PKCε信号通路介导的葛根素抗心肌细胞缺氧/复氧损伤作用  

Role of PKCε signal pathway in puerarin preconditioning against cardiomyocytes anoxia / reoxygenation injury

作　　者：汤蕾[1,2] 胥甜甜[2] 易小清[2] 刘瑛[2] 罗喻超[2] 尹东[3] 何明[1,2] 

机构地区：[1]南昌大学食品科学与技术国家重点实验室,江西南昌330006 [2]南昌大学江西省基础药理学重点实验室,江西南昌330006 [3]南昌大学第二附属医院分子医学中心实验室,江西南昌330006

出　　处：《中国药理学通报》2014年第1期77-81,共5页Chinese Pharmacological Bulletin

基　　金：国家重点基础研究发展计划(973计划)资助项目(No2009CB526405);国家自然科学基金资助项目(No81260492)

摘　　要：目的探讨葛根素(puerarin,Pue)抗心肌细胞缺氧/复氧(A/R)损伤保护作用与PKCε蛋白表达的关系。方法培养新生SD大鼠原代心肌细胞,分为正常对照(Con)组、A/R组、葛根素+A/R(Pue+A/R)组、葛根素+抑制剂+A/R(Pue+εV1-2+A/R)组、抑制剂+A/R(εV1-2+A/R)组。Western blot测定PKCε蛋白表达水平,MTT法检测细胞存活率,比色法测定培养液肌酸磷酸酶(CPK)、乳酸脱氢酶(LDH)活性、细胞内丙二醛(MDA)的含量和超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)的活性,流式细胞仪测定线粒体膜电位和细胞凋亡。结果葛根素预处理24h后,心肌细胞PKCε表达呈剂量依赖性上调(P<0.01);培养液CPK、LDH活性降低,细胞内SOD、GSH-Px活性升高,MDA含量减少,细胞存活率升高,细胞凋亡减少(P<0.01);PKCε抑制剂εV1-2则可明显消弱葛根素的上述心肌保护作用。结论葛根素的抗心肌缺血/再灌注损伤保护作用涉及PKCε信号通路,至少部分依赖于其对PKCε表达水平的上调。Aim To investigate the relationship be- tween the cardioprotective effect of puerarin from anoxi- a/reoxygenation （A/R） injury and PKCe pathway. Methods Primary neonatal SD rat cardiomyocytes were cultured and divided into normal control group, A/R group, puerarin ＋ A/R group（Pur ＋ A/R） , pu- erarin ＋ inhibitor group （ Pur ＋ eV1-2 ＋ A/R） , and in- hibitor group （eV1-2 ＋ A/R）. Expression of PKCe was determined by Western blot, and cell viability was measured by MTT method. CPK, LDH, SOD, GSH- Px, MDA activity was determined by chromometry. Apoptosis and mitochondrial membrane potential were determined by flow cytometry. Results 24h after pu- erarin precondition, the expression of PKCe was upreg- ulated in cardiomyocytes （P 〈 0. 01 ）. In the group pretreated with 320 μmol · L^-1 puerarin before A/R, the activity of CPK and LDH in culture medium de- creased; the activity of intracellular SOD, GSH-Px in- creased; the content of MDA decreased; cell viability increased and cell apoptosis decreased （ P 〈 0.01 ）. However, all these protective effects were attenuated significantly in the group pretreated with puerarin and PKCe inhibitor eVI-2. Conclusion The effect of pu- erarin against A/R injury in eardiomyocytes involues PKCe pathway, and at least partly depends on its effect of upregulating the expression of PKCe.

关 键 词：葛根素 PKCe 缺氧 复氧 心肌保护 凋亡 线粒体 膜电位 

分 类 号：R-332[医药卫生] R284.1