大黄素对肠缺血再灌注小鼠肝损伤的影响及HO-1介导的自噬在其中的作用
Effect of emodin on liver injury in a mouse model of intestinal ischemia-reperfusion and role of heme oxygenase-1-mediated autophagy
刘晚丽 1季拓 1陈萌 1徐五星 2刘印印 1何祥虎 1张宗泽1
作者信息
- 1. 武汉大学中南医院麻醉科,武汉 430071
- 2. 武汉大学中南医院嘉鱼医院麻醉科,咸宁 437299
- 折叠
摘要
目的 评价大黄素对肠缺血再灌注小鼠肝损伤的影响及血红素加氧酶-1(HO-1)介导的自噬在其中的作用.方法 SPF级健康雄性C57BL/6小鼠24只,6～8周龄,体质量18～22 g,采用随机数字表法分为4组(n=6):假手术组(Sham组)、肠缺血再灌注组(I/R组)、大黄素组(E组)和大黄素+HO-1抑制剂锡原卟啉Ⅸ组(ES组).采用夹闭肠系膜上动脉45 min再灌注120 min的方法制备小鼠肠缺血再灌注损伤模型.E组缺血前5 d每天灌胃给予大黄素40mg/kg;ES组缺血前5 d每天灌胃给予大黄素40 mg/kg,缺血前12 h尾静脉注射锡原卟啉Ⅸ 7.5 mg/kg.于再灌注120 min时采集眶静脉血样,采用比色法检测血清ALT和AST浓度,随后处死小鼠取肝组织,观察肝组织病理学变化,分别采用WST-1法和TBA法检测SOD活性和MDA含量,荧光定量PCR法测定IL-6 mRNA和TNF-α mRNA的表达,Western blot法检测HO-1、自噬蛋白-苄氯素1(Beclin1)和微管相关蛋白1轻链3(LC3)的表达,计算LC3-Ⅱ/Ⅰ比值.结果 与Sham组比较,I/R组肝组织SOD活性降低,MDA含量、血清ALT和AST浓度升高,IL-6 mRNA、TNF-α mRNA和HO-1表达上调,Beclin1表达下调,LC3-Ⅱ/Ⅰ比值降低(P＜0.05),出现明显肝组织病理学损伤;与I/R组比较,E组肝组织SOD活性升高,MDA含量、血清ALT和AST浓度降低,IL-6 mRNA和TNF-α mRNA表达下调,HO-1和Beclin1表达上调,LC3-Ⅱ/Ⅰ比值升高(P＜0.05),肝组织病理学损伤减轻;与E组比较,ES组肝组织SOD活性降低,MDA含量、血清ALT和AST浓度升高,IL-6 mRNA和TNF-α mRNA表达上调,HO-1和Beclin1表达下调,LC3-Ⅱ/Ⅰ比值降低(P＜0.05),肝组织病理学损伤加重.结论 大黄素可减轻小鼠肠缺血再灌注诱导的肝损伤,其机制可能与激活HO-1介导的自噬有关.
Abstract
Objective To evaluate the effect of emodin on liver injury in a mouse model of intesti-nal ischemia-reperfusion(I/R)and the role of heme oxygenase-1-mediated autophagy.Methods Twenty-four SPF-grade healthy male C57BL/6 mice,aged 6-8 weeks,weighing 18-22 g,were divided into 4 groups(n=6 each)using a random number table method:sham operation group(Sham group),I/R group,emodin group(E group)and emodin plus HO-1 inhibitor Zinc Protoporphyrin Ⅸ(ZnPP)group(ES group).The intestinal I/R injury model was established by clamping the superior mesenteric artery for 45 min followed by 120 min of reperfusion.Emodin 40 mg/kg dissolved in 5％methylcellulose sodium was given by gastric gavage once a day for 5 days before ischemia in E group.Emodin 40 mg/kg dissolved in 5％methylcellulose sodium was given by gastric gavage once a day for 5 days before intestinal I/R,and ZnPP 7.5 mg/kg was injected via the tail vein at 12 h before ischemia in ES group.Orbital venous blood samples were collected at the end of reperfusion for determination of serum alanine aminotransferase(ALT)and as-partate aminotransferase(AST)concentrations.Then the mice were sacrificed,and liver tissues were ob-tained for microscopic examination of the pathological changes(after HE staining)and for determination of the activity of superoxide dismutase(SOD),content of malondialdehyde(MDA),expression of interleukin-6(IL-6)and tumor necrosis factor-alpha(TNF-α)mRNA(by fluorescent quantitative polymerase chain reaction),the expression of HO-1,autophagy-related protein Beclin1 and microtubule-associated protein 1 light chain 3(LC3)(by Western blot).The LC3-Ⅱ/Ⅰ ratio was calculated.Results Compared with Sham group,the activity of SOD was significantly decreased,the content of MDA and serum ALT and AST con-centrations were increased,the expression of IL-6 and TNF-α mRNA and HO-1 was up-regulated,the ex-pression of Beclin1 was down-regulated,the LC3-Ⅱ/Ⅰ ratio was decreased(P＜0.05),and the pathological changes of liver tissues were found in I/R group.Compared with I/R group,the activity of SOD was signifi-cantly increased,the content of MDA and serum ALT and AST concentrations were decreased,the expres-sion of IL-6 and TNF-α mRNA was down-regulated,the expression of HO-1 and Beclin1 was up-regulated,the LC3-Ⅱ/Ⅰ ratio was increased(P＜0.05),and the pathological changes of liver tissues were significantly attenuated in E group(P＜0.05).Compared with E group,the activity of SOD was significantly decreased,the content of MDA and serum ALT and AST concentrations were increased,the expression of IL-6 and TNF-α mRNA was up-regulated,the expression of HO-1 and Beclin1 was down-regulated,the LC3-Ⅱ/Ⅰ ra-tio was decreased(P＜0.05),and the pathological changes of liver tissues were aggravated in ES group.Conclusions Emodin can alleviate liver injury induced by intestinal I/R in mice,and the mechanism may be related to the activation of HO-1-mediated autophagy.
关键词
大黄素/血红素加氧酶-1/自噬/肠/再灌注损伤/肝Key words
Emodin/Heme oxygenase-1/Autophagy/Intestines/Reperfusion injury/Liver引用本文复制引用
基金项目
国家自然科学基金(81000849)
武汉大学中南医院科技创新培育基金(znpy2018022)
出版年
2024
NETL
NSTL
万方数据