Inhibition of osteoclastogenesis by prolyl hydroxylase inhibitor dimethyloxallyl glycine

Abstract

Studies examining the effects of hypoxia upon osteoclast biology have consistently revealed a stimulatory effect; both osteoclast differentiation and resorption activity have been shown to be enhanced in the presence of hypoxia. In the present study we examined the effects of the hypoxia mimetics dimethyloxallyl glycine (DMOG) and desferrioxamine (DFO) upon osteoclastogenesis. In contrast to hypoxia, our studies revealed a dose-dependent inhibition of osteoclast formation from macrophages treated with DMOG and DFO. Moreover, expression of a constitutively active form of hypoxia-inducible factor 1alpha (HIF-1alpha) did not enhance osteoclastogenesis and actually attenuated the differentiation process. DMOG did not affect cell viability or receptor activator of nuclear factor kappaB ligand (RANKL)-dependent phosphorylation of mitogen-activated protein (MAP) kinases. However, RANKL-dependent transcription of tartrate-resistant acid phosphatase (TRAP) was reduced in the presence of DMOG. Additionally, DMOG promoted transcription of the pro-apoptotic mediator B-Nip3. These studies suggest that a hypoxia-responsive factor other than HIF-1alpha is necessary for enhancing the formation of osteoclasts in hypoxic settings.

Bone substitute materials supplemented with prolyl hydroxylase inhibitors decrease osteoclastogenesis in vitro

“…[28][29][30] However, prolyl hydroxylase inhibitors at concentrations below 0.3 mM did not induce toxic effects in osteoclast progenitor cells such as RAW 264.7 and primary hematopoietic stem cells from rats. 25 Our results from supernatants of bone substitute preparations suggest that the concentration released is below the threshold of toxicity. DFO at high concentrations can give false-positive readings in the MTT assay (data not shown).…”

“…Although we and Leger et al added prolyl hydroxylase inhibitors in the beginning of the osteoclastogenesis assay with cells of rodent origin, Knowles et al used the prolyl hydroxylase inhibitor for the last day of the human osteoclast assays. 24,25 The inhibition of osteoclast numbers and their TRAP activity as observed by us and Leger et al might therefore be due to effects on osteoclast progenitor cells.…”

“…In addition to this potential antiresorptive effect, prolyl hydroxylase inhibitors stabilize HIF-1 in the osteoclast progenitor cells and increase their vascular endothelial growth factor production, suggesting a contribution of this cell population to angiogenesis, which is crucial during bone regeneration. 25 However, in addition to the available in vitro studies, further in vivo studies are required that address the effect of supplementation of bone substitute materials with prolyl hydroxylase inhibitors on the resorption kinetics and angiogenesis of during bone regeneration.…”

“…27 Data from previous studies where the mouse cell line RAW 264.7 and rat primary hematopoietic stem cells were exposed to DMOG and DFO in osteoclastogenesis assays support our observations. 25 However, hypoxia and prolyl hydroxylase inhibitors have also been shown to enhance resorption activity in human culture systems. 24 In this system, numbers of osteoclasts remained unchanged in the groups treated with the prolyl hydroxylase inhibitors DFO and L-MIM, whereas DMOG decreased their numbers.…”

“…25 When prolyl hydroxylase inhibitors are released from biomaterials, migrating mesenchymal and hematopoietic progenitor cells are exposed to a continuum of concentrations of the inhibitors. 10 The current data from bone substitute materials loaded with prolyl hydroxylase inhibitors focuses on the pro-angiogenic and therefore anabolic effects of the biomaterial.…”