Automated Video-Based Analysis of Contractility and Calcium Flux in Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes Cultured over Different Spatial Scales
Automated Video-Based Analysis of Contractility and Calcium Flux in Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes Cultured over Different Spatial Scales
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DOI：
10.1089/ten.tec.2014.0283
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发表时间：
2015-05-01
影响因子：
3
通讯作者：
Healy, Kevin E.
中科院分区：
文献类型：
--
作者：
Huebsch, Nathaniel;Loskill, Peter;Healy, Kevin E.
在体外，收缩运动是衡量心肌细胞健康的最简单的指标，但无偏量化是具有挑战性的。我们描述了一种快速的自动化方法，只需要标准的视频显微镜，就可以分析人类诱导的多能干细胞来源的心肌细胞(iPS-CM)的收缩性能。用于生成和过滤运动矢量的新算法与新开发的含有遗传编码钙指示剂GCaMP6f的等基因iPSC系列相结合，允许同时测量和分析钙通量和收缩性能之间的耦合。测试了这些算法在改善信噪比方面的相对性能。应用这些算法可以分析从单个细胞到三维结构的多个空间尺度上培养的iPS-CM的收缩能力。这一开源软件通过分析异丙肾上腺素在这些细胞中的反应得到了验证，可以应用于未来在组织工程背景下比较不同微环境中培养的iPS-CM的药物反应性的研究。
Contractile motion is the simplest metric of cardiomyocyte health in vitro, but unbiased quantification is challenging. We describe a rapid automated method, requiring only standard video microscopy, to analyze the contractility of human-induced pluripotent stem cell-derived cardiomyocytes (iPS-CM). New algorithms for generating and filtering motion vectors combined with a newly developed isogenic iPSC line harboring genetically encoded calcium indicator, GCaMP6f, allow simultaneous user-independent measurement and analysis of the coupling between calcium flux and contractility. The relative performance of these algorithms, in terms of improving signal to noise, was tested. Applying these algorithms allowed analysis of contractility in iPS-CM cultured over multiple spatial scales from single cells to three-dimensional constructs. This open source software was validated with analysis of isoproterenol response in these cells, and can be applied in future studies comparing the drug responsiveness of iPS-CM cultured in different microenvironments in the context of tissue engineering.