Systems pharmacology and molecular docking elucidate multi-target anti-osteoarthritic mechanisms of LC–MS-identified constituents from ethanolic Tinospora crispa stem extract

Osteoarthritis is a multifactorial degenerative joint disease characterized by chronic inflammation, oxidative stress, extracellular matrix degradation, and chondrocyte apoptosis. In our previous study, Tinospora crispa demonstrated significant antioxidant and chondroprotective properties; however, its systems-level molecular mechanisms remain poorly defined. This study aimed to elucidate the multi-target anti-osteoarthritic mechanisms of LC–MS-identified constituents from ethanolic T. crispa extract using an integrated in silico approach. Seven bioactive compounds (N-trans-caffeoyltyramine, magnoflorine, reticuline, stylopine, columbamine, physalin I, and pseudolaric acid B) were subjected to target prediction analyses, while osteoarthritis-associated genes were retrieved from GeneCards and OMIM databases. Overlapping targets were analyzed through protein–protein interaction network construction, hub gene identification, and Gene Ontology and KEGG pathway enrichment analyses. Drug-likeness and pharmacokinetic properties were evaluated using ADMETlab 3.0. Molecular docking was performed against key targets (MAPK3, CASP3, BCL2, HSP90AA1, and SRC), followed by molecular dynamics simulation of the top-ranked complex. A total of 326 overlapping targets were identified, including key hub genes such as NFKB1, MAPK3, JUN, STAT3, BCL2, CASP3, SRC, and HSP90 isoforms. Enrichment analysis highlighted the involvement of IL-17, TNF, apoptosis, PI3K-Akt, MAPK, Toll-like receptor, and NF-κB signaling pathways. Molecular docking demonstrated favorable binding affinities, with stylopine showing strong interactions with MAPK3 and SRC. Molecular dynamics simulation further supported the stability of the stylopine–MAPK3 complex under dynamic conditions. ADMET profiling identified alkaloid constituents, particularly reticuline and stylopine, as promising compounds with favorable drug-like properties. These findings suggest that T. crispa constituents may exert anti-osteoarthritic effects through multi-target modulation of inflammatory and apoptosis-related pathways.