大鼠心肌缺血/再灌注损伤心肌细胞凋亡与Fas、FasL基因表达的变化

Changes of apoptosis and expression of Fas and FasL genes in rats with experimental myocardial ischemia/reperfusion injury

目的 探讨大鼠实验性心肌缺血再灌注时心肌细胞凋亡与Fas及Fas蛋白配体 (FasLigand ,FasL)基因表达的变化及与心肌组织损伤的关系。方法 以穿线结扎或松扎左冠状动脉制备大鼠心肌缺血再灌注模型。 6 4只大鼠随机分成假手术组 (假手术 2 4h)、缺血再灌注Ⅰ组 (缺血30min、再灌注 2 4h)、缺血再灌注Ⅱ组 (缺血 30min、再灌注 72h)及缺血再灌注Ⅲ组 (缺血 3h、再灌注2 4h)。以缺口末端标记法检测心肌细胞凋亡的变化 ,S P免疫组化法分别检测Fas与FasL蛋白水平变化 ,采用逆转录聚合酶链反应法检测Fas基因mRNA的表达改变 ,并分析心肌组织病理学损伤程度。结果 心肌缺血再灌注后心肌细胞凋亡指数及Fas蛋白阳性染色指数与炎性细胞FasL蛋白阳性染色指数均增加 ,且均随缺血或再灌注时间延长而进一步增高 ;Fas基因的mRNA表达也上调 ,但以再灌注2 4h时达高峰 ;心肌缺血再灌注后心肌组织呈大小不一的灶性坏死 ,坏死周围有炎性细胞浸润。结论心肌缺血再灌注时心肌细胞凋亡、Fas基因的蛋白与mRNA表达水平及炎性细胞的FasL蛋白表达量均增加 ,心肌细胞凋亡与Fas/FasL系统参与了心肌缺血再灌注损伤过程。

Objective To investigate the changes of apoptosis and the expression of Fas and FasL genes in rats myocardium with myocardial ischemia/reperfusion(MIR)and its relation with myocardium injury. Methods Rat models with MIR were established by occluding left coronary artery(LCA)and followed by release of it with ligation.Sixty four rats were divided randomly into sham group(sham operated for 24h),MIR group Ⅰ(30min of ischemia followed by 24h of reperfusion),MIR group Ⅱ(30min ischemia followed by 72h of reperfusion)and MIR group Ⅲ(3h of ischemia followed by 24h of reperfusion).Terminal deoxynucleotidyl transferase-mediated dUTP-fluorescein nick end labeling(TUNEL)and S-P immunohistochemical staining were used respectively to detect the changes of apoptosis and protein expression of Fas/FasL genes, and mRNA expression of Fas gene was evaluated by RT-PCR analysis. Histopathological changes in myocardium was also observed. Results Apoptotic index(AI) and possitive index(PI) of Fas protein in myocytes and PI of FasL protein in inflammatory cells were increased in myocardium with MIR,and gradually upregulated with duration of ischemia or reperfusion.mRNA induction of Fas gene was increased following MIR and peaked at 24h of reperfusion.Local myocyte necrosis and inflammatory cell infiltration around infarcted area were observed in myocardium with MIR. Conclusions The increase of mentioned parameters suggests that apoptosis and Fas/FasL system are involved in the process of MIR injury.