Hyperoxia and iron homeostasis in developing human airway smooth muscle

Premature infants exposed to supplemental oxygen (O₂) are at increased risk of developing airway diseases such as asthma, hyperoxic lung injury (HLI), and bronchopulmonary dysplasia (BPD). Therefore, it is important to understand how O₂ detrimentally impacts developing airways. Previous studies found that severe (80%-90%) O₂ exposure increases reactive oxygen species (ROS) and lipid peroxidation, inducing ferroptosis in models of HLI. However, the impact of clinically relevant moderate (<60%) O₂ exposure is less understood. Recognizing the importance of smooth muscle in airway dysfunction, the present study uses human fetal airway smooth muscle (fASM) as a model to investigate whether hyperoxia contributes to the establishment of a ferroptotic phenotype. fASM pretreated with or without deferoxamine (DFO; 100 µM) or ferrostatin (Fer-1; 10 µM) was exposed for 48 h to normoxia (21% O₂) versus moderate to severe hyperoxia (50%, 70%, or 90% O₂). The effects of hyperoxia on antioxidant systems, iron metabolism, and lipid peroxidation, and the alleviating effect of DFO or Fer-1 were examined. Moderate hyperoxia impaired antioxidant systems involved in preventing ferroptosis and dysregulated iron metabolism. Interestingly, only severe hyperoxia (90% O₂) induced negative effects on downstream mechanisms involving early onset of ferroptosis such as increased labile iron and lipid peroxidation. DFO and Fer-1 showed no rescue effect on antioxidant systems. However, DFO decreased cytosolic iron, and Fer-1 decreased lipid peroxidation byproducts. Together, these data highlight the impact of supplemental oxygen on premature airways and introduce the concept of a dose-dependent effect of hyperoxia in the context of iron metabolism, lipid peroxidation, and ultimately ferroptosis.NEW & NOTEWORTHY Recognizing that antioxidant systems are impaired in the airways of premature infants, we used human fetal airway cells to explore the impact of oxygen on iron regulation and iron-mediated cell death (ferroptosis). We find moderate hyperoxia impairs antioxidant systems that prevent ferroptosis and dysregulates iron metabolism, while severe hyperoxia has a negative effect on mechanisms driving the early onset of ferroptosis. Inhibitors of ferroptosis decrease iron and lipid peroxidation, demonstrating links between oxygen and iron regulation in developing airways.