Physiological calcium combined with electrical pacing accelerates maturation of human engineered heart tissue

Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have wide potential application in basic research, drug discovery, and regenerative medicine, but functional maturation remains challenging. Here, we present a method whereby maturation of hiPSC-CMs can be accelerated by simultaneous application of physiological Ca²⁺ and frequency-ramped electrical pacing in culture. This combination produces positive force-frequency behavior, physiological twitch kinetics, robust β-adrenergic response, improved Ca²⁺ handling, and cardiac troponin I expression within 25 days. This study provides insights into the role of Ca²⁺ in hiPSC-CM maturation and offers a scalable platform for translational and clinical research.