Cerignoli Fabio, Charlot David, Whittaker Ross, Ingermanson Randy, Gehalot Piyush, Savchenko Alex, Gallacher David J, Towart Rob, Price Jeffrey H, McDonough Patrick M, Mercola Mark
Sanford-Burnham Medical Research Institute, 10901N. Torrey Pines Road, La Jolla, CA 92037, USA.
J Pharmacol Toxicol Methods. 2012 Nov-Dec;66(3):246-56. doi: 10.1016/j.vascn.2012.08.167. Epub 2012 Aug 25.
Current methods to measure physiological properties of cardiomyocytes and predict fatal arrhythmias that can cause sudden death, such as Torsade de Pointes, lack either the automation and throughput needed for early-stage drug discovery and/or have poor predictive value. To increase throughput and predictive power of in vitro assays, we developed kinetic imaging cytometry (KIC) for automated cell-by-cell analyses via intracellular fluorescence Ca²⁺ indicators. The KIC instrument simultaneously records and analyzes intracellular calcium concentration Ca²⁺ at 30-ms resolution from hundreds of individual cells/well of 96-well plates in seconds, providing kinetic details not previously possible with well averaging technologies such as plate readers. Analyses of human embryonic stem cell and induced pluripotent stem cell-derived cardiomyocytes revealed effects of known cardiotoxic and arrhythmogenic drugs on kinetic parameters of Ca²⁺ dynamics, suggesting that KIC will aid in the assessment of cardiotoxic risk and in the elucidation of pathogenic mechanisms of heart disease associated with drugs treatment and/or genetic background.
目前用于测量心肌细胞生理特性以及预测可能导致猝死的致命性心律失常（如尖端扭转型室速）的方法，要么缺乏早期药物发现所需的自动化和通量，要么预测价值不佳。为了提高体外测定的通量和预测能力，我们开发了动力学成像细胞术（KIC），用于通过细胞内荧光Ca²⁺指示剂进行逐个细胞的自动化分析。KIC仪器能在数秒内以30毫秒的分辨率同时记录和分析96孔板中数百个单个细胞/孔的细胞内钙浓度[Ca²⁺]（i），提供了以往诸如酶标仪等孔平均技术无法实现的动力学细节。对人类胚胎干细胞和诱导多能干细胞衍生的心肌细胞的分析揭示了已知心脏毒性和致心律失常药物对Ca²⁺动力学参数的影响，这表明KIC将有助于评估心脏毒性风险，并阐明与药物治疗和/或遗传背景相关的心脏病致病机制。
