Aryl Hydrocarbon Receptor Activation Prevents Lung Ischemia Reperfusion Injury by Inhibiting ROS-triggered Ferroptosis in Mice.

INTRODUCTION

Lung ischemia-reperfusion injury (LIRI) is a serious complication of lung transplantation that causes respiratory distress and is associated with high mortality. Ferroptosis, a novel form of programmed cell death, contributes to the pathogenesis of LIRI. The Aryl Hydrocarbon Receptor (AhR) is a ligand-activated transcription factor that regulates a variety of physiological functions. However, the role and mechanism of AhR in ferroptosis during LIRI after lung transplantation remain to be further investigated.

MATERIALS AND

METHODS

AhR signaling and ferroptosis were examined in a mouse LIRI model and a pulmonary vascular endothelial cell line following hypoxiareoxygenation (H/R) culture. The severity of LIRI was assessed based on histological architecture and cytokine levels. Cell viability and damage were evaluated using the CCK-8 and LDH assays. Ferroptosis biomarkers, including Fe²⁷, malondialdehyde (MDA), reactive oxygen species (ROS), and glutathione (GSH), were measured using respective assay kits. PTGS2 (Prostaglandin-Endoperoxide Synthase 2), SLC7A11 (Solute Carrier Family 7 Member 11), and GPX4 (Glutathione Peroxidase 4) were analyzed by western blot. In vitro, lipid peroxidation was assessed by flow cytometry after DCFH-DA labeling. Transmission electron microscopy was used to observe mitochondrial ultrastructure.

RESULTS

Enhanced transcription of AhR and its downstream targets, CYP1A1 (Cytochrome P450 1A1) and CYP1B1 (Cytochrome P450 1B1), was observed. AhR deficiency exacerbated LIRI-induced ferroptosis and lung injury, whereas AhR activation mitigated lung injury and ferroptosis. H/R-induced cell injury and ferroptosis were alleviated by AhR activation. Increased ROS formation, primarily mitochondrial ROS (Mito-ROS), was triggered by AhR ablation or antagonism, and ROS inhibition subsequently reduced cell damage and ferroptosis.

DISCUSSION: Our findings indicate that AhR activation functions upstream by directly inhibiting excessive ROS accumulation triggered by I/R. This reduction in cellular oxidative stress prevents the lethal lipid peroxidation that drives ferroptosis.

CONCLUSION

This study demonstrates the protective effect of AhR activation in LIRI by inhibiting ROS-induced ferroptosis, suggesting it may be a promising target for clinical prevention of LIRI.