Nrf2/ARE通路在饱和氢盐水减轻自体肝移植大鼠肝缺血再灌注损伤中的作用

Role of Nrf2/ARE pathway in hydrogen-rich saline-induced reduction of liver ischemia-reperfusion injury in rats undergoing liver autotransplantation

目的 探讨核转录因子E2相关因子2（Nrf2）/反应元件（ARE）通路在饱和氢盐水减轻自体肝移植大鼠肝缺血再灌注损伤中的作用.方法 SPF级成年健康雄性SD大鼠32只,8～10周龄,体重200～ 250 g,采用随机数字表法分为4组（n=8）：假手术组（S组）、自体肝移植组（T组）、饱和氢盐水组（H组）和Nrf2抑制剂全反式维甲酸＋饱和氢盐水组（A组）.A组腹腔注射全反式维甲酸7 mg/kg,1次/d,连续2d,最后1次给药后10 h建立自体肝移植模型.H组和A组在建立自体肝移植模型前5 min,肝下下腔静脉注射4℃饱和氢盐水6 ml/kg.门静脉开放后6h（S组术毕后6h）时,取血标本,测定血清ALT、AST、TNF-α和IL-10浓度,随后取肝组织,行肝组织病理学评分,测定肝组织MDA含量、SOD活性和Bcl-2、Bax、Nrf2和血红素氧合酶-1（HO-1）的mRNA表达水平.结果 与S组比较,T组血清ALT和AST浓度、肝组织病理学评分、MDA含量和血清TNF-α浓度升高,SOD活性和血清IL-10浓度降低,肝组织Bax、Nrf2、HO-1的mRNA表达上调,Bcl-2 mRNA表达下调（P＜0.05）;与T组比较,H组血清ALT和AST浓度、肝组织病理学评分、MDA含量和血清TNF-α浓度降低,SOD活性和血清IL-10浓度升高,肝组织Bcl-2、Nrf2、HO-1的mRNA表达上调,Bax mRNA表达下调（P＜0.05）;与H组比较,A组血清ALT和AST浓度、肝组织病理学评分、MDA含量和血清TNF-α浓度升高,SOD活性和血清IL-10浓度降低,肝组织Bcl-2、HO-1 mRNA表达下调,Bax mRNA表达上调（P＜0.05）.结论 饱和氢盐水减轻自体肝移植大鼠肝缺血再灌注损伤的机制与激活Nrf2/ARE通路,上调HO-1表达有关.

Objective To investigate the role of nuclear factor E2-related factor 2 （Nrf2）/antioxidant response element （ARE） pathway in hydrogen-rich saline-induced reduction of liver ischemiareperfusion （I/R） injury in rats undergoing liver autotransplantation.Methods Thirty-two SPF adult male Sprague-Dawley rats, aged 8-10 weeks, weighing 200-250 g, were randomly divided into 4 groups （n=8each） using a random number table ： sham operation group （group S）;liver autotransplantation group （group T）;hydrogen-rich saline group （group H）;Nrf2 inhibitor all-trans retinoic acid ＋ hydrogen-rich saline group （group A）.In group A, all-trans retinoic acid 7 mg/kg was injected intraperitioneally once a day for 2 consecutive days, and the model of liver autotransplantation was established at 10 h after the last administration.In H and A groups, hydrogen-rich saline 6 ml/kg was injected through the infrahepatic inferior vena cava at 5 min before establishment of the model.At 6 h after the portal vein was unclamped （at 6 h after the end of operation in group S）, blood samples were obtained for determination of serum alanine aminotransferase （ALT）, aspartate aminotransferase （AST）, tumor necrosis factor-alpha （TNFα） , and interleukin-10 （IL-10） concentrations.The rats were then sacrificed, and livers were removed for microscopic examination of pathologic changes which were scored and for determination of malondialdehyde （MDA） content, superoxide dismutase （SOD） activity, and expression of Bcl-2, Bax, Nrf2 and heme oxygenase-1 （HO-1） mRNA.Results Compared with group S, the serum ALT, AST and TNF-α concentrations, pathologic score, and MDA content were significantly increased, the SOD activity and serum IL-10 concentrations were decreased, the expression of Bax, Nrf2 and HO-1 mRNA was up-regulated, and the expression of Bcl-2 mRNA was down-regulated in group T （P〈0.05）.Compared with group T, the serum ALT, AST and TNF-α concentrations, pathologic score, and MDA content were significantly decreased, the SOD activity and serum IL-10 concentrations were increased, the expression of Bcl-2, Nrf2 and HO-1 mRNA was up-regulated, and the expression of Bax mRNA was down-regulated in group H （P 〈 0.05）.Compared with group H, the serum ALT, AST and TNF-α concentrations, pathologic score, and MDA content were significantly increased, the SOD activity and serum IL-10 concentrations were decreased, the expression of Bcl-2 and HO-1 mRNA was down-regulated, and the expression of Bax mRNA was up-regulated in group A （P〈0.05）.Conclusion The mechanism by which hydrogen-rich saline reduces liver I/R injury is associated with activated Nrf2/ARE pathway and upregulated expression of HO-1 in rats undergoing liver autotransplantation.