Mechanical and Electrical Phenotype of hiPSC‐Cardiomyocytes on Fibronectin‐Based Hydrogels

A major challenge in cardiac research is the limited translatability of drug screening and toxicity assays due to the use of in vitro models that poorly mimic the native cardiac environment. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) offer a promising route forward, but conventional 2D culture on rigid substrates hinders their functional maturation and predictive accuracy. This study addresses this problem by investigating the effect of hybrid fibronectin-based hydrogels with tunable stiffness on the mechanical and electrical properties of hiPSC-CMs. We engineered hydrogels with stiffness mimicking the lowest range of neonatal heart tissue stiffness (2-4 kPa) and compared hiPSC-CM behavior on these substrates to that on standard fibronectin-coated glass. Our results demonstrate that hydrogel culture promotes more uniform and stable cardiomyocyte contractions, as evidenced by increased single peak percentages and altered contraction duration. Electrophysiological analysis revealed that hydrogel stiffness influences action potential duration and signal amplitude. Furthermore, hiPSC-CMs on hydrogels exhibited enhanced cell-matrix and cell-cell adhesion, indicating improved structural and functional connectivity. Drug testing with known cardioactive compounds, including isoproterenol and nifedipine, revealed distinct differences in drug responses between hydrogel and glass cultures, suggesting that hydrogels provide a more physiologically relevant platform for assessing drug effects. This work highlights the potential of engineered hydrogel substrates to enhance the functional maturity and predictive accuracy of hiPSC-CMs for cardiac research and drug development.