摘要
目的观察瑞芬太尼对大鼠全脑缺血再灌注损伤后海马神经细胞凋亡和半胱天冬酶(Caspase)-3表达的影响。方法80只大鼠随机分为假手术(sham)、NS、REM2、REM6和REM20组,每组16只。NS、REM2、REM6和REM20组分别于缺血再灌注前静脉滴注0.9%氯化钠溶液及瑞芬太尼2、6、20μg.kg-1.min-1。采用双侧颈总动脉阻断+低血压法建立大鼠短暂性全脑缺血模型。于全脑缺血前30 min由靶控微量注射泵经股静脉注射瑞芬太尼,并于双侧颈总动脉阻断前及开放后10 min进行动脉血血气分析。再灌注后24 h,每组各取8只大鼠,取其新鲜海马组织,采用实时逆转录-聚合酶链反应(RT-PCR)检测各组Caspase-3 mRNA的表达。再灌注后72 h,取剩余大鼠,灌注取脑,制作脑组织切片,采用苏木精-伊红(H-E)染色和原位末端标记法(TUNEL法)观察各组大鼠海马退变的神经元数和凋亡细胞数,采用免疫组织化学法检测各组Caspase-3蛋白的表达。结果①H-E染色结果:sham组偶可见退变的锥体细胞,缺血再灌注72 h后,NS组及各瑞芬太尼预处理组海马退变的神经元数较sham组显著增加(P值均<0.05),REM6及REM20组海马退变的神经元数较NS组显著减少(P值均<0.05)。②TUNEL法检测结果:sham组未见TUNEL阳性细胞,缺血再灌注72 h后,NS组及各瑞芬太尼预处理组海马CAl区的凋亡锥体细胞数较sham组显著增加(P值均<0.05),REM6和REM20组海马CAl区的凋亡锥体细胞数较NS组显著降低(P值均<0.05)。③免疫组织化学法检测结果:sham组可见少量Caspase-3免疫阳性细胞,缺血再灌注72 h后,NS组及各瑞芬太尼预处理组海马区Caspase-3蛋白的免疫阳性细胞数较sham组显著增多(P值均<0.05),各瑞芬太尼预处理组Caspase-3蛋白的免疫阳性细胞数显著低于NS组(P值均<0.05),REM6和REM20组Caspase-3蛋白的免疫阳性细胞数显著低于REM2组(P值均<0.05)。④实时RT-PCR结果:sham组大鼠海马组织中Caspase-3 mRNA呈低水平表达,�
Objective To study the effect of remifentanil preconditioning on neuron apoptosis and Caspase-3 expression after forebrain ischemia/reperfusion（I/R） injury in rats.Methods Eighty male Sprague-Dawlay rats were randomly divided into 5 groups： sham group（n=16）;control group（NS,n=16）;REM2 group（remifentanil 2 μg·kg^-1·min^-1,n=16）;REM6 group（remifentanil 6 μg·kg-1·min-1,n=16）;and REM20 group（remifentanil 20 μg·kg^-1·min^-1,n=16）.Transient forebrain ischemia was induced by bilateral common carotid artery（CCA） occlusion for 10 min combined with hypotension.Rats received preconditioning with intravenous NS,remifentanil 2,6 or 20μg·kg^-1·min^-1 30 min before occlusion.The right femoral artery was cannulated to collect 0.5 mL blood samples for analysis blood gas 10 min before ischemia and 10 min after ischemia.Half of rats in each group were killed 24 h after reperfusion,and real-time reverse transcriptase polymerase chain reaction（RT-PCR） was used to detect the expression of Caspase-3 mRNA in the hippocampus.The rest rats were killed 72 h after reperfusion,and brain paraffin sections were prepared.The total degenerated neurons in hippocampal CA1 were counted blindly under high power field.The number of neuronal apoptosis was assessed by terminal oxynucleotidyl transferase mediated dUTP biotin nick end labeling（TUNEL）.The apoptosis-related proteins caspase-3 were analyzed,and immunohistochemical was used to detect caspase-3 protein.Results ① The degenerated neurons in hippocampal CA1 were significantly increased in NS,REM2,REM6,and REM20 groups compared with that in the sham group after 72 hours reperfusion.Degenerated neurons in hippocampal CA1 were significantly reduced in REM6 and REM20 groups compared with that in the NS group（P〈0.05）.② TUNEL showed no positive cells in the sham group.The neuron apoptosis increased significantly in the hippocampus CA1 in REM groups 72 h after reperfusion.The numbers of neuronal apoptosis in hippocampus CAl were significantly
出处
《上海医学》
CAS
CSCD
北大核心
2010年第2期123-127,I0001,共6页
Shanghai Medical Journal
