衔接蛋白p66shc在雌激素抑制AngⅡ诱导的心肌细胞凋亡中的作用  

Effects of p66shc Adapter Protein and Estrogen on Cardiomyocyte Apoptosis Induced by Angiotensin Ⅱ

作　　者：张明[1] 魏瑾[1] 单虎[1] 闫蕊[1] 林琳[1] 朱延河[1] 

机构地区：[1]西安交通大学医学院第二附属医院心内科,西安710004

出　　处：《四川大学学报（医学版）》2014年第2期202-206,211,共6页Journal of Sichuan University(Medical Sciences)

基　　金：国家自然科学基金(No.30771862和No.81170209)资助

摘　　要：目的探讨衔接蛋白p66shc在血管紧张素Ⅱ(AngⅡ)诱导的心肌细胞凋亡中的作用及雌激素的干预影响。方法原代培养SD乳鼠心肌细胞,将心肌细胞分为5组:正常对照组、10-11 mol/L AngⅡ组、10-9 mol/L AngⅡ组、10-7 mol/L AngⅡ组、10-7 mol/L AngⅡ+雌二醇组;经药物干预后应用MTT法测定细胞存活率、流式细胞仪测定活性氧含量和细胞凋亡率、荧光酶标仪检测线粒体膜电位水平、Western blot检测p66shc和磷酸化(pp66shc)蛋白的表达水平。结果随着AngⅡ浓度的升高,心肌细胞存活率和线粒体膜电位水平逐渐下降,而细胞内活性氧含量和凋亡率逐渐升高(P<0.05);且雌激素预处理可减轻AngⅡ对心肌细胞的损伤(P<0.05)。AngⅡ可使心肌细胞内p-p66shc和线粒体内的p66shc的表达呈剂量依赖性升高(P<0.05),雌激素预处理可减弱AngⅡ对细胞内p-p66shc和线粒体内p66shc表达水平的影响(P<0.05)。结论 p66shc参与了AngⅡ诱导的心肌细胞凋亡反应,且雌激素可通过下调线粒体内p66shc的表达而减轻AngⅡ对心肌细胞的损伤。Objective To explore the role of p66shc in cardiomyocyte apoptosis induced by angiotensin （Ang） II and the effect of estrogen pretreatment. Methods Neonatal rat cardiomyocytes were randomly divided into five groups： normal control, 10-11 mol/L Angl] , 10-9 mol/L Angl] , 10-7 mol/L Ang Ⅱ , and 10-7 mol/L Angll ＋ estrogen treated groups. The cell viability was measured by MTT. The level of reactive oxygen species （ROS） and cell apoptosis rate were measured by flow cytometry. Mitochondrial membrane potential （MMP） was detected using a fluorescence microplate reader, and the protein expression of phosphorylated and total p66shc were detected using Western blot. Results With the increase of Ang 11 concentrations, cell viabilities and MMP levels decreased, whereas, the levels of ROS and cell apoptosis rates increased （P〈0.05）. Pretreatment with estrogen significantly attenuated the cardiomyocyte injury induced by Ang 11 （P〈0.05）. The protein expression of phosphorylated p66shc in the whole cell lysates and total p66shc in the mitochondria increased in a dose-dependent manner when cardiomyocytes were exposed to Ang II （P〈0. 05）. Pretreatment with estrogen significantly down-regulated the protein expression of phosphorylated p66shc in the whole cell lysates and total p66shc in the mitocbondria （P〈0.05）. Conclusion p66shc is involved in cardiomyocyte apoptosis induced by Ang II , and estrogen could attenuate Ang 11 induced cardiomyocyte injury through down-regulating the protein expression of p66shc.

关 键 词：雌激素 心肌细胞 血管紧张素Ⅱ P66SHC 

分 类 号：R363[医药卫生—病理学]