Hypoxic Preconditioning Prevents the Neonatal Lung Against Hyperoxia-Induced Ferroptosis Through the Nrf2–CBS Axis

Aims: Hyperoxia-induced oxidative injury contributes to ferroptosis and abnormal lung remodeling in preterm infants receiving oxygen therapy. We previously demonstrated that postnatal hypoxic preconditioning (HP) reduces hyperoxia-induced lung injury in a neonatal mouse model. In the present study, we investigated whether HP protects the neonatal lung against ferroptosis through a nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent mechanism involving transsulfuration and glutathione (GSH) metabolism. Results: In a neonatal mouse model, exposure to HP (FiO 2 = 0.14) prior to hyperoxic challenge (FiO 2 = 1.0) markedly reduced lipid peroxidation, pulmonary ferrous iron accumulation, ferroptosis-associated lipid remodeling, inflammation, and early fibrotic changes in the lung. Mechanistically, HP enhanced antioxidant defenses by increasing Nrf2 activity and promoting transsulfuration-dependent GSH synthesis through upregulation of cystathionine β-synthase (CBS) and xCT, leading to preservation of GSH homeostasis. To determine the functional role of Nrf2, Nrf2 knockout mice were subjected to the same HP/hyperoxia protocol. Nrf2 deficiency abolished the protective effects induced by HP, resulting in increased pulmonary ferrous iron levels, depletion of downstream transsulfuration metabolites and GSH, increased lipid peroxidation, reduced CBS and GPX4 expression, and persistent induction of fibrosis-associated genes. Innovation: This study demonstrates that postnatal HP protects the neonatal lung against hyperoxia-induced ferroptosis through an Nrf2-dependent transsulfuration program. Our findings identify the Nrf2–CBS axis as a key regulator of GSH homeostasis, ferroptosis resistance, and fibrotic remodeling in the developing lung. Conclusion: HP effectively attenuates hyperoxia-induced oxidative stress, ferroptosis, and profibrotic remodeling in neonatal lungs by preserving Nrf2-dependent GSH metabolism and redox homeostasis. These findings support HP as a promising therapeutic strategy to mitigate oxygen-induced pulmonary injury in preterm infants. Antioxid. Redox Signal. 00, 000–000.