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01-01-2012 | Original Article

FGF and ERK signaling coordinately regulate mineralization-related genes and play essential roles in osteocyte differentiation

Authors: Ai Kyono, Nanthawan Avishai, Zhufeng Ouyang, Gary E. Landreth, Shunichi Murakami

Published in: Journal of Bone and Mineral Metabolism | Issue 1/2012

Abstract

To examine the roles of FGF and ERK MAPK signaling in osteocyte differentiation and function, we performed microarray analyses using the osteocyte cell line MLO-Y4. This experiment identified a number of mineralization-related genes that were regulated by FGF2 in an ERK MAPK-dependent manner. Real-time PCR analysis indicated that FGF2 upregulates Ank, Enpp1, Mgp, Slc20a1, and Dmp1 in MLO-Y4 cells. Consistent with this observation, the selective FGF receptor inhibitor PD173074 decreased Ank, Enpp1, Slc20a1, and Dmp1 mRNA expression in mouse calvaria in organ culture. Since Dmp1 plays a central role in osteocyte differentiation and mineral homeostasis, we further analyzed FGF regulation of Dmp1. Similar to FGF2, FGF23 upregulated Dmp1 expression in MLO-Y4 cells in the presence of Klotho. Furthermore, increased extracellular phosphate levels partially inhibited FGF2-induced upregulation of Dmp1 mRNA expression, suggesting a coordinated regulation of Dmp1 expression by FGF signaling and extracellular phosphate. In MLO-Y4 osteocytes and in MC3T3E1 and primary calvaria osteoblasts, U0126 strongly inhibited both basal expression of Dmp1 mRNA and FGF2-induced upregulation. Consistent with the in vitro observations, real-time PCR and immunohistochemical analysis showed a strong decrease in Dmp1 expression in the skeletal elements of ERK1 ^−/− ; ERK2 ^flox/flox ; Prx1-Cre mice. Furthermore, scanning electron microscopic analysis revealed that no osteocytes with characteristic dendritic processes develop in the limbs of ERK1 ^−/− ; ERK2 ^flox/flox ; Prx1-Cre mice. Collectively, our observations indicate that FGF signaling coordinately regulates mineralization-related genes in the osteoblast lineage and that ERK signaling is essential for Dmp1 expression and osteocyte differentiation.

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Title: FGF and ERK signaling coordinately regulate mineralization-related genes and play essential roles in osteocyte differentiation
Authors: Ai Kyono
Nanthawan Avishai
Zhufeng Ouyang
Gary E. Landreth
Shunichi Murakami
Publication date: 01-01-2012
Publisher: Springer Japan
Published in: Journal of Bone and Mineral Metabolism / Issue 1/2012
Print ISSN: 0914-8779
Electronic ISSN: 1435-5604
DOI: https://doi.org/10.1007/s00774-011-0288-2

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