Eupatilin suppresses osteoclastogenesis and periodontal bone loss by inhibiting the MAPKs/Siglec-15 pathway

Periodontitis is a widely prevalent oral disease around the world characterized by the disruption of the periodontal ligament and the subsequent development of periodontal pockets, as well as the loss of alveolar bone, and may eventually lead to tooth loss. This research aims to assess the suppressive impact of Eupatilin, a flavone obtained from Artemisia argyi, on osteoclastogenesis in vitro and periodontitis in vivo. We found that Eupatilin can efficiently obstruct the differentiation of Raw264.7 and bone marrow-derived macrophages (BMDMs) induced by RANKL, leading to the formation of mature osteoclasts. Consistently, bone slice resorption assay showed that Eupatilin significantly inhibited osteoclast-mediated bone resorption in a dose-dependent manner. Eupatilin also downregulated the expression of osteoclast-specific genes and proteins in Raw264.7 and BMDMs. RNA sequencing showed that Eupatilin notably downregulated the expression of Siglec-15. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses identified significantly enriched pathways in DEGs, including MAPK signaling pathway. And further mechanistic investigations confirmed that Eupatilin repressed MAPKs/NF-κBsignaling pathways. It was found that Siglec-15 overexpression reversed the inhibitory impact of Eupatilin on the differentiation of osteoclasts. Furthermore, activating MAPK signaling pathway reversed the downregulation of Siglec-15 and the inhibition of osteoclastogenesis by Eupatilin. To sum up, Eupatilin reduced the expression of Siglec-15 by suppressing MAPK signaling pathway, ultimately leading to the inhibition of osteoclastogenesis. Meanwhile, Eupatilin suppressed the alveolar bone resorption caused by experimentalperiodontitis in vivo. Eupatilin exhibits potential therapeutic effects in the treatment of periodontitis, rendering it a promising pharmaceutical agent.