异氟烷预处理联合浅低温对大鼠离体心脏缺血再灌注损伤的影响

Effects of isoflurane preconditioning plus mild hypothermia on ischemia-reperfusion injury to isolated rat hearts

目的 评价异氟烷预处理联合浅低温对大鼠离体心脏缺血再灌注损伤的影响.方法 雄性SD大鼠50只,体重250～300 g,随机分为5组（n=10）：正常对照组（C组）、缺血再灌注组（I/R组）、异氟烷预处理组（P组）、浅低温组（M组）和异氟烷预处理联合浅低温组（PM组）.制备离体心脏Langendorff灌注模型,各组经主动脉灌注K-H液,平衡灌注20 min后,C组继续灌注K-H液120 min,其余各组制备缺血再灌注模型;I/R组和P组常温（37 ℃）缺血30 min时进行再灌注;M组和PM组浅低温（31 ℃）缺血30 min时进行再灌注.P组和PM组于缺血前用1.0%异氟烷预充饱和的K-H液灌注15 min,再用K-H液冲洗15 min.分别于平衡灌注20 min、缺血前即刻、再灌注30、60 min时记录左室舒张末期压、左心室收缩压、左室压最大上升速率、左室压最大下降速率和HR.再灌注60min时取心肌组织,测定心肌梗死体积、心肌细胞线粒体和胞浆的细胞色素C水平.电镜下观察心肌细胞线粒体形态.结果 与I/R组比较,P组、M组和PM组再灌注期间心功能改善,心肌梗死体积缩小,心肌细胞胞浆细胞色素C水平降低,心肌细胞线粒体细胞色素C水平升高（P＜0.05或0.01）;与P组比较,PM组再灌注期间心功能改善,心肌梗死体积缩小,心肌细胞胞浆细胞色素C水平降低,心肌细胞线粒体胞色素C水平升高（P＜0.05）;与M组比较,PM组心肌细胞胞浆细胞色素C水平降低,心肌细胞线粒体细胞色素C水平升高（P＜0.05或0.01）.电镜下PM组线粒体损伤轻于I/R组、P组和M组.结论 异氟烷预处理联合浅低温减轻大鼠心肌缺血再灌注损伤的效应强于异氟烷预处理或浅低温,其机制与减少心肌细胞线粒体细胞色素C的扩散,保护心肌细胞有关.

Objective To investigate the protective effects of isoflurane preconditioning plus mild hypothermia on isolated rat hearts against ischemia-reperfusion （I/R） injury. Methods Fifty male SD rats weighing 250-300 g were randomly divided into 5 groups （ n = 10 each）： group Ⅰ control （group C）; group Ⅱ I/R; group Ⅲ isoflurane preconditioning （group P）; group Ⅳ mild hypothermia （group M） and group Ⅴ isoflurane preconditioning ＋ mild hypothermia （group PM）. The animals were anesthetized with intraperitoneal pentobarbital 40 mg/kg. The hearts were immediately removed and perfused with Krebs-Hensleit （K-H） solution aerated with 95% O2 and 5% CO2 at 10 kPa via aorta at 37℃ in a Langendorff apparatus. The hearts were made globally ischemic for 30 min followed by 60 min reperfusion in group Ⅱ-Ⅴ. In group P and PM the hearts were perfused with K-H solution saturated with 1.0% isoflurane for 15 min followed by 15 min washout before ischemia.In group M and PM the hearts were made ischemic at 31 ℃ and perfused at 37℃. LVEDP, LVSP, dp/dtmax,dp/dtmm and HR were measured after equilibration （baseline）, immediately before ischemia, and at 30 and 60 min reperfusion. The infarct size and cytochrome C level in cytoplasm and mitochondria of myocytes were measured.Motochondrial ultrastructure was examined using electron microscope. Results Cardiac function was significantly better, the infarct size significantly smaller, the cytochrome C level in cytoplasm significantly lower, while the cytochrome C level in mitochondria of myocytes significantly higher in group Ⅲ-Ⅴ than in group Ⅱ and in group Ⅴ than in group Ⅲ. The cytochrome C level in cytoplasm was significantly lower, while the cytochrome C level in mitochondria of myocytes significantly higher in group Ⅴ than in group Ⅳ. Less damage to mitochondria was observed in group PM than in group I/R, P and M. Conclusion Isoflurane preconditioning combined with mild hypothermia provides better protection against myocardial I/R injury by attenuating the release of cytochrome C from mitochrondria.