Saikosaponin D Alleviates Liver Ischemia–Reperfusion Injury by Inhibiting Mitophagy‐Associated Ferroptosis via the STAT3 / PINK1 Pathway

Liver ischemia-reperfusion injury (LIRI) may cause severe complications in liver surgery, primarily involving damage to hepatocytes and endothelial cells, as well as an inflammatory response from immune cells. Saikosaponin D, a plant widely used in traditional Chinese medicine for treating inflammation, fever, and liver diseases. This research was intended to elucidate the function and mechanism of Saikosaponin D (SSD) in ameliorating liver ischemia-reperfusion injury. The effect of SSD on liver ischaemia-reperfusion injury was studied in mice following oral gavage administration. Liver injury was assessed using haematoxylin and eosin (H&E) staining and serum biochemical analysis. To evaluate ferroptosis levels, various methods were used, including reactive oxygen species (ROS), lipid peroxidation, glutathione (GSH) and oxidized glutathione (GSSG) levels, Malondialdehyde (MDA), Fe2+ indicator FerroOrange, Western blotting and transmission electron microscopy (TEM). The effect of Saikosaponin D in combination with signal transducer and activator of transcription 3 (STAT3) overexpression or PTEN-induced putative kinase 1 (PINK1) knockdown on ferroptosis in a hypoxia-reoxygenation model was studied in vitro. SSD alleviated liver ischemia-reperfusion injury by activating mitophagy and inhibiting ferroptosis. The mitochondrial autophagy inhibitor Mdivi-1 reversed the pharmacodynamic effects of SSD. Importantly, in hypoxia-reoxygenation of AML12 cells, STAT3 overexpression inhibited PINK1-associated mitophagy, aggravated Fe2+ accumulation, lipid peroxidation and malondialdehyde levels, accelerated GSH depletion, aggravated the downregulation of Glutathione Peroxidase 4 (GPX4) and Solute carrier family 7 membrane 11 (SLC7A11) and upregulation of Transferrin receptor (TFR), while PINK1 silencing produced similar results. The inhibitory effect of SSD on ferroptosis in a hypoxia-reoxygenation model was counteracted by STAT3 overexpression and PINK1 silencing. SSD protects the liver against ischemia-reperfusion injury by inhibiting ferroptosis through STAT3/PINK1/PARKIN-mediated mitophagy, highlighting its potential as a therapeutic agent in liver ischemia-reperfusion injury treatment.