TRANCE (tumor necrosis factor–related activation-induced cytokine) is a recently described member of the tumor necrosis factor superfamily that stimulates dendritic cell survival and has also been found to induce osteoclastic differentiation from hemopoietic precursors. However, its effects on mature osteoclasts have not been defined. It has long been recognized that stimulation of osteoclasts by agents such as parathyroid hormone (PTH) occurs through a hormonal interaction with osteoblastic cells, which are thereby induced to activate osteoclasts. To determine whether TRANCE accounts for this activity, we tested its effects on mature osteoclasts. TRANCE rapidly induced a dramatic change in osteoclast motility and spreading and inhibited apoptosis. In populations of osteoclasts that were unresponsive to PTH, TRANCE caused activation of bone resorption equivalent to that induced by PTH in the presence of osteoblastic cells. Moreover, osteoblast-mediated stimulation of bone resorption was abrogated by soluble TRANCE receptor and by the soluble decoy receptor osteoprotegerin (OPG), and stimulation of isolated osteoclasts by TRANCE was neutralized by OPG. Thus, TRANCE expression by osteoblasts appears to be both necessary and sufficient for hormone-mediated activation of mature osteoclasts, and TRANCE-R is likely to be a receptor for signal transduction for activation of the osteoclast and its survival.
Colony-stimulating-factor-1 (CSF-1) signaling through cFMS receptor kinase is increased in several diseases. To help investigate the role of cFMS kinase in disease, we identified GW2580, an orally bioavailable inhibitor of cFMS kinase. GW2580 completely inhibited human cFMS kinase in vitro at 0.06 M and was inactive against 26 other kinases. GW2580 at 1 M completely inhibited CSF-1-induced growth of mouse M-NFS-60 myeloid cells and human monocytes and completely inhibited bone degradation in cultures of human osteoclasts, rat calvaria, and rat fetal long bone. In contrast, GW2580 did not affect the growth of mouse NS0 lymphoblastoid cells, human endothelial cells, human fibroblasts, or five human tumor cell lines. GW2580 also did not affect lipopolysaccharide (LPS)-induced TNF, IL-6, and prostaglandin E2 production in freshly isolated human monocytes and mouse macrophages. After oral administration, GW2580 blocked the ability of exogenous CSF-1 to increase LPS-induced IL-6 production in mice, inhibited the growth of CSF-1-dependent M-NFS-60 tumor cells in the peritoneal cavity, and diminished the accumulation of macrophages in the peritoneal cavity after thioglycolate injection. Unexpectedly, GW2580 inhibited LPS-induced TNF production in mice, in contrast to effects on monocytes and macrophages in vitro. In conclusion, GW2580's selective inhibition of monocyte growth and bone degradation is consistent with cFMS kinase inhibition. The ability of GW2580 to chronically inhibit CSF-1 signaling through cFMS kinase in normal and tumor cells in vivo makes GW2580 a useful tool in assessing the role of cFMS kinase in normal and disease processes.CSF-1 ͉ macrophage colony-stimulating factor
SummaryMacrophage colony-stimulating factor (M-C3F) is known to play an important role in osteoclast formation. However, its actions on mature cells have not been fully characterized. We now report that M-CSF dramatically stimulates osteoclastic motility and spreading; osteoclasts responded to a gradient of M-CSF with orientation, and random cell polarization occurred after isotropic exposure. M-CSF also supported the survival of osteoclasts by preventing apoptosis. Paradoxically, M-CSF inhibits bone resorption by isolated osteoclasts. We found that this was effected predominantly by reduction in the number of excavations. Thus, M-CSF showed a propensity to suppress resorption through a reduction in the proportion of cells that were resorbing bone. Our data suggest that apart from the established role of M-CSF in the provision of precursors for osteoclastic induction, a major role for M-CSF in bone resorption is to enhance osteoclastic survival, migration, and chemotaxis. It seems appropriate that during these processes resorptive functions should be suppressed. We suggest that M-CSF continues to modulate osteoclastic activity once osteoclasts are on resorptive sites, through regulation of the balance between resorption and migration, such that not only the quantity, but the spatial pattern of resorption can be controlled by adjacent M-CSF-secreting cells of osteoblastic lineage.
