Identification of the active constituents and molecular mechanism of Eulophia nuda extract in the treatment of osteoarthritis by network pharmacology, molecular modelling and experimental assays.

Osteoarthritis is a degenerative joint disease that worsens over time, often resulting in chronic pain. Eulophia nuda (Orchidaceae), a medicinal herb widely used by folklore and indigenous healers for treating arthritis but the active ingredients and the molecular mechanisms of action are yet to be explored. The present study systematically investigates the underlying anti-osteoarthritic mechanism of ENE through network pharmacology, molecular dynamics simulation and experimental assays. A comprehensive search on IMPPAT, KNApSAcK and Pubchem databases resulted 26 active compounds from E. nuda, of which 23 passed the drug-likeness criteria. A total of 2344 compound targets, 1370 osteoarthritis targets and 81 overlapping compound-disease targets were identified. The compound-disease target network resulted in five active constituents with degree > 23. Topological analysis of the protein-protein interaction network revealed six hub target genes. KEGG analysis revealed IL-17, TNF and AGE-RAGE signalling pathways as the enriched pathways involved in osteoarthritis. Molecular docking showed eulophiol had the good binding energy (>8.0 kcal/mol) with MMP9, JNK1, p38 and NF-kβ. The molecular dynamics simulations and the MMPBSA analysis indicate high stability and greater binding energy of eulophiol with the target proteins. ENE did not show cytotoxicity on SW982 cells up to a concentration of 100 μg/ml. ENE exhibited considerable anti-inflammatory effect by reducing PGE2, IL-6 and IL8 levels as well as reducing the mRNA expression of matrix metalloproteinases (MMP2 and MMP9). Furthermore, ENE effectively inhibited the NF-kβ nuclear translocation and phosphorylation of ERK2, p38 and JNK in SW982 cells. The current study showed that ENE may act as a potential drug candidate for treating osteoarthritis.