Irilin D suppresses RANKL-induced osteoclastogenesis and prevents inflammation-induced bone loss by disrupting the NF-κB and MAPK signaling pathways.

Excessive activity of osteoclasts(OCs) lead to bone resorption in chronic inflammatory conditions. The use of natural compounds to target OCs offers significant promise in the treatment or prevention of OC-associated diseases. Irilin D (IRD), a natural isoflavone derived from Belamcanda chinensis (L.) DC., has potential effects on OC differentiation both in vitro and in vivo that have yet to be thoroughly explored. In our study, we found that IRD inhibited receptor activator of nuclear factor-κB ligand (RANKL)-induced OC differentiation, actin ring formation, and bone resorption in vitro without compromising cell viability. However, IRD did not exhibit anti-inflammatory effects in lipopolysaccharide (LPS)-stimulated macrophages. Furthermore, IRD reduced LPS-induced inflammatory bone loss by blocking osteoclastogenesis in a mouse model. Mechanistically, IRD disrupted RANKL-induced activation of mitogen-activated protein kinases (MAPKs) and nuclear factor-κB (NF-κB), leading to the inhibition of c-Fos and nuclear factor of activated T cells cytoplasmic 1 (NFATc1) activation. We also demonstrated that IRD inhibited RANKL-induced osteoclastic NFATc1 target genes, including DC-STAMP, ACP5, and CtsK. Our results indicate that IRD mitigates LPS-induced inflammatory bone resorption in mice by inhibiting RANKL-activated MAPKs and NF-κB signaling pathways, suggesting its potential as a natural isoflavone for preventing or treating OC-associated diseases.