Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) can model heritable arrhythmias to personalize therapies for individual patients. Although atrial fibrillation (AF) is a leading cause of cardiovascular morbidity and mortality, current platforms to generate iPSC-atrial(a)CMs are inadequate for modeling AF. We applied a combinatorial engineering approach, which integrated multiple physiological cues including metabolic conditioning and electrical stimulation to generate mature iPSC-aCMs. Using the patient’s own atrial tissue as a gold standard benchmark, we assessed the electrophysiological, structural, metabolic and molecular maturation of iPSC-aCMs. Unbiased transcriptomic analysis and inference from gene regulatory networks identified key gene expression pathways and transcription factors mediating atrial development and maturation. Only mature human iPSC-aCMs generated from heritable AF patients carrying the non-ion channel gene (NPPA) mutation showed enhanced expression and function of a cardiac potassium channel and revealed mitochondrial electron transport chain dysfunction. Collectively, we propose that ion channel remodeling in conjunction with metabolic defects created an electrophysiological substrate for AF. Overall, our electro-metabolic maturation (EMM) approach generated mature human iPSC-aCMs that unmasked the underlying mechanism of the first non-ion channel gene (NPPA) that causes AF. We then synergistically combined our EMM protocol with two additional maturation approaches – micropatterning and co-culture with atrial fibroblasts (cMPCC) to generate the EMM-cMPCC protocol. The EMM-cMPCC approach improves recapitulation of native myocardium by incorporating crucial supporting cells in the atrium (atrial fibroblasts) and controlling the topographical geometry of iPSC-aCM maturation. We used EMM-cMPCC to investigate the underlying cellular mechanisms and pathophysiology by which loss-of-function TTN mutations generate a substrate for AF. Our maturation approach will not only elucidate the cellular mechanisms of AF but also identify new therapeutic targets that will pave the way for personalized therapy targeting cardiomyocyte biology and ion channel remodeling as current pharmacological therapy has limited efficacy
