High extracellular inorganic phosphate concentration inhibits RANK–RANKL signaling in osteoclast‐like cells

Abstract

In this work, we investigated the effect of inorganic phosphate (Pi) on the differentiation of monocyte/macrophage precursors into an "osteoclastic" phenotype, and we delineated the molecular mechanisms which could be involved in this phenomenon. This was achieved by stimulating human peripheral blood monocytic cells and RAW 264.7 monocyte-macrophage precursor cells to differentiate into osteoclast-like cells in the presence of receptor activator of NF-kappaB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). RANKL has been previously reported to stimulate the signaling kinases ERK 1/2, p38, Akt, JNK, and the DNA-binding activity of the transcription factors AP-1 and NF-kappaB. Increase in extracellular Pi concentration (1.5-4.5 mM) dose-dependently inhibits both osteoclastic differentiation and bone resorption activity induced by RANKL and M-CSF. Pi was found to specifically inhibit the RANKL-induced JNK and Akt activation, while RANKL-induced p38 and ERK 1/2 phosphorylation were not significantly affected. Moreover, we found that Pi significantly reduced the RANKL-stimulated DNA-binding activity of NF-kappaB. The effects of Pi on osteoclast differentiation and DNA-binding activity of NF-kappaB were prevented by Foscarnet, a sodium-phosphate cotransport inhibitor, suggesting that the effects of Pi occur subsequently to its intake. These results demonstrate that Pi downregulates the differentiation of osteoclasts via a negative feedback exerted on RANK-RANKL signaling.

PKA-induced Receptor Activator of NF-κB Ligand (RANKL) Expression in Vascular Cells Mediates Osteoclastogenesis but Not Matrix Calcification

“…For example, oxidized phospholipids did not induce RANKL expression in vascular smooth muscle cells. Non-atherogenic factors also inhibit osteoclasts in the vasculature; Mozar et al (42) found that inorganic phosphate inhibits osteoclastic activity by inhibiting JNK and Akt activation. Combined with low expression of osteoclast activators, and secretion of osteoclast inhibitory factors, vascular cells may limit osteoclast formation, thereby promoting a milieu favoring progression of ossification.…”

Human Alternative Macrophages Populate Calcified Areas of Atherosclerotic Lesions and Display Impaired RANKL-Induced Osteoclastic Bone Resorption Activity

“…Monocytes were differentiated into osteoclasts by growth on cortical bone slices obtained from bovine femora, 23 in the absence or in the presence of RANKL/MCSF, with or without IL-4. Mature osteoclasts were stained for TRAP expression using a commercially available kit (leukocyte acid phosphatase staining kit; Sigma).…”

Phosphate (Pi)-regulated heterodimerization of the high-affinity sodium-dependent Pi transporters PiT1/Slc20a1 and PiT2/Slc20a2 underlies extracellular Pi sensing independently of Pi uptake

“…Interestingly, the Pi-dependent activation of the ERK1/2 pathway up-regulated the gene expression of the mineralization inhibitors matrix gla protein (Mgp) and osteopontin (Opn), most likely setting off a feed-back mechanism to control Pi-induced mineralization (7,16,18,19). The elevated extracellular Pi level was also shown to be important in osteoblast proliferation and differentiation (16,(20)(21)(22)(23)(24), cementoblast formation (25), odontoblast differentiation (26,27) and osteoclast differentiation (28)(29)(30).…”