Rev-erbα/Bmal1信号通路在糖尿病大鼠心肌缺血再灌注损伤中的作用及其与自噬的关系  

Role of Rev-erbα/Bmal1 signaling pathway in myocardial ischemia-reperfusion injury and relationship with autophagy in diabetic rats

作　　者：邱珍[1] 张艺[1] 明浩 雷少青[1] 夏中元[1] Qiu Zhen;Zhang Yi;Ming Hao;Lei Shaoqing;Xia Zhongyuan(Department of Anesthesiology,Renmin Hospital of Wuhan University,Wuhan 430060,China)

机构地区：[1]武汉大学人民医院麻醉科,430060

出　　处：《中华麻醉学杂志》2021年第2期234-238,共5页Chinese Journal of Anesthesiology

基　　金：国家自然科学基金(81970722)。

摘　　要：目的评价Nr1D1核受体亚家族1,组D,成员1(Rev-erbα)/芳香烃受体核转位蛋白样1受体(Bmal1)信号通路在糖尿病大鼠心肌缺血再灌注损伤中的作用及其与自噬的关系。方法SPF级成年雄性SD大鼠,6~8周龄,体重200~220 g,腹腔注射链脲佐菌霉素60 mg/kg建立1型糖尿病模型,糖尿病造模成功后继续饲养8周。取糖尿病大鼠30只,采用随机数字表法分为3组:糖尿病假手术组(DS组,n=6)、糖尿病心肌缺血再灌注组(DI/R组,n=12)和糖尿病心肌缺血再灌注+Rev-erbα拮抗剂SR-8278组(DI/R+SR组,n=12)。另取非糖尿病大鼠18只,采用随机数字表法分为2组:非糖尿病假手术组(NS组,n=6)和非糖尿病心肌缺血再灌注组(NI/R组,n=12)。采用结扎左冠状动脉前降支30 min再灌注120 min的方法建立心肌缺血再灌注损伤模型。DI/R+SR组于缺血前1 h时经股静脉注射SR-82782 mg/kg。再灌注结束即刻采集颈动脉血标本,采用ELISA法检测血清cTnI、CK-MB和LDH水平;然后处死大鼠,取心肌组织,采用TTC法测定心肌梗死面积百分比,透射电镜下计数自噬小体,采用RT-PCR检测Rev-erbα和Bmal1的mRNA表达水平,Western blot法检测Rev-erbα、Bmal1、微管相关蛋白1轻链3(LC3)Ⅱ和LC3Ⅰ的表达水平,并计算LC3Ⅱ/LC3Ⅰ比值。结果与NS组比较,NI/R组心肌梗死体积百分比、血清cTnI、CK-MB和LDH水平和自噬小体计数升高,心肌组织Rev-erbα及其mRNA表达上调,Bmal1及其mRNA表达下调,LC3Ⅱ/LC3Ⅰ比值升高,DS组血清cTnI、CK-MB和LDH水平升高,自噬小体计数降低,心肌组织Rev-erbα及其mRNA表达上调,Bmal1及其mRNA表达下调,LC3Ⅱ/LC3Ⅰ比值降低(P<0.05);与NI/R组比较,DI/R组心肌梗死体积百分比、血清cTnI、CK-MB和LDH水平升高,自噬小体计数降低,心肌组织Rev-erbα及其mRNA表达上调,Bmal1及其mRNA表达下调,LC3Ⅱ/LC3Ⅰ比值降低(P<0.05);与DS组比较,DI/R组心肌梗死体积百分、血清cTnI、CK-MB和LDH水平和自噬小体计数升高,心肌组织RObjective To evaluate the role of nuclear receptor subfamily 1,group D,member 1/brain and muscle Arnt-like 1(Rev-erbα/Bmal1)signaling pathway in myocardial ischemia-reperfusion(I/R)injury in diabetic rats and its relationship with autophagy.Methods SPF-grade adult male Sprague-Dawley rats,aged 6-8 weeks,weighing 200-220 g,were used in this study.Type I diabetes mellitus was induced by intraperitoneal streptozotocin 60 mg/kg.The rats were continuously fed for 8 weeks after successful establishment of the model.Thirty rats with type 1 diabetes mellitus were divided into 3 groups by a random number table method:diabetic sham operation group(DS group,n=6),diabetic myocardial I/R group(DI/R group,n=12)and diabetic myocardial I/R plus Rev-erbαantagonist SR-8278 group(DI/R+SR group,n=12).Eighteen non-diabetic rats were divided into 2 groups by a random number table method:non-diabetic sham operation group(NS group,n=6)and non-diabetic myocardial I/R group(NI/R group,n=12).The myocardial I/R model was established by ligation of the left anterior descending coronary artery for 30 min followed by 120-min reperfusion in anesthetized rats.SR-82782 mg/kg was intravenously injected via the femoral vein at 1 h before ischemia in group DI/R+SR.Blood samples were collected from the carotid artery immediately after the end of reperfusion for determination of serum troponin I(cTnI),creatine kinase-MB(CK-MB)and lactic dehydrogenase(LDH)levels(by enzyme-linked immunosorbent assay).Then the rats were sacrificed,and myocardial tissues were obtained for determination of myocardial infarct size(by TTC method),expression of Rev-erbαand Bmal1 mRNA(by real-time polymerase chain reaction)and expression of Rev-erbα,Bmal1,microtubule-associated protein 1 light chain(LC3)Ⅱand LC3Ⅰ(by Western blot)and for calculation of the ratio of LC3Ⅱ/LC3Ⅰand the number of autophagosomes(with a transmission electron microscope).Results Compared with group NS,the percentage of myocardial infarct size,serum levels of cTnI,CK-MB and LDH and the numbe

关 键 词：核受体亚家族1 D组 成员1 ARNTL转录因子类 糖尿病 心肌再灌注损伤 自噬 

分 类 号：R587.1[医药卫生—内分泌]