ERK信号通路介导的EP300过表达在苯肾上腺素诱导小鼠心肌细胞肥大中的作用  

Over-expression of EP300 mediated by ERK signaling pathway is involved in phenylephrine-induced mouse cardiomyocyte hypertrophy

作　　者：黄丽欣 彭波辉 彭昌 冯玉松 罗孝美 吴书琪 张焕婷 HUANG Li-xin;PENG Bo-hui;PENG Chang;FENG Yu-song;LUO Xiao-mei;WU Shu-qi;ZHANG Huan-ting(Department of Pediatrics,The Affiliated Hospital,Zunyi Medical University,Zunyi 563000,China;First Clinical College,Zunyi Medical University,Zunyi 563000,China;Department of Physiology,School of Basic Medical Sciences,Zunyi Medical University,Zunyi 563000,China)

机构地区：[1]遵义医科大学附属医院儿内一科,贵州遵义563000 [2]遵义医科大学第一临床学院,贵州遵义563000 [3]遵义医科大学基础医学院生理教研室,贵州遵义563000

出　　处：《中国病理生理杂志》2021年第5期818-824,共7页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.81560040,No.82060046);遵义医科大学博士启动基金项目[院字(2015)4号];遵义医科大学大学生创新创业训练项目(No.ZYDC2018053)。

摘　　要：目的:探讨细胞外信号调节激酶(ERK)信号通路介导的E1A结合蛋白p300(EP300)过表达在苯肾上腺素(PE)诱导的小鼠心肌细胞肥大中的作用。方法:原代培养新生小鼠心肌细胞,按照随机数字表法分为:正常组、生理盐水(NS)组、PE组、溶剂对照组、漆树酸(AA)组、ERK抑制剂组和AA+ERK抑制剂组。收集干预48 h的小鼠心肌细胞,采用Western blot检测ERK、第9位赖氨酸乙酰化的组蛋白H3(H3K9ac)和β-肌球蛋白重链(β-MHC)的蛋白表达水平;RT-qPCR检测心肌细胞肥大标志物β-MHC的mRNA表达水平;免疫共沉淀验证EP300与H3K9ac之间的调控关系;免疫荧光染色及Western blot检测心肌细胞中EP300的表达水平。结果:Western blot结果表明小鼠心肌细胞中H3K9ac水平在PE组显著高于生理盐水对照组(P<0.05);Western blot及免疫荧光结果表明PE组EP300表达水平显著高于生理盐水对照组(P<0.05);免疫共沉淀结果表明EP300与H3K9ac之间能够相互结合;PE组p-ERK蛋白水平及β-MHC的mRNA和蛋白水平均显著高于NS组(P<0.05);而ERK抑制剂组及AA组EP300、H3K9ac、β-MHC及p-ERK水平均显著低于PE组(P<0.05)。结论:EP300介导的H3K9ac高乙酰化参与了PE诱导的小鼠心肌细胞肥大,而ERK信号通路可能是AA减轻PE诱导的心肌肥厚的信号通路之一。AIM:To investigate the role of over-expression of E1 A-binding protein p300(EP300)mediated by the extracellular signal-regulated kinase(ERK)signaling pathway in phenylephrine(PE)-induced mouse cardiomyo-cyte hypertrophy.METHODS:Cardiomyocyte hypertrophy model was induced by PE in primary neonatal mouse myocar-dial cells.According to random number table method,the experiments were designed in 7 groups as following:normal group,normal saline group,PE group,vehicle group,anacardic acid(AA)group,ERK inhibitor group and AA+ERK in-hibitor group.Kunming mouse myocardial cells were collected after intervention for 48 h.The protein levels of p-ERK,acetylated histone H3 at lysine 9(H3 K9 ac)andβ-myosin heavy chain(β-MHC)were assayed by Western blot.The mRNA level ofβ-MHC was tested by RT-qPCR.The interaction between EP300 and H3 K9 ac were verified by co-immuno-precipitation.The expression of EP300 was assayed by immunofluorescence staining and Western blot.RESULTS:The results of Western blot showed that the level of H3 K9 ac in PE group was significantly increased compared with normal sa-line group(P<0.05).The results of Western blot and immunofluorescence staining showed that the level of EP300 in PE group was significantly increased compared with normal saline group(P<0.05).The results of co-immunoprecipitation showed that EP300 and H3 K9 accombined with each other.The protein level of p-ERK in PE group was significantly in-creased compared with normal saline group(P<0.05).The mRNA and protein levels ofβ-MHC in PE group were appar-ently increased compared with normal saline group(P<0.05).However,ERK inhibitor U0126 and histone acetylase in-hibitor AA attenuated the over-expression of EP300,H3 K9 ac,β-MHC and p-ERK in the myocardial cells treated with PE(P<0.05).CONCLUSION:Hyperacetylation of H3 K9 ac mediated by EP300 is involved in PE-induced cardiomyocyte hypertrophy,and ERK signaling pathway may be one of the signaling pathways for AA to attenuate PE-induced cardiomyo-cyte hypertrophy in mice.

关 键 词：ERK信号通路 心肌细胞肥大 组蛋白乙酰化 E1A结合蛋白p300 漆树酸 

分 类 号：R542.2[医药卫生—心血管疾病] R363.2[医药卫生—内科学]