We recently found that estrogen deficiency leads to a lowering of thiol antioxidant defenses in rodent bone. Moreover, administration of agents that increase the concentration in bone of glutathione, the main intracellular antioxidant, prevented estrogen-deficiency bone loss, whereas depletion of glutathione by buthionine sulfoximine administration provoked substantial bone loss. To analyze further the mechanism by which antioxidant defenses modulate bone loss, we have now compared expression of the known antioxidant enzymes in osteoclasts. We found that glutathione peroxidase 1 (Gpx), the enzyme primarily responsible for the intracellular degradation of hydrogen peroxide, is overwhelmingly the predominant antioxidant enzyme expressed by osteoclasts and that its expression was increased in bone marrow macrophages by receptor activator of nuclear factor-kappaB ligand (RANKL) and in osteoclasts by 17beta-estradiol. We therefore tested the effect of overexpression of Gpx in osteoclasts by stable transfection of RAW 264.7 (RAW) cells, which are capable of osteoclastic differentiation in response to RANKL, with a Gpx-expression construct. Osteoclast formation was abolished. The Gpx expression construct also suppressed RANKL-induced nuclear factor-kappaB activation and increased resistance to oxidation of dihydrodichlorofluorescein by exogenous hydrogen peroxide. We therefore tested the role of hydrogen peroxide in the loss of bone caused by estrogen deficiency by administering pegylated catalase to mice. We found that catalase prevented ovariectomy-induced bone loss. These results suggest that hydrogen peroxide is the reactive oxygen species responsible for signaling the bone loss of estrogen deficiency.
The mechanisms through which estrogen prevents bone loss are uncertain. Elsewhere, estrogen exerts beneficial actions by suppression of reactive oxygen species (ROS). ROS stimulate osteoclasts, the cells that resorb bone. Thus, estrogen might prevent bone loss by enhancing oxidant defenses in bone. We found that glutathione and thioredoxin, the major thiol antioxidants, and glutathione and thioredoxin reductases, the enzymes responsible for maintaining them in a reduced state, fell substantially in rodent bone marrow after ovariectomy and were rapidly normalized by exogenous 17-β estradiol. Moreover, administration of N-acetyl cysteine (NAC) or ascorbate, antioxidants that increase tissue glutathione levels, abolished ovariectomy-induced bone loss, while L-buthionine-(S,R)-sulphoximine (BSO), a specific inhibitor of glutathione synthesis, caused substantial bone loss. The 17-β estradiol increased glutathione and glutathione and thioredoxin reductases in osteoclast-like cells in vitro. Furthermore, in vitro NAC prevented osteoclast formation and NF-κB activation. BSO and hydrogen peroxide did the opposite. Expression of TNF-α, a target for NF-κB and a cytokine strongly implicated in estrogen-deficiency bone loss, was suppressed in osteoclasts by 17-β estradiol and NAC. These observations strongly suggest that estrogen deficiency causes bone loss by lowering thiol antioxidants in osteoclasts. This directly sensitizes osteoclasts to osteoclastogenic signals and entrains ROS-enhanced expression of cytokines that promote osteoclastic bone resorption
It has previously only been possible to assess osteoclastic bone resorption in intact bone, where other cell types may modify or mediate osteoclastic responses to environmental agents. We have recently developed techniques which enable us to measure bone resorption by osteoclasts extracted from bone and have used these techniques to assess the effects of prostaglandins (PGs) and calcium-regulating hormones on bone resorption by these cells. Osteoclasts were mechanically disaggregated from neonatal rabbit long bones and cultured on slices of devitalized cortical bone for 8 or 24 h. After this time, osteoclasts were associated with the appearance in the scanning electron microscope of characteristic resorption pits, the volume of which was calculated by computer-assisted morphometric and stereophotogrammetric techniques after removal of cells. Salmon calcitonin inhibited osteoclastic bone resorption at concentrations of 1 pg/ml and above, while PTH and 1,25-dihydroxyvitamin D3 were without significant effect. This suggests that the latter hormones do not increase bone resorption in intact bone through a direct effect on osteoclasts. PGI2, PGE1, and PGE2, all of which are known to stimulate resorption when added to intact bone, paradoxically reduced resorption in our cultures. It appears likely that PGs act as direct inhibitors of osteoclastic bone resorption but have an additional effect on other cells in bone, which are induced by PGs to cause osteoclastic stimulation.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.