PEP-1介导过氧化氢酶对大鼠离体心肌缺血再灌注损伤的保护作用

目的:研究细胞穿透肽PEP-1携带过氧化氢酶(CAT)穿透大鼠离体心肌的能力,并探讨PEP-1-CAT融合蛋白预处理对大鼠离体心肌缺血-再灌注损伤的保护作用及其机制。方法:建立大鼠离体心脏Langendorff灌流模型,随机分为5组(n=10):缺血-再灌注组(I/R组)行平衡30 min、停灌30 min与再灌注60 min,CAT组、PEP-1-CAT融合蛋白预处理组(A、B、C、D组)在平衡15 min后依次以10,5,10,20μmol/L的融合蛋白预处理15 min,停灌、复灌与I/R组相同。记录各组左室舒张末压(LVEDP)、左室收缩压(LVSP)、左室内压最大变化速率(±dp/dtmax)及冠脉流量(CF)变化,测定平衡15 min时、再灌注15 min时冠脉流出液乳酸脱氢(LDH)活性。测定再灌注60 min时心肌丙二醛(MDA)含量。结果:PEP-1-CAT融合蛋白能高效转导入离体心肌组织并呈现剂量依赖性。在平衡15 min末,各组LVEDP、LVSP、±dp/dt max以及LDH活性比较差异无统计学差异(P>0.05)。与I/R组比较,B、C、D组在再灌注后各时间点LVEDP降低(P<0.01),LVSP、±dp/dt max以及CF升高(P<0.01),灌注15 min时冠脉流出液LDH活性降低(P<0.01),再灌注后60 min心肌MDA含量降低(P<0.01)。CAT处理组所有结果和对照组相比无统计学意义(P>0.05)。结论:PEP-1-CAT融合蛋白能高效转导入离体心肌织并对大鼠离体心肌缺血-再灌注损伤具有明显的保护作用,将为其治疗心肌缺血再灌注损伤奠定坚实的实验基础。

Protective Effect of PEP-1-CAT Fusion Protein on the Ischemia-Reperfusion Injury in the Isolated Rat Heart

Objective: To investigate the transduction ability of PEP-1-CAT fusion protein into the isolated rat heart and the effects on ischemia-reperfusion injury in the isolated rat hearts.Methods: Male rat hearts were isolated and perfused with Langendorff apparatus.The isolated hearts were randomly divided into five groups(n=10) as an ischemia-reperfusion group(I/R),and a CAT group(group A) and three CAT+ PEP-1-CAT fusion protein pretreatment groups(B,C,and D),according to the perfusion with 10,5,10,and 20 μmol/L CAT or fusion protein for 15 min before ischemia-reperfusion respectively.Left ventricular systolic pressure(LVSP),left ventricular and diastolic pressure(LVEDP),LV dp/dt max and coronary flow(CF) were monitored continuously.Coronary effluent was collected during 15 min stabilization and during 15 min reperfusion for determination of LDH activity.At the end of 60 min reperfusion,the hearts were removed for determination of myocardial MDA content.Results: The PEP-1-CAT fusion protein could be transducted into the isolated rat hearts in a dose-dependent manner.At the end of 15 min stabilization,LVEDP,LVSP,dp/dt max,and LDH activity were not significantly changed in each group.During reperfusion,LVEDP,LVSP,and dp/dt max recovered significantly sooner in PEP-1-CAT fusion protein group than in I/R group.The myocardial MDA content,and LDH release were significantly lower in PEP-1-CAT fusion protein groups than in I/R group.Under fluorescent microscope,the number of apoptotic myocytes were decreased significantly in PEP-1-CAT groups.All the results of CAT group had no significant change compared with those of I/R group(P>0.05).Conclusion: The PEP-1-CAT fusion protein could efficiently penetrate myocardial tissue and attenuate the ischemia-reperfusion injury.It provides a more efficient strategy for the treatment of myocardial ischemia-reperfusion in future.