Transforming Growth Factor-β Stimulates the Production of Osteoprotegerin/Osteoclastogenesis Inhibitory Factor by Bone Marrow Stromal Cells

doi:10.1074/jbc.273.42.27091

Transforming Growth Factor-β Stimulates the Production of Osteoprotegerin/Osteoclastogenesis Inhibitory Factor by Bone Marrow Stromal Cells*

From the ^‡Department of Geriatric Research, National Institute for Longevity Sciences, Obu, Aichi 474-8522, Japan and ^§Research Institute of Life Science, Snow Brand Milk Products Co., Ltd., Shimotsuga, Tochigi 329-0521, Japan

Abstract

Osteoprotegerin (OPG)/osteoclastogenesis inhibitory factor (OCIF) is a recently identified cytokine that belongs to the tumor necrosis factor receptor superfamily and regulates bone mass by inhibiting osteoclastic bone resorption. The present study was undertaken to determine whether OPG/OCIF is produced in bone microenvironment and how the expression is regulated. A transcript for OPG/OCIF at 3.1 kilobases was detected in bone marrow stromal cells (ST2 and MC3T3-G2/PA6) as well as in osteoblastic cells (MC3T3-E1). Transforming growth factor-β1 (TGF-β1) markedly increased the steady-state level of OPG/OCIF mRNA in a dose-dependent manner, while TGF-β1 suppressed the mRNA expression of tumor necrosis factor-related activation-induced cytokine (TRANCE)/receptor activator of NF-κB ligand (RANKL), a positive regulator of osteoclastogenesis to which OPG/OCIF binds.

The effect of TGF-β1 on the expression of OPG/OCIF mRNA was transient, with a peak level at 3–6 h. The up-regulation of OPG/OCIF mRNA by TGF-β1 in ST2 cells did not require de novoprotein synthesis and involved both a transcriptional and a post-transcriptional mechanism. Western blot analysis and an enzyme-linked immunosorbent assay revealed that TGF-β1 significantly increased the secretion of OPG/OCIF protein by ST2 cells at 6–24 h. In murine bone marrow cultures, TGF-β1 markedly inhibited the formation of tartrate-resistant acid phosphatase-positive multinucleated osteoclast-like cells in the presence of 1,25-dihydroxyvitamin D₃, whose effect was significantly reversed by a neutralizing antibody against OPG/OCIF.

These results suggest that TGF-β1 negatively regulates osteoclastogenesis, at least in part, through the induction of OPG/OCIF by bone marrow stromal cells and that the balance between OPG/OCIF and TRANCE/RANKL in local environment may be an important determinant of osteoclastic bone resorption.

Footnotes

↵* This work was supported in part by Research Grants for Longevity Sciences from the Ministry of Health and Welfare of Japan (to K. I. and Y. Y.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¶ To whom correspondence should be addressed: Dept. of Geriatric Research, National Institute for Longevity Sciences, 36-3 Gengo, Morioka, Obu, Aichi 474-8522, Japan. Tel./Fax: 81-562-44-6595; E-mail:yoyamada{at}nils.go.jp.
↵2 We therefore refer to this cytokine as OPG/OCIF in this paper.
↵3 K. Yano, E. Tsuda, N. Washida, F. Kobayashi, M. Goto, A. Harada, K. Ikeda, K. Higashio, and Y. Yamada, submitted for publication.
Abbreviations:

TNF

tumor necrosis factor

OPG

osteoprotegerin

OCIF

osteoclastogenesis inhibitory factor

TGF-β1

transforming growth factor-β1

1

25(OH)₂D₃, 1α,25-dihydroxyvitamin D₃

CHX

cycloheximide

DRB

5,6-dichloro-1-β-d-ribofuranosylbenzimidazol

PCR

polymerase chain reaction

TRANCE

tumor necrosis factor-related activation-induced cytokine

RANKL

receptor activator of NF-κB ligand

EF1α

elongation factor 1 α

ELISA

enzyme-linked immunosorbent assay.
- Received April 7, 1998.
- Revision received July 28, 1998.