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胰岛素保护缺血/再灌注心脏:PI3--K/Akt和JNKs信号通路间的交互作用

刘海涛, 张海锋, 司瑞, 张全江, 张昆茹, 郭文仪, 王海昌, 高峰

第四军医大学西京医院心血管内科.陕西,西安 710032；第四军医大学生理学教研室.陕西,西安 710032

摘要

作者前期研究表明胰岛素可激活细胞内信号转导机制如磷脂酰肌醇3--激酶--蛋白激酶B--内皮型一氧化氮合酶--一氧化氮(PI3--K--Akt--eNOS--NO)信号通路,减轻心肌缺血/再灌注(ischemia/reperfusion,I/R)损伤,改善缺血后心肌功能恢复。然而c--Jun氨基末端激酶(c--Jun NH_(2)--terminal kinase,JNK)信号通路在胰岛素保护I/R心肌中的作用尚不清楚,该研究旨在探讨JNK信号通路在胰岛素保护I/R心肌中的作用及其与PI3--K/Akt信号通路间的相互关系。离体Sprague--Dawley大鼠心脏缺血30min后施行2h或4h的再灌注,缺血前用LY294002(15mmol/L)和SP600125(10mmol/L)灌注15min,分别阻断PI3--K/Akt和磷酸化JNK(phosphorylated--JNK,p--JNK)活化,观测心脏功能、心肌梗死、细胞凋亡和蛋白磷酸化水平。与对照组相比,胰岛素再灌注2h后,心率、左心室发展压和左心室收缩/舒张最大速率均明显增加,梗死面积减少约16.1%[(28.9±2.0)%vs(45.0±4.0)%,n=6,{sl P}<0.01],细胞凋亡指数从(27.6±1.3)%减少到(16.0±0.7)%({sl n}=6,{sl P}<0.01),Akt的活性增加1.7倍({sl n}=6,{sl P}<0.05),同时JNK活性增加1.5倍({sl n}=6,{sl P}<0.05)。用LY294002处理后,胰岛素对I/R心肌的保护作用消失;而用SP600125处理可增强胰岛素的保护作用,且可部分逆转LY294002的抑制作用。进一步观察发现SP600125减弱了Akt的磷酸化({sl n}=6,{sl P}<0.05)。上述结果表明,在I/R心肌中,胰岛素可同时激活PI3--K/Akt及JNK信号通路,且通过后者进一步增加Akt活化,从而减轻I/R损伤,改善心肌功能。这种PI3--K/Akt与JNK信号通路交互机制对胰岛素保护I/R心肌有重要意义。

关键词： 缺血/再灌注损伤; 胰岛素; 凋亡; 蛋白激酶B; c-Jun氨基未端激酶; 交互机制

Insulin protects isolated hearts from ischemia/reperfusion injury: cross--talk between PI3--K/Akt and JNKs

Liu Haitao, Zhang Haifeng, Si Rui, Zhang Quanjiang, Zhang Kunru, Guo Wenyi, Wang Haichang, Gao Feng

Department of Cardiology, Xijing Hospital, the Fourth Military Medical University.Xi'an 710032,Shaanxi；China

Abstract

Our previous results have demonstrated that insulin reduces myocardial ischemia/reperfusion (MI/R) injury and increases the postischemic myocardial functions via activating the cellular survival signaling, i.e., phosphatidylinositol 3-kinase (PI3-K)-Akt-endothelial nitric oxide synthase (eNOS)-nitric oxide (NO) cascade. However, it remains largely controversial whether c-Jun NH_(2)-terminal kinase (JNK) is involved in the effects of insulin on MI/R injury. Therefore, the aims of the present study were to investigate the role of JNK, especially the cross-talk between JNK and previously expatiated Akt signaling, in the protective effect of insulin on I/R myocardium. Isolated hearts from adult Sprague-Dawley rats were subjected to 30 min of regional ischemia and followed by 2 or 4 h of reperfusion (n=6). The hearts were pretreated with PI3-K inhibitor LY294002, or phosphorylated-JNK inhibitor SP600125,respectively, then perfused retrogradely with insulin, and the mechanical functions of hearts, including the heart rate (HR), left ventricular developed pressure (LVDP) and instantaneous first derivation of left ventricular pressure (±LVdp/dtmax) were measured. At the end of reperfusion, the infarct size (IS) and apoptotic index (AI) were examined. MI/R caused significant cardiac dysfunction and myocardial apoptosis (strong TUNEL-positive staining). Compared with the control group, insulin treatment in MI/R rats exerted protective effects as evidenced by reduced myocardial IS [(28.9±2.0)% vs (45.0±4.0)%, n=6, P＜0.01 ], inhibited cardiomyocyte apoptosis [decreased AI: (16.0±0.7)% vs (27.6±1.3)%, n=6, P＜0.01] and improved recovery of cardiac systolic/diastolic function (including LVDP and ±LVdp/dtmax) at the end of reperfusion. Moreover, insulin resulted in 1.7-fold and 1.5-fold increases in Akt and JNK phosphorylation in I/R myocardium, respectively (n=6, P＜0.05). Inhibition of Akt activation with LY294002 abolished, and inhibition of JNK activation with SP600125 enhanced the cardioprotection by insulin, respectively. And the abolishment by LY294002could be partly converted by SP600125 pretreatment. In addition, SP600125 also decreased the Akt phosphorylation (n=6, P＜0.05).These results demonstrate that insulin simultaneously activates both Akt and JNK, and the latter further increases the phosphorylation of Akt which attenuates MI/R injury and improves heart function; this cross-talk between Akt and JNK in the insulin signaling is involved in insulin-induced cardioprotective effect.

Key words: ischemia/reperfusion injury;Insulin;Apoptosis;Akt;JNK;cross-talk

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引用本文：

刘海涛, 张海锋, 司瑞, 张全江, 张昆茹, 郭文仪, 王海昌, 高峰. 胰岛素保护缺血/再灌注心脏:PI3--K/Akt和JNKs信号通路间的交互作用[J]. 生理学报 2007; 59 (5): 651-659.

Liu Haitao, Zhang Haifeng, Si Rui, Zhang Quanjiang, Zhang Kunru, Guo Wenyi, Wang Haichang, Gao Feng. Insulin protects isolated hearts from ischemia/reperfusion injury: cross--talk between PI3--K/Akt and JNKs. Acta Physiol Sin 2007; 59 (5): 651-659 (in Chinese with English abstract).