二氮嗪后处理对大鼠离体心脏缺血再灌注损伤的影响

Effects of diazoxide postconditioning on myocardial ischemia-reperfusion injury in isolated rat hearts

目的 评价二氮嗪后处理对大鼠离体心脏缺血再灌注损伤的影响.方法 雄性SD大鼠,体重250～300 g,成功建立Langendorff再灌注模型的64个心脏随机分为4组（n=16）：正常对照组（C组）、缺血再灌注组（I/R组）、二氮嗪后处理组（D组）和线粒体ATP敏感性钾通道阻断剂5-羟葵酸＋二氮嗪后处理组（5-HD＋D组）.采用K-H液平衡灌注20 min时,C组继续灌注K-H液70 min;I/R组、D组和5-HD＋D组进行心肌缺血40 min,I/R组缺血前灌注4 ℃ ST.Thomas停跳液10 ml/kg;D组再灌注5 min时灌注含50μmol/L二氮嗪的K-H液5 min,然后再灌注20 min;5-HD＋D组灌注二氮嗪前灌注含100 μmol/L 5-羟葵酸的K-H液5 min,再灌注20 min.分别于平衡灌注末与再灌注末时取8个心脏,记录心功能指标,然后提取线粒体,测定心肌细胞线粒体膜电位（MMP）、氧自由基（ROS）生成量和呼吸功能指标.结果 各组平衡灌注末时各指标差异无统计学意义（P＞0.05）.与C组比较,再灌注末时其余3组心功能和线粒体呼吸功能减退,MMP降低,ROS生成量增加（P＜0.05或0.01）;与I/R组和5-HD＋D组比较,D组心功能和线粒体呼吸功能改善,MMP升高,ROS水平降低（P＜0.01）.结论二氮嗪后处理可减轻大鼠心肌缺血再灌注损伤,其机制与开放线粒体ATP敏感性钾通道而改善线粒体功能有关.

Objective To investigate the effects of diazoxide postconditioning on myocardial ischemiareperfusion （I/R） injury in isolated rat hearts. Methods Male SD rats weighing 250-300 g were anesthetized with intraperitoneal pentobarbital 40 mg/kg. Their hearts were excised and perfused in a Langendorff apparatus with K-H solution saturated with 95% O2-5% CO2 at 37 ℃. Sixty-four isolated rat hearts were randomized into 4 groups after 20 min of equilibration （n = 16 each）： group control （group C）, group I/R, group diazoxide postconditioning （group D） and group mito-KATP channel blocker 5-hydroxydecanoate （5-HD） ＋ diazoxide postconditioning （group 5-HD ＋ D）. Group C received continuous perfusion for another 70 min. In group I/R, D and 5-HD ＋ D, the hearts underwent 40 min of iscbemia followed by 30 min of reperfusion. 4 ℃ ST. Thomas cardioplegic solution was administered prior to ischemia in group I/R. Group D received 5 min of perfusion with K-H solution containing 50μ mol/L diazoxide at 5 min of reperfusion. Group 5-HD ＋ D received 5 min of perfusion with K-H solution containing 50 μmol/L 5-HD before reperfusion with diazoxide. Eight hearts were taken at the end of equilibration and reperfusion and the indexes of cardiac function were recorded. Then the mitochondria were extracted for determination of mitochondrial membrane potential （MMP）, reactive oxygen species （ROS） release, and respiratory function indexes. Results Compared with group C, MMP was significantly decreased, ROS release was significantly increased, mitochondrial respiratory function and cardiac function declined at the end of reperfusion in the other three groups （ P 〈 0.05 or 0.01 ）. MMP was significantly increased, ROS release was significantly decreased, mitochondrial respiratory function and cardiac function were improved in group D compared with group I/R and 5-HD ＋ D.Conclusion Diazoxide postconditioning can reduce myocardial I/R injury in rats via the opening of mito-KATPchannels and improving the mitochondrial function.