摘要
目的评价磷酸甘油酸变位酶5(PGAM5)在糖尿病大鼠心肌缺血再灌注损伤中的作用及其与线粒体质量的关系。方法SPF级健康雄性SD大鼠,6~8周龄,体重200~220 g,采用腹腔注射1%链脲佐菌素-柠檬酸盐缓冲液60 mg/kg的方法制备大鼠1型糖尿病模型。糖尿病造模成功后继续饲养8周,取1型糖尿病大鼠72只,采用随机数字表法分为4组(n=18):糖尿病假手术组(DS组)、糖尿病心肌缺血再灌注组(DIR组)、糖尿病心肌缺血再灌注+AAV9-PGAM5 shRNA组(DIR+PGAM5 shRNA组)和糖尿病心肌缺血再灌注+AAV9-GFP组(DIR+GFP组)。糖尿病建模后8周,采用结扎左冠状动脉前降支30 min后再灌注2 h的方法制备大鼠心肌缺血再灌注损伤模型。DIR+PGAM5 shRNA组和DIR+GFP组缺血前3周尾静脉缓慢注射AAV9-PGAM5 shRNA和AAV9-GFP 2×10^(12)μg/kg。AAV9-PGAM5 shRNA组再灌注2 h时记录左心室收缩压(LVSP)和左心室收缩压上升和下降最大速率(±dp/d_(tmax)),随后采集右颈动脉血样,采用ELISA法检测血清cTnI、CK-MB和LDH水平,随后处死大鼠取心脏,双重染色法确定心肌梗死面积,HE染色法观察心肌组织病理学结果,Western blot法检测PGAM5、自噬相关蛋白LC3B、p62、动力相关蛋白1(Drp1)及线粒体自噬受体蛋白FUNDC1的表达。结果与DS组比较,DIR组和DIR+GFP组LVSP和±dp/d_(tmax)降低,血清cTnI、CK-MB和LDH水平升高,心肌梗死面积增加,PGAM5、LC3B、Drp1和FUNDC1表达上调,p62表达下调(P<0.05);与DIR组比较,DIR+PGAM5 shRNA组LVSP和±dp/d_(tmax)升高,血清cTnI、CK-MB和LDH水平降低,心肌梗死面积减小,PGAM5、LC3B、Drp1和FUNDC1表达下调,p62表达上调(P<0.05),DIR+GFP组上述指标差异无统计学意义(P>0.05)。结论PGAM5参与了糖尿病大鼠心肌缺血再灌注损伤的过程,与降低线粒体质量有关。
Objective To evaluate the role of phosphoglycerate mutase 5(phosphoglycerate mutase family member 5,PGAM5)in myocardial ischemia-reperfusion(I/R)injury in diabetic rats and the relationship with mitochondrial quality.Methods SPF healthy male Sprague-Dawley rats,aged 6-8 weeks,weighing 200-220 g,were used in this study.Type 1 diabetes mellitus was induced by 1%streptozotocin diluted in citrate buffer solution 60 mg/kg.The rats were continuously fed for 8 weeks after successful establishment of the model.Seventy-two rats with type 1 diabetes mellitus were divided into 4 groups(n=18 each)by a random number table method:diabetic sham operation group(DS group),diabetic myocardial I/R group(DIR group),diabetic myocardial I/R plus AAV9-PGAM5 shRNA group(DIR+PGAM5 shRNA group),and diabetic myocardial I/R plus AAV9-GFP group(DIR+GFP group).The myocardial I/R model was established by ligation of the left anterior descending coronary artery for 30 min followed by reperfusion for 2 h starting from 8 weeks after establishment of type 1 diabetes mellitus model.AAV9-PGAM5 shRNA and AAV9-GFP 2×10^(12)μg/kg were slowly injected via tail vein 3 weeks before ischemia.In group AAV9-PGAM5 shRNA,left ventricular systolic pressure(LVSP)and the maximum rate of increase or decrease in left ventricular systolic pressure(±dp/d_(tmax))were monitored and recorded at the end of reperfusion,and then blood samples from the the right carotid artery were collected for determination of serum troponinⅠ(cTnI),creatine kinase-MB(CK-MB)and lactate dehydrogenase(LDH)levels(by enzyme-linked immunosorbent assay).The animals were sacrificed and hearts were obtained for determination of myocardial infarct size(by Evans Blue plus TTC double staining method)and expression of PGAM5,autophagy-related proteins(LC3B,p62),dynamin-related protein 1(Drp1),and mitochondrial autophagy receptor protein(FUNDC1)(by Western blot)and for microscopic examination of pathological changes of myocardial tissues(by HE staining).Results Compared with group DS,the LVSP and±
作者
陶洁
熊永红
田浩
夏中元
Tao Jie;Xiong Yonghong;Tian Hao;Xia Zhongyuan(Department of Anesthesiology,Renmin Hospital of Wuhan University,Wuhan 430060,China)
出处
《中华麻醉学杂志》
CAS
CSCD
北大核心
2021年第11期1382-1386,共5页
Chinese Journal of Anesthesiology
基金
国家自然科学基金(81970722)。
关键词
磷酸甘油酸酯变位酶
糖尿病
心肌再灌注损伤
线粒体
Phosphoglycerate mutase
Diabetes mellitus
Myocardial reperfusion injury
Mitochondria
