ERK1/2介导的p70S6K信号通路在七氟醚后处理离体大鼠缺血-再灌注损伤中的作用

Role of p70S6K signaling pathway induced by ERK1/2 in sevoflurane postconditioning protecting isolated rats against ischemia-reperfusion injury

目的 评价七氟醚后处理对离体大鼠缺血-再灌注损伤时心肌细胞胀亡的影响,探讨ERK1/2介导的p70S6K信号通路在其中的作用。方法 取Langendorff离体灌注模型成功的大鼠心脏,随机均分为6组：假手术组（S组）、缺血-再灌注组（IR组）、七氟醚后处理组（SP组）、PD98059溶剂二甲基亚砜（DMSO）组（DM组）、ERK1/2抑制剂PD98059组（PD组）、七氟醚后处理组＋PD98059（PP组）。除S组外,其它各组均缺血30min后再灌注2h。SP组、DM组和PD组分别于缺血末至再灌注15min灌注经3.0%七氟醚、DMSO（〈0.2%）和PD98059（20μmol/L）饱和的K-H液,PP组于缺血末至再灌注15min灌注经3.0%七氟醚和PD98059（20μmol/L）饱和的K-H液,随即更换为正常K-H液再灌注105min。记录缺血前即刻（T0）、再灌注30min（T1）、60min（T2）、90min（T3）、2h（T4）时的HR、左室收缩压（LVSP）和左室舒张末压（LVEDP）;再灌注末测定心肌梗死范围;采用Western blot法检测p-ERK1/2/t-ERK1/2、p-p70S6K/t-p70S6K、Porimin和Calpain-I蛋白表达水平。结果 与T0时和S组比较,T2-T4时IR、SP、DM、PD、PP组HR明显减慢,LVSP明显降低,LVEDP明显升高（P〈0.05）。与IR组比较,T1-T4时SP组LVSP明显升高,LVEDP明显降低,心肌梗死范围明显减少（P〈0.05）。与S组比较,IR、SP和DM组p-ERK1/2/t-ERK1/2、pp70S6K/t-p70S6K、Porimin和Calpain-I表达均明显上调（P〈0.05）,PD、PP组p-p70S6K/t-p70S6K、Porimin和Calpain-I表达均明显上调（P〈0.05）。与IR组比较,SP组p-ERK1/2/t-ERK1/2和pp70S6K/t-p70S6K表达明显上调,Porimin和Calpain-I表达明显下调（P〈0.05）。结论 七氟醚后处理对离体大鼠心肌缺血-再灌注损伤具有保护作用,可能与其激活ERK1/2介导的p70S6K表达,下调胀亡蛋白Porimin和Calpain-I的表达,进而减少心肌细胞胀亡有关。

Objective To evaluate the effects of sevoflurane postconditioning on myocardial ischemia-reperfusion injury induced cardiomyocytes oncosis in vitro rats,and to investigate the role of ERK1/2mediated p70S6 Ksignaling pathway in it.Methods Langendorff isolated perfused rat hearts were randomly divided into 6groups（n=12）：sham operation（group S）,ischemia-reperfusion group（group IR）,sevoflurane postconditioning group（group SP）,the PD98059 vehicle DMSO group（group DM）,ERK inhibitor PD98059group（group PD）,PD98059＋sevoflurane postconditioning group（group PP）.With the exception of group S,each group was subjected to occlusion for 30 min followed by reperfusion for 2h.In groups SP,DM and PD,the hearts were perfused with K-H solution saturated with 3.0% sevoflurane,DMSO（ 0.2%）and PD98059（20μmol/L）,respectively,for 15 min starting from the end of ischemia until 15 min of reperfusion,and then with plain K-H solution for 105 min.In group PP,the hearts were perfused with K-H solution saturated with 3.0%sevoflurane and PD98059 for 15min starting from the end of ischemia until 15 min of reperfusion.Recorded before ischemia（T0）and again 30 min of reperfusion（T1）,60min（T2）,90min（T3）,2h（T4）of HR,left ventricular systolic pressure（LVSP）and left ventricular end-diastolic pressure（LVEDP）.Myocardial infarct size percentage was measured at the end of reperfusion,the expressions of proteins（p-ERK1/2/t-ERK1/2,p-p70S6K/t-p70S6 K,Porimin and Calpain-I）were measured by Western blot at the end of reperfusion.Results Compared with T0 and group S,HR and LVSP were significantly slower and LVEDP was significantly higher at T2-T4,in groups IR,SP,DM,PD,PP（P〈0.05）.Compared with group IR,the LVSP of group SP was significantly higher,LVEDP were significantly slower at T1-T4,and the range of myocardial infarction was decreased significantly（P〈0.05）.Compared with group S,p-ERK1/2/t-ERK1/2,p-p70S6K/t-p70S6 K,Porimin and Calpain-I expression were significantly increased in groups IR,SP and DM（P〈0.05）.And pp70S6K/t-p70S6 K,Porimin and Calpain-I expression were significantly increased groups PD,PP（P〈0.05）.Compared with group IR,p-ERK1/2/t-ERK1/2and p-p70S6K/t-p70S6 Kexpression was significantly up-regulated,and Calpain-I Porimin expression was significantly reduced in group SP（P〈0.05）.Conclusion Sevoflurane postconditioning protects isolated rat hearts against myocardial ischemia-reperfusion injury,which may be related to activating the expression of ERK1/2 mediated p70S6 K,inhibiting the up-regulation of oncosis protein Porimin and Calpain-I,and thus decreasing cardiomyocytes oncosis.