摘要
目的 评价磷酸肌醇3激酶/蛋白激酶B/糖原合成酶激酶3β（PI3K/Akt/GSK-3β）信号通路在二氮嗪后处理减轻大鼠心肌缺血再灌注损伤中的作用.方法 取清洁级SD成年大鼠,制备离体灌注心脏30个,采用随机数字表法分为5组（n=6）：正常对照组（C组）、缺血再灌注组（I/R组）、二氮嗪后处理组（DZ组）、PI3K抑制剂LY294002组（LY组）和二氮嗪后处理＋LY294002组（DZ＋ LY组）.C组采用K-H液平衡灌注70 min;I/R组灌注4℃ST-Thomas停跳液10 ml/kg,继之停止灌注泵造成全心缺血,40 min后再次灌注K-H液30 min;DZ组于再灌注开始即刻经主动脉逆行灌注50 μmol/L二氮嗪5 min; LY组于再灌注开始即刻经主动逆行脉灌注15μmol/L LY294002 5 min;DZ＋ LY组于再灌注开始即刻经主动脉逆行灌注15 μmol/L LY294002 5 min,随后逆行灌注50 μmol/L二氮嗪5 min.于平衡灌注20 min时（T1）和再灌注30 min时（T2）记录HR、冠状动脉流量（CF）、左心室发展压（LVDP）、左室舒张末压（LVEDP）和压力瞬时最大变化率（±dp/dtmax）.采用Western blot法测定心肌组织总Akt（t-Akt）和总GSK-3β（t-GSK-3β）表达水平及其磷酸化水平.结果 与C组比较,其余4组T2时HR、LVDP和±dp/dtmax降低,LVEDP升高,I/R组、LY组和DZ＋ LY组CF降低,DZ组心肌组织Akt和GSK-3β磷酸化水平升高（P＜0.01）;与I/R组比较,DZ组T2时HR、CF、LVDP和±dp/dtmax升高,LVEDP降低,心肌组织Akt和GSK-3β磷酸化水平升高（P＜0.01）,LY组和DZ＋ LY组心肌组织Akt和GSK-3β磷酸化水平差异无统计学意义（P＞0.05）;与DZ组比较,LY组和DZ＋ LY组HR、CF、LVDP和±dp/dtmax降低,LVEDP升高,心肌组织Akt和GSK-3β磷酸化水平降低（P＜0.01）.结论 PI3K/Akt/GSK-3β信号通路参与了二氮嗪后处理减轻大鼠心肌缺血再灌注损伤.
Objective To evaluate the role of phosphoinositide 3 kinase/protein kinase B/glycogen synthase kinase 3β （PI3K/Akt/GSK-3β） signaling pathway in mitigation of ischemia-reperfusion （I/R） injury by diazoxide postconditioning in isolated rat hearts.Methods Pathogen-free Sprague-Dawley rats were used in the study.Thirty hearts were excised and passively perfused in a Langendorff apparatus with oxygenated K-H solution at 37 ℃.The hearts were randomly divided into 5 groups （n =6 each） using a random number table：control group （group C）,I/R group,diazoxide postconditioning group （group DZ）,PI3K inhibitor LY294002 group （group LY）,and diazoxide postconditioning ＋ LY294002 group （group DZ ＋ LY）.In group C,the hearts were continuously perfused with K-H solution for 70 min.In group I/R,the hearts were perfused with cardioplegic solution 4 ℃ ST-Thomas 10 ml/kg,the perfusion pump was then stopped to induce global ischemia,and 40 min later the hearts were perfused with K-H solution for 30 min.In DZ group,5 min of retrograde perfusion with diazoxide 50μmol/L was performed through the aorta starting from the onset of reperfusion.In LY group,5 min of retrograde perfusion with LY294002 15 μnol/L was performed through the aorta starting from the onset of reperfusion.In LY ＋ DZ group,5 min of retrograde perfusion with LY294002 15 μmol/L was performed through the aorta starting from the onset of reperfusion,followed by 5 min of retrograde perfusion with diazoxide 50 μmol/L.At 20 min of stabilization （T1） and 30 min of reperfusion （T2）,heart rate （HR）,coronary flow （CF）,left ventricular developed pressure （LVDP）,left ventricular developed pressure （LVEDP） and ± dp/dtmax were measured.The expression of total Akt （t-Akt） and total GSK-3β （t-GSK-3β） and phosphorylation of Akt and GSK-3β in myocardial tissues were determined by Western blot.Results Compared with C group,HR,LVDP and ± dp/dtmax were significantly decreased,and LVEDP was increased at T2 in the other four groups,CF was decreased in I/R,LY and DZ ＋ LY groups,and the phosphorylation of Akt and GSK-3β in myocardial tissues was increased in DZ group.Compared with I/R group,HR,CF,LVDP and ± dp/dtmax were significantly increased,and LVEDP was decreased at T2,and the phosphorylation of Akt and GSK-3β in myocardial tissues was increased in DZ group,and no significant changes were found in the phosphorylation of Akt and GSK-3 β in LY and DZ ＋ LY groups.Compared with DZ group,HR,CF,LVDP and ± dp/dtmax were significantly decreased,LVEDP was increased,and the phosphorylation of Akt and GSK-3β in myocardial tissues was decreased in LY and DZ ＋ LY groups.Conclusion PI3K/Akt/GSK-3β signaling pathway is involved in the mechanism by which diazoxide postconditioning mitigates I/R injury in isolated rat hearts.
出处
《中华麻醉学杂志》
CAS
CSCD
北大核心
2014年第10期1237-1240,共4页
Chinese Journal of Anesthesiology
基金
国家自然科学基金（30740044）
