Open‐source bioreactor delivers electrical and perfusion stimulation supporting <scp>3D</scp> cardiac engineered tissue maturation

Abstract Cardiac tissue engineering requires control over physical stimuli, such as mechanical and electrical, to promote the maturation and functionality of cardiomyocytes. While perfusion bioreactors and electrical stimulation systems have been employed before, their synergistic impact, specifically when using direct perfusion in soft hydrogel environments, remains underexplored. We developed a cost‐effective, modified perfusion bioreactor that is commercially available, to integrate electrical stimulation to support the culture of fibrin‐based cardiac constructs. The system delivers continuous unidirectional flow (0.3 mL/min) and controlled electrical pulses (3 V/cm, 1 Hz), with validated flow and field uniformity via computational fluid dynamics and finite element analysis simulations. Neonatal rat cardiac cell‐based constructs were cultured either under static or perfusion conditions in presence or not of electrical stimulation. Cellular outcomes were evaluated by immunofluorescence, gene expression, and live‐cell functional analyses. Perfusion significantly improved cell retention and cardiomyocyte yield, while electrical stimulation promoted cardiomyocyte elongation, sarcomere organization, and maturation. The combination of perfusion and electrical stimulation led to the highest proportion of mature cardiomyocytes (51.4%), significantly outperforming perfusion alone (12.3%), static combined with electrical stimulation (22.2%), and static alone (7.5%). A reduced fibroblast activation was observed, along with enhanced tissue remodeling, as shown by increased extracellular matrix density and upregulation of remodeling genes. Functionally, constructs under perfusion with electrical stimulation exhibited the lowest excitation thresholds, highest maximum capture rates, and the greatest contraction displacement (2.2 ± 0.49 V, 4.2 ± 0.71 Hz, 13.35 ± 3.6 μm respectively), confirming superior functional performance. Our integrated bioreactor system enables efficient culture of soft 3D cardiac tissues and demonstrates that combining perfusion with electrical stimulation synergistically enhances cardiomyocyte maturation, construct remodeling, and functional performance. Beyond being a promising tool, this platform represents a powerful and scalable solution for cardiac tissue engineering, with potential for applications in disease modeling, drug screening, and regenerative medicine.