The Molecular Mechanism of Silymarin in Relieving Myocardial Fibrosis and Reducing Oxidative Stress Levels in Adjuvant‐Induced Rheumatoid Arthritis Rats by Inhibiting IRAK4

Rheumatoid arthritis (RA) is associated with myocardial fibrosis and elevated oxidative stress, contributing to cardiovascular complications. Silymarin (SIL), a natural flavonoid compound with potent antioxidant and anti-inflammatory properties, has shown therapeutic potential in inflammatory diseases. This study aimed to investigate the effects and underlying mechanism of SIL on myocardial fibrosis and oxidative stress in adjuvant-induced RA rats. An adjuvant-induced RA rat model and an in vitro myocardial injury model were established, followed by SIL treatment. Rats were randomly divided into five groups: healthy control (Control), untreated RA model (Model), SIL (25 mg/kg/day), SIL (50 mg/kg/day), and methotrexate (7.6 mg/kg/day) groups, with 10 mice in each group. Cardiac function was assessed by echocardiography. Myocardial inflammation, oxidative stress, and fibrosis were evaluated using enzyme-linked immunosorbent assay (ELISA), H&E, and Masson's trichrome staining, respectively. Western blot analysis and real-time PCR were used to analyze the expression of IRAK4, collagen I, Alpha-Smooth Muscle Actin (α-SMA), Bcl-2-Associated X Protein (Bax), B-Cell Lymphoma/Leukemia-2 Protein (Bcl2), and Interleukin-1 Receptor-Associated Kinase 4 (IRAK4). In vitro, H9c2 cardiomyocytes were used to assess cell viability, proliferation, and apoptosis via using cell counting kit-8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU), and flow cytometry assays. SIL treatment markedly decreased the levels of pro-inflammatory cytokines (TNF-α, IL-6, and IL-1β), fibrosis markers (collagen I and α-SMA), and oxidative stress indicators, while improving cardiac function in RA rats. The higher dose (50 mg/kg) produced a more pronounced therapeutic effect, suggesting a dose-dependent response. In vitro, SIL significantly alleviated LPS-induced myocardial injury by suppressing IRAK4 expression, thereby attenuating oxidative stress, apoptosis, and fibrosis. SIL effectively mitigates myocardial fibrosis and oxidative stress in adjuvant-induced RA rats, primarily through inhibition of IRAK4-mediated inflammatory signaling. These findings highlight SIL as a promising therapeutic candidate for RA-related cardiac injury.