Targeted inactivation of Npt2 in mice leads to severe renal phosphate wasting, hypercalciuria, and skeletal abnormalities

Abstract

Npt2 encodes a renal-specific, brush-border membrane Na ؉ -phosphate (P i ) cotransporter that is expressed in the proximal tubule where the bulk of filtered P i is reabsorbed. Mice deficient in the Npt2 gene were generated by targeted mutagenesis to define the role of Npt2 in the overall maintenance of P i homeostasis, determine its impact on skeletal development, and clarify its relationship to autosomal disorders of renal P i reabsorption in humans. Homozygous mutants (Npt2 ؊/؊ ) exhibit increased urinary P i excretion, hypophosphatemia, an appropriate elevation in the serum concentration of 1,25-dihydroxyvitamin D with attendant hypercalcemia, hypercalciuria and decreased serum parathyroid hormone levels, and increased serum alkaline phosphatase activity. These biochemical features are typical of patients with hereditary hypophosphatemic rickets with hypercalciuria (HHRH), a Mendelian disorder of renal P i reabsorption. However, unlike HHRH patients, Npt2؊/؊ mice do not have rickets or osteomalacia. At weaning, Npt2 ؊/؊ mice have poorly developed trabecular bone and retarded secondary ossification, but, with increasing age, there is a dramatic reversal and eventual overcompensation of the skeletal phenotype. Our findings demonstrate that Npt2 is a major regulator of P i homeostasis and necessary for normal skeletal development.Inorganic phosphate (P i ) is essential for a variety of cellular processes including skeletal mineralization. Because a disturbance in P i availability can affect the functional integrity of many organ systems, specialized tissues have evolved to maintain the extracellular P i concentration. In mammals, the regulation of P i homeostasis is largely determined by the kidney, with 60-70% of the filtered P i load reclaimed in the proximal segment of the nephron (1). Transepithelial transport of P i from the renal lumen to the blood compartment involves uptake across the brushborder membrane (BBM), translocation across the cell, and efflux across the basolateral membrane (2). Evidence suggests that P i transport across the BBM is the rate-limiting step in the overall P i reabsorptive process and the major site for its regulation (2, 3). Kinetics studies demonstrated that the transport is mediated by high capacity, low affinity, and low capacity, high affinity Na ϩ -P i cotransport systems (4).Recently, cDNAs encoding two distinct low capacity, high affinity, renal BBM Na ϩ -P i cotransporters (NPT1 and NPT2) that share only 20% identity have been identified in several mammalian species by expression and homology cloning (5-13). NPT2 is expressed exclusively in the proximal convoluted tubule and is regulated by P i intake (14) and parathyroid hormone (PTH) (15), factors known to regulate renal BBM Na ϩ -P i cotransport. In contrast, NPT1 is expressed throughout the proximal tubule and is not subject to regulation by dietary P i (10).X-chromosome-linked and autosomal disorders of renal P i reabsorption have been described in humans (16). Both are characterized by growth retardation, ...

Loss of function of NaPiIIa causes nephrocalcinosis and possibly kidney insufficiency

“…In mice, a homozygous single amino acid substitution in NaPiIIa altered phosphate (Pi) homeostasis and led to renal calcification [5]. NaPiIIa knock-out mice show increased urinary Pi excretion, hypophosphatemia, elevated serum 1,25 dihydroxyvitamin D levels, decreased serum PTH levels, hypercalcemia and hypercalciuria [6].…”

“…Extensive studies in murine models have shown that either complete deletion of the SLC34A1 gene or homozygosity loss-offunction NaPiIIa mutations reduce renal reabsorption of phosphate, alter phosphate homeostasis and lead to renal calcification [5], [6].…”

Disorders of Renal Tubular Phosphate Transport

“…The V max decrease is secondary to a corresponding decrement in the renal abundance of type IIa Na/P i cotransporter (Npt2) mRNA and protein in kidneys of Hyp and Gy mice (52,53), consistent with the central role of the type IIa transporter in renal P i reabsorption (14). In addition, type IIc protein abundance is decreased in renal brush border membranes of Hyp mice (Ͻ10% of normal) (17).…”

“…A key role for the type IIa transporter in renal P i handling in adult animals is supported by the following observations: (1) exclusive proximal tubular brush border membrane localization (13); (2) disruption of the gene encoding type IIa in mice (Npt2-/-) leads to impaired renal P i reabsorption and a 70 to 80% loss of brush border membrane Na/P i cotransport (14); (3) abundance of type IIa protein in the brush border membrane correlates with Na/P i cotransport activity under a variety of physiologic/pathophysiologic conditions (for review, see reference 1); (4) residual brush border membrane Na/P i cotransport in Npt2-/-mice is not responsive to regulatory phenomena (e.g., parathyroid hormone [PTH] and dietary P i intake [15,16]; see below). A candidate for residual Na/P i cotransport activity in brush border membranes of type IIa knockout mice is the type IIc cotransporter (12).…”

Phosphate transporters of the SLC20 and SLC34 families

“…(Villa-Bellosta and Sorribas)). NaPi-IIa seems to be the major transporter in murine kidneys, as mice deficient for this cotransporter are hypophosphatemic due to a major loss of P i in urine (Beck et al, 1998). In contrast, urinary excretion and circulating levels of P i are normal in NaPi-IIc -/-animals ).…”