Human heart-macrophage assembloids mimic immune-cardiac interactions and enable arrhythmia disease modeling

Yolk-sac-derived embryonic cardiac tissue-resident macrophages (TRMPs) colonize the heart early in development and are essential for proper heart development, supporting tissue remodeling, angiogenesis, electrical conduction, efferocytosis, and immune regulation. We present here a human heart-macrophage assembloid (hHMA) model by integrating autologous human pluripotent stem cell (hPSC)-derived embryonic monocytes into heart organoids to generate physiologically relevant TRMPs that persist long-term and contribute to cardiogenesis. Using single-cell transcriptomics, live imaging, and proteomics, we demonstrate that TRMPs modulate cardiac paracrine signaling, perform efferocytosis, and regulate extracellular matrix remodeling and electrical conduction. In a proof-of-concept maturated hHMA model of chronic inflammation, TRMPs adopt pro-inflammatory phenotypes that promote arrhythmogenic activity, consistent with atrial fibrillation through activation of the NOD-like receptor pyrin domain-containing 3 (NLRP3) inflammasome. This system enables detailed mechanistic studies of immune-cardiac interactions and provides a powerful in vitro platform for modeling human heart development and inflammation-driven arrhythmias. • Human heart-macrophage assembloids model embryonic cardiac macrophage integration • Tissue-resident macrophages regulate heart morphogenesis and electrical conduction • Macrophages communicate with cardiac cells via EVs and efferocytosis pathways • Chronic inflammasome activation in matured hHMAs models arrhythmia phenotypes O’Hern et al. present a novel human heart-macrophage assembloid platform generated by integrating hPSC-derived embryonic monocytes into heart organoids. These tissue-resident macrophages regulate cardiac development and conduction and, under chronic inflammation, recapitulate atrial fibrillation features, offering a powerful model to study immune-cardiac interactions and arrhythmia mechanisms.