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TNF-? Directly Enhances Osteocyte RANKL Expression and Promotes Osteoclast Formation.

ABSTRACT: Osteoimmunology peeks into the interaction of bone and the immune system, which has largely proved to be a multiplex reaction. Osteocytes have been shown to regulate bone resorption through the expression of RANKL in physiologic and pathologic conditions. TNF-?, a product of the immune system, is an important cytokine regulating bone resorption in inflammatory conditions either directly or by increasing RANKL and M-CSF expressions by osteoblasts and stromal cells. The effect of TNF-? on a wide range of cell types has been documented; however, the direct effect of TNF-? on osteocytes has not been established yet. In this study, primary osteocytes were isolated by cell sorting from neonatal calvaria of Dmp1-Topaz mice, which express the green fluorescent protein under the influence of dentin matrix protein 1 promoter. The results show that osteocytes have a significantly higher RANKL mRNA expression when cultured with TNF-?. A co-culture system of osteocytes and TNF receptors I and II deficient osteoclast precursors treated with TNF-? show a significant increase in TRAP-positive cells while cultures without TNF-? failed to show TRAP-positive cells. Additionally, in vivo experiments of TNF-? injected to mouse calvaria show an increase in TRAP-positive cell number in the suture mesenchyme and an increase in the percentage of RANKL-positive osteocytes compared to PBS-injected calvaria. Osteocytes cultured with TNF-? show up-regulation of MAPKs phosphorylation measured by western blot, and adding MAPKs inhibitors to osteocytes cultured with TNF-? significantly decreases RANKL mRNA expression compared to osteocytes cultured with TNF-? alone. We also found that TNF-? activates the NF-?B pathway in osteocytes measured as a function of p65 subunit nuclear translocation. TNF-? directly affects osteocyte RANKL expression and increases osteoclastogenesis; our results demonstrate that osteocytes guard an important role in inflammatory bone resorption mediated by TNF-?.

SUBMITTER: Marahleh A

PROVIDER: S-EPMC6923682 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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TNF-α Directly Enhances Osteocyte RANKL Expression and Promotes Osteoclast Formation.

Marahleh Aseel A Kitaura Hideki H Ohori Fumitoshi F Kishikawa Akiko A Ogawa Saika S Shen Wei-Ren WR Qi Jiawei J Noguchi Takahiro T Nara Yasuhiko Y Mizoguchi Itaru I

Frontiers in immunology 20191213

Osteoimmunology peeks into the interaction of bone and the immune system, which has largely proved to be a multiplex reaction. Osteocytes have been shown to regulate bone resorption through the expression of RANKL in physiologic and pathologic conditions. TNF-α, a product of the immune system, is an important cytokine regulating bone resorption in inflammatory conditions either directly or by increasing RANKL and M-CSF expressions by osteoblasts and stromal cells. The effect of TNF-α on a wide r ...[more]

PMID: 31921183

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Project description:Docosahexaenoic acid (DHA) is an n-3 fatty acid that is an important structural component of the cell membrane. DHA exerts potent anti-inflammatory effects through G protein-coupled receptor 120 (GPR120), which is a functional receptor for n-3 fatty acids. DHA also regulates osteoclast formation and function. However, no studies have investigated the effect of DHA on inflammation-induced osteoclast formation in vivo. In the present study, we investigated whether DHA influences osteoclast formation, bone resorption and the expression of osteoclast-associated cytokines during lipopolysaccharide (LPS)-induced inflammation in vivo, and then we elucidated the underlying mechanisms by using in vitro experiments. In vitro experiments revealed both receptor activator of NF-kB ligand (RANKL)- and tumor necrosis factor-α (TNF-α)-induced osteoclast formation was inhibited by DHA. Supracalvarial administration of LPS with or without DHA was carried out for 5 days and then the number of osteoclasts, ratio of bone resorption pits and the level of type I collagen C-terminal cross-linked telopeptide were measured. All measurements were significantly lower in LPS+DHA-co-administered mice than LPS-administered mice. However, this DHA-induced inhibition was not observed in LPS-, DHA-, and selective GPR120 antagonist AH7614-co-administered mice. Furthermore, the expression of RANKL and TNF-α mRNAs was lower in the LPS+DHA-co-administered group than in the LPS-administered group in vivo. TNF-α mRNA levels were decreased in macrophages co-treated with LPS+DHA compared with cells treated with LPS in vitro. In contrast, RANKL mRNA expression levels from osteoblasts co-treated with DHA and LPS in vitro were equal to that in cells treated with LPS alone. Finally, the inhibitory effects of DHA on osteoclast formation in vitro were not observed by using osteoclast precursors from GPR120-deficient mice, and inhibition of LPS-induced osteoclast formation and bone resorption by DHA in vivo was not observed in GPR120-deficient mice. These results suggest that DHA inhibits LPS-induced osteoclast formation and bone resorption in vivo via GPR120 by inhibiting LPS-induced TNF-α production in macrophages along with direct inhibition of osteoclast formation.

Dataset Information

TNF-? Directly Enhances Osteocyte RANKL Expression and Promotes Osteoclast Formation.

Publications

TNF-α Directly Enhances Osteocyte RANKL Expression and Promotes Osteoclast Formation.

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