Leonurine alleviates lung ischemia–reperfusion injury through suppression of ferroptosis via RORα in male mice

Lung ischemia-reperfusion injury (LIRI) is a complex pathological condition that significantly impairs clinical outcomes following lung transplantation and thoracic surgery. Leonurine (LEO), an alkaloid derived from Leonurus japonicus, which has known anti-inflammatory and antioxidant properties, has shown therapeutic potential in various oxidative stress-related diseases. However, the effects of LEO on LIRI and its underlying mechanisms remain unclear. In the present study, a murine model of LIRI was established using wild-type mice. LEO treatment significantly improved lung histopathology, reduced oxidative stress, decreased pulmonary edema, and enhanced survival. Bioinformatics analyses - including volcano plot, KEGG enrichment, and GSEA - identified ferroptosis as a key regulatory pathway. In vivo and in vitro assays (HE, 4-HNE, and DHE labeling; immunofluorescence; and immunoblotting) confirmed that LEO inhibited ferroptosis in lung tissue and in MLE-12 cells. Mechanistically, LEO upregulated the RORα/Nrf2/GPX4 axis, thereby reducing lipid peroxidation and iron overload, as validated by BODIPY581/591 C11 and FeRhoNox-1 staining. Moreover, RORα inhibition abolished the anti-ferroptotic effects of LEO, indicating that its protective function is RORα dependent. Molecular docking further supported a potential direct interaction between LEO and RORα. Collectively, LEO alleviates LIRI by inhibiting ferroptosis through activation of the RORα/Nrf2/GPX4 signaling pathway. These findings suggest that LEO may serve as a promising therapeutic agent for the treatment of LIRI.