Calcitonin Gene-related Peptide Inhibits Osteoclast Differentiation by Inducing the Negative Regulators <i>MafB</i> and <i>Bcl6</i>

Abstract

Calcitonin gene-related peptide (CGRP), a neuropeptide derived from sensory nerves, suppresses osteoclast differentiation; however, the detailed mechanism is unknown. Therefore, in this study, we investigated the mechanism underlying the CGRP-mediated suppression of osteoclast differentiation using a model in which mouse monocyte/macrophage RAW264.7 cells were induced to differentiate into osteoclasts with recombinant mouse soluble receptor activator of nuclear factor-kappa B ligand (s-RANKL). RAW264.7 cells were cultured with s-RANKL in the presence or absence of CGRP for 24–72 h and then quantitative real-time PCR was performed. The mRNA expression of the positive regulators of osteoclast differentiation [i.e., nuclear factor of activated T-cells 1 (Nfatc1) and B lymphocyte-induced maturation protein-1 (Blimp1)], as well as that of the negative regulators of osteoclast differentiation [i.e., MAF bZIP transcription factor B (MafB), B-cell lymphoma 6 (Bcl6), and interferon regulatory factor-8 (Irf8)], was analyzed. In RAW264.7 cells treated with s-RANKL, CGRP significantly suppressed Nfatc1 and Blimp1 mRNA expression at 24 and 48 h. On the contrary, CGRP significantly increased the mRNA expression of MafB at 24 and 48 h of culture and of Bcl6 at 48 and 72 h of culture. However, CGRP did not affect Irf8 mRNA expression. In s-RANKL-untreated RAW264.7 cells, MafB mRNA expression significantly increased from 2 to 8 h, whereas Bcl6 mRNA expression significantly increased from 4 to 24 h of culture. These results suggest that CGRP inhibits osteoclast differentiation by increasing the mRNA expression of MafB and Bcl6, which are negative regulators of osteoclast differentiation.