人α1-抗胰蛋白酶对小鼠心肌缺血再灌注损伤的作用及相关机制  

Effects of Humanα1-antitrypsin on Myocardial Ischemia-reperfusion Injury in Mice and Related Mechanism

作　　者：吴成鑫 吴婷婷[1] 郑颖颖[1] 闫拓 马依彤[1] 谢翔[1] WU Chengxin;WU Tingting;ZHENG Yingying;YAN Tuo;MA Yitong;XIE Xiang(Department of Hypertension,The First Affiliated Hospital of Xinjiang Medical University,Urumqi 830011,China)

机构地区：[1]新疆医科大学第一附属医院高血压病科,乌鲁木齐830011

出　　处：《中国循环杂志》2023年第5期560-567,共8页Chinese Circulation Journal

基　　金：国家自然科学基金(82170345、80000238)。

摘　　要：目的:探讨人α1-抗胰蛋白酶(hAAT)对小鼠心肌缺血再灌注损伤(MIRI)的作用及相关机制。方法:采用随机数字表法将SPF级健康雄性C57BL/6野生型小鼠和hAAT转基因过表达小鼠分为四组:野生型假手术组(Sham组)、野生型缺血再灌注组(I/R组)、hAAT转基因假手术组(hAAT-Tg+Sham组)、hAAT转基因缺血再灌注组(hAAT-Tg+I/R组),每组18只。建立小鼠MIRI模型,假手术组只开胸不结扎。再灌注24 h后,每组随机取6只小鼠,通过伊文思蓝和2, 3, 5-氯化三苯基四氮唑(TTC)双染色法测定心肌梗死面积。再灌注7 d后,应用小动物超声系统测量各组小鼠心功能指标,苏木素-伊红(HE)染色观察心肌组织病理学改变,Masson染色观察心肌纤维化程度。采用酶联免疫吸附试验(ELISA)法检测各组小鼠血清标志物心肌肌钙蛋白I (cTnI)、肌酸激酶同工酶(CK-MB)及白细胞介素-1β(IL-1β)的浓度变化。蛋白免疫印迹法和免疫组化染色法检测各组心肌组织中核苷酸寡聚化结构域样受体蛋白3(NLRP3)、活化半胱氨酸蛋白酶-1(caspase-1)、IL-1β蛋白表达。结果:(1)与Sham组比较,I/R组和h AAT-Tg+I/R组心肌梗死面积均显著增加(P均<0.01),与I/R组比较,hAAT-Tg+I/R组心肌梗死面积显著减小(P<0.05)。(2)hAAT-Tg+I/R组小鼠左心室射血分数和左心室短轴缩短率均明显高于I/R组(P均<0.05),左心室舒张末期内径和左心室收缩末期内径均明显低于I/R组(P均<0.05)。(3)与Sham组比较,I/R组炎性细胞浸润明显、心肌纤维排列紊乱、纤维化程度较重;与I/R组比较,hAAT-Tg+I/R组炎性细胞浸润减少、心肌纤维排列相对整齐、纤维化程度较轻。(4)与Sham组比较,I/R组和hAAT-Tg+I/R组再灌注后24 h和7 d血清中cTnI、CK-MB及IL-1β水平均明显升高(P均<0.05);hAAT-Tg+I/R组血清cTnI、CK-MB及IL-1β水平均明显低于I/R组(P均<0.05)。(5)与Sham组比较,I/R组心肌组织中NLRP3、caspase-1、IL-1β蛋白水平均明显上调,而hAAT-Tg+I/R�Objectives:To investigate the effects of humanα1-antitrypsin(hAAT)on myocardial ischemia-reperfusion injury(MIRI)in mice and related mechanism.Methods:SPF grade healthy male C57BL/6 wild-type mice and hAAT transgenic overexpression mice were randomly divided into four groups(n=18 each):wild type sham operation(sham)group,wild type ischemia-reperfusion(I/R)group,hAAT transgenic sham operation(hAAT-Tg+sham)group,hAAT transgenic ischemia-reperfusion(hAAT-Tg+I/R)group.The myocardial ischemia-reperfusion model was established by ligating the left anterior descending coronary artery for 60 min followed by reperfusion.After 24 h of reperfusion,six mice in each group were randomly selected and the myocardial infarction area was measured by Evans blue and 2,3,5-triphenyl tetrazolium chloride double staining.After 7 d of reperfusion,cardiac function of mice in each group was measured by small animal ultrasound system,the pathological changes of myocardial tissue were observed by hematoxylin eosin staining,the degree of myocardial fibrosis was observed by Masson staining.Cardiac troponin I(cTnI),creatine kinase MB isoenzyme(CK-MB)and inflammatory factor interleukin-1β(IL-1β)were detected by enzyme-linked immunosorbent assay.Protein expression levels of nucleotide oligomerization domain like receptor protein 3(NLRP3),cysteine proteinase-1(caspase-1)and IL-1βin myocardial tissue of each group were detected by Western blot and immunohistochemical staining.Results:(1)Compared with I/R group,the myocardial infarction area in hAAT-Tg+I/R group decreased significantly(P<0.05).(2)The left ventricular ejection fraction and left ventricular short axis shortening were significantly higher,and the left ventricular end diastolic diameter and left ventricular end systolic diameter were significantly smaller in the hAATTg+I/R group than in the I/R group(all P<0.05).(3)Compared with sham group,I/R group had obvious inflammatory cell infiltration,disordered arrangement of myocardial fibers and severe fibrosis;compared with I/R group,hAA

关 键 词：Α1-抗胰蛋白酶 心肌缺血再灌注损伤 核苷酸寡聚化结构域样受体蛋白3 炎性小体 

分 类 号：R54[医药卫生—心血管疾病]