摘要
目的探讨肠缺血/再灌注(VR)损伤后乙醛脱氢酶-2(ALDH2)对线粒体应激的影响。方法32只雄性SD大鼠,随机分为假手术组、VR模型组、基因沉默(ALDH2-siRNA)组和空载体(Scrambled-siRNA)组(每组均8只)。结扎肠系膜上动脉血流1h、再灌注2h,建立肠VR损伤大鼠模型;假手术组不进行结扎。ALDH2-siRNA组和Scrambled-siRNA组于术前24、6、1h分别采用经尾静脉注射ALDH2-siRNA和Scrambled-siRNA混合液各1mL;假手术组、VR模型组则分别注射等量0.9%生理盐水注射液。再灌注2h后取血并处死大鼠取肠组织。采用酶联免疫吸附试验(ELISA)检测白细胞介素(IL-6)、肿瘤坏死因子受体相关因子6(TRAF6)水平;用Western Blot和RT-PCR分别检测肠组织ALDH2及线粒体应激相关因子,如热休克蛋白(HSP60、HSP70)、NIP3蛋白(Nix)及其mRNA的表达。结果与假手术组比较,VR模型组血清炎症因子明显升高[IL-6(ng/L):172.6±38.7比450.3±101.8,P<0.01;TRAF6(ng/L):499.5±142.1比962.5±213.8,P<0.01]及线粒体应激因子蛋白水平明显升高(HSP60:1.00±0.15比4.19±0.38,P<0.01;HSP70:1.00±0.12比3.22±0.41,P<0.01;Nix:1.00±0.12比3.03±0.31,P<0.01),且其mRNA水平明显升高(HSP60:1.00±0.23比11.16±0.40,P<0.01;HSP70:1.00±0.14比5.13±0.61,P<0.01;Nix:1.00±0.11比8.41±0.52,P<0.01),而ALDH2呈低水平表达;与VR模型组比较,ALDH2-siRNA组血清炎性因子水平明显升高[IL-6(ng/L):450.3±101.8比613.3±134.5,P<0.05;TRAF6(ng/L):962.5±213.8比1383.4±238.3,P<0.01)及线粒体应激因子蛋白水平明显升高(HSP60:4.19±0.38比6.02±0.51,P<0.01;HSP70:3.22±0.41比5.11±0.52,P<0.01;Nix:3.03±0.31比4.48±0.50,P<0.01),且其mRNA水平明显升高(HSP60:11.16±0.40比14.87±0.58,P<0.01;HSP70:5.13±0.61比7.17±0.65,P<0.01;Nix:8.41±0.52比9.97±0.78,P<0.01),而ALDH2水平明显降低。结论ALDH2可能介导炎症及线粒体应激信号通路起到抑制炎症反应及调控线粒体应激的稳态作用,从而保护大鼠肠VR损伤。
Objective To explore the effect of acetaldehyde dehydrogenase-2(ALDH2)on mitochondrial stress after the injury of intestinal ischemia/repeafusion(VR).Methods Thirty-two male Sprague-Dawley rats were randomly divided into pseudo-surgical group,IVR model group,ALDH2-siRNA group and Srambled-siRNA group.And each group had eight rats.Rat model of intestinal I/R injury was established by ligation the superior mesenteric artery blood flow for 1 h and reperfusion for 2 h.However,no ligation was performed in the pseudo-surgical group.ALDH2-siRNA group was injected ALDH2-siRNA 1 ml in preoperative 24,6,1 hour by tail vein.And scrambled-siRNA group was injected scrambled-siRNA.However,pseudo-surgical group and VR model group were injected with the same amount of sodium chloride injection.After 2 h reperfusion,blood was taken and intestinal tissue was sacrificed.The contents of interleukin(IL-6)and tumor necrosis factor receptor-associated factor 6(TRAF6)were detected by ELISA.The levels of ALDH2 and mitochondrial stress related factors,such as heat shock protein(HSP60,HSP70),NIP3 protein(Nix)and mRNA expression in intestinal tissues were detected by Western blot and reverse-transcription PCR(RT-PCR).Results Compared with the pseudo-surgical group,the levels of inflammatory cytokines were significantly increased in the IR model group[IL-6(ng/L):172.6土38.7vs.450.3±101.8,P<0.01;TRAF6(ng/L):499.5±142.1 vs.962.5±213.8,P<0.01].And the protein levels of mitochondrial stress factors were significantly increased in the I/R model group(HSP60:1.00±0.15 vs.4.19±0.38,P<0.01;HSP70:1.00±0.12 vs.3.22±0.41,P<0.01;Nix:1.00±0.12vs.3.03±0.31,P<0.01).In addition,mRNA levels of mitochondrial stress factors were significantly increased in the LR model group(HSP60:1.00±0.23 vs.11.16±0.40,P<0.01;HSP70:1.00±0.14 vs.5.13±0.61,P<0.01;Nix:1.00±0.11 vs.8.41±0.52,P<0.01).While the expression of ALDH2 was low.Compared with the IVR model group,the levels of serum inflammatory factors were significantly increased in the ALDH2-siRNA group[I
作者
王益群
姚平波
赵红
Wang Yi-qun;Yao Ping-bo;Zhao Hong(Department of Anesthesiology,Afiliated Nanhua Hospital,Nanhua University,Hengyang 421002,China)
出处
《中国急救医学》
CAS
CSCD
北大核心
2019年第5期467-470,共4页
Chinese Journal of Critical Care Medicine
基金
国家自然科学基金资助项目(81460433)
湖南省自然科学基金资助项目(2018JJ2339,2018JJ6122)
湖南省教育科学规划资助项目(XJK17BGD070).
