Astragaloside IV attenuates H₂O₂-induced oxidative stress and ferroptosis-like injury in HBZY-1 mesangial cells via the Nrf2/HO-1 signaling pathway

Ferroptosis has recently been demonstrated to play a critical role in the initiation and progression of diabetic nephropathy (DN). Emerging evidence indicates that ferroptosis is tightly regulated by the Nrf2/HO-1 signaling pathway. Astragaloside IV (As-IV) exhibits potent antioxidant effects in various disease models. However, whether As-IV inhibits ferroptosis by modulating the Nrf2/HO-1 pathway remains unclear. An in vitro model of oxidative stress-induced ferroptosis-like injury was established by exposing HBZY-1 mesangial cells to H₂O₂ (400 µmol/L) for 4 h. Intracellular ferrous ion (Fe²⁺) levels were measured using a specific fluorescent probe. The expression of ferroptosis-related proteins was detected by Western blotting. Reactive oxygen species (ROS) production was assessed by flow cytometry. Superoxide dismutase (SOD) activity and malondialdehyde (MDA) content were determined using commercial assay kits. The expression levels of Nrf2/HO-1 pathway-related proteins were detected by Western blotting. H₂O₂ significantly increased ROS levels and MDA content, decreased SOD activity, downregulated the expression of Nrf2, HO-1, GPX4, and NQO1, and induced changes in iron metabolism and lipid peroxidation consistent with ferroptosis. As-IV attenuated H₂O₂-induced oxidative stress and ferroptosis-like changes in vitro. Further mechanistic investigation revealed that the Nrf2 inhibitor all-trans retinoic acid (ATRA) reversed the aforementioned effects of As-IV, suggesting that As-IV acts through activation of the Nrf2/HO-1 signaling pathway. The results from the present study suggest that As-IV inhibits ferroptosis, at least in part, through activation of the Nrf2 pathway, with HO-1 identified as a potential key downstream mediator.