Hirsutine mitigates high glucose-induced cell injury via autophagy activation and NRF2/GPX4-mediated ferroptosis inhibition.

Hirsutine, a potent indole alkaloid extracted from Uncaria rhynchophylla (Miq.) Miq. ex Havil., family Rubiaceae., exhibits antioxidant and cytoprotective activities. However, its potential role in diabetic nephropathy (DN) remains unclear. DN is the leading cause of end-stage renal disease, and podocyte injury is a key event in the progression of DN. MPC5 cells were exposed to 30 mM HG with or without hirsutine (5-20 μM). Hirsutine significantly improved cell viability. Hirsutine significantly improved cell viability, accompanied by a marked reduction in intracellular ROS levels and apoptosis, collectively indicating that hirsutine exerts a pronounced protective effect and effectively enhances podocyte viability under hyperglycemic stress. Autophagy-related proteins LC3 and p62 were examined by Western blotting and immunofluorescence, showing that hirsutine significantly modulated autophagy-related protein expression under hyperglycemic conditions. Apoptosis and ROS formation were evaluated using the Western blotting, and immunofluorescence, indicating that hirsutine attenuated high glucose -induced apoptosis and oxidative stress. LC3 and p62 expression patterns were altered in a manner consistent with autophagy-associated changes. Hirsutine increased the markers of NRF2 pathway activation and also reduced the biochemical signatures consistent with ferroptosis.This study provides new evidence that hirsutine protects against HG-induced cell injury by simultaneously modulating apoptosis, autophagy and ferroptosis, highlighting its potential as a candidate for future DN therapy. However, these results are limited to an in vitro model and lack direct autophagic flux assessment and genetic validation of NRF2 dependency, which warrant further investigation.