七氟醚预处理对大鼠心肌缺血再灌注时wnt/GSK-3β/β-catenin信号通路的影响：离体实验

Effects of sevoflurane preconditioning on wnt/GSK3-β/β-catenin signaling pathway during myocardial ischemia-reperfusion injury in rats in vitro

目的 评价七氟醚预处理对大鼠心肌缺血再灌注时wnt/糖原合成酶激酶-3β（GSK-3β）/β-连环蛋白（β-catenin）信号通路的影响.方法 成年健康雄性Wistar大鼠,体重220～ 280 g,建立Langendorff离体心脏灌注模型.取离体心脏模型36个,采用随机数字法,将其随机分为3组（n=12）：假手术组（S组）、心肌缺血再灌注组（I/R组）和七氟醚预处理组（SP组）.平衡灌注30 min后,S组继续灌注150 min;I/R组缺血30 min,再灌注120 min;SP组用含2.4％七氟醚的K-H液灌注15min,然后洗脱5 min,缺血30 min,再灌注120 min.分别于平衡灌注末和再灌注30 min时,记录HR、左室舒张末压（LVEDP）、左室发展压（LVDP）、左心室内压最大上升速率（＋dp/dtmax）和左心室内压最大下降速率（-dp/dtmax）.再灌注期间行心律失常评分.于再灌注60 min时,每组取3个心脏,采用Western blot法测定心肌组织wnt3a、磷酸化GSK-3β（p-GSK-3β）和β-catenin的表达水平.再灌注120min时,每组取6个心脏,采用TTC染色法测定心肌梗死体积.结果 与S组比较,I/R组再灌注30min时HR、LVDP、＋dp/dtmax和-dp/dtmax降低,LVEDP升高,心律失常评分升高,心肌梗死体积百分比增加,心肌组织wnt3a、p-GSK-3β和β-catenin的表达下调（P＜0.05）;与I/R组比较,SP组再灌注30min时HR、LVDP、＋dp/dtmax和-dp/dtmax升高,LVEDP降低,心律失常评分降低,心肌梗死体积百分比减小,心肌组织wnt3a、p-GSK-3β和β-catenin的表达上调（P＜0.05）.结论 七氟醚预处理可通过激活wnt/GSK-3β/β-catenin信号通路减轻大鼠心肌缺血再灌注损伤.

Objective To evaluate the effects of sevoflurane preconditioning on wnt/glycogen synthase kinase-3 beta （GSK-3β）/β-catenin signaling pathway during myocardial ischemia-reperfusion （I/R） injury in rats in vitro.Methods Ault male Wistar rats,weighing 220-280 g,were heparinized and anesthetized with intraperitoneal 3% pentobarbital 30 mg/kg.Their hearts were rapidly excised and perfused in a langendorff apparatus with oxygenated （95% O2-5% CO2） K-H solution at 37 ℃.After 15 min of equilibration,36 isolated hearts were randomly divided into 3 groups （n=12 each） using a random number table：sham operation group （group S）,group I/R and sevoflurane preconditioning group （group SP）.After 30 min of equilibration,the hearts were continuously perfused for 150 min in group S.The isolated hearts were subjected to 30 min of ischemia followed by 120 min of reperfusion.In SP group,the hearts were perfused for 15 min with K-H solution containing 2.4% sevoflurane,followed by 5 min washout before reperfusion.At the end of equilibration and 30 min of reperfusion,HR,left ventricular end-diastolic pressure （LVEDP）,left ventricular developed pressure （LVDP） and ＆#177; dp/dtmax were recorded.The severity of arrhythmias was assessed during reperfusion.At 60 min of reperfusion,3 hearts in each group were chosen for measurement of expression of wnt3a,phosphor-GSK-3β （p-GSK-3β） and β-catenin （by Western blot）.At 120 min of reperfusion,6 hearts in each group were chosen for determination of myocardial infarct size by TTC staining.Results Compared with group S,HR,LVDP,＋dp/dtmax and -dp/dtmax were significantly decreased,and LVEDP was increased at 30 min of reperfusion,arrhythmia scores and the percentage of myocardial infarct size were increased,and the expression of wnt3a,p-GSK-3β and β-catenin was down-regulated in I/R group.Compared with group I/R,HR,LVDP,＋dp/dtmax and-dp/dtmax were significantly increased,and LVEDP was decreased at 30 min of reperfusion,arrhythmia scores and the percentage of myocardial infarct size were decreased,and the expression of wnt3a,p-GSK-3β and β-catenin was up-regulated in group SP.Conclusion Sevoflurane preconditioning attenuates myocardial I/R injury by activating wnt/GSK-3β/β-catenin signaling pathway in isolated rat hearts.