Establishing a Single-cell and High-throughput Phenotypical Platform to Identify Novel Arrhythmia-Causing Genes

Here we describe an original phenotypical platform to identify novel regulators of cardiac rhythm and arrhythmia-causing genes in hPSC-derived atrial-like cardiomyocytes (hPSC-ACM).The system applies well-characterized atrial subtype cardiomyocytes generated from Id1-induced cardiogenic mesoderm progenitors. The platform integrates automated high-throughput/high-content kinetic imaging system with fluorescence quantification and single-cell resolution trace analysis algorithm to retrieve physiological metrics of hPSC-ACMs biology. This generic platform enables the high-throughput phenotypical evaluation of functional genomics and small molecules on cardiac physiological parameters in both healthy or disease contexts. The system is also functionally validated with genes previously associated with atrial fibrillation (AF) and confirmed the occurrence of arrhythmia-like activity in hPSC-ACMs induced by siRNA-mediated knockdown of the genes along with AF-associated perturbagens. We then modeled and categorized different arrhythmia-subtypes arise from distinct molecular events. Overall, the hPSC-ACM differentiation protocol provides a robust resource to study atrial-specific cardiac diseases and development; the platform and single-cell analysis generate higher resolutions of information without sacrificing the throughout; and the modeling and characterization of arrhythmia-like activity shed light upon this unique approach to identify novel regulators of cardiac rhythm and arrhythmia-causing genes in an atrial-specific manner