Revealing the Pharmacological Mechanism of Tibetan Medicine Wugeng San in Treating Rheumatoid Arthritis Through an Integrated Strategy of Chemical Composition Analysis, Network Pharmacology, Machine Learning, and In Vivo Experiments.

Background: Wugeng San (WGS) is a traditional Tibetan medicinal preparation that has long been used to treat inflammatory and arthritic conditions. However, its contemporary pharmacological validation and the mechanisms underlying its action in rheumatoid arthritis (RA) have not been fully investigated. Objective: For the first time, this study aimed to systematically investigate the therapeutic effects of WGS on RA, identify its potential targets, and elucidate its action mechanisms. Methods: This study, as the first comprehensive investigation of WGS in RA, employed integrated multiple approaches including chemical component identification via UPLC-Q-TOF/MS, network pharmacology, bioinformatics, machine learning, and in vivo efficacy assessment and mechanism verification in a collagen-induced arthritis (CIA) rat model, a widely accepted experimental model that mimics the key pathological features of RA. Results: The results demonstrated that WGS reduced the severity of arthritis in a dose-dependent manner, as evidenced by decreased paw swelling, normalized body weight, and restored levels of pro- and anti-inflammatory cytokines. The high dose of WGS (252 mg/kg) showed an effect comparable to that of methotrexate (0.2 mg/kg). Histological analysis revealed that WGS reduced synovial hyperplasia, cartilage erosion and bone destruction, decreased osteoclast numbers, and promoted osteoblast activity. Eighty-four compounds were identified using UPLC-Q-TOF/MS. Network pharmacology and machine learning analyses indicated SYK as a key target enriched in the NF-κB signaling and osteoclast differentiation pathways. Experimental validation confirmed that WGS suppressed the phosphorylation of SYK and NF-κB pathway components (p65, IκBα, and IKKα/β), decreased MMP1/MMP3 levels, and modulated the Bax/Bcl-2 ratio to promote apoptosis. Conclusions: In conclusion, WGS exhibits strong anti-arthritic effects through "multi-component, multi-target, and multi-pathway" mechanisms, likely attributable to the inhibition of the SYK/NF-κB signaling axis, suppression of matrix degradation, and regulation of cellular apoptosis. This research offers a pharmacological basis for repurposing WGS as a promising natural candidate for RA therapy.