SIRT1在小鼠肠缺血再灌注损伤中的作用分析

Role of SIRT1 in intestinal ischemia-reperfusion injury in rats

摘要:

目的：探讨SIRT1在小鼠肠缺血再灌注损伤中的作用。方法：45只大鼠分为假手术组、模型组和SIRT1组。假手术组和模型组经腹腔注射空载体腺相关病毒，SIRT1组大鼠经腹腔注射过表达SIRT1的腺相关病毒。注射21 d后，模型组和SIRT1组大鼠采用肠系膜上动脉夹闭-松夹闭方式建立小肠缺血再灌注模型，假手术组大鼠仅做肠系膜上动脉分离。再灌注后24 h后处死大鼠，收集血清和小肠标本。采用ELISA试剂盒检测3组大鼠外周血炎症因子白细胞介素（interleukin，IL）-1β、IL-6和肿瘤坏死因子（tumor necrosis factor，TNF）-α水平，采用试剂盒检测小肠组织中谷胱甘肽过氧化物酶（glutathione peroxidase，GSH-PX）、超氧化物歧化酶（superoxide dismutase，SOD）和丙二醛（malondialdehyde，MDA）的水平；采用Western blot分析3组大鼠小肠组织凋亡蛋白的表达水平；采用TUNEL染色分析小肠绒毛细胞凋亡指数。结果：与假手术组比较，模型组和SIRT1组大鼠外周血炎症因子水平（IL-1β、IL-6和TNF-α）和MDA水平明显上调，抗氧化物酶水平（GSH-PX和SOD）明显下调（P<0.05）。与模型组比较，SIRT1组大鼠外周血炎症因子水平（IL-1β、IL-6和TNF-α）和MDA水平明显下调，抗氧化物酶水平（GSH-PX和SOD）明显升高（P<0.05）。与假手术组比较，模型组和SIRT1组大鼠小肠组织中Caspase-3和Bax表达明显上调，而Bcl-2表达明显下调（P<0.05）。与模型组比较，SIRT1组大鼠小肠组织中Caspase-3和Bax表达明显下调，而Bcl-2表达明显上调（P<0.05）。与模型组比较，SIRT1组大鼠小肠上皮细胞凋亡指数明显下降（P<0.05）。结论：过表达SIRT1可明显降低小肠缺血再灌注大鼠炎症、氧化应激和细胞凋亡水平，进而保护小肠组织，为临床提供一定的指导意义。

Abstract:

Objective：To investigate the role of SIRT1 in intestinal ischemia-reperfusion injury in rats. Methods：45 rats were divided into sham-operated group，model group and SIRT1 group. The sham-operated group and model group were intraperitoneally injected with adeno-associated virus of empty vector and the SIRT1 group was intraperitoneally injected with adeno-associated virus of over－expression SIRT1. After 21 days of injection，rats in model group and SIRT1 group were treated with superior mesenteric artery oc－clusion-relaxation occlusion to establish small intestinal ischemia-reperfusion model，while rats in sham-operated group were only treated with superior mesenteric artery separation. 24 hours after reperfusion，rats were killed. Serum and small intestine samples were collected. The levels of inflammatory factors of interleukin-1β（IL-1β），interleukin-6（IL-6） and tumor necrosis factor-α（TNF-α） in peripheral blood of three groups of rats were detected by ELISA kit. The levels of glutathione peroxidase（GSH-PX），superoxide dis－mutase（SOD） and malondialdehyde（MDA） in small intestine tissue were detected by ELISA kit. The expression level of apoptotic protein in small intestine tissue of three groups of rats was analyzed by Western blot. The apoptotic index of small intestinal villus cells was analyzed by TUNEL staining. Results：Compared with sham-operated group，the levels of inflammatory factors（IL-1β，IL-6 and TNF-α） and MDA in peripheral blood significantly increased，but GSH-PX and SOD level decreased significantly in small in－testine of rats in model group and SIRT1 group（P<0.05）. Compared with the model group，the levels of inflammatory factors（IL-1β，IL-6 and TNF-α） and MDA in peripheral blood in SIRT1 group decreased significantly，but GSH-PX and SOD level increased sig－nificantly（P<0.05）. Compared with sham-operated group，the expression of Caspase-3 and Bax in intestinal tissue of model group and SIRT1 group increased significantly，while the expression of Bcl-2 decreased significantly（P<0.05）. Compared with model group，the expression of Caspase-3 and Bax in small intestine tissue of SIRT1 group decreased significantly，while the expres－sion of Bcl-2 increased significantly（P<0.05）. Compared with the model group，the apoptotic index of intestinal epithelial cells in SIRT1 group decreased significantly（P<0.05）. Conclusion：Overexpression of SIRT1 can protect small intestinal tissues by reducing the levels of inflammation，oxidative stress and cell apoptosis in rats with small intestinal ischemia-reperfusion，which provides some guidance for clinical practice.