FGF23 decreases renal NaPi-2a and NaPi-2c expression and induces hypophosphatemia in vivo predominantly via FGF receptor 1

Abstract

Fibroblast growth factor-23 (FGF23) is a phosphaturic hormone that contributes to several hypophosphatemic disorders by reducing the expression of the type II sodium-phosphate cotransporters (NaPi-2a and NaPi-2c) in the kidney proximal tubule and by reducing serum 1,25-dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] levels. The FGF receptor(s) mediating the hypophosphatemic action of FGF23 in vivo have remained elusive. In this study, we show that proximal tubules express FGFR1, -3, and -4 but not FGFR2 mRNA. To determine which of these three FGFRs mediates FGF23's hypophosphatemic actions, we characterized phosphate homeostasis in FGFR3(-/-) and FGFR4(-/-) null mice, and in conditional FGFR1(-/-) mice, with targeted deletion of FGFR1 expression in the metanephric mesenchyme. Basal serum phosphorus levels and renal cortical brush-border membrane (BBM) NaPi-2a and NaPi-2c expression were comparable between FGFR1(-/-), FGFR3(-/-), and FGFR4(-/-) mice and their wild-type counterparts. Administration of FGF23 to FGFR3(-/-) mice induced hypophosphatemia in these mice (8.0 +/- 0.4 vs. 5.4 +/- 0.3 mg/dl; p < or = 0.001) and a decrease in renal BBM NaPi-2a and NaPi-2c protein expression. Similarly, in FGFR4(-/-) mice, administration of FGF23 caused a small but significant decrease in serum phosphorus levels (8.7 +/- 0.3 vs. 7.6 +/- 0.4 mg/dl; p < or = 0.001) and in renal BBM NaPi-2a and NaPi-2c protein abundance. In contrast, injection of FGF23 into FGFR1(-/-) mice had no effects on serum phosphorus levels (5.6 +/- 0.3 vs. 5.2 +/- 0.5 mg/dl) or BBM NaPi-2a and NaPi-2c expression. These data show that FGFR1 is the predominant receptor for the hypophosphatemic action of FGF23 in vivo, with FGFR4 likely playing a minor role.

Fig. 1.

A: mRNA expression of different FGF receptors (FGFRs) in the proximal tubule. Proximal tubules were dissected, total RNA was isolated, and the products of RT-PCR were fractionated on a 1% agarose gel. FGFR1, FGFR3, and FGFR4 but not FGFR2 mRNA are expressed in the proximal tubule. The details of the primer sequences are described in methods. Either of the FGFR3 primer sets was used. The negative control for each primer set was a RT-PCR in which RT (reverse transcriptase) was omitted. B: absence of FGFR1 transcript in the proximal tubule of FGFR1^−/− mice. Proximal tubules were dissected, total RNA was isolated, and the products of RT-PCR were fractionated on a 1% agarose gel. FGFR1 mRNA was not detected in the proximal tubules of FGFR1^−/− mice, whereas FGFR3 and FGFR4 mRNA are expressed in the proximal tubule. RT-PCR in which RT was omitted served as a negative control, and mRNA from a wild-type mouse was used as a positive control.

Fig. 2.

Renal cortical brush-border membrane (BBM) type II sodium-phosphate cotransporter (NaPi-2a) protein expression in FGFR^−/− mice at baseline. NaPi-2a expression relative to β-actin from renal cortical BBMV of FGFR^−/− mice was analyzed by immunoblotting. NaPi-2a expression at baseline did not differ between FGFR^−/− mice and their respective controls. Values are means ± SE.

Fig. 3.

Renal cortical BBM NaPi-2c protein expression in FGFR^−/− mice at baseline. NaPi-2c expression relative to β-actin from renal cortical BBM vesicles (BBMV) of FGFR^−/− mice was analyzed by immunoblotting. NaPi-2c expression at baseline did not differ between FGFR^−/− mice and their respective controls. Values are means ± SE.

Fig. 4.

Effect of exogenous FGF23 on renal cortical BBMV NaPi-2a expression in FGFR1^−/− mice. Shown are immunoblots of renal cortical BBM NaPi-2a and β-actin. NaPi-2a protein expression (relative to β-actin expression) did not change in FGFR1^−/− mice in response to FGF23 treatment, whereas, as expected, it decreased significantly in wild-type mice. Values are means ± SE.

Fig. 5.

Effect of exogenous FGF23 on renal cortical BBMV NaPi-2c expression in FGFR1^−/− mice. Shown are immunoblots of renal cortical BBM NaPi-2c and β-actin. NaPi-2c protein expression (relative to β-actin expression) did not change in FGFR1^−/− mice in response to FGF23 treatment, while, as expected, it decreased significantly in wild-type mice. Values are means ± SE.

Fig. 6.

Effect of exogenous FGF23 on renal cortical BBM NaPi-2a protein expression in FGFR3^−/− and FGFR4^−/− Mice. Shown are immunoblots of renal cortical BBM NaPi-2a and β-actin. After FGF23 administration, NaPi-2a (relative to β-actin expression) significantly decreased in FGFR3^−/− and FGFR4^−/− mice similar to that seen in their wild-type counterparts. Values are means ± SE.

Fig. 7.

Effect of Exogenous FGF23 on renal cortical BBM NaPi-2c protein expression in FGFR3^−/− and FGFR4^−/− mice. Shown are immunoblots of renal cortical BBM NaPi-2c and β-actin. After FGF23 administration, NaPi-2c (relative to β-actin expression) significantly decreased in FGFR3^−/− and FGFR4^−/− mice similar to that seen in their wild-type counterparts. Values are means ± SE.