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J Endocrinol Invest. 2013 Nov;36(10):876-82. doi: 10.3275/8975. Epub 2013 May 22.

Afamin stimulates osteoclastogenesis and bone resorption via Gi-coupled receptor and Ca2+/calmodulin-dependent protein kinase (CaMK) pathways.

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  • 1Division of Endocrinology and Metabolism, Asan Medical Center, University of Ulsan College of Medicine, 388-1 Poongnap2- Dong, Songpa-Gu, Seoul 138-736, Korea.

Abstract

BACKGROUND:

Afamin was recently identified as a novel osteoclast-derived coupling factor that can stimulate the in vitro and in vivo migration of preosteoblasts.

AIM:

In order to understand in more detail the biological roles of afamin in bone metabolism, we investigated its effects on osteoclastic differentiation and bone resorption.

METHODS:

Osteoclasts were differentiated from mouse bone marrow cells. Tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells were considered as osteoclasts, and the resorption area was determined by incubating the cells on dentine discs. The intracellular cAMP level was determined using a direct enzyme immunoassay. Signaling pathways were investigated using western blot and RT-PCR. Recombinant afamin was administered exogenously to bone cell cultures.

RESULTS:

Afamin stimulated both osteoclastogenesis and in vitro bone resorption. Consistently, the expressions of osteoclast differentiation markers were significantly increased by afamin. Although afamin mainly affected the late-differentiation stages of osteoclastogenesis, the expression levels of receptor activator of nuclear factor-κB ligand (RANKL)-dependent signals were not changed. Afamin markedly decreased the levels of intracellular cAMP with reversal by pretreatment with pertussis toxin (PTX), a specific inhibitor of Gi-coupled receptor signaling. In addition, PTX almost completely blocked afamin-stimulated osteoclastogenesis. Furthermore, pretreatment with KN93 and STO609 - Ca2+/cal - mo dulin-dependent protein kinase (CaMK) and CaMK kinase inhibitors, respectively - significantly prevented decreases in the intracellular cAMP level by afamin while attenuating afamin-stimulated osteoclastogenesis.

CONCLUSION:

Afamin enhances osteoclastogenesis by decreasing intracellular cAMP levels via Gi-coupled receptor and CaMK pathways.

PMID:
23698732
[PubMed - indexed for MEDLINE]
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