精胺预处理对离体灌流大鼠心肌缺血/再灌注损伤及细胞凋亡的影响

目的探讨精胺预处理对离体灌流大鼠心肌缺血/再灌注损伤的影响及其可能的抗凋亡作用。方法应用Langehdorff离体灌流大鼠心脏复制模拟心肌缺血/再灌注损伤模型。24只大鼠随机分为3组,每组8只:对照组(Con-trol)、缺血/再灌注组(IR)、精胺干预组(Spermine)。比色法测定冠脉流出液中乳酸脱氢酶(LDH)活力,心肌组织中超氧化物歧化酶(SOD)活性及心肌丙二醛(MDA)含量;多导生理记录系统记录心脏功能;HE染色光镜下观察心肌形态学变化;三苯基氯化四氮唑(TTC)染色检测心肌梗死面积;透射电镜和TUNEL方法检测细胞凋亡;免疫组化检测心肌Fas与Bcl-2蛋白的表达。结果 (1)与对照组比,IR组冠脉LDH漏出明显增多、心肌组织中MDA水平增加、SOD活性降低(P<0.01);心功能明显下降(LVDP,HR,CF均低于对照组,P<0.05);光镜下可见心肌细胞呈凝固性或带状坏死。与Control组比,IR组心肌梗死面积及心肌细胞凋亡率明显增加(P<0.01);心肌Fas蛋白表达上调,Bcl-2蛋白表达下调。透射电镜下心肌细胞核染色质浓缩、凝聚且边集,染色质在核膜周边聚集形成新月形,线粒体嵴排列紊乱。(2)与IR组比,Spermine组冠脉LDH漏出减少,心肌组织中MDA减少,SOD增加(P<0.01);心功能有明显改善(LVDP,HR,CF均明显高于IR组,P<0.05);光镜下心肌细胞结构清晰。Spermine组与IR组比心肌梗死面积及心肌细胞凋亡率均明显降低(P<0.01);心肌Fas蛋白表达下调,Bcl-2蛋白表达上调;电镜下可见心肌肌节结构清晰,线粒体嵴完整、基质致密,未见核染色质凝聚。结论外源性精胺能明显减轻离体灌流大鼠心肌缺血/再灌注损伤,其机制可能与精胺抑制细胞凋亡有关。

Effect of spermine preconditioning on myocardial ischemia/reperfusion injury and cardiomyocyte apoptosis in isolated perfused rat heart

Aim To explore the effect of spermine preconditioning on myocardial ischemia/reperfusion injury and its potential antiapoptotic action in isolated perfused rat heart.Methods The model of myocardial ischemia-reperfusion injury was replicated with Langehdorff isolated perfused rat heart.24 rats were randomly divided into 3 groups: control,IR and spermine,with 8 rats in each group.LDH activity in coronary effluent,SOD activity and MDA content in myocardium were determined with colorimetry.Cardiac performances were monitored on multichannel physiologic recorder.Morphological changes of myocardium were observed by light microscope with HE staining.Myocardial infarction size was measured with TTC staining.Cell apoptosis was detected with transmission electron microscope and TUNEL assay.Fas and Bcl-2 protein expressions were tested with immunohistochemistry.Results(1) Compared with the control group,in IR group,LDH activity in coronary effluent increased distinctly,MDA in myocardial tissues rose,SOD activity decreased(P < 0.01),and heart function declined obviously(LVDP,HR,CF were all lower than those in the control group,P < 0.05).Myocardial cells were found to show coagulated zonal necrosis under light microscope.In comparison to the control,in IR group,myocardial infarction size and myocardial cells apoptotic rate increased evidently(P < 0.05).Fas protein expression was up-regulated while Bcl-2 expression downregulated.Under transmission electron microscope,myocardium cells displayed chromatin condensation and aggregation at the periphery of the nucleons,with chromatins aggregating like a crescent around nuclear membrane and mitochondria crista disorder.(2) Compared with IR group,in spermine group,LDH activity decreased,MDA content declined,while SOD activity increased(P < 0.01),heart function improved greatly(LVDP,HR and CF were all higher than those in IR group,P < 0.05).Cardiac myocytes showed clear structure under light microscope.In contrast to IR group,in spermine group,myocardial infarction size and cells apoptotic rate decreased evidently(P < 0.01).Fas protein expression was down-regulated while Bcl-2 expression up-regulated.Electron microscope showed clearly defined sarcomere in myocardium,complete mitochondria crista,dense substrate but no condensation of chromatin.Conclusion Exogenous spermine can greatly decrease the myocardial ischemia/reperfusion injury in isolated perfused rat heart and the mechanisms may be related with its antiapoptotic action.