Rationale Human induced pluripotent stem cell derived cardiomyocytes (hiPSC-CM) are increasingly being used for modeling heart disease and are under development for regeneration of the injured heart. However, incomplete structural and functional maturation of hiPSC-CM including lack of t-tubules, immature excitation-contraction (EC) coupling, and inefficient Ca-induced Ca release (CICR) remain major limitations. Objective Thyroid and glucocorticoid hormones are critical for heart maturation. We hypothesized that their addition to standard protocols would promote t-tubule development and mature EC coupling of hiPSC-CM when cultured on extracellular matrix with physiological stiffness (Matrigel mattress). Methods and Results HiPSC-CM were generated using a standard chemical differentiation method supplemented with triiodo-L-thyronine (T3) and/or dexamethasone (Dex) during days 16–30 followed by single-cell culture for 5 days on Matrigel mattress. HiPSC-CM treated with T3+Dex, but not with either T3 or Dex alone, developed an extensive t-tubule network. Notably, Matrigel mattress was necessary for t-tubule formation. Compared to adult human ventricular CM, t-tubules in T3+Dex-treated hiPSC-CM were less organized and had more longitudinal elements. Confocal line scans demonstrated spatially and temporally uniform Ca release that is characteristic of EC coupling in the heart ventricle. T3+Dex enhanced elementary Ca release measured by Ca sparks as well as promoted ryanodine receptor (RyR2) structural organization. Simultaneous measurements of L-type Ca current and intracellular Ca release confirmed enhanced functional coupling between L-type Ca channels and RyR2 in T3+Dex cells. Conclusions Our results suggest a permissive role of combined thyroid and glucocorticoid hormones during the cardiac differentiation process which, when coupled with further maturation on Matrigel mattress, is sufficient for t-tubule development, enhanced CICR, and more ventricular-like EC coupling. This new hormone maturation method could advance the utility of hiPSC-CM for disease modeling and cell-based therapy.

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Heart transplant (HTx) recipients usually have reduced exercise capacity with reported VO 2peak levels of 50-70% predicted value. Our hypothesis was that highintensity interval training (HIIT) is an applicable and safe form of exercise in HTx recipients and that it would markedly improve VO 2peak.Secondarily, we wanted to evaluate central and peripheral mechanisms behind a potential VO 2peak increase. Forty-eight clinically stable HTx recipients >18 years old and 1-8 years after HTx underwent maximal exercise testing on a treadmill and were randomized to either exercise group (a 1-year HIIT-program) or control group (usual care). The mean ± SD age was 51 ± 16 years, 71% were male and time from HTx was 4.1 ± 2.2 years. The mean VO 2peak difference between groups at follow-up was 3.6 [2.0, 5.2] mL/kg/min (p < 0.001). The exercise group had 89.0 ± 17.5% of predicted VO 2peak versus 82.5 ± 20.0 in the control group (p < 0.001). There were no changes in cardiac function measured by echocardiography. We have demonstrated that a long-term, partly supervised and community-based HIIT-program is an applicable, effective and safe way to improve VO 2peak , muscular exercise capacity and general health in HTx recipients. The results indicate that HIIT should be more frequently used among stable HTx recipients in the future.Key words: Aerobic exercise, chronotropic response, heart transplantation, maximum oxygen uptake, muscle strength, VO 2peak Abbreviations: % HR max , percent of age-predicted maximum heart rate; AT, anaerobic threshold (ventilatory threshold); BIA, bioelectrical impedance analysis; CG, control group; CO, cardiac output; CRI, chronotropic response index; CRP, C-reactive protein; DXA, dual-emission X-ray absorptiometry; EG, exercise group; eGFR, estimated glomerular filtration rate; Hb, hemoglobin; HF, heart failure; HIIT, high-intensity interval training; HR, heart rate; HR max , maximum heart rate; HRQoL, health-related quality of life; HTx, heart transplant; J, Joule; LV, left ventricle; LVe', left ventricle early diastolic mitral annular velocity; LVEF, left ventricle ejection fraction; Nm, Newtonmeter; NT-proBNP, N-terminal prohormone of brain natriuretic peptide; RER, respiratory exchange ratio; RPE, rated perceived exertion; VAS scale, visual analog scale; VE max , maximum ventilation; VO 2peak , peak oxygen uptake.