Study On The Mechanism Of Probenecid Ameliorating Cerebral Ischemic Injury By Inhibiting Brain Inflammation And Edema

Stroke causes inflammation,cerebral edema and neuronal death.However,there is currently no effective neuroprotective agent available for treatment.Recent studies have shown that the Pannexin 1(Panxl)channel is involved in ischemic brain injury and cellular inflammatory responses,while the Panxl inhibitor,probenecid,will reverse this damage,but the underlying mechanism is still unclear.In this study,we examined whether probenecid,the pannexin 1-channel inhibitor,alleviates focal ischemic brain damage by inhibiting brain inflammation and edema.Transient focal ischemia induced by middle cerebral artery occlusion(MCAO)was performed in C57BL/6J mice for 1 hour.Probenecid(lmg/ml)was injected intraperitoneally at the dose of lml/kg before MCAO and 2 hours after reperfusion,respectively.And infarct volume,neurological score and brain water content were assessed 48 hours after MCAO.Subsequently,the cerebral cortex was isolated,the meninges were removed and astrocytes were extracted from the cerebral cortex digested with trypsin and cultured in medium containing with different concentrations of probenecid.By the immunostaining,western blot analysis and ELISA,we assess the effects of probenecid on cellular inflammatory response,astrocyte activation and ROS production,HMGB1,IL-1? release and AQP4 expression.The experimental data showed that the probenecid diminished infarct size,reduced brain water content,inhibited neuronal death,and alleviated intracerebral inflammation 48 hours after stroke.In addition,the expression of AQP4 protein was significantly down-regulated in the ischemic penumbra after the treatment with probenecid.These results indicate that probenecid is a potent pannexin 1-channel inhibitor that prevents ischemic brain damage by inhibiting brain inflammation and edema.