神经调节蛋白-1原核表达及其对阿霉素致大鼠心肌细胞毒性的保护作用  

Prokaryotic expression of neuregulin-1 and its protective effect on doxorubicin-induced cardiomyocytes toxicity in rats

作　　者：刘永强[1] 韩培立[1] 刘辉[2] LIU Yongqiang;HAN Peili;LIU Hui(Department of Cardiovascular Surgery,the First Affiliated Hospital of Xinxiang Medical University,Henan Key Laboratory of Neural Rehabilitation,Weihui 453100,Henan Province,China;Department of Cardiology,the First Affiliated Hospital of Xinxiang Medical University,Weihui 453100,Henan Province,China)

机构地区：[1]新乡医学院第一附属医院心外科,河南省神经修复重点实验室,河南卫辉453100 [2]新乡医学院第一附属医院心内科,河南卫辉453100

出　　处：《新乡医学院学报》2020年第2期101-106,共6页Journal of Xinxiang Medical University

基　　金：河南省医学科技攻关计划项目(编号:201602155)。

摘　　要：目的探讨神经调节蛋白-1(NRG-1)对减轻阿霉素(DOX)所致大鼠心肌细胞毒性的作用及其机制。方法提取Sprague Dawley(SD)胎鼠的心室肌细胞总RNA并原核表达NRG-1蛋白;分离并培养SD乳鼠原代心肌细胞,应用细胞计数试剂盒(CCK-8)测定不同浓度DOX作用下大鼠原代心肌细胞的活性;并将心肌细胞分为正常对照组、DOX(5.0μmol·L^-1)组、DOX(5.0μmol·L^-1)+NRG-1(11.0 nmol·L^-1)组和NRG-1(11.0 nmol·L^-1)组,培养7 d后分别采用Western blot法和荧光定量聚合酶链反应检测大鼠原代心肌细胞的Na+-Ca2+交换体(NCX-1)和心肌球蛋白轻链激酶(cMLCK)蛋白和mRNA表达。结果原核表达NRG-1蛋白成功;DOX的心肌细胞毒性随其浓度增高而加重;NRG-1对不同浓度DOX干预的大鼠心肌细胞活力均有恢复作用(P<0.05),DOX浓度继续增高时其恢复作用受限(P<0.05),同时NRG-1浓度越高其细胞活力恢复越好(F=3606.28、2010.60、215.41,P<0.05);5.0μmol·L^-1DOX能抑制cMLCK蛋白和mRNA的表达(P<0.05),也能提高NCX-1蛋白和mRNA的表达(P<0.05),而11.0 nmol·L^-1NRG-1能够逆转DOX的上述作用(P<0.05)。结论NRG-1能够改善DOX所致大鼠心肌细胞毒性,其机制可能与cMLCK表达上调及NCX-1表达下调有关。Objective To investigate the effect of neuregulin-1(NRG-1)on reducing cardiomyocyte toxicity of rats induced by doxorubicin(DOX)and its mechanism.Methods Total RNA was extracted from ventricular myocytes of Sprague Dawley(SD)fetal rats and NRG-1 was expressed in prokaryotic cells.Primary cardiomyocytes of SD rats were isolated and cultured.The cell counting kit-8(CCK-8)assay was used for determining the activity of rat cardiomyocytes at different DOX concentrations.Cardiomyocytes were divided into normal control group,DOX(5.0μmol·L^-1)group,DOX(5.0μmol·L^-1)+NRG-1(11.0 nmol·L^-1)group,and NRG-1(11.0 nmol·L^-1)group.All the cells in each group were cultured for seven days.The protein expression of Na+-Ca2+exchanger(NCX-1)and cardiac myosin light chain kinase(cMLCK)was detected by Western blot,and the mRNA expression level was measured by fluorescence quantitative polymerase chain reaction.Results NRG-1 was obtained by prokaryotic expression.DOX-induced cardiomyocytes toxicity became more significant with the increasing dose of DOX.The NRG-1 could restore the viability of cardiomyocytes under different does of DOX(P<0.05),However,the protective effect of NRG-1 was limited when the DOX concentration continued to incresase(P<0.05);meanwhile,the cell viability recovery became more significant with higher degree of NRG-1 concentrations(F=3606.28,2010.60,215.41;P<0.05).The 5.0μmol·L^-1 DOX could inhibit the expression of cMLCK protein and mRNA(P<0.05),and increase the expression of NCX-1 protein and mRNA(P<0.05),while the 11.0 nmol·L^-1 NRG-1 could reverse the above effects of DOX(P<0.05).Conclusion NRG-1 can improve DOX-induced cardiomyocyte toxicity in rats,and its mechanism may be related to the up-regulation of the cMLCK expression and the down-regulation of the NCX-1 expression.

关 键 词：神经调节蛋白-1 阿霉素 心肌球蛋白轻链激酶 Na^+-Ca^2+交换体 

分 类 号：R965[医药卫生—药理学] Q816[医药卫生—药学]