Rationale: Bronchopulmonary dysplasia (BPD) is a leading complication of preterm birth that affects infants born in the saccular stage of lung development at,32 weeks of gestation. Although the mechanisms driving BPD remain uncertain, exposure to hyperoxia is thought to contribute to disease pathogenesis. Objectives: To determine the effects of hyperoxia on epithelial-mesenchymal interactions and to define the mediators of activated Wnt/b-catenin signaling after hyperoxia injury. Methods: Three hyperoxia models were used: A three-dimensional organotypic coculture using primary human lung cells, precision-cut lung slices (PCLS), and a murine in vivo hyperoxia model. Comparisons of normoxia- and hyperoxia-exposed samples were made by real-time quantitative PCR, RNA in situ hybridization, quantitative confocal microscopy, and lung morphometry. Measurements and Main Results: Examination of an array of Wnt ligands in the three-dimensional organotypic coculture revealed increased mesenchymal expression of WNT5A. Inhibition of Wnt5A abrogated the BPD transcriptomic phenotype induced by hyperoxia. In the PCLS model, Wnt5A inhibition improved alveolarization following hyperoxia exposure, and treatment with recombinant Wnt5a reproduced features of the BPD phenotype in PCLS cultured in normoxic conditions. Chemical inhibition of NF-kB with BAY11-7082 reduced Wnt5a expression in the PCLS hyperoxia model and in vivo mouse hyperoxia model, with improved alveolarization in the PCLS model. Conclusions: Increased mesenchymal Wnt5A during saccular-stage hyperoxia injury contributes to the impaired alveolarization and septal thickening observed in BPD. Precise targeting of Wnt5A may represent a potential therapeutic strategy for the treatment of BPD.