Excessive Osteocytic Fgf23 Secretion Contributes to Pyrophosphate Accumulation and Mineralization Defect in Hyp Mice

Abstract

X-linked hypophosphatemia (XLH) is the most frequent form of inherited rickets in humans caused by mutations in the phosphate-regulating gene with homologies to endopeptidases on the X-chromosome (PHEX). Hyp mice, a murine homologue of XLH, are characterized by hypophosphatemia, inappropriately low serum vitamin D levels, increased serum fibroblast growth factor-23 (Fgf23), and osteomalacia. Although Fgf23 is known to be responsible for hypophosphatemia and reduced vitamin D hormone levels in Hyp mice, its putative role as an auto-/paracrine osteomalacia-causing factor has not been explored. We recently reported that Fgf23 is a suppressor of tissue nonspecific alkaline phosphatase (Tnap) transcription via FGF receptor-3 (FGFR3) signaling, leading to inhibition of mineralization through accumulation of the TNAP substrate pyrophosphate. Here, we report that the pyrophosphate concentration is increased in Hyp bones, and that Tnap expression is decreased in Hyp-derived osteocyte-like cells but not in Hyp-derived osteoblasts ex vivo and in vitro. In situ mRNA expression profiling in bone cryosections revealed a ~70-fold up-regulation of Fgfr3 mRNA in osteocytes versus osteoblasts of Hyp mice. In addition, we show that blocking of increased Fgf23-FGFR3 signaling with anti-Fgf23 antibodies or an FGFR3 inhibitor partially restored the suppression of Tnap expression, phosphate production, and mineralization, and decreased pyrophosphate concentration in Hyp-derived osteocyte-like cells in vitro. In vivo, bone-specific deletion of Fgf23 in Hyp mice rescued the suppressed TNAP activity in osteocytes of Hyp mice. Moreover, treatment of wild-type osteoblasts or mice with recombinant FGF23 suppressed Tnap mRNA expression and increased pyrophosphate concentrations in the culture medium and in bone, respectively. In conclusion, we found that the cell autonomous increase in Fgf23 secretion in Hyp osteocytes drives the accumulation of pyrophosphate through auto-/paracrine suppression of TNAP. Hence, we have identified a novel mechanism contributing to the mineralization defect in Hyp mice.

Osteocyte Death and Bone Overgrowth in Mice Lacking Fibroblast Growth Factor Receptors 1 and 2 in Mature Osteoblasts and Osteocytes

“…Fgfr3 is expressed in mature osteoblasts and osteocytes . Because most osteocytes are lost in OC‐Fgfr1/2 DCKO mice at 12 weeks, it is likely that Fgfr3 expression is increased in mature osteoblasts or other cell types.…”

“…Although it is not known if FGFR3 signaling is increased, the combination of increased FGF23 and increased FGFR3 provides a potentially active ligand‐receptor pair. One function of FGF23‐FGFR3 signaling in bone is to suppress tissue nonspecific alkaline phosphatase (TNAP) expression and reduce mineralization . In the case of OC‐Fgfr1/2 DCKO mice, this could be a compensatory response to the massive increase in bone formation at 12 weeks.…”

“…Mechanical unloading and PTH signaling stimulate RANKL expression in osteocytes . Osteocytes are also a major source of FGF23, an endocrine member of the fibroblast growth factor (FGF) family that primarily signals to the kidney proximal tubules to suppress phosphate resorption but also signals to FGF receptor 3 (FGFR3) in osteocytes to inhibit mineralization …”

“…In the periosteum of mature cortical bone, FGFR2 expression is lost, while FGFR1 expression persists in a subpopulation of perivascular cells . Additionally, cultured calvarial osteoblasts and osteocytes and osteocyte‐enriched fractions from cortical bone express FGFR1 and FGFR3 . During fracture repair, expression of both FGFR1 and FGFR2 are increased .…”

Biology and Mechanisms of Action of the Vitamin D Hormone

“…In the case of calcium, the level of this ion is continually monitored by the calcium-sensing receptor in the parathyroid gland, which responds by increasing parathyroid gland secretion of PTH when calcium content decreases. While the consequence of calcium liberation from the skeleton increases phosphate levels as well, both PTH and FGF23 function collectively in the kidney to promote phosphate diuresis through a mechanism involving cellular relocation of multiple sodium-phosphate transporters (29,67). FGF23 represents the long sought-after phosphate regulating hormone or phosphatonin that is not only induced by phosphate when it is in abundance but by 1,25(OH) 2 D 3 as well, although neither of these mechanisms are currently understood (28,68,69).…”