Typhaneoside suppresses osteoclastogenesis and osteoporosis by stabilizing ADORA1

Osteoporosis, characterized by impaired bone remodeling and increased bone fragility, is increasingly prevalent among postmenopausal women. Current pharmacological treatments exhibit efficacy but are associated with adverse effects, prompting interest in natural flavonoids like Typhaneoside, known for anti-inflammatory and antioxidant properties. However, the molecular mechanism by which TYP influences osteoclast differentiation remains unclear.

This study aims to investigate the anti-osteoporotic effect of TYP and elucidate its underlying molecular mechanisms involving osteoclast differentiation and activity.

RANKL-induced osteoclast differentiation models in mouse bone marrow-derived monocytes (BMMCs) and ovariectomy (OVX)-induced osteoporosis mouse models were utilized to assess the effect of TYP on bone metabolism in vitro and in vivo.

The effects of TYP on osteoclast differentiation and function were evaluated by TRAP staining, qRT-PCR, Western blot, immunoprecipitation, molecular docking, and mass spectrometry. Micro-CT, histological staining, and biochemical assays were performed to evaluate bone structure and metabolism in OVX mice.

TYP significantly inhibited RANKL-induced osteoclast differentiation and bone resorption activity without cytotoxic effects in vitro. In OVX mice, TYP treatment dose-dependently improved bone microarchitecture, reduced osteoclast numbers, and decreased serum bone resorption markers. Mechanistically, TYP directly bound ADORA1 receptor at the LYS265 site, competitively inhibiting NEDD4-1-mediated ubiquitination, thus stabilizing ADORA1 and suppressing osteoclast-specific gene expression (TRAP, CTSK, NFATc1).

These findings reveal that TYP effectively attenuates osteoclast differentiation and activity by stabilizing ADORA1 via inhibition of NEDD4-1-mediated ubiquitination, suggesting its therapeutic potential for osteoporosis treatment.