线粒体ATP敏感性钾通道在七氟醚预处理对大鼠离体缺血再灌注心脏延迟性保护中的作用

Role of mitochondrial ATP-sensitive potassium channel in sevoflurnne preconditioning-induced delayed cardioprotection against ischemia-reperfusion injury in isolated rat hearts

目的评价线粒体ATP敏感性钾通道（mito—KATP通道）在七氟醚预处理对大鼠离体缺血再灌注心脏延迟性保护中的作用。方法健康成年雄性SD大鼠72只，体重270～350g，采用随机数字表法，将其随机分为6组（n=12）：对照组（CON组）、缺血再灌注组（I／R组）各吸入33％氧气1h；七氟醚对照组（SEVO组）、七氟醚预处理组（SWOP组）各吸入2．5％七氟醚1h；5-羟葵酸＋七氟醚预处理组（5-HD＋SWOP组）于七氟醚预处理前30min腹腔注射mito-KATP阻断剂5-羟葵酸10mg／kg；5-羟葵酸对照组（5-HD组）腹腔注射5-羟葵酸10mg／kg，24h后除CON组和SEVO组，其余组建立大鼠Langendorff离体心脏灌注模型，停止灌注30min，再灌注120min。于缺血前即刻（T0）和再灌注120min（T1）时采用Western blot法检测心肌组织磷酸化的ε型蛋白激酶C（p-PKC-ε）、caspase-8蛋白的表达水平；于再灌注120min时采用TFC法测定心肌梗死范围，计算心肌梗死体积百分比。结果与CON组比较，I／R组、SWOP组、5-HD＋SWOP组和5-HD组T1时心肌梗死体积百分比升高，SEVO组和SWOP组T0时，I／R组、SWOP组、5-HD＋SWOP组和5-HD组T1时心肌p-PKC-ε蛋白表达上调，SEVO组L时caspase-8蛋白表达下调（P〈0．05）；与I／R组比较，SWOP组和5-HD＋SWOP组T1时心肌梗死体积百分比降低，SEVO组和SWOP组T0时p-PKC-ε蛋白表达上调，SWOP组T1时caspase-8蛋白表达下调（P〈0．05）；与SWOP组比较，5-HD＋SWOP组心肌梗死体积百分比升高，T0时p-PKC-ε蛋白表达下调，T1时caspase-8蛋白表达上调（P〈0．05）。结论mito-KATP通道参与了七氟醚预处理减轻大鼠离体心脏缺血再灌注损伤的过程，可能与上调缺血前p-PKC-ε蛋白表达水平，从而抑制再灌注时细胞凋亡有关。

Objective To evaluate the role of the mitochondrial ATP-sensitive potassium （mito-KATP） channel in sevoflurane preconditioning-induced delayed cardioprotection against ischemia-reperfusion （I/R） injury in isolated rat hearts. Methods Seventy-two adult male Sprague-Dawley rats were randomly divided into 6 groups （n = 12 each）： control group （group CON）, I/R group, sevoflurane control group （group SEVO）, sevoflurane preconditioning group （group SWOP）, 5-hydroxydeconoate （5-HD） ＋ sevoflurane preconditioning group （group 5- HD＋ SWOP） and 5-HD control group （group 5-HD）. The rats were exposed to 33% pure oxygen for 1 h in groups CON and I/R. The rats were exposed to 2.5% sevoflurane for 1 h in groups SEVO and SWOP. 5-HD （amit-KATP channel inhibitor） 10 mg/kg was injected intraperitoneally 30 min before sevoflurane preconditioning in group 5-HD ＋ SWOP. 5-HD 10 mg/kg was injected intraperitoneally in group 5-HD. The hearts were immediately removed and perfused in a Langendorff apparatus. The hearts were made globally ischemic for 30 min followed by 120 min reperfusion in groups I/R, SWOP, 5-HD ＋ SWOP and 5-HD. The expression of phosphorylated protein kinase C-epsilon （p-PKC-ε） and caspase-8 was measured by Western blot immediately before ischemia （T0） and at 120 min of reperfusion （T1）. The myocardial infarct volume was measured by TI＇C staining. Results Compared with group CON, the myocardial infarct volume was significantly increased at TI in groups I/R, SWOP, 5-HD ＋ SWOP and 5-HD, p-PKC-ε expression was up-regulated at To in groups SEVO and SWOP and at T1 in groups I/R, SWOP, 5-HD ＋ SWOP and 5-HD, and caspase-8 expression was down-regulated at TI in group SEVO （P 〈 0.05）. Compared with group I/R, the myocardial infarct volume was significantly decreased at T1 in groups SWOP and 5-HD ＋ SWOP, p-PKC-ε expression was up-regulated at To in groups SEVO and SWOP, and caspase-8 expression was down-regulated at T1 in group SWOP （P 〈 0.05）. Compared with group SWOP, the myocardial infarct volume was significantly increased, p-PKC-ε expression was down-regulated at To, and caspae-8 expression was up-regulated at T1 in group 5-HD ＋ SWOP （ P 〈 0.05）. Cortelusion The mito-KATP channel is involved in sevoflurane preconditioning-induced delayed cardioprotection against I/R injury in isolated rat hearts through upregulation of p-PKC-ε expression before ischemia and inhibition of cell apoptosis during reperfusion.