PFASs Disrupt Airway Epithelial Homeostasis in Human Organoids: Evidence Relevant to Obstructive Airway Disease

Per- and polyfluoroalkyl substances (PFASs) are implicated in respiratory diseases, yet systematic toxicity data and mechanistic understanding for most compounds remain critically lacking. To address this gap, we established human 3D airway organoids that recapitulate native epithelial architecture and function to assess the respiratory toxicity of 10 PFAS compounds. At 0.1 μM, a concentration relevant to high-exposure, PFASs induced a cascade of effects, including mitochondrial dysfunction, barrier disruption, and inflammation. These events converged on epithelial differentiation imbalance, characterized by a 21-66% reduction in ciliated cells marker FOXJ1 and a 1.2-2.3-fold upregulation of goblet cells marker MUC5AC. Multiend point Toxicological Priority Index ranking revealed that emerging substitutes, particularly sodium p-perfluorinated noneoxybenzenesulfonate (OBS) and fluorotelomer alcohols (FTOHs), exhibited higher toxicity potency than legacy compounds. In vivo validation confirmed perfluorooctanesulfonic acid (PFOS)-induced airway obstruction and mucus hypersecretion, supporting the in vivo relevance of organoid-derived airway injury phenotypes. Single-cell transcriptomics implicated Notch-related signaling in PFOS-associated differentiation disturbance, supported by inhibitor intervention. These findings provide functional and mechanistic evidence relevant to obstructive airway disease; establish an adverse outcome pathway for PFASs' respiratory toxicity; and provide a sensitive, predictive platform for hazard identification and regulatory prioritization of emerging PFASs.