HIF-1α在糖尿病大鼠心肌缺血再灌注诱发肾损伤中的作用:与SLC7A11的关系

Role of HIF-1αin renal injury induced by myocardial ischemia-reperfusion in diabetic rats:relationship with SLC7A11

目的:评价缺氧诱导因子-1α(HIF-1α)在糖尿病大鼠心肌缺血再灌注诱发肾损伤中的作用及其与溶质运载蛋白7家族成员11(SLC7A11)的关系。方法:SPF级健康雄性SD大鼠,4周龄,体重100~130 g,高脂高糖饲料自由饮食,每周测1次体重,体重达到240 g后腹腔注射1%链脲佐菌素(STZ)-柠檬酸盐缓冲液35 mg/kg诱导2型糖尿病模型,STZ注射后继续予以高脂高糖饲料饲养,1周后尾静脉取血检测血糖,3次随机血糖≥16.7 mmol/L表明2型糖尿病模型制备成功。造模成功后继续高脂高糖饲养6周,取2型糖尿病大鼠18只,采用随机数字表法分为3组(n=6):糖尿病假手术组(DS组)、糖尿病心肌缺血再灌注组(DIR组)和糖尿病心肌缺血再灌注+HIF-1α激动剂DMOG组(DIR+DMOG组)。取非糖尿病大鼠12只,采用随机数字表法分为2组(n=6):非糖尿病假手术组(NS组)和非糖尿病心肌缺血再灌注组(NIR组)。采用结扎左冠状动脉前降支30 min再灌注120 min的方法建立大鼠心肌缺血再灌注损伤模型。于再灌注120 min时分离右侧颈内动脉取血,采用ELISA法检测血清肌酐(Cr)、尿素氮(BUN)、中性粒细胞明胶酶相关脂质运载蛋白(NGAL)浓度;取部分肾组织,采用HE染色法观察肾脏组织病理学结果,进行肾小管损伤评分;Western blot法检测肾组织HIF-1α和SLC7A11的表达。结果:与NS组比较,DS组和NIR组血清Cr、BUN、NGAL浓度和肾小管损伤评分升高,DS组肾组织HIF-1α和SLC7A11表达下调,NIR组肾组织HIF-1α和SLC7A11表达上调(P<0.05);与DS组比较,DIR组血清Cr、BUN、NGAL浓度和肾小管损伤评分升高,肾组织HIF-1α和SLC7A11表达上调(P<0.05);与NIR组比较,DIR组血清Cr、BUN、NGAL浓度和肾小管损伤评分升高,肾组织HIF-1α和SLC7A11表达下调(P<0.05);与DIR组比较,DIR+DMOG组血清Cr、BUN、NGAL浓度和肾小管损伤评分降低,肾组织HIF-1α和SLC7A11表达上调(P<0.05)。结论:HIF-1α参与了心肌缺血再灌注诱发糖尿病大鼠肾损伤的过程,与调控SLC7A11表达有关。

Objective To evaluate the role of hypoxia-inducible factor-1α(HIF-1α)in the renal injury induced by myocardial ischemia-reperfusion(I/R)in diabetic rats and its relationship with solute carrier family7 member11(SLC7A11).Methods SPF-grade healthy male Sprague-Dawley rats,aged 4 weeks,weighing 100-130 g,were fed with high-fat and high-sucrose diet freely.The weight of the rats was measured once a week.After the weight of the animals reached 240 g,1%streptozotocin(STZ)-citrate buffer 35 mg/kg was injected intraperitoneally to induce type 2 diabetes mellitus.After injection of STZ,the animals were fed with high-fat and high-sucrose diet continuously.Blood samples were collected from the tail vein for determination of blood glucose concentrations 1 week later.When random blood glucose was≥16.7 mmol/L for 3 times,the model of type 2 diabetes mellitus was considered to be established successfully.After the model was established successfully,the animals were fed with high-fat and high-sucrose diet continuously for 6 weeks.Eighteen rats with type 2 diabetes mellitus were selected and divided into 3 groups(n=6 each)using a random number table method:diabetic sham operation group(group DS),diabetic myocardial I/R group(group DIR)and diabetic myocardial I/R+HIF-1αagonist DMOG group(DIR+DMOG group).Twelve non-diabetic rats were divided into 2 groups(n=6 each)using a random number table method:non-diabetic sham operation group(NS group)and non-diabetic myocardial I/R group(NIR group).The rat myocardial I/R injury model was established by ligating the anterior descending branch of the left coronary artery for 30 min followed by 120 min reperfusion in anesthetized rats.Blood samples were collected from the right internal carotid artery at 120 min of reperfusion for determination of the serum creatinine(Cr),urea nitrogen(BUN)and neutrophil gelatinase-associated lipocalin(NGAL)concentrations(by enzyme-linked immunosorbent assay).Renal tissues were obtained for examination of the pathological changes(by HE staining method)and for determination of the expression of HIF-1αand SLC7A11(by Western blot).The damage to the renal tubules was scored.Results Compared with group NS,the concentrations of serum Cr,BUN and NGAL and renal tubular damage score were significantly increased in group DS and group NIR,the expression of HIF-1αand SLC7A11 was down-regulated in group DS,and the expression of HIF-1αand SLC7A11 was up-regulated in group NIR(P<0.05).Compared with group DS,the concentrations of serum Cr,BUN and NGAL and renal tubular damage score were significantly increased,and the expression of HIF-1αand SLC7A11 was up-regulated in group DIR(P<0.05).Compared with group NIR,the concentrations of serum Cr,BUN and NGAL and renal tubular damage score were significantly increased,and the expression of HIF-1αand SLC7A11 was down-regulated in group DIR(P<0.05).Compared with group DIR,the concentrations of serum Cr,BUN and NGAL and renal tubular damage score were significantly decreased,and the expression of HIF-1αand SLC7A11 was up-regulated in group DIR+DMOG(P<0.05).Conclusion HIF-1αis involved in the renal injury induced by myocardial I/R,which is related to regulation of the expression of SLC7A11 in rats.