Normal Pi Diet Research Articles

High-phosphate diet causes atrial remodeling and increases atrial fibrillation vulnerability via STAT3/NF-κB signaling and oxidative stress.

Hyperphosphatemia is associated with adverse cardiovascular outcomes in both the general population and patients with end-stage renal disease. We evaluated whether high inorganic phosphate (Pi) intake causes atrial remodeling and increased atrial fibrillation (AF) risk. The 5/6 nephrectomized chronic kidney disease (CKD) mice were fed a high-Pi (2%) diet for 10 weeks. AF vulnerability was evaluated through transesophageal burst atrial pacing. Phosphoproteomic, Western blotting, and immunohistochemistry were used to evaluate the effects of high Pi in atrial fibroblasts, atrial myocytes, and HL-1 myocytes. CKD and sham mice fed a high-Pi diet exhibited increased AF vulnerability, atrial fibrosis, and oxidative stress compared with mice fed a normal diet. Compared with normal (1 mM) Pi, high (2 mM) Pi significantly increased the activity of atrial fibroblasts and mitochondrial oxidative stress. Phosphoproteomic analysis revealed that compared with normal Pi, high Pi considerably increased the phosphorylation of intracellular proteins in atrial fibroblasts, including proteins related to NF-κB signaling and STAT3. Inhibition of NF-κB, STAT3, and Nox4 by small interfering RNA reduced the high-Pi-induced expression of collagen. In HL-1 myocytes, the high Pi induced the degradation of myofibril proteins and hyperphosphorylation of RyR2, which was abolished by Nox4 and CaMKII inhibition. Switching back to a normal-Pi diet improved the atrial abnormalities induced by high-Pi diet. High-Pi intake causes atrial structural and electrical remodeling and increases AF vulnerability, which is mediated through STAT3/NF-κB signaling and oxidative stress. High dietary Pi intake can exert detrimental effects on atria and may increase AF risk.

High Phosphate Induces and Klotho Attenuates Kidney Epithelial Senescence and Fibrosis

Cellular senescence is an irreversible cell growth arrest and is associated with aging and age-related diseases. High plasma phosphate (Pi) and deficiency of Klotho contribute to aging and kidney fibrosis, a pathological feature in the aging kidney and chronic kidney disease. This study examined the interactive role of Pi and Klotho in kidney senescence and fibrosis. Homozygous Klotho hypomorphic mice had high plasma Pi, undetectable Klotho in plasma and kidney, high senescence with massive collagen accumulation in kidney tubules, and fibrin deposits in peritubular capillaries. To examine the Pi effect on kidney senescence, a high (2%) Pi diet was given to wild-type mice. One week of high dietary Pi mildly increased plasma Pi, and upregulated kidney p16/p21 expression, but did not significantly decrease Klotho. Two weeks of high Pi intake led to increase in plasminogen activator inhibitor (PAI)-1, and decrease in kidney Klotho, but still without detectable increase in kidney fibrosis. More prolonged dietary Pi for 12 weeks exacerbated kidney senescence and fibrosis; more so in heterozygous Klotho hypomorphic mice compared to wild-type mice, and in mice with chronic kidney disease (CKD) on high Pi diet compared to CKD mice fed a normal Pi diet. In cultured kidney tubular cells, high Pi directly induced cellular senescence, injury and epithelial-mesenchymal transition, and enhanced H2O2-induced cellular senescence and injury, which were abrogated by Klotho. Fucoidan, a bioactive molecule with multiple biologic functions including senescence inhibition, blunted Pi-induced cellular senescence, oxidation, injury, epithelial-mesenchymal transition, and senescence-associated secretary phenotype. In conclusion, high Pi activates senescence through distinct but interconnected mechanisms: upregulating p16/p21 (early), and elevating plasminogen activator inhibitor-1 and downregulating Klotho (late). Klotho may be a promising agent to attenuate senescence and ameliorate age-associated, and Pi-induced kidney