Secreted frizzled-related protein 4 is a potent tumor-derived phosphaturic agent

Abstract

Tumors associated with osteomalacia elaborate the novel factor(s), phosphatonin(s), which causes phosphaturia and hypophosphatemia by cAMP-independent pathways. We show that secreted frizzled-related protein-4 (sFRP-4), a protein highly expressed in such tumors, is a circulating phosphaturic factor that antagonizes renal Wnt-signaling. In cultured opossum renal epithelial cells, sFRP-4 specifically inhibited sodium-dependent phosphate transport. Infusions of sFRP-4 in normal rats over 2 hours specifically increased renal fractional excretion of inorganic phosphate (FEPi) from 14% +/- 2% to 34% +/- 5% (mean +/- SEM, P < 0.01). Urinary cAMP and calcium excretion were unchanged. In thyro-parathyroidectomized rats, sFRP-4 increased FEPi from 0.7% +/- 0.2% to 3.8% +/- 1.2% (P < 0.05), demonstrating that sFRP-4 inhibits renal inorganic phosphate reabsorption by PTH-independent mechanisms. Administration of sFRP-4 to intact rats over 8 hours increased FEPi, decreased serum phosphate (1.95 +/- 0.1 to 1.53 +/- 0.09 mmol/l, P < 0.05) but did not alter serum 1alpha, 25-dihydroxyvitamin D, renal 25-hydroxyvitamin D 1alpha-hydroxylase cytochrome P450, and sodium-phosphate cotransporter mRNA concentrations. Infusion of sFRP-4 antagonizes Wnt action as demonstrated by reduced renal beta-catenin and increased phosphorylated beta-catenin concentrations. The sFRP-4 is detectable in normal human serum and in the serum of a patient with tumor-induced osteomalacia. Thus, sFRP-4 displays phosphatonin-like properties, because it is a circulating protein that promotes phosphaturia and hypophosphatemia and blunts compensatory increases in 1alpha, 25-dihydroxyvitamin D.

Figure 1

Effect of sFRP-4 on sodium-dependent phosphate uptake in OK cells maintained in culture. *P < 0.05.

Figure 2

Effect of the infusion of sFRP-4 on solute excretion in intact rats. Intact rats were administered sFRP-4 (black bars; group 2) at a dose of 0.3 μg/kg/h or vehicle (white bars; group 1) by intravenous infusion over a period of 2 hours. C₁ indicates equilibration period prior to the infusion of sFRP-4 or vehicle. C₂ indicates experimental period during which sFRP-4 or vehicle was infused. Fractional excretion of inorganic phosphate, sodium, and calcium were measured as described in the text. *P < 0.05.

Figure 3

Effect of the infusion of sFRP-4 on solute excretion in TPTX rats. TPTX rats were administered sFRP-4 (black bars; group 4) at a dose of 0.3 μg/kg/h or vehicle (white bars; group 3) by intravenous infusion over a period of 2 hours. C₁ indicates collection period prior to the infusion of sFRP-4 or vehicle. C₂ indicates collection period during which sFRP-4 or vehicle was infused. Fractional excretion of inorganic phosphate, sodium, and calcium were measured as described in the text. *P < 0.05.

Figure 4

The effect infusion of vehicle (VEH) or sFRP-4 (0.3 μg/kg/h) for 8 hours, or the effect of phosphate deprivation induced by a low-phosphate diet phosphate (LPD) compared with a normal-phosphate diet (NPD), on the concentration of 25-hydroxyvitamin D 1α-hydroxylase cytochrome P450 mRNA concentrations in kidney.*P < 0.05. 25(OH)D, 25-hydroxyvitamin D.

Figure 5

Western blots of renal homogenates obtained from rats infused with vehicle (control) or sFRP-4 for 8 hours. Ab’s against β-catenin (upper panel) or phosphorylated β-catenin (phospho β-catenin) (lower panel) were used to detect proteins as described in Methods.