脑缺血再灌注后大鼠缺血侧脑组织中SIRT1的表达

摘要

目的:探讨脑缺血再灌注后大鼠缺血侧脑组织中沉默信息调节因子2同系物1(SIRT1)的表达和分布.方法:84只健康雄性SD大鼠随机分为假手术组和脑缺血再灌注组,脑缺血再灌注组再分为再灌注0、4、8、24、48及72 h共6个观察时间点亚组(n=12);脑缺血再灌注组采用插入线栓制作短暂性大脑中动脉栓塞(M CAO)模型(缺血2h),假手术组仅分离颈总、颈外和颈内动脉,不插入线栓;分别于术前、再灌注0、4、8、24、48及72 h,对大鼠进行神经功能缺损评分,采用免疫荧光单标染色法观察缺血侧脑组织中SIRT1的表达,采用Westem-Blot方法检测缺血侧脑组织匀浆中SIRT1蛋白表达.结果:假手术组大鼠神经功能缺损评分0分;再灌注大鼠2h恢复血流最初的24h内,大鼠神经功能缺损评分呈明显升高趋势,与假手术组比较差异有统计学意义(P＜0.05),但0h组与4h组、8h组与24 h组比较差异无统计学意义(P＞0.05),再灌注48 h组和72 h组大鼠神经功能缺损评分低于24 h组明显下降,差异有统计学意义(P＜0.05);假手术组和再灌注各时间点组大鼠缺血侧脑组织SIRT1主要表达于细胞质中;与假手术组比较,再灌注0h组、4h组、8h组和24 h组大鼠缺血侧脑组织中SIRT1蛋白表达显著减少,随着时间的推移呈下降趋势,两两比较差异有统计学意义(P＜0.05);与再灌注24 h组比较,在再灌注48h组和72 h组,大鼠缺血侧脑组织中SIRT1蛋白表达有所回升,组间两两比较,差异有统计学意义(P＜0.05).结论:脑缺血再灌注损伤时SIRT1表达水平发生变化,SIRT1可能参与脑缺血再灌注损伤的发生过程.%Objective:To investigate the expression and distribution of SIRT1 in rat ischemic brain after cerebral ischemia-reperfusion.Methods:84 healthy male SD rats were randomly divided into sham group and MCAO/R group.The rats in MCAO/R group were randomly assigned into reperfusion at 0 h,4 h,8 h,24 h,48 h and 72 h subgroups(n =12).The transient focal cerebral ischemiareperfusion model was established by modified thread-occlusion method (ischemia 2 h).Sham group only split CCA,external carotid and internal carotid without thread-occlusion method;Neurological deficit score was performed to evaluate rats' neurologic symptoms.Immunofluorescence staining were applied to analyze the expression and distribution of SIRT1 in rat ischemic brain,and Western blotting were applied to detect the SIRT1 protein expression in ischemic homogenate of brain tissue.Results:Rats' neurological deficit scores were 0 point in sham group.During reperfusion 24 h,rats' neurological deficit scores increased significantly,compared with sham group showed statistical significance (P ＜ 0.05).While the comparison between 0 h group and 4 h group,8 h group and 24 group showed no statistical significance (P ＞ 0.05);for reperfusion 48 h group and 72 h group,rats' neurological deficit scores were lower than 24 h group,differences were statistically significant (P ＜ 0.05).Sham group and reperfusion groups showed ischemic brain tissue SIRT1 mainly located in the cytoplasm.Compared with sham group,SIRT1 protein expression of reperfusion 0 h group,4 h group,8 h group and 24 h group decreased significantly,which further decreased with the time moved on,pairwise comparison showed statistical significance (P ＜ 0.05);protein expression of SIRT1 rise again in reperfusion 48 h and 72 h group compared with 24 h group;pairwise comparison within group showed statistical significance (P ＜ 0.05).Conclusions:SIRT1 expression changes may be associated with the occurrence of cerebral ischemia-reperfusion injury,SIRT1 might involve in the process of cerebral ischemia-reperfusion injury.