PERK通路参与了缺氧心肌细胞的内质网应激及凋亡  

PERK signal pathway is involved in hypoxia-induced endoplasmic reti-culum stress and apoptosis in cultured cardiac myocytes

作　　者：刘春蕾[1,2] 李鑫[1] 李蕊君[1] 何云云[1,2] 何昆仑[1] 王莉莉[3] 

机构地区：[1]中国人民解放军总医院心内科,北京100853 [2]南开大学医学院,天津300071 [3]军事医学科学院毒物药物研究所,北京100850

出　　处：《中国病理生理杂志》2012年第8期1392-1398,共7页Chinese Journal of Pathophysiology

基　　金：国家科技部国际科技合作重大专项课题(No.2006DFB32210)

摘　　要：目的:观察缺氧对原代培养的Wistar乳鼠心肌细胞的损伤,探讨内质网应激在缺氧心肌损伤发生发展过程中起的作用及PERK通路是否参与其信号转导过程。方法:将原代培养的乳鼠心肌细胞随机分为正常对照组和缺氧1 h、4 h、8 h、12 h、24 h组,通过测定细胞ATP含量反映细胞活力;高内涵分析细胞成像系统检测多参数凋亡;采用免疫细胞化学和蛋白印迹方法检测以内质网为靶点的分子伴侣(GRP78和钙网蛋白)的表达,PERK通路(PERK和eIF2α)的磷酸化水平,以及其下游分子(ATF4和CHOP)在缺氧不同时点蛋白的表达变化特征。采用PERK通路激活型药物salubrinal处理原代培养的Wistar乳鼠心肌细胞,观察药物是否对缺氧损伤的心肌细胞有保护作用。结果:缺氧引起心肌细胞凋亡,缺氧早期(约1 h)钙网蛋白和GPR78的表达上调;缺氧中期(4 h)p-PERK、p-eIF2α和ATF4的表达上调;缺氧后期(12 h)CHOP的表达上调。Salubrinal对缺氧心肌有保护作用。结论:在培养的心肌细胞中,缺氧可激发内质网应激。在缺氧早期激活PERK通路保护机体对抗缺氧损伤,后期激活细胞凋亡通路。To investigate the role of endoplasmic reticulum （ER） stress in the process of hypoxia - in- duced neonatal rat myocardial injury through PERK signal pathway. METHODS： Neonatal rat cardiac myocytes were ran- domly divided into control group and hypoxia 1 h, 4 h, 8 h, 12 h and 24 h groups. Cell viability was evaluated by determi- ning the intracellular content of ATP. Apoptosis was measured by high - content analysis （HCA） cell imaging system. The protein levels of GRP78, calreticulin, p - PERK, p - eIF2ct, ATF4 and CHOP were detected by Western blotting at differ- ent time points. The primary cultured neonatal rat cardiac myocytes were treated with an agonist of PERK pathway salu- brinal and the cell apoptosis was observed under hypoxia. RESULTS： In the early phase, hypoxia induced an increase in the expression of calreticulin and GPR78. In the middle phase of hypoxia, the levels of p - PERK, p - eIF2a and ATF4 were increased. In the later phase of hypoxia, increased CHOP level was also observed. Salubrinal effectively protected the cardiac myocytes from hypoxic injury. CONCLUSION： Hypoxia activates ER stress in cardiac myocytes and also activates PERK signal pathway. PERK signaling protects cardiac myocytes from hypoxic damage in the early stage and triggers apop- tosis of the cells in the later phase.

关 键 词：心肌细胞 内质网应激 缺氧 未折叠蛋白反应 细胞凋亡 

分 类 号：R363.2[医药卫生—病理学]