Dataset Information

IKKbeta activation is sufficient for RANK-independent osteoclast differentiation and osteolysis.

ABSTRACT: Monocytes differentiate into osteoclasts through stimulation of receptor activator of NF-kappaB (RANK). Many downstream effectors of RANK play a positive role in osteoclastogenesis, but their relative importance in osteoclast differentiation is unclear. We report the discovery that activation of a single pathway downstream of RANK is sufficient for osteoclast differentiation. In this regard, introduction of constitutively activated IKKbeta (IKKbeta(SSEE)) but not wild-type IKKbeta into monocytes stimulates differentiation of bona fide osteoclasts in the absence of RANK ligand (RANKL). This phenomenon is independent of upstream signals because IKKbeta(SSEE) induced the development of bone-resorbing osteoclasts from RANK and IKKalpha knockout monocytes and in conditions in which NEMO-IKKbeta association was inhibited. NF-kappaB p100 and p105, but not RelB, were critical mediators of this effect. Inflammatory autocrine signaling by tumor necrosis factor alpha (TNF-alpha) and interleukin 1 (IL-1) were dispensable for the spontaneous osteoclastogenesis driven by IKKbeta(SSEE). More important, adenoviral gene transfer of IKKbeta(SSEE) induced osteoclasts and osteolysis in calvariae and knees of mice. Our data establish the sufficiency of IKKbeta activation for osteolysis and suggest that IKKbeta hyperactivation may play a role in conditions of pathologic bone destruction refractory to RANK/RANKL proximal therapeutic interventions.

SUBMITTER: Otero JE

PROVIDER: S-EPMC3153134 | biostudies-other | 2010 Jun

REPOSITORIES: biostudies-other

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IKKbeta activation is sufficient for RANK-independent osteoclast differentiation and osteolysis.

Otero Jesse E JE Dai Simon S Alhawagri Muhammad A MA Darwech Isra I Abu-Amer Yousef Y

Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research 20100601 6

Monocytes differentiate into osteoclasts through stimulation of receptor activator of NF-kappaB (RANK). Many downstream effectors of RANK play a positive role in osteoclastogenesis, but their relative importance in osteoclast differentiation is unclear. We report the discovery that activation of a single pathway downstream of RANK is sufficient for osteoclast differentiation. In this regard, introduction of constitutively activated IKKbeta (IKKbeta(SSEE)) but not wild-type IKKbeta into monocytes ...[more]

PMID: 20200955

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Project description:It has been shown that high vitamin A intake is associated with bone fragility and fractures in both animals and humans. However, the mechanism by which vitamin A affects bones is unclear. In the present study, the direct effects of retinoic acid (RA) on human and murine osteoclastogenesis were evaluated using cultured peripheral blood CD14(+) monocytes and RAW264.7 cells. Both the activity of the osteoclast marker tartrate resistant acid phosphatase (TRAP) in culture supernatant and the expression of the genes involved in osteoclast differentiation together with bone resorption were measured. To our knowledge, this is the first time that the effects of RA on human osteoclast progenitors and mature osteoclasts have been studied in vitro. RA stimulated proliferation of osteoclast progenitors both from humans and mice. In contrast, RA inhibited differentiation of the receptor activator of nuclear factor κB ligand (RANKL)-induced osteoclastogenesis of human and murine osteoclast progenitors via retinoic acid receptors (RARs). We also show that the mRNA levels of receptor activator of nuclear factor κB (RANK), the key initiating factor and osteoclast associated receptor for RANKL, were potently suppressed by RA in osteoclast progenitors. More importantly, RA abolished the RANK protein in osteoclast progenitors. This inhibition could be partially reversed by a RAR pan-antagonist. Furthermore, RA treatment suppressed the expression of the transcription factor nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) and increased the expression of interferon regulatory factor-8 (IRF-8) in osteoclast progenitors via RARs. Also, RA demonstrated differential effects depending on the material supporting the cell culture. RA did not affect TRAP activity in the culture supernatant in the bone slice culture system, but inhibited the release of TRAP activity if cells were cultured on plastic. In conclusion, our results suggest that retinoic acid increases proliferation of human osteoclast progenitors and that it inhibits RANK-stimulated osteoclast differentiation by suppressing RANK.

Project description:Excessive osteoclast-mediated bone resorption is a critical cause of osteoporosis affecting many aging people worldwide. 5'-Methylthioadenosine (MTA) is a natural sulfur-containing nucleoside normally produced in prokaryotes, plants, yeast, and higher eukaryotes via polyamine metabolism. MTA affects various physiological responses particularly the inflammatory pathway in both normal and cancerous cells and modulates the activation of nuclear factor-κB involved in the osteoclastogenesis signalling process. While several studies have reported that natural products possess anti-osteoclastogenesis phenolics and flavonoids, the effect of nucleoside derivatives on osteoclastogenesis remains limited. Therefore, this study aimed to explore the molecular mechanisms by which MTA affects pre-osteoclastic RAW 264.7 cells as a potential alleviation compound for inflammation-mediated bone loss. Osteoclasts were established by incubating RAW264.7 macrophage cells with receptor activator of nuclear factor kappa B ligand (RANKL) and macrophage colony-stimulating factor, the vital cytokines for activation of osteoclast differentiation. Cell viability was measured using MTT assays at 24, 48, and 72 h. The suppressive effect of MTA on RANKL-induced osteoclast differentiation and function was assessed using tartrate-resistant acid phosphatase (TRAP) analysis, qRT-PCR, and pit formation, Western blot, and immunofluorescence assays. MTA showed dose-dependent anti-osteoclastogenic activity by inhibiting TRAP-positive cell and pit formation and reducing essential digestive enzymes, including TRAP, cathepsin K, and matrix metallopeptidase 9. MTA was observed to suppress the osteoclast transduction pathway through (RANKL)-induced nuclear factor kappa-light-chain-enhancer of activated B cells (NFƘB); it attenuated NFƘB-P65 expression and down-regulated cFos proto-oncogene and nuclear factor of activated T cell c1 (NFATc1), the main regulators of osteoclasts. Moreover, the suppression of RANK (the initial receptor triggering several osteoclastogenic transduction pathways) was observed. Thus, this study highlights the potential of MTA as an effective therapeutic compound for restoring bone metabolic disease by inhibiting the RANK-NFATc1 signal pathway.

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IKKbeta activation is sufficient for RANK-independent osteoclast differentiation and osteolysis.

Publications

IKKbeta activation is sufficient for RANK-independent osteoclast differentiation and osteolysis.

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