自噬抑制剂3-MA对“饥饿状态”下心肌细胞自噬和凋亡的影响  

Effect of Autophagy Inhibitor 3-MA on Autophagy and Apoptosis of“Starving”Myocardial Cells

作　　者：杨翼[1] 李章华[2] 贺磊磊[3] 姚宇琴 柳华[1] 王凌骁[3] 

机构地区：[1]武汉体育学院健康科学学院,湖北武汉430079 [2]武汉大学人民医院,湖北武汉430060 [3]武汉体育学院研究生院,湖北武汉430079

出　　处：《武汉体育学院学报》2014年第11期61-66,共6页Journal of Wuhan Sports University

基　　金：国家自然科学基金项目(30700388);湖北省自然科学基金项目(2011CDB313);湖北省教育厅科研计划重点项目(D20144101)

摘　　要：目的:观察自噬抑制剂3-甲基腺嘌呤(3-MA)对"饥饿状态"心肌细胞自噬和凋亡的影响,以探讨调控"饥饿状态"下心肌细胞自噬与凋亡的方法。方法:新生1-3日龄Wistar雄性大鼠乳鼠30只,随机平均分为三组:正常对照组、无糖无血清组和3-MA组,每组10只。取出心脏分离心肌细胞,培养5d进行相应处理后,通过流式细胞仪和透射电镜观察心肌细胞自噬和凋亡,免疫荧光和Western-blot检测心肌细胞Beclin1蛋白的表达。结果:1正常培养的心肌细胞较少发生自噬和凋亡,"饥饿状态"下心肌细胞凋亡率无明显上升,但自噬数量明显增加;2与正常培养的心肌细胞相比,"饥饿状态"下心肌细胞Beclin1表达表现为显著性的上升(P<0.01);而3-MA预处理后心肌细胞Beclin1表达呈现显著性的下降(P<0.01)。结论:"饥饿状态"可促使心肌细胞自噬发生而对凋亡无明显影响;自噬抑制剂3-MA则明显抑制"饥饿状态"下心肌细胞发生自噬,从而间接引起心肌细胞凋亡的上升,造成心肌细胞死亡。The purpose of this study was to investigate the effect of autophagy inhibitor, 3-methyladenine （3-MA）, on au- tophagy and apoptosis of myocardial cells, and explore the regulation methods of autophagy and apoptosis of ＂starving＂ myocardial cells. 30 newborn （1-3 days old） male Wistar rats were randomly divided into three groups including normal control group, serum and glucose deprivation group and 3-MA pretreatment group. Ten rats were in each group. After myocardial cells were isolated and cultured for 5 days, autophagy and apoptosis of myocardial cells were examined by flow eytometry and transmission electron microscope, and the expression of Beclinl protein were evaluated by immunofluorescence and Western-blotting. It was shown that less autophagy and apoptosis were observed in normal cultured myocardial cells while a significant increase of autophagy was observed in serum and glucose deprivation group although no obvious apoptosis was observed. The expression of Beclinl in ＂starving＂ myocardial cells revealed a significant increase （P〈0. 01）, while it exhibited a significant reduction in the presence of 3-MA preconditioning （P〈0. 01）. Starvation could promote autophagy activation of myocardial cells instead of apoptosis; in contrast, 3-MA revealed the obvious inhibitory effect on the autophagy of the ＂starving＂ myocardial cells, which indirectly resulted in the enhanced apoptosis myocardial cells.

关 键 词：3-甲基腺嘌呤 无糖无血清培养 心肌细胞 自噬 凋亡 

分 类 号：G804.5[文化科学—运动人体科学]