δ阿片受体激动剂对脓毒症大鼠肝细胞凋亡及肝组织bcl-2和caspase-3表达的影响

Effects of δ-Opioid Receptor Agonists on Hepatocyte Apoptosis and Expressions of bcl-2 and caspase-3 in Septic Rat

目的观察δ阿片受体激动剂D-丙2,D-亮5脑啡肽(D-Ala2,D-Leu5-enkephali,DADLE)对脓毒症大鼠肝细胞凋亡及肝组织bcl-2和caspase-3表达的影响,探讨DADLE对肝脏保护作用的可能机理。方法采用盲肠结扎加穿孔(CLP)法制作大鼠脓毒症模型,SD大鼠54只(雌雄不限),采用随机数字表法随机分为CLP组(n=18)、DADLE组(n=18)和假手术组(n=18)。在不同时间点(2、4及6 h)处死大鼠,原位末端标记法检测肝细胞凋亡情况,免疫组化法检测肝组织中bcl-2及caspase-3蛋白表达的动态变化并观察肝脏的病理改变。结果CLP组大鼠肝组织病理损害明显较假手术组严重,DADLE组肝脏的病理变化明显改善;CLP组大鼠肝组织细胞凋亡指数明显较假手术组升高(P<0.01),以4 h最显著(P<0.01),DADLE组各时相肝细胞凋亡指数均明显降低(P<0.01);CLP组大鼠肝组织caspase-3蛋白的表达强度比假手术组明显增强(P<0.01),而bcl-2蛋白的表达则明显减弱(P<0.05);DADLE组肝组织caspase-3蛋白的表达强度较CLP组明显减弱(P<0.01),而bcl-2蛋白的表达则明显增强(P<0.05)。肝组织caspase-3的表达与肝细胞凋亡指数呈正相关(r=0.83,P<0.01),bcl-2的表达与肝细胞凋亡指数呈负相关(r=-0.65,P<0.01)。结论DADLE能明显改善脓毒症大鼠肝脏的病理变化,其作用机理可能与DADLE下调caspase-3表达、上调bcl-2表达,从而抑制肝细胞凋亡有关。

Objective To observe the effects of δ-opioid receptor agonists D-Ala^2-D-Leu^5-enkephali （DADLE） on hepatocyte apoptosis and expressions of bcl-2 and caspase-3 in septic rat, and to investigate the possible mechanism by which DADLE protects the liver in sepsis. Methods Sepsis was reproduced in rats by cecum libation and puncture （CLP）. Fifty-four SD rats （either male or female） were randomly divided into CLP group （n= 18）, DADLE group （n=18） and sham operation （SO） group （n=18）. The rats were respectively killed at different time （2 h, 4 h and 6 h after operation）. Hepatoeyte apoptosis was detected by TdT-mediated dUTP Nick End Labeling （TUNEL）. The expressions of bcl-2 and caspase-3 protein were detected by immunohistochemistry. And the changes of pathology in hepatic tissue were detected by light microscope. Results The hepatic pathological lesion of rats in CLP group was obviously serious compared with SO group, while it was obviously improved in DADLE group. The apoptosis index of rat hepatocytes in CLP group significantly increased compared with SO group, and further it was prominent at 4 h （P〈0.01）. The apoptosis index of rat hepatocytes at each time of DADLE group was significantly decreased compared with CLP group （P〈0.01）. Expression of caspase-3 protein in liver tissues of CLP group significantly increased compared with SO group （P〈0. 01）, while the expression of bcl-2 protein significantly decreased （P〈0.05）. Expression of caspase-3 protein in liver tissues of DADLE group significantly decreased