Effect Of Selective Brain Hypothermia On SUMOylation Of Cerebral Cortex During Cerebral Ischemia-reperfusion Injury In Rats

Objective:To investigate the effect of selective brain hypothermia induced by intra-carotid artery infusion of cold normal saline on SUMOylation of cerebral cortex during cerebral ischemia-reperfusion injury in rats and to provide new research targets for hypothermic brain protection.Methods:100 healthy clean-grade male Sprague-Dawley rats,aged 8 weeks,weighing 200-250g,were divided into 4 groups according to random number table:sham operation group?group S?,cerebral ischemia-reperfusion injury group?group I/R?,infusion with 37?normal saline group?group N?,selective brain hypothermia group?group H??n=25 in each group?.A rat model of focal cerebral ischemia-reperfusion injury was prepared by occlusion of the right middle cerebral artery?MCAO?using a nylon thread with rounded tip in rats.The right middle cerebral artery was occluded for 2h removing the nylon thread before reperfusion in group I/R.10-13?normal saline was infused at the rate of 100 ml�kg^-1�h^-1 for 20 minutes starting from the time point immediately after removing the nylon thread in group H.37?normal saline was infused instead at the same rate for 20 minutes in group N.In group S,only the right common carotid artery,internal carotid artery and external carotid artery of the rat were isolated,and the middle cerebral artery was not occluded by a nylon thread.Detection index:the cerebral water contents was measured by dry-wet weight method to evaluate the degree of cerebral edema at 24h after reperfusion.The neurological deficit score was assessed by using the Zea Longa neurological deficit score at 24h and 48h after reperfusion.HE staining was used to observe pathological changes of ischemic cerebral cortex after cerebral ischemia-reperfusion injury.TUNEL staining was used to calculate the apoptosis rate of ischemic cerebral cortex after cerebral ischemia-reperfusion injury to evaluate the apoptosis.Western Blot was used to detect the expression of SUMO specific binding enzyme?Ubc9?,SUMO specific proteases 3?SENP3?and SUMO2/3 conjugated protein in cerebral cortex.Results:?1?Compared with group S,cerebral water contents was significantly increased at 24h after reperfusion in the other three groups?P<0.01?;compared with group I/R and group N,cerebral water contents was decreased at 24h after reperfusion in group H?P<0.05?,and there was no significant difference between group I/R and group N?P>0.05?.?2?Compared with group S,neurological deficit score was significantly increased at 24h and 48h after reperfusion in the other three groups?P<0.01?;compared with group I/R and group N,neurological deficit score was decreased at 24h and 48h after reperfusion in group H?P<0.05?,and there was no significant difference between group I/R and group N?P>0.05?.?3?HE staining showed that compared with group S,there was different degrees of nuclear pyknosis and fragmentation at 24h and 48h after reperfusion in the other three groups;compared with group I/R and group N,the degree of nuclear pyknosis and fragmentation was reduced in group H.?4?TUNEL staining showed that compared with group S the apoptosis rate was increased at 24h and 48h after reperfusion in the other three groups?P<0.01?;compared with group I/R and group N,the apoptotic rate was decreased at 24h and 48h after reperfusion in group H?P<0.05?,and there was no significant difference between group I/R and group N?P>0.05?.?5?Western blot showed that compared with group S,the expression of Ubc9 and SUMO2/3 conjugates were up-regulated in the other three groups,the expression of SENP3 was up-regulated in group I/R and group N,and the expression of SENP3 was down-regulated in group H?P<0.01?;compared with group I/R and group N,the expression of Ubc9 and SUMO2/3conjugates were up-regulated,the expression of SENP3 was down-regulated in group H?P<0.05?,there was no significant difference between group I/R and group N?P>0.05?.Conclusions:?1?Under the conditions of this experiment,Selective brain hypothermia induced by intra-arterial infusion of cold normal saline can reduce degree of cerebral edema,neurological impairment,pathological damage of brain tissue and degree of apoptosis in rats after focal cerebral ischemia-reperfusion injury,reduce brain damage and play the role of brain protection.?2?Its possible mechanism is related to up-regulation the expression of Ubc9 and SUMO2/3 conjugates in cerebral cortex,down-regulate the expression of SENP3,and enhancement of SUMOylation in cerebral cortex.