A Novel Adult Three-Dimensional (3D) Human Cardiac Organoid Model of Cardiac Fibrosis in CKD

Background Cardiac fibrosis is a major complication of chronic kidney disease (CKD) lacking effective targeted therapies. Progress has been limited by animal models that incompletely recapitulate human cardiorenal pathology and by conventional 2D cultures that lack tissue architecture and physiological maturity. Recent initiatives from the NIH and the FDA promote modernization of drug development using physiologically relevant human systems. We developed a human cardiac organoid model that recapitulates CKD-associated cardiac fibrosis with advanced structural and functional maturation. Methods Human iPSC-derived embryoid bodies were sequentially exposed to Activin A, BMP4, and CHIR to induce cardiac specification and generate fetal cardiac organoids. After induction, control fetal organoids were maintained in RPMI+B27 medium for 30 days. Parallel organoids underwent metabolic maturation to generate adult-like cardiac organoids via AMPK and ERR activation (DCN, EX229, DY131), glucose reduction to 1 mM, and BSA–palmitate supplementation to promote adult-like oxidative phosphorylation. CKD-associated fibrogenesis was induced by mineral stress (2 mM Ca 2+ , 3.8 mM PO 4 3- , Ca/Pi) for 3 weeks. Organoids underwent structural, metabolic, and electrophysiological interrogation. Results Matured organoids exhibited increased adult cardiac structural markers including *MYH7/6* ratio, *BIN1* , *TNNI3* , and *MYL2v* (all *P* <0.0001). Light-sheet microscopy revealed development of sophisticated internal chambers, atrioventricular specification and complex vasculature. Organoids also developed well-organized multi-lineage cardiac cell types as confirmed by pan-cardiomyocyte marker positivity. Metabolic maturation towards oxidative phosphorylation was confirmed by increased *COX7C* and *COX6A2* expression ( *P* <0.0001). Ca/Pi exposure induced fibrogenesis with elevated COL1A1 (P<0.05), TGF-β (P<0.001), and α-SMA (P<0.01). Microelectrode array analysis demonstrated reduced beating rate, shortened field potential duration, and decreased spike amplitude with Ca/Pi exposure (all P<0.0001). Conclusion This reproducible adult human cardiac organoid model recapitulates structural, metabolic, and electrophysiological features of CKD-associated cardiac fibrosis. This versatile platform enables mechanistic discovery and scalable drug screening to accelerate therapeutic development for cardiac complications in CKD.