BioMediTech Institute and Faculty of Biomedical Sciences and Engineering, Tampere University of Technology, Korkeakoulunkatu 10, 33720, Tampere, Finland.
BioMediTech Institute and Faculty of Medicine and Life Sciences, University of Tampere, Tampere, Finland.
Ann Biomed Eng. 2018 Jan;46(1):148-158. doi: 10.1007/s10439-017-1933-2. Epub 2017 Oct 3.
Induced pluripotent stem cell derived cardiomyocytes (iPSC-CM) provide a powerful platform for disease modeling and drug development in vitro. Traditionally, electrophysiological methods or fluorescent dyes (e.g. calcium) have been used in their functional characterization. Recently, video microscopy has enabled non-invasive analysis of CM contractile motion. Simultaneous assessments of motion and calcium transients have not been generally conducted, as motion detection methods are affected by changing pixel intensities in calcium imaging. Here, we present for the first time a protocol for simultaneous video-based measurement of contraction and calcium with fluorescent dye Fluo-4 videos without corrections, providing data on both ionic and mechanic activity. The method and its accuracy are assessed by measuring the effect of fluorescence and background light on transient widths and contraction velocity amplitudes. We demonstrate the method by showing the contraction-calcium relation and measuring the transient time intervals in catecholaminergic polymorphic ventricular tachycardia patient specific iPSC-CMs and healthy controls. Our validation shows that the simultaneous method provides comparable data to combined individual measurements, providing a new tool for measuring CM biomechanics and calcium simultaneously. Our results with calcium sensitive dyes suggest the method could be expanded to use with other fluorescent reporters as well.
诱导多能干细胞衍生的心肌细胞（iPSC-CM）为体外疾病建模和药物开发提供了强大的平台。传统上，电生理学方法或荧光染料（如钙）已被用于其功能表征。最近，视频显微镜使 CM 收缩运动的非侵入性分析成为可能。由于运动检测方法受钙成像中像素强度变化的影响，因此通常不会同时进行运动和钙瞬变的评估。在这里，我们首次提出了一种无需校正即可同时基于视频测量荧光染料 Fluo-4 视频的收缩和钙的方案，提供了离子和力学活动的数据。该方法及其准确性通过测量荧光和背景光对瞬变宽度和收缩速度幅度的影响来评估。我们通过显示儿茶酚胺多形性室性心动过速患者特异性 iPSC-CM 和健康对照的收缩-钙关系并测量瞬变时间间隔来证明该方法。我们的验证表明，同时测量方法可提供与组合单独测量相当的数据，为同时测量 CM 生物力学和钙提供了一种新工具。我们使用钙敏感染料的结果表明，该方法也可以扩展到使用其他荧光报告基因。
