I. Department of Medicine (Cardiology), Klinikum rechts der Isar, Technische Universität München, Munich 81675, Germany.
Institute for Molecular Cell Biology, Medical Faculty, University Homburg/Saar, Universität des Saarlandes, Homburg/Saar 66421, Germany.
Eur Heart J. 2017 Jan 21;38(4):292-301. doi: 10.1093/eurheartj/ehw189.
Cardiomyocytes (CMs) generated from human induced pluripotent stem cells (hiPSCs) are increasingly used in disease modelling and drug evaluation. However, they are typically a heterogeneous mix of ventricular-, atrial-, and nodal-like cells based on action potentials (APs) and gene expression. This heterogeneity and the paucity of methods for high-throughput functional phenotyping hinder the full exploitation of their potential. We aimed at developing a method for rapid, sequential, and subtype-specific phenotyping of hiPSC-CMs with respect to AP morphology and single-cell arrhythmias.
We used cardiac lineage-specific promoters to drive the expression of a voltage-sensitive fluorescent protein (VSFP-CR) in hiPSC-CMs, enabling subtype-specific optical AP recordings. In a patient-specific hiPSC model of long-QT syndrome type 1, AP prolongation and frequent early afterdepolarizations were evident in mutant ventricular- and atrial like, but not in nodal-like hiPSC-CMs compared with their isogenic controls, consistent with the selective expression of the disease-causing gene. Furthermore, we demonstrate the feasibility of sequentially probing a cell over several days to investigate genetic rescue of the disease phenotype and to discern CM subtype-specific drug effects.
By combining a genetically encoded membrane voltage sensor with promoters that drive its expression in the major subtypes of hiPSC-CMs, we developed a convenient system for disease modelling and drug evaluation in the relevant cell type, which has the potential to advance the emerging utility of hiPSCs in cardiovascular medicine.
源自人诱导多能干细胞（hiPSC）的心肌细胞（CMs）越来越多地用于疾病建模和药物评估。然而，基于动作电位（APs）和基因表达，它们通常是心室、心房和结样细胞的异质混合物。这种异质性以及高通量功能表型分析方法的缺乏阻碍了其潜力的充分发挥。我们旨在开发一种用于快速、连续和亚型特异性表型分析 hiPSC-CMs 的方法，以研究其 AP 形态和单细胞心律失常。
我们使用心脏谱系特异性启动子驱动 hiPSC-CMs 中电压敏感荧光蛋白（VSFP-CR）的表达，从而实现亚型特异性的光学 AP 记录。在 1 型长 QT 综合征的患者特异性 hiPSC 模型中，与同基因对照相比，突变型心室和心房样 hiPSC-CMs 中可见 AP 延长和频繁的早期后除极，而结样 hiPSC-CMs 中则没有，这与致病基因的选择性表达一致。此外，我们还证明了在几天内连续探测一个细胞以研究疾病表型的基因矫正并辨别 CM 亚型特异性药物作用的可行性。
通过将基因编码的膜电压传感器与在 hiPSC-CMs 的主要亚型中驱动其表达的启动子相结合，我们开发了一种方便的疾病建模和药物评估系统，该系统有可能推进 hiPSC 在心血管医学中的新兴应用。
