Molecular Mechanisms of Curdlan-Induced Suppression of NFATc1 Expression in Osteoclasts.

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.

Koga A ，Nagai-Yoshioka Y ，Yamasaki R ，Adachi Y ，Fujii W ，Ariyoshi W ... -  《-》

Radiofrequency field inhibits RANKL-induced osteoclast differentiation in RAW264.7 cells via modulating the NF-κB signaling pathway.

Ding C ，Wang H ，Yang C ，Hang Y ，Zhu S ，Cao Y ... -  《-》

S6K/FLNC/ITGβ3 signaling pathway regulates osteoclastogenesis and the inhibition of osteoclastogenesis by columbianadin.

Over-activation of osteoclastogenesis is a significant factor contributing to bone loss, leading to increased resorption of bone. Columbianadin (CBN), a compound derived from Angelicae Pubescentis Radix, has traditionally been used in Chinese medicine to treat bone-related disorders. However, the specific effects of CBN on bone loss are still poorly understood. This study aims to identify a novel target for inhibiting osteoclast differentiation and to elucidate the effects and underlying mechanisms of CBN on osteoclastogenesis. We employed a transcriptomics approach to identify genes that undergo significant changes during osteoclast differentiation. These findings were validated using reverse transcription-polymerase chain reaction (RT-PCR) and Western blot (WB) analysis. Subsequently, we utilized adenoviral transfection to investigate the effects of target genes on osteoclast differentiation. Additionally, we employed proteomics to elucidate the signaling pathways that regulate osteoclast differentiation. We examined the effect of CBN on S6K/FLNC/ITGβ3 signaling pathway and osteoclast differentiation. Our results revealed a dramatic increase in filamin C (FLNC) levels during osteoclast differentiation. Inhibition of FLNC expression significantly suppressed markers of osteoclast differentiation such as tartrate-resistant acid phosphatase (TRAP), nuclear factor-activated T cell 1 (NFATc1), and c-Fos, as well as inhibited the activity of bone resorption. We further conducted a proteomic analysis and found S6 K protein might be involved in this process. Then we utilized an S6K-specific inhibitor (PF-4,708,671) and demonstrated that inhibiting the S6 K protein reduced FLNC expression and the interaction between FLNC and integrin β3 (ITGβ3) in osteoclasts. Finally, we found that CBN inhibited osteoclast differentiation and bone loss in ovariectomized mice by targeting the S6K/FLNC/ITGβ3 signaling pathway. FLNC was identified as a critical protein in osteoclastogenesis. The S6K/FLNC/ITGβ3 signaling pathway played a significant role in osteoclast differentiation and bone loss. Furthermore, CBN exhibited anti-osteoporotic effects by inhibiting the S6K/FLNC/ITGβ3 signaling pathway.

Wei Q ，Niu ZC ，Fu XY ，OuYang MH ，Guo XT ，Li J ，Chang YX ，Mao HP ，Gao XM ... -  《-》

Hypoxia Promotes Osteoclast Differentiation by Weakening USP18-Mediated Suppression on the NF-κB Signaling Pathway.

Fan X ，Li B ，Chai S ，Zhang R ，Cai C ，Ge R ... -  《INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES》

The dectin 1 agonist curdlan regulates osteoclastogenesis by inhibiting nuclear factor of activated T cells cytoplasmic 1 (NFATc1) through Syk kinase.

Several immune system cell surface receptors are reported to be associated with osteoclastogenesis. Dectin 1, a lectin receptor for β-glucan, is found predominantly on cells of the myeloid lineage. In this study, we examined the effect of the dectin 1 agonist curdlan on osteoclastogenesis. In mouse bone marrow cells and dectin 1-overexpressing RAW 264.7 cells (d-RAWs), curdlan suppressed receptor activator of NF-κB ligand (RANKL)-induced osteoclast differentiation, bone resorption, and actin ring formation in a dose-dependent manner. This was achieved within non-growth inhibitory concentrations at the early stage. Conversely, curdlan had no effect on macrophage colony-stimulating factor-induced differentiation. Furthermore, curdlan inhibited RANKL-induced nuclear factor of activated T cell cytoplasmic 1 (NFATc1) expression, thereby decreasing osteoclastogenesis-related marker gene expression, including tartrate-resistant acid phosphatase, osteoclast stimulatory transmembrane protein, cathepsin K, and matrix metallopeptidase 9. Curdlan inhibited RANKL-induced c-fos expression, followed by suppression of NFATc1 autoamplification, without significantly affecting the NF-κB signaling pathway. We also observed that curdlan treatment decreased Syk protein in d-RAWs. Inhibition of the dectin 1-Syk kinase pathway by Syk-specific siRNA or chemical inhibitors suppressed osteoclast formation and NFATc1 expression stimulated by RANKL. In conclusion, our results demonstrate that curdlan potentially inhibits osteoclast differentiation, especially NFATc1 expression, and that Syk kinase plays a crucial role in the transcriptional pathways. This suggests that the activation of dectin 1-Syk kinase interaction critically regulates the genes required for osteoclastogenesis.

Yamasaki T ，Ariyoshi W ，Okinaga T ，Adachi Y ，Hosokawa R ，Mochizuki S ，Sakurai K ，Nishihara T ... -  《-》