摘要
目的 探讨高迁移率族蛋白B1（HMGB1）对心肌缺血/再灌注（I/R）损伤后内质网应激（ERS）的影响.方法 按随机数字表法将40只雄性SD大鼠分为假手术组、I/R模型组、基因沉默（HMGB1-siRNA）组和空载体（Scrambled-siRNA）组,每组10只.结扎冠状动脉（冠状）血流30 min、再灌注2 h建立心肌I/R损伤大鼠模型;假手术组不进行结扎.各组分别于术前24、12、0 h采用水压法经尾静脉相应注射磷酸盐缓冲液（PBS）、HMGB1-siRNA或Scrambled-siRNA混合液各1 mL进行预处理;再灌注2 h后取血并处死大鼠取心肌组织.采用酶联免疫吸附试验（ELISA）检测血清肿瘤坏死因子-α（TNF-α）、白细胞介素（IL-6、IL-8）水平;用免疫组化法检测心肌损伤部位HMGB1蛋白表达;用蛋白质免疫印迹试验（Western Blot）和实时荧光定量反转录-聚合酶链反应（RT-PCR）分别检测心肌组织HMGB1以及ERS相关因子,如糖调节蛋白78（GRP78）、增强子结合蛋白同源蛋白（CHOP）、天冬氨酸特异性半胱氨酸蛋白酶12（caspase-12）的蛋白和mRNA表达.结果 I/R可使大鼠血清炎性因子水平和HMGB1阳性细胞以及心肌组织GRP78、CHOP、caspase-12的蛋白和mRNA表达较假手术组明显增加.HMGB1-siRNA预处理后可致大鼠血清炎性因子水平较I/R模型组明显降低〔TNF-α（ng/L）：783.4±203.4比963.9±214.1,IL-6（ng/L）：358.8±94.8比452.3±103.7,IL-8（ng/L）：180.5±73.6比347.3±90.3,均P〈0.05〕,HMGB1阳性细胞以及心肌组织GRP78、CHOP、caspase-12的蛋白和mRNA表达也明显低于I/R模型组（HMGB1蛋白：1.59±0.26比3.21±0.40,GRP78蛋白：2.59±0.28比4.21±0.42,CHOP蛋白：2.01±0.23比3.21±0.43,caspase-12蛋白：1.48±0.22比3.01±0.48;HMGB1 mRNA：2.35±0.26比4.67±0.45,GRP78 mRNA：6.59±0.26比11.21±0.40,CHOP mRNA：2.01±0.43比5.21±0.63,caspase-12 mRNA：4.48±0.32比8.41±0.52,均P〈0.05）.Scrambled-siRNA组各指标与I/R模型组比较差异均无统计学意义.�
Objective To investigate the effect of high mobility group protein B1 （HMGB1） inhibition on endoplasmic reticulum stress （ERS） after myocardial ischemia/reperfusion （I/R） in rats.Methods Forty male Sprague-Dawley （SD） rats were randomly divided into four groups （n = 10）： sham operation group, I/R model group, Gene silencing （HMGB1-siRNA） group, and empty vector （Scrambled-siRNA） group. Coronary blood flow of the rats were ligated for 30 minutes, relaxed the ligament line for 2 hours, to establish I/R injury model; not ligation with the sham operation group. Each group was injected 1 mL phosphate buffer （PBS）, HMGB1-siRNA mixture or Scrambled-siRNA mixture preoperative by tail vein 0 hour, 12 hours, and 24 hours before surgery. After 2 hours reperfusion, the levels of tumor necrosis factor-α （TNF-α）, interleukins （IL-6, IL-8） in the serum were detected by enzyme linked immunosorbent assay （ELISA）; the expression of HMGB1 protein in myocardium was detected by immunohistochemistry; the protein and mRNA expressions of HMGB1, GRP78, CHOP and caspase-12 in myocardium were detected by Western Blot and real-time fluorescence quantitative reverse transcription-polymerase chain reaction （RT-PCR）.Results Compared with sham operation group, the levels of serum inflammatory factor, HMGB1 positive cells, and the protein and mRNA expressions of GRP78, CHOP, caspase-12 were significantly increased in I/R model group. The levels of serum inflammatory factor in HMGB1-siRNA group were significantly lower than those in the I/R model group [TNF-α （ng/L）： 783.4±203.4 vs. 963.9±214.1, IL-6 （ng/L）： 358.8±94.8 vs. 452.3±103.7, IL-8 （ng/L）： 180.5±73.6 vs. 347.3±90.3, allP 〈 0.05], HMGB1 positive cells, and the protein and mRNA expressions of GRP78, CHOP, caspase-12 in HMGB1-siRNA group were significantly lower than I/R model group （HMGB1 protein：1.59±0.26 vs. 3.21±0.40, GRP78 protein： 2.59±0.28 vs. 4.21±0.42, CHOP protein： 2.01±0.23 vs. 3.21±0.43,
作者
鄂璐莎
南景龙
E Lusha Nan Jinglong(Department of Cardiology, the Inner Mongolia Autonomous Region People＇s Hospital, Hohhot 010017, Inner Mongolia, China)
出处
《中华危重病急救医学》
CAS
CSCD
北大核心
2017年第10期916-920,共5页
Chinese Critical Care Medicine
基金
内蒙古自治区自然科学基金（2016MS0867）
关键词
高迁移率族蛋白B1
缺血/再灌注
心
炎性因子
内质网应激
High mobility group protein B1
Myocardial ischemia/reperfusion
Inflammatory factor
Endoplasmic retieulum stress
