Neuro-protective Effect And The Potential Mechanism Of Serpina3n On Cerebral Ischemia Reperfusion Injury In Mice

Objective: Ischemic stroke is an important cause of premature death and disability.At present,the strategy for the treatment of acute ischemic stroke is to perform revascularization within a strict time window.After revascularization treatment cerebral ischemia-reperfusion injury is an unavoidable problem.A series of biochemical cascades occur in ischemic brain tissue during cerebral ischemia-reperfusion injury.In our previou study of transcriptome of rat four-vessel occlusion model,it is found that serpina3 n dramaticly increased.The dramatic up-regulation suggests that it may play an important role in cerebral ischemia-reperfusion injury.Existing research on serpina3 n in the central nervous system has focused on the its expression as a reactive astrocyte marker and neuroinflammatory marker,but its functional research is limited and its neuroprotective effects are still controversial.The role of serpina3 n in cerebral ischemia reperfusion injury is currently unknown.Therefore,this study focuses on the expression pattern and function of serpina3 n in cerebral ischemia-reperfusion injury,and hopes to explore its mechanism of action.Methods: In this study,the expression pattern of serpina3 n in a mouse model of transient middle cerebral artery occlusion was detected by immunofluorescence,Western Blot analysis and real-time quantitative PCR.To investigate the function of serpina3 n in cerebral ischemia-reperfusion injury and its effects on the motor function and infarct size of mice after transient middle cerebral artery occlusion,adeno-associated virus are use to infect mouse brain to perform over-expression and interference of serpina3 n.Coimmunoprecipitation-mass spectrometry and immunofluorescence co-staining are used to identify the interaction proteins with serpina3 n and Western Blot is used to analyze possible downstream pathways.Results: Our study found that serpina3 n is expressed in astrocytes and neurons in penumbra after reperfusion.The expression pattern of serpina3 n gradually increased from 6h after reperfusion,and reached a peak on the second to third day after reperfusion,then decreased until the seventh day,when the level of serpina3 n still higher than the sham operation group.In mice with over-expression of serpina3 n undergoing transient middle cerebral artery occlusion,it is found that their infarct size shrank,with improvd motor function,alleviated inflammation and oxidative stress;while in serpina3 n interfered mice experienced transient middle cerebral artery occlusion,the infarct size increased,with exacerbated inflammatory response and oxidative stress.These results indicate that serpina3 n plays a neuroprotective role in cerebral ischemia-reperfusion injury.Coimmunoprecipitation-mass spectrometry and used Western Blot identified clusterin interact with serpina3 n during cerebral ischemia-reperfusion injury in mice.Studies on downstream mechanisms have found that the the level of activation of the JAK2-STAT3 pathway and Akt pathway in cortex of mice overexpressing serpina3 n is significantly higher than that in mice injected with control virus,while in striatum the level of activation of the JAK2-STAT3 pathway was significantly lower than mice injected with control virus.Conclusions: Serpina3 n is expressed in astrocytes and penumbra neurons after cerebral ischemia-reperfusion injury in mice,and reduce brain tissue damage possibly via interacting with clusterin to affect the JAK2-STAT3 pathway and Akt pathway.