Network pharmacology-guided identification of IL-1β as a target for dihydroartemisinin-mediated anti-osteoarthritic effects: validation in chondrocyte and ACLT mouse models

Osteoarthritis (OA) is a highly prevalent degenerative joint disorder with limited disease-modifying treatment options. Dihydroartemisinin (DHA), an artemisinin derivative with anti-inflammatory activity, has shown therapeutic potential in several inflammatory settings. Whether DHA engages interleukin-1β (IL-1β), a major inflammatory mediator in OA, and how this relates to inflammasome signaling remain unclear. Network pharmacology and molecular docking were used to identify candidate DHA targets relevant to OA. Mouse primary chondrocytes were stimulated with IL-1β (10 μg/mL) and treated with DHA (40 μM). An anterior cruciate ligament transection (ACLT)-induced mouse OA model was treated with DHA (10 mg/kg, intraperitoneally, every other day for six weeks). Inflammatory and cartilage-matrix markers were assessed by western blotting, ELISA and immunohistochemistry, and cartilage degeneration by H&E and Safranin O-Fast Green staining. A lysate-based cellular thermal shift assay (CETSA) was used to test DHA-IL-1β target engagement, and NLRP3, ASC and cleaved caspase−1 were examined to assess inflammasome signaling. Network pharmacology identified 239 DHA-related targets and 2077 OA-related targets, with 72 overlapping targets. IL-1β was prioritized among the hub candidates, and molecular docking predicted a favorable DHA-IL-1β interaction (binding energy, − 6.5 kcal/mol). In IL-1β-stimulated chondrocytes, DHA reduced iNOS, COX-2, TNF-α, IL-6, MMP-13 and ADAMTS-5 while restoring aggrecan and collagen-2. In ACLT-induced OA mice, DHA improved cartilage histology and reduced inflammatory and catabolic protein changes. DHA also reduced IL-1β protein abundance among the screened hub proteins. In lysate-based CETSA, DHA increased the thermal stability of IL-1β, supporting direct target engagement. DHA additionally reduced NLRP3 and cleaved caspase−1 and lowered ASC abundance in IL-1β-stimulated chondrocytes, consistent with attenuation of NLRP3 inflammasome signaling. The data support an IL-1β-centered mechanism for the anti-inflammatory and cartilage-protective effects of DHA, involving IL-1β target engagement together with attenuation of NLRP3 inflammasome signaling. These findings provide a mechanistic basis for further evaluation of DHA as a repurposing candidate for OA.