Orexins Facilitates Osteogenic Differentiation of MC3T3-E1 Cells.

Dysfunction of osteoblastic bone formation and matrix mineralization plays a key role in the pathological development of osteoporosis. The orexin peptide orexin-A, a highly excitatory neuropeptide hormone, possesses various biological functions by activating its specific G protein-coupled receptors, orexin-1 receptor (OX1R) and orexin-2 receptor (OX2R). Here, we report that OX1R but not OX2R was expressed in MC3T3-E1 cells. Importantly, we found that orexin-A accelerated osteoblast differentiation and matrix mineralization in MC3T3-E1 cells, as manifested by elevation of physiological markers of osteoblastic differentiation [alkaline phosphatase (ALP) and osteogenic genes] and Alizarin Red staining, respectively. Importantly, our findings indicated that orexin-A significantly increased the expression of runt-related transcription factor 2 (Runx-2), which is the central transcriptional factor. Orexin-A treatment phosphorylated the kinase p38 mitogen-activated protein kinase (MAPK) in a dose- and time-dependent manner. Also, orexin-induced increase in gene expression (Runx-2, ALP, osteocalcin, and osterix) and matrix mineralization were prevented by the p38 MAPK specific inhibitor SB203580. Additionally, we also revealed that protein kinase D (PKD) is involved in the effects of Orexin-A on p38 MAPK activation and Runx-2 expression. Finally, we found that Orexin-A-induced osteoblastic formation and matrix mineralization and the activation of the PKD/p38 MAPK pathway are mediated by OX1R. Based on these findings, we concluded that activation of OX1R by orexin-A might possess a therapeutic strategy for bone disease.