Elucidating the Multi-Target Therapeutic Mechanisms of Quercetin in Rheumatoid Arthritis: A Network Pharmacology, Molecular Docking, and In-Vitro Study.

AbstractBackground: Rheumatoid arthritis (RA) is a prevalent autoimmune disorder that causeschronic inflammation, stiffness, and progressive destruction of synovial joints. TraditionalAyurvedic formulations contain multi-target phytoconstituents that can modulate complexinflammatory pathways. Network pharmacology provides a holistic, systems-based approachto understanding these synergistic therapeutic mechanisms.Objective: This study aims to elucidate the multi-target therapeutic mechanisms of quercetin,a prominent Ayurvedic phytoconstituent, against RA by integrating network pharmacology,molecular docking, and in-vitro experimental validation.Methodology: Quercetin’s pharmacokinetic properties and targets were retrieved usingTCMSP and SwissTargetPrediction. RA-related genes were collected from GeneCards,OMIM, and DisGeNET. Protein-protein interaction (PPI) networks were constructed viaSTRING and Cytoscape, followed by GO and KEGG enrichment analyses. Binding affinitieswere simulated using molecular docking. Finally, in-vitro validation was conducted usingTNF-α-induced Rheumatoid Arthritis Fibroblast-Like Synoviocytes (RAFLS), evaluatedthrough MTT assays, ELISA, RT-PCR, Western blotting, and flow cytometry.Results: Network analysis revealed 58 intersecting therapeutic targets, with TNF, IL6, AKT1,and MAPK1 identified as central hub genes. KEGG analysis demonstrated significantenrichment in TNF and NF-κBsignaling pathways. Molecular docking showed strong bindingenergies, notably with COX-2 (-9.0 kcal/mol) and TNF-α (-8.7 kcal/mol). In-vitroexperiments confirmed that quercetin dose-dependently reduced abnormal RAFLSproliferation, significantly suppressed inflammatory cytokines (IL-1β, IL-6, TNF-α),downregulated NF-κB and MAPK protein expression, reduced oxidative stress, and promotedcellular apoptosis.Conclusion: Quercetin demonstrates significant potential as a multi-target therapeutic agentfor rheumatoid arthritis. By integrating computational predictions with in-vitro validation, thestudy concludes that quercetin effectively mitigates disease progression by synergisticallymodulating key inflammatory pathways, suppressing cytokine production, and reducingabnormal cellular proliferation and oxidative stress.