Multicellular human cardiac organoids with real-time calcium imaging enable functional modeling of heart failure and myocardial infarction.

Cardiovascular diseases remain leading causes of mortality, yet progress in therapy development is limited by inadequate human-relevant models. Here, we develop a simplified, multicellular human iPSC-derived cardiac organoid system that enables functional and translational disease modeling. Using a streamlined two-component differentiation strategy, we generate robust beating cardiomyocytes and assemble three- and four-cell-type organoids incorporating endothelial cells, fibroblasts, and macrophages. Integration of a genetically encoded calcium reporter allows real-time, non-invasive functional assessment. Multicellular organoids exhibit enhanced maturation and viability, with macrophages contributing to improved functional properties. Optimized oxygen-permeable culture further enhances organoid performance. This platform recapitulates key features of overnutrition, heart failure, and myocardial infarction, including altered contractility, calcium dynamics, and biomarker expression. Together, this cost-effective and scalable system provides a physiologically relevant platform for studying cardiac disease mechanisms, drug responses, and cardiotoxicity.