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KCI등재 학술저널

Effects of Inositol 1,4,5-triphosphate on Osteoclast Differentiation in RANKL-induced Osteoclastogenesis

Effects of Inositol 1,4,5-triphosphate on Osteoclast Differentiation in RANKL-induced Osteoclastogenesis

The receptor activator of NF-&#1354;B ligand (RANKL) signal is an activator of tumor necrosis factor receptor-associated factor 6 (TRAF6), which leads to the activation of NF-&#1354;B and other signal transduction pathways essential for osteoclastogenesis, such as Ca<sup>2+</sup> signaling. However, the intra&lt;/SUP&gt;cellular levels of inositol 1,4,5-trisphosphate (IP<sub>3</sub>) and IP<sub>3</sub>-mediated cellular function of RANKL during osteoclastogenesis are not known. In the present study, we determined the levels of IP<sub>3</sub> and evaluated IP<sub>3</sub>-mediated osteoclast differentiation and osteoclast activity by RANKL treatment of mouse leukemic macrophage cells (RAW 264.7) and mouse bone marrow-derived monocyte/macrophage precursor cells (BMMs). During osteoclastogenesis, the expression levels of Ca<sup>2+</sup> signaling proteins such as IP<sub>3</sub> receptors (IP<sub>3</sub>Rs), plasma membrane Ca<sup>2+</sup> ATPase, and sarco/endoplasmic reticulum Ca<sup>2+</sup> ATPase type2 did not change by RANKL treatment for up to 6 days in both cell types. At 24 h after RANKL treatment, a higher steady-state level of IP<sub>3</sub> was observed in RAW264.7 cells transfected with green fluorescent protein (GFP)-tagged pleckstrin homology (PH) domains of phospholipase C (PLC) &#1348;, a probe specifically detecting intracellular IP<sub>3</sub> levels. In BMMs, the inhibition of PLC with U73122 [a specific inhibitor of phospholipase C (PLC)] and of IP<sub>3</sub>Rs with 2-aminoethoxydiphenyl borate (2APB; a non-specific inhibitor of IP<sub>3</sub>Rs) inhibited the generation of RANKL-induced multinucleated cells and decreased the bone-resorption rate in dentin slice, respectively. These results suggest that intracellular IP<sub>3</sub> levels and the IP<sub>3</sub>-mediated signaling pathway play an important role in RANKL-induced osteoclastogenesis.

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