Molecular Mechanisms of Curdlan‐Induced Suppression of NFATc1 Expression in Osteoclasts

Abstract

Osteoclasts derived from hematopoietic stem cells express immunoreceptors on their cell surface. Previously, we showed that the β‐glucan curdlan suppressed osteoclastogenesis via binding to dectin‐1, a pattern recognition receptor. Curdlan negatively regulates osteoclast differentiation and bone resorption capacity by suppressing the expression of nuclear factor of activated T cells 1 (NFATc1), a master factor for osteoclast differentiation, in a dectin‐1‐dependent manner; however, the mechanism involved in this process has not yet been fully elucidated. In this study, we aimed to elucidate the molecular mechanism involved in the suppression of RANKL‐induced osteoclast differentiation by curdlan. Real‐time RT‐qPCR results showed that curdlan suppressed the expression of NFATc1 in cells of the osteoclast progenitor cell line RAW264.7 overexpressing dectin‐1 (d‐RAW cells), without altering the expression of negative regulators. Therefore, we examined the effect of curdlan on the NF‐κB pathway, which is important for the induction of NFATc1 expression. Western blot analysis results showed that curdlan addition suppressed RANKL‐induced NF‐κB activation in the vector control line (c‐RAW) cells with low expression of dectin‐1, in d‐RAW cells, and the parental RAW264.7 (RAW) cells. The results of tartrate‐resistant alkaline phosphatase staining and real‐time RT‐qPCR showed that curdlan addition suppressed osteoclast differentiation in RAW cells, suggesting the presence of a dectin‐1‐independent modification system. Finally, we focused on the complement receptor 3 (CR3), which binds β‐glucan, and revealed that blocking the binding of β‐glucan to the CD11b molecule, a component of CR3, by neutralizing antibody, recovered the suppression of IκBα degradation by curdlan. These results suggest that the suppression of osteoclast differentiation by curdlan involves not only the dectin‐1‐dependent pathway but also the negative regulation of NFATc1 via modification of the NF‐κB pathway via CR3 recognition. The results of this study may aid to establish treatment methods for metabolic bone diseases and inflammatory bone destruction and to clarify their pathogenesis.