Hesperidin Suppresses Complete Freund's Adjuvant (CFA)-Induced Rheumatoid Arthritis (RA) in Wistar Albino Rats by Targeting TNF-α/IL-6/IL-1β and NF-κB/COX-2 Pathways

Objective Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disorder for which current therapeutic options are often insufficient. This study aimed to evaluate the therapeutic potential of hesperidin (HSP), in suppressing complete Freund's adjuvant (CFA)-induced RA in Wistar albino rats, through modulation of key pro-inflammatory markers (TNF-α, IL-6, IL-1β, NF-κB, and COX-2), supported by in silico docking and pharmacokinetic profiling. Methods Forty-eight female Wistar albino rats were randomized into six groups (n = 8). Group 1 received normal saline; Group 2 was induced with RA via 0.1 ml CFA and left untreated; Group 3 received CFA + naproxen (8 mg/kg). Groups 4, 5, and 6 received hesperidin at 100, 200, and 400 mg/kg, respectively, following RA induction. Serum pro-inflammatory markers were quantified using standard biochemical assays. Molecular docking was performed to assess HSP binding affinity toward COX-2, TNF-α, IL-1β, and IL-6, while ADMET profiling was carried out to predict pharmacokinetic and toxicity attributes. Results In vivo results revealed that RA induction significantly elevated TNF-α, IL-6, IL-1β, and NF-κB levels ( p < .05) across the treated groups. Whereas, hesperidin treatment produced a dose-dependent and significant ( p < .05 ) reduction in these markers, restoring them to levels comparable to naproxen-treated rats. In silico docking analysis revealed that hesperidin has multi-target binding capability inhibitor to COX-2, TNF-α, IL-1β, and IL-6. Protein-ligand binding was predominantly surface-oriented except for COX-2, which showed interior pocket binding. ADMET predictions indicated that HSP is a P-glycoprotein substrate with low gastrointestinal absorption, does not cross the blood–brain barrier, shows no cytochrome P450 enzyme subunit inhibition, and lacks carcinogenic or hepatotoxic potential. Conclusion Combined in vivo and in silico findings demonstrate that hesperidin exhibits strong anti-inflammatory effects by inhibiting key mediators of RA pathogenesis, alongside a favorable safety pharmacokinetic profile.