Supplementary MaterialsVideo_1. cell heterogeneity under high regularity stimuli undergo conduction disturbance utilizing an two-dimensional (2D) monolayer preparation consisting of atrial-like CMs derived from human induced pluripotent stem cells (hiPSCs) and atrial fibroblasts (Fbs). We induced hiPSCs into atrial-like CMs using a directed cardiac differentiation protocol with the addition of all-retinoic acid (ATRA). The atrial-like hiPSC-derived CMs (hiPSC-CMs) and atrial Fbs were transferred in defined ratios (CMs/Fbs: 100%/0% or 70%/30%) on manually fabricated plates with or without geometrical patterning imitating the PVs-LA Abscisic Acid junction. High frequency field activation emulating repetitive ectopic foci originated in PVs were delivered, and the electrical propagation was assessed by optical mapping. We generated high purity CMs with or without the ATRA application. ATRA-treated hiPSC-CMs exhibited significantly higher atrial-specific properties by immunofluorescence staining, gene expression patterns, and optical action potential parameters than those of ATRA-untreated hiPSC-CMs. Electrical stimuli at a higher frequency preferentially induced impaired electrical conduction on atrial-like hiPSC-CMs monolayer preparations with an abrupt geometrical transition than on Abscisic Acid those with uniform geometry. Additionally, the application of human atrial Fbs to the geometrically patterned atrial-like hiPSC-CMs tended to further deteriorate the integrity of electrical conduction compared with those using the atrial-like hiPSC-CM alone preparations. Thus, geometrical narrow-to-wide patterning under high frequency stimuli preferentially jeopardized electrical conduction within atrial-like hiPSC-CM monolayers. Constituent cell heterogeneity represented by atrial Fbs also contributed to the further deterioration of conduction stability. neonatal rat cardiomyocyte monolayer as a result of source-to-sink mismatch (Rohr and Kucera, 1997; Rohr et al., 1997; Kondratyev et al., 2007; Auerbach et al., 2011). On the other hand, the electrophysiological and pharmacological properties of atrial-like CMs derived from hESCs or hiPSCs have recently been elucidated (Leyton-Mange et al., 2014; Laksman et al., 2017; Lee et al., 2017). Therefore, it is important to develop a platform of human Abscisic Acid atrial arrhythmias using human-based atrial CMs with geometrical characteristics of PVs or PVs-LA junction. In addition, constituent cell heterogeneity represented by non-CMs including fibroblasts (Fbs) may also deteriorate the stability of electrical conduction. However, to our knowledge, little is known regarding the effects of geometrical patterning and constituent cell heterogeneity on electrical conduction in atrial-like hESC/hiPSC-CM preparations. In the current study, we focused on an abrupt switch in 3D sleeve muscle mass thickness at PVs-LA interface, simplified it into a precipitous alteration in 2D strand width for the geometrical discontinuity. Furthermore, we utilized individual atrial Fbs as non-CMs for constituent cell heterogeneity. The goal of the present research was therefore to see whether geometrical patterning and constituent cell heterogeneity under electric stimuli at high regularity provoked impaired electric conduction, a prerequisite for the initiation of AF, within an 2D monolayer comprising atrial-like hiPSC-CMs and individual atrial Fbs. Outcomes Cardiac Differentiation From hiPSCs as well as the Purity of hiPSC-Derived CMs Using the technique as proven in Body 1, we differentiated the hiPSCs within a pluripotent condition through the mesoderm stage into CMs (Body 2ACC and Supplementary Number 1). Quantitative immunofluorescence analysis using a high-content imaging system demonstrated the TGFB2 protocol in the current study generated a high purity of CMs, positive for cardiac troponin T (cTnT), with no relation to the application of ATRA (cTnT positive cells; 99.6 0.3 (ATRA-untreated) vs. 99.5 0.3%.