乌头碱致斑马鱼和人心肌细胞心率失常模型的研究
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摘要
目的研究天然产物乌头碱致斑马鱼和人心肌细胞心率失常模型建立的可行性。方法将2 d大的心脏表达绿色荧光的斑马鱼幼鱼随机分为4组:对照组(0.1%DMSO)和低、中、高3个剂量实验组(1,3,10μmol·L~(-1)乌头碱),每组12条。作用1 d后,用活细胞工作站系统成像并分析斑马鱼心室跳动相对荧光的强度和心室的心搏量。将人胚胎干细胞诱导成具有跳动功能的心肌细胞,经1μmol·L~(-1)乌头碱作用后,分析其自动跳动时钙离子的闪烁情况。结果经乌头碱作用1 d后的斑马鱼幼鱼,对照组心博量为(233.89±30.09)p L,低、中、高剂量实验组的心博量分别为(177.23±29.05),(136.32±43.34),(42.63±23.34)p L,其中,高剂量实验组心搏量显著减少(P<0.05)。作用10min后的人心肌细胞,对照组钙离子振荡频率和振荡幅度分别为21.67±2.86,102.20±7.35,低剂量实验组钙离子振荡频率和振荡幅度分别为46.66±6.66,57.06±6.63,药物处理后,钙离子振荡频率显著增加(P<0.05),振幅显著降低(P<0.05)。结论经乌头碱作用后,斑马鱼幼鱼和人心肌细胞均表现出心率失常的特征,为抗心律失常药物开发提供了体内外药物筛选模型。
Objective To study the feasibility of establishing arrhythmia model induced by natural product aconitine in zebrafish and human cardiomyocytes.Methods Two-day old zebrafish juveniles expressing green fluorescence were randomly divided into 4 groups:control group(0.1%DMSO)and low,middle and high dose experimental groups(1,3,10μmol·L~(-1)Aconitine),12 in each group.One day later,the live cell workstation system was used to image and analyze the relative fluorescence intensity of zebrafish ventricular beat and the ventricular stroke volume.The human embryonic stem cells were induced into beating cardiomyocytes,and after the action of 1μmol·L~(-1)aconitine,the calcium ion scintillation was automatically analyzed.Results After 1 d of aconitine in zebrafish juveniles,the heart rate of control group was(233.89±30.09)p L,and those of low,middle and high experimental groups were(177.23±29.05),(136.32±43.34)and(42.63±23.34)p L,the stroke volume in high dose experimental group had significantly decreased(P<0.05).The calcium ion oscillation frequency and oscillation amplitude of human myocardial cells and control group after 10 min were 21.67±2.86,102.20±7.35,and were46.66±6.66,57.06±6.63 in low dose experiment group,and the frequency of calcium ion oscillation was significantly increased after drug treatment(P<0.05),the amplitude was significantly reduced(P<0.05).Conclusion After the action of aconitine,zebrafish juvenile fish and human cardiomyocytes exhibit arrhythmia,which provides screening model for the development of antiarrhythmic drugs in vitro and in vivo.
Objective To study the feasibility of establishing arrhythmia model induced by natural product aconitine in zebrafish and human cardiomyocytes.Methods Two-day old zebrafish juveniles expressing green fluorescence were randomly divided into 4 groups:control group(0.1%DMSO)and low,middle and high dose experimental groups(1,3,10μmol·L~(-1)Aconitine),12 in each group.One day later,the live cell workstation system was used to image and analyze the relative fluorescence intensity of zebrafish ventricular beat and the ventricular stroke volume.The human embryonic stem cells were induced into beating cardiomyocytes,and after the action of 1μmol·L~(-1)aconitine,the calcium ion scintillation was automatically analyzed.Results After 1 d of aconitine in zebrafish juveniles,the heart rate of control group was(233.89±30.09)p L,and those of low,middle and high experimental groups were(177.23±29.05),(136.32±43.34)and(42.63±23.34)p L,the stroke volume in high dose experimental group had significantly decreased(P<0.05).The calcium ion oscillation frequency and oscillation amplitude of human myocardial cells and control group after 10 min were 21.67±2.86,102.20±7.35,and were46.66±6.66,57.06±6.63 in low dose experiment group,and the frequency of calcium ion oscillation was significantly increased after drug treatment(P<0.05),the amplitude was significantly reduced(P<0.05).Conclusion After the action of aconitine,zebrafish juvenile fish and human cardiomyocytes exhibit arrhythmia,which provides screening model for the development of antiarrhythmic drugs in vitro and in vivo.
引文
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[3]崔国祯,徐燕玲,孙安露,等.丹参素衍生物对斑马鱼促血管新生作用的研究[J].中国药理学通报,2016,32(6):795-800.
[4]崔国祯,周鹤峰,陈焕娴,等.阿奇霉素对斑马鱼心脏毒性的初步研究[J].遵义医学院学报,2014,37(5):495-498.
[5]CUI G,LUK S C,LI R A,et al.Cytoprotection of baicalein against oxidative stress-induced cardiomyocytes injury through the Nrf2/Keap1 pathway[J].J Cardiovasc Pharmacol,2015,65(1):39-46.
[6]YANG L,SOONPAA M H,ADLER E D,et al.Human cardiovascular progenitor cells develop from a KDR plus embryonic-stemcell-derived population[J].Nature,2008,453(7194):524-526.
[7]LEI S W,CUI G,LEUNG G P H,et al.Icaritin protects against oxidative stress-induced injury in cardiac H9c2 cells via Akt/Nrf2/HO-1 and calcium signalling pathways[J/OL].J Funct Foods,2015,18 213-223.2015-10-01[2017-02-23].https://ac.els-cdn.com/S1756464615003485/1-s2.0-S1756464615003485-main.pdf?_tid=dc1cf59f-4621-4532-aeed-f11bb940e29a&acdnat=1520934180_6a294fe67b830bf9986aeb314f9ffbe0.