乌头碱对人诱导多能干细胞衍生心肌细胞中钙信号的影响

Effect of Aconitine on Calcium Signal in Human Induced Pluripotent Stem Cells-Derived Cardiomyocytes

该文旨在探究乌头碱对人诱导多能干细胞衍生心肌细胞(human induced pluripotent stem cells-derived cardiomyocytes,hiPSCs-CMs)中钙信号的影响。用CCK-8法测定乌头碱对hiPSCsCMs细胞存活率的影响;通过共聚焦显微镜的明场模式记录不同加药时间和浓度条件下乌头碱对心肌细胞的搏动频率的影响;通过共聚焦显微镜荧光系统记录乌头碱对细胞自发及1、2、3 Hz电刺激条件下钙信号的影响;使用咖啡因诱导心肌细胞肌质网中的钙离子,令其全部被释放出来,以探究乌头碱对钙库的影响。乌头碱在加药浓度为9μmol/L、加药时间为6 h时将产生毒性作用,可对hiPSCs-CMs细胞存活率造成显著影响;乌头碱能够明显加快hiPSCs-CMs搏动频率,最大搏动频率与加药时间和浓度相关;乌头碱能够明显降低hiPSCs-CMs自发及1、2、3 Hz电刺激条件下钙瞬变的振幅;经0.3μmol/L乌头碱作用3 h的hiPSCs-CMs的钙库总钙量下降;且乌头碱对hiPSCs-CMs搏动频率及钙信号的影响具有洗脱作用。乌头碱能够加快hiPSCs-CMs搏动频率,降低钙瞬变振幅与钙库总钙量,并由此影响心肌功能。

This study aims to investigate the effect of aconitine on calcium signal in hiPSCs-CMs(human induced pluripotent stem cells-derived cardiomyocytes).CCK-8 assay was used to determine the effect of aconitine on the survival rate of human cardiomyocytes.The effect of aconitine on the beating frequency of cardiomyocytes under different dosage time and concentration conditions was recorded by confocal microscope light field mode.The effect of aconitine on calcium signal was recorded by confocal microscope fluorescence system under spontaneous and 1,2,3 Hz electrical stimulation conditions.To investigate the effect of aconitine on calcium pool,caffeine was used to induce the release of calcium ions from the sarcoplasmic reticulum of cardiomyocytes.Aconitine had toxic effects when the addition dosage was 9μmol/L and the dosage time was 6 h,which significantly affected the survival rate of human cardiomyocytes.Aconitine could significantly accelerate the beat frequency of hiPSCsCMs,which was related to the drug treatment time and concentration.Aconitine could significantly reduce the amplitude of spontaneous calcium transients and calcium transients under 1,2,3 Hz electrical stimulation in human cardiomyocytes.Sarcoplasmic reticulum calcium content of human cardiomyocytes treated with 0.3μmol/L aconitine for 3 h was decreased.Aconitine had elution effect on the pulsatile frequency and calcium signal of hiPSCsCMs.Aconitine can accelerate the beat frequency of hiPSCs-CMs,reduce the amplitude of calcium transient and the sarcoplasmic reticulum calcium content,and thus affect the myocardial function.