To investigate the mechanism by which Yiqi Huoxue Lishui Decoction ameliorates hepatic fibrosis (HF) in rats by inhibiting hepatocyte ferroptosis via activation of the nuclear factor-E2-related factor 2 (Nrf2)/glutathione peroxidase 4 (GPX4) signaling pathway.

Forty Sprague-Dawley (SD) rats were randomly divided into five groups (n =8 per group) : the control group (Control), model group (Model), traditional Chinese medicine group (TCM), traditional Chinese medicine + ferroptosis inhibitor group (TCM + Fer-1), and traditional Chinese medicine + ferroptosis inhibitor + autophagy inhibitor group (TCM + Fer-1 + 3-MA). The model was established by intraperitoneal injection of a 50% carbon tetrachloride (CCl₄) solution (1.5 mL/kg), administered twice weekly for 4 weeks. Treatment groups received their respective interventions: the Yiqi Huoxue Lishui Decoction was administered via oral gavage (2 mL/100 g), while the ferroptosis inhibitor Ferrostatin-1(Fer-1) (1 mg/kg) and the autophagy inhibitor 3-Methyladenine (3-MA) (15 mg/kg) were administered via intraperitoneal injection. All administrations were performed once daily for 4 weeks, while the other groups were given an equivalent volume of normal saline. Hepatic histopathological changes were evaluated using hematoxylin-eosin (HE) and Masson staining. Serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT), tumor necrosis factor-α (TNF-α), superoxide dismutase (SOD), malondialdehyde (MDA), and Fe²⁺ content in serum and liver tissue were measured. Western blot (WB) was employed to assess protein expression of GPX4, Nrf2, solute carrier family 7 member 11 (SLC7A11), transferring receptor 1 (TFR1), Beclin-1, and the LC3 II/I ratio. Quantitative real-time polymerase chain reaction (qPCR) was utilized to determine mRNA expression levels of Nrf2 and GPX4.

Compared with the Control group, the Model group exhibited severe hepatic steatosis, significant fibrosis, elevated collagen deposition, increased serum AST, ALT, MDA, TNF-α, and Fe²⁺ levels, and reduced SOD activity (P < 0.05). Protein expression of GPX4, Nrf2, and SLC7A11 was upregulated, while TFR1 and Beclin-1 expression increased, and mRNA levels of GPX4 and Nrf2 were markedly suppressed (P < 0. 05). Intervention with Yiqi Huoxue Lishui Decoction and/or inhibitors significantly attenuated histopathological damage, reduced AST, ALT, MDA, TNF-α, Fe²⁺ levels, and downregulated TFR1 and Beclin-1 expression (P < 0.05). Concurrently, SOD activity, GPX4 and Nrf2 protein/mRNA expression, and SLC7A11 protein levels were elevated (P <0. 05). No significant difference was observed in the LC3 Ⅱ/Ⅰ ratio across groups(P > 0.05).

Yiqi Huoxue Lishui Decoction may ameliorate hepatic fibrosis by inhibiting ferroptosis through activation of the Nrf2/GPX4 signaling pathway.