细胞外信号调节蛋白激酶1/2信号通路介导丙酮酸乙酯抗大鼠离体心脏缺血再灌注损伤的研究

Role of extracellular regulated protein kinase 1/2 signaling in the protective effect of ethyl pyruvate against ischemia reperfusion injury in isolated rat heart

目的:研究细胞外信号调节蛋白激酶1/2(extracellular regulated protein kinase 1/2,ERK1/2)信号通路在丙酮酸乙酯(ethyl pyruvate,EP)抗大鼠离体心脏缺血再灌注损伤(ischemia reperfusion injury,IRI)中的作用。方法:采用离体大鼠心脏Langendoff逆行灌注模型,缺血45min,再灌注60min。实验分为3组(每组8只),分别为IRI组、EP(2mmol/L)+IRI组、EP(2mmol/L)+PD98059(ERK1/2阻断剂,20μmol/L)+IRI组。记录各组处理后血流动力学指标,包括:左心室发展压(LVDP)、左心室内压最大变化速率(+dp/dtmax)、心率(HR)、冠脉流量(CF);ELISA法检测冠脉流出液中乳酸脱氢酶(LDH)含量;TTC法测定各组心肌梗死面积(infarct size,MI);Western法检测ERK1/2通路相关分子表达(p-ERK1/2、total-ERK1/2、p-p70S6K、total-p70S6K)。结果:与IRI组相比,EP+IRI组心脏IRI后各项血流动力学指标(LVDP、+dp/dtmax、HR、CF)显著改善,其数值分别为(68.1.4±4.5)mmHg,(1 130±93)mmHg/s,(253.6±6.5)次/min和(9.37±0.65)ml/min(P<0.05),冠脉流出液中LDH显著减少到(55.8±4.3)%(P<0.05),心肌MI显著减少到(57.1±4.8)%(P<0.05),p-ERK1/2和p-p70S6K蛋白表达分别显著增加到(308.90±17.81)%和(375.04±20.60)%(P<0.05)。与EP+IRI组比较,PD98059处理可以显著逆转EP对离体心脏血流动力学(P<0.05)、LDH释放量(P<0.05)和心肌MI(P<0.05)的保护作用,同时可以有效下调p-ERK1/2和p-p70S6K蛋白表达(P<0.05)。结论:EP对大鼠离体心脏IRI损伤有明确的保护作用,该作用与激活ERK1/2通路有关。

Objective：To explore the role of extracellular regulated protein kinase 1/2（ERK1/2）signaling in the protective effect of ethyl pyruvate（EP）against ischemia reperfusion injury（IRI）in isolated rat heart.Method：The isolated rat hearts were isolated and treated with Langendroff perfusion.The rat heart injury were induced by ischemia for 45 min and followed by 60 min reperfusion.The hearts were divided into 3groups（n=8）：IRI,EP（2mmol/L）＋IRI,EP（2mmol/L）＋ PD98059（20μmol/L）＋IRI.Then,the haemodynamics indications were recorded before ischemia and after reperfusion,including left ventricular peak developing pressure（LVDP）,the maximum rate of pressure change in the ventricle（±dp/dt max）,heart rate（HR）and coronary flow（CF）.The lactic dehydrogenase（LDH）level in coronary flow was detected with special ELISA kit.The myocardial infarct size（MI）was detected by TTC method.The molecules of ERK1/2signaling（p-ERK1/2,total-ERK1/2,p-p70S6 K,total- p70S6K）were detected by Western Blot.Result：Compared with IRI group,EP treatment significantly improved the haemodynamics indications（LVDP,＋dp/dtmax,HR,CF）,the values are（68.1.4±4.5）mmHg,（1 130±93）mmHg/s,（253.6±6.5）beats/min and（9.37±0.65）ml/min,separately（P〈0.05）.EP treatment also decreased the LDH release to 55.8±4.3%（P〈0.05）,reduced the myocardial MI to57.1±4.8%（P〈0.05）and increased the p-ERK1/2and p-p70S6 Kexpression to 308.90±17.81% and 375.04±20.60%（P〈0.05）.Compared with the EP＋IRI group,PD98059 treatment reversed the protection effects of EP on the haemodynamics indicates（P〈0.05）,LDH release（P 0.05）and myocardial MI（P 0.05）.PD98059 also down-regulated the p-ERK1/2and p-p70S6 Kexpression induced by EP（P〈0.05）.Conclusion：EP treatment displays a protective effect against myocardial IRI in isolated rat heart,and this effect may be dependent on the activation of ERK1/2signaling.