δ阿片受体激动剂对脓毒症大鼠心肌细胞凋亡及心肌组织Bcl-2、Caspase-3表达的影响

Effects of δ-opioid Receptor Agonists on Cardiocyte Apoptosis and Expression 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)对脓毒症大鼠心肌细胞凋亡及心肌组织Bcl-2、Caspase-3表达的影响,探讨DADLE对心肌组织保护的可能机理。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(雌雄不拘),利用随机数字表随机分为CLP组(n=18)、DADLE组(n=18)和假手术(SHAM)组(n=18)。在不同时间点(6h、12h、24h)处死大鼠,原位末端标记检测心肌细胞凋亡情况,免疫组化法检测心肌组织中Bcl-2、Caspase-3蛋白表达的动态变化并比较心肌组织的病理改变。结果 CLP组大鼠心肌组织病理损害较SHAM组明显严重,DADLE组心肌组织的病理变化明显改善;CLP组大鼠心肌组织细胞凋亡指数较SHAM组明显升高(P<0.01),以12h组最明显(P<0.01),各DADLE组心肌细胞凋亡指数明显降低(P<0.01);CLP组大鼠心肌组织Caspase-3的表达比SHAM组明显增加(P<0.01)、Bcl-2的表达明显降低(P<0.05);DADLE组心肌组织Caspase-3的表达较CLP组明显降低(P<0.01)、Bcl-2的表达明显增加(P<0.05)。心肌组织Caspase-3的表达与心肌细胞凋亡指数正相关(r=0.92,P<0.01),Bcl-2的表达与心肌细胞凋亡指数负相关(r=-0.94,P<0.01)。结论 DADLE明显改善脓毒症大鼠心肌组织的病理变化,其作用机理可能与DADLE下调Caspase-3表达、上调Bcl-2表达,从而抑制心肌细胞凋亡有关。

OBJECTIVE To observe the effects of δ-opioid receptor agonists DADLE（（D-Ala2,D-Leu5）-enkephali） on cardiocyte apoptosis and expressions of Bcl-2 and Caspase-3 in septic rat,and to investigate the possible mechanism by which DADLE protects the myocardial tissue in sepsis.METHODS Sepsis was reproduced in rats by cecun ligation and puncture（CLP）.Forty-two SD rats（either male or female） were randomly divided into CLP group（n=18）,DADLE group（n=18） and sham-operation（SHAM） group（n=18）.The rats were respectively killed at different time（6h,12h and 24h after operation）.Cardiocyte 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 myocardial tissue were detected by light microscope.RESULTS The myocardial pathological lesion of rats in CLP groups was obviously serious compared with SHAM group,while it was improved obviously in DADLE group.The apoptosis index of rat cardiocytes in CLP group significantly increased compared with SHAM group（P0.01）,and further it was prominently at 12h（P0.01）.The apoptosis index of rat cardiocytes at each time of DADLE group was significantly decreased compared with CLP group（P0.01）.Expression of Caspase-3 protein in myocardial tissues of CLP group significantly increased compared with the control group（P0.01）,while the expression of Bcl-2 protein significantly decreased（P0.05）.Expression of Caspase-3 protein in myocardial tissues of DADLE group significantly decreased compared with the CLP group（P0.01）,while the expression of Bcl-2 protein significantly decreased（P0.05）.There was positive correlation between expression of Caspase-3 in myocardial tissues and apoptosis index of cardiocyte（r=0.92,P0.01）and negative correlation between expression of Bcl-2 in myocardial tissues and apoptosis index of cardiocyte（r=-0.94,P0.01）.CONCLUSION The findings indicate that δ-opioid receptor agonists DADLE can obviously improve myocardial pathological changes of septic rats.And its protective mechanism contains downregulation of Caspase-3 expression,upregulation of Bcl-2 expression and thus the apoptosis of cardiocyte is repressed.