotting were used to observed apoptotic protein associated with HIF-lα.In vivo,30 healthy rabbits were divided into 5 groups,including sham operation group,the solvent control group,the AG-490group,the OGD/R group,and the OGD/R+AG-490group.The marginal veins were treated with corresponding drugs,sodium nitroprusside was used to control blood pressure after the start of the experiment,the MAP was reduced to 50%of the basic value for 30 minutes,and the MAP was restored to the basic value after maintaining for 2 hours.Then the damage of hippocampal of rabbits in different groups was judged,the apoptosis of hippocampal was judged by TUNEL,and the ratio of BCL-2/BAX and HIF-1α protein expression in different groups were detected by Western blotting and Immunohistochemistry.Results:1.Compared with the OGD/R group,the ratio of BCL-2/BAX in the OGD/R+AG-490 group were decreased,the expression of HIF-1α were decreased,and the leakage rate of LDH were increased(p<0.05).2.In vivo,HE staining in CH group and CH+AG-490 group,the cells were loosely arranged,nuclear pyknosis,nuclear fragmentation,nuclear edge aggregation and other phenomena appeared,among which was more obvious in CH+AG-490 group;Compared with the CH group,the CH+AG-490 group had increased apoptosis,the expression of apoptotic protein were increased,and the expression of HIF-1α protein were decreased(p<0.05).Conclusion:It was verified by in vitro and in vivo that inhibiting HIF-1α protein aggravated the cerebral ischemia-reperfusion injury in intraoperative hypotension;inhibiting JAK2/STAT3 pathway would reduce HIF-1α protein expression and aggravate intraoperative hypotension-induced cerebral ischemia reperfusion injury.