Dual epithelial-mesenchymal induction of Sfrp1/Frzb constrains alveolar epithelial differentiation in neonatal hyperoxia-induced lung injury.

Abstract

Bronchopulmonary dysplasia (BPD) is a major complication of prematurity, characterized by impaired alveolar epithelium regeneration and long-term respiratory morbidity. The cellular basis of this defect and the underlying epithelial-mesenchymal signals remain unclear. To define alveolar type 2 (AT2) cell heterogeneity and regenerative capacity, we analyzed lineage-traced AT2 cells in neonatal mice exposed to hyperoxia (85% O2, postnatal days 1-14). We assessed cellular states and function by flow cytometry, transcriptomics, and organoid assays. During normal lung development, AT2 cells gradually segregated into two transcriptionally distinct subpopulations, TomHigh and TomLow. TomHigh cells exhibited canonical AT2 identity, lipid metabolism, and enhanced mitochondrial function, showing strong organoid-forming and differentiation potential. TomLow cells displayed metabolic restriction and limited regenerative capacity. Hyperoxia caused alveolar simplification, expanding TomLow cells at the expense of TomHigh cells. AT2 cells entered a proliferative state and became Krt8+ transitional cells, but RNA velocity and lineage analyses showed impaired maturation toward AT1 cells, leading to persistent AT1 deficiency. Hyperoxia induced Sfrp1 and Frzb expression in a rare but detectable Sftpc+Acta2+ AT2 subset, and more prominently in Pdgfra+ fibroblasts, which adopted a myofibroblast-like phenotype. Fibroblast-epithelial co-culture and fibroblast-specific Sfrp1/Frzb overexpression impaired organoid formation and reduced AT2-to-AT1 differentiation. These findings identify a fibroblast-dominated Sfrp1/Frzb-mediated niche that constrains AT2 differentiation into AT1 cells, providing a cellular basis for impaired alveolar regeneration in BPD and suggesting potential therapeutic targets.