Dysregulated Gene Expression in the Primary Osteoblasts and Osteocytes Isolated from Hypophosphatemic Hyp Mice

Abstract

Osteocytes express multiple genes involved in mineral metabolism including PHEX, FGF23, DMP1 and FAM20C. In Hyp mice, a murine model for X-linked hypophosphatemia (XLH), Phex deficiency results in the overproduction of FGF23 in osteocytes, which leads to hypophosphatemia and impaired vitamin D metabolism. In this study, to further clarify the abnormality in osteocytes of Hyp mice, we obtained detailed gene expression profiles in osteoblasts and osteocytes isolated from the long bones of 20-week-old Hyp mice and wild-type (WT) control mice. The expression of Fgf23, Dmp1, and Fam20c was higher in osteocytic cells than in osteoblastic cells in both genotypes, and was up-regulated in Hyp cells. Interestingly, the up-regulation of these genes in Hyp bones began before birth. On the other hand, the expression of Slc20a1 encoding the sodium/phosphate (Na+/Pi) co-transporter Pit1 was increased in osteoblasts and osteocytes from adult Hyp mice, but not in Hyp fetal bones. The direct effects of extracellular Pi and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] on isolated osteoblastic and osteocytic cells were also investigated. Twenty-four-hour treatment with 10−8 M 1,25(OH)2D3 increased the expression of Fgf23 in WT osteoblastic cells but not in osteocytic cells. Dmp1 expression in osteocytic cells was increased due to the 24-hour treatment with 10 mM Pi and was suppressed by 10−8 M 1,25(OH)2D3 in WT osteocytic cells. We also found the up-regulation of the genes for FGF1, FGF2, their receptors, and Egr-1 which is a target of FGF signaling, in Hyp osteocytic cells, suggesting the activation of FGF/FGFR signaling. These results implicate the complex gene dysregulation in osteoblasts and osteocytes of Hyp mice, which might contribute to the pathogenesis.

Interleukin-1-induced acute bone resorption facilitates the secretion of fibroblast growth factor 23 into the circulation

“…The expression profiles of the osteoblastic marker Kera and osteocytic marker Sost in the freshly isolated cells of each fraction suggested that Fr 6/7 and 8/9 were osteocyte-rich. As we previously reported [32], the expression of Fgf23 was higher in Fr 6/7 and 8/9 than in the earlier fractions, confirming its higher expression in osteocytes than in osteoblasts (Fig. 7a).…”

“…Kera, which encodes the proteoglycan keratocan, was previously reported to be highly expressed in osteoblasts in vivo [31]. In parallel with our previous report [32], we found that the expression of Kera was the strongest in Fr 3 and was almost undetectable in Fr 6/7 and 8/9. On the other hand, the expression of Sost, encoding sclerostin, a marker for mature osteocytes, was the highest in Fr 8/9 (Fig.…”

“…Data are expressed as the mean ± SEM J Bone Miner Metab 1b (10 ng/ml) or vehicle for 24 h, before RNA was extracted to examine the expression of Fgf23. Since we have recently reported that 1,25(OH) 2 D 3 up-regulated Fgf23 expression in the primary osteoblasts isolated by this method [32], we also examined the effects of 1,25(OH) 2 D 3 for comparison. The expression of Fgf23 was significantly increased by 24-h treatment with 1,25(OH) 2 D 3 in the osteoblastic cells, and a similar tendency was observed in the osteocytic cells.…”

Conditional Deletion of Murine Fgf23: Interruption of the Normal Skeletal Responses to Phosphate Challenge and Rescue of Genetic Hypophosphatemia

“…It is unclear why rescue of specific portions of the skeletal phenotype, such as shape, occurs when FGF23 bioactivity is absent during global deletion of Fgf23 (28), but bone length is not. Although not directly tested herein, with similarly reduced baseline serum FGF23 concentrations between the flox- Fgf23 /Col2.3-cre and flox- Fgf23 /Dmp1-cre mice, taken together with early expression of Dmp1 in Hyp osteoblasts/osteocytes (59), it is anticipated that a Hyp /flox- Fgf23 /Dmp1-cre cross could produce a similar rescue. A circulating form of αKL (‘cKL’) may interact with osteoblasts (60), but whether blood αKL affects bone development in the Fgf23 -null environment is currently unknown.…”

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

“…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 .…”