异叶青兰总黄酮对去甲肾上腺素诱导大鼠心肌细胞肥大的抑制作用
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摘要
目的观察异叶青兰总黄酮(DHBF)对去甲肾上腺素(NE)诱导的心肌细胞肥大的抑制作用,并探讨其作用机制。方法体外培养SD乳鼠的心肌细胞,实验分为正常对照组,模型对照组(给予2μmol·L-1NE造模),哌唑嗪组(NE造模后给予哌唑嗪50μmol·L-1),DHBF低浓度组(NE造模后给予DHBF 10μmol·L-1),DHBF中浓度组(NE造模后给予DHBF 25μmol·L-1),DHBF高浓度组(NE造模后给予DHBF 50μmol·L-1)。用NE诱导心肌细胞肥大,DHBF和哌唑嗪分别对其进行干预,CCK-8法观察心肌细胞活性,RT-PCR技术检测心肌的原癌基因(c-jun)和肥大基因脑钠肽(BNP) mRNA的表达水平,Western blotting蛋白检测心肌细胞钙调素蛋白酶(Ca N)、T细胞核活化因子-3(NFAT-3)蛋白的表达水平;激光共聚焦法检测心肌细胞的表面积,观察上述干预因素对心肌细胞肥大的作用。结果与正常对照组比较,模型对照组可使心肌细胞相对生存率明显下降,心肌肥大标志物基因c-jun和BNP的mRNA表达明显上调,Ca N和NFAT-3的蛋白表达明显增加,相对表面积明显增大(P<0.05);给予不同浓度的DHBF对NE引起的心肌细胞生存率下降,表面积增大产生拮抗作用,对NE诱导的c-jun和BNP mRNA表达的增加产生拮抗作用,同时对Ca N和NFAT-3的蛋白表达增加也具有一定拮抗作用(P<0.05)。结论 DHBF能改善NE诱导的心肌细胞肥大的生存率,下调原癌基因c-jun和心肌肥大因子BNP的mRNA表达,降低心肌细胞内Ca N和NFAT-3的蛋白表达水平,减小由NE诱导的心肌细胞的表面积。
Objective To probe into effect of Dracocephalum heterophyllum Benth flavonoids( DHBF) on noradrenaline( NE)-induced myocardial cell hypertrophy,and to study the mechanism. Methods SD neonatal rat myocardial cells were cultured in vitro. The experiment was divided into normal control group,model control group( NE 2 μmol·L-1),prazosin group( prazosin 50 μmol·L-1),low-,medium-and high-dose DHBF group( DHBF 10,25,50 μmol·L-1).Cardiomyocyte hypertrophy were induced by NE( 2 μmol·L-1). DHBF and prazosin were intervened respectively. CCK-8 method was used to observe the activity of myocardial cells. RT-PCR technique was used to detect the expression of mRNA of cardiac hypertrophy c-jun and brain natriuretic peptide( BNP),and Western blotting was used to detect the protein expression of Ca N and NFAT-3 in myocardial cells. Confocal laser scanning was used to detect the surface area of myocardial cell. Results Compared with the normal control group,survival rate of cardiomyocytes was significantly decreased,expression of mRNA of c-jun and BNP significantly upregulated,protein expression of Ca N and NFAT-3 decreased,and surface area increased in model control group( P< 0.05).Compared with the cell of model control group,low-,medium-and high-dose DHBF significantly reversed NE-induced decrease of the survival rate,increase of surface area,increase of c-jun and BNP mRNA,and increase of Ca N and NFAT-3 protein expression( P<0.05). Conclusion DHBF can improve the survival rate of cardiac hypertrophy patients,down-regulate c-jun and BNP mRNA expression, decrease Ca N and NFAT-3 protein expression, and decrease NE-induced surface area of cardiomyocytes.
Objective To probe into effect of Dracocephalum heterophyllum Benth flavonoids( DHBF) on noradrenaline( NE)-induced myocardial cell hypertrophy,and to study the mechanism. Methods SD neonatal rat myocardial cells were cultured in vitro. The experiment was divided into normal control group,model control group( NE 2 μmol·L-1),prazosin group( prazosin 50 μmol·L-1),low-,medium-and high-dose DHBF group( DHBF 10,25,50 μmol·L-1).Cardiomyocyte hypertrophy were induced by NE( 2 μmol·L-1). DHBF and prazosin were intervened respectively. CCK-8 method was used to observe the activity of myocardial cells. RT-PCR technique was used to detect the expression of mRNA of cardiac hypertrophy c-jun and brain natriuretic peptide( BNP),and Western blotting was used to detect the protein expression of Ca N and NFAT-3 in myocardial cells. Confocal laser scanning was used to detect the surface area of myocardial cell. Results Compared with the normal control group,survival rate of cardiomyocytes was significantly decreased,expression of mRNA of c-jun and BNP significantly upregulated,protein expression of Ca N and NFAT-3 decreased,and surface area increased in model control group( P< 0.05).Compared with the cell of model control group,low-,medium-and high-dose DHBF significantly reversed NE-induced decrease of the survival rate,increase of surface area,increase of c-jun and BNP mRNA,and increase of Ca N and NFAT-3 protein expression( P<0.05). Conclusion DHBF can improve the survival rate of cardiac hypertrophy patients,down-regulate c-jun and BNP mRNA expression, decrease Ca N and NFAT-3 protein expression, and decrease NE-induced surface area of cardiomyocytes.
引文
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[14]刘思宁,栾天竺.阿托伐他汀对异丙肾上腺素诱发的鼠心肌细胞肥大的抑制作用[J].哈尔滨医科大学学报,2015,29(4):305-308.
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[17] SHIROSHITA-TAKESHITA A,BRUNDEL B J,LAVOIE J,et al. Prednisone prevents atrial fibrillation promotion by atrial tachycardia remodeling in dogs[J]. Cardiovasc Res,2006,69(4):865-875.
[2] MULLER BRUNOTTE R,KAHAN T,LOPEZ B,et al.Myocardial fibrosis and diastolic dysfunction in patients with hypertension:results from the Swedish Irbesartan Left Ventricular Hypertrophy Investigation versus Atenolol(SILVHIA)[J].J Hypertens,2007,25(9):1958-1966.
[3]刘晓丽,邢晓为,黄利华,等.β1肾上腺素受体与心衰、高血压和心肌纤维化之间的关系研究进展[J].中国药理学通报,2013,29(12):1640-1644.
[4] BARRY S P,DAVIDSON S M,TOWNSEND P A.Molecular regulation of cardiac hypertrophy[J]. Int J Biochem Cell Biol[J],2008,40(10):2023-2039.
[5]何雯,邬利娅·伊明,司丽君,等.异叶青兰总黄酮对高血压大鼠心肌肥厚的影响[J].中国药理学通报,2013,29(1):1008-1011.
[6]陶静,马依彤,李晓梅,等.原代乳鼠心肌细胞培养方法的改进[J].中华心血管病杂志,2014,42(1):53-56.
[7]周庆峰,王洪新,王桂君,等.高糖对去甲肾上腺素诱导的心肌细胞肥大的促进作用[J].中国药理学通报,2003,19(9):1054-1057.
[8] MOSTERD A,HOES A W.Clinical epidemiology of heart failure[J].Heart,2007,93(9):1137.
[9] BROUWERS F P,DE BOER R A,VANDER HARST P,et al,Incidence and epidemiology of new onset heart failure with preserved vs. reduced ejection fraction in a communitybased cohort:11-year follow-up of PREVEND[J].Eur Heart J,2013,34(19):1424-1431.
[10] RODILLA E,PASCUAL J M,COSTA J A,et al. Regression of left ventricular hypertrophy and microalbuminuria changes during antihypertensive treatment[J].J Hypertension,2013,31(8):1683-1691.
[11]张婧怡,林淑娴,李利生,等.吴茱萸次碱对腹主动脉缩窄大鼠左室肥厚的抑制作用[J].医药导报,2018,38(2):152-156.
[12] LIOU S F,HSU J H,CHEN Y T,et al. KMUP-1 attenuates endothelin-1-induced cardiomyocyte hypertrophy through activation of heme oxygenase-1 and suppression of the Akt/GSK-3β,calcineurin/NFATc4 and Rho A/ROCK Pathways[J].Molecules,2015,20(6):1043-1045.
[13] SHAH M S,BRPWNLEE M.Molecular and cellular mechanisms of cardiovascular disorders in diabetes[J].Circul Res,2016,118(11):1808.
[14]刘思宁,栾天竺.阿托伐他汀对异丙肾上腺素诱发的鼠心肌细胞肥大的抑制作用[J].哈尔滨医科大学学报,2015,29(4):305-308.
[15] FELKIN L E,NARITA T,GERMACK R,et al. Calcineurin splicing variant calcineurin Abeta1 improves cardiac function after myocardial infarction without inducing hypertrophy[J].Circulation,2011,123(24):2838-2847.
[16] SHEN X,ZHANG J,CHENG B.Construction of rat calcineurin A alpha c DNA recombinant adenovirus vector and its identification[J].J Huazhong University Sci:Technolog Med Sci,2006,26(1):9-12.
[17] SHIROSHITA-TAKESHITA A,BRUNDEL B J,LAVOIE J,et al. Prednisone prevents atrial fibrillation promotion by atrial tachycardia remodeling in dogs[J]. Cardiovasc Res,2006,69(4):865-875.