Exploring and validating the molecular targets and potential mechanisms of shikonin against osteomyelitis through computational analyses and animal experiments.

Osteomyelitis is a severe bone infection caused by bacterial pathogens, involving inflammation and tissue destruction. Shikonin, a naphthoquinone compound extracted from the traditional Chinese medicinal herb Lithospermum erythrorhizon, exhibits anti-inflammatory, antibacterial, antitumor, and wound-healing properties. Nonetheless, the efficacy and specific mechanisms of shikonin in treating osteomyelitis remain unexplored. In this study, the potential mechanism of shikonin in the treatment of osteomyelitis was systematically explored using network pharmacology, molecular docking, molecular dynamics simulation, and experimental validation in a Staphylococcus aureus (S. aureus) -induced mouse osteomyelitis model. Microcomputed tomography and Hematoxylin-Eosin staining demonstrated that shikonin alleviated bone destruction and inflammatory infiltration. Through network pharmacology, eight potential core targets of shikonin were identified: MET, MMP9, EGFR, SRC, PTGS2, CASP3, MMP2, and CCND1. Molecular docking and dynamics simulations indicated that shikonin had high binding affinity and stable interactions with MET, MMP9, and EGFR, with binding energies of -9.18 kcal/mol, -9.73 kcal/mol, and -9.58 kcal/mol, respectively. Furthermore, Western blot analysis confirmed that shikonin significantly downregulated the expression of MMP9 and EGFR. KEGG enrichment analysis suggested that shikonin may exert anti-inflammatory and antibacterial effects via pathways such as PI3K-Akt, VEGF, chemokine, and Toll-like receptor signaling. These findings reveal the multi-target mechanism of shikonin against osteomyelitis and provide a theoretical basis for its further therapeutic development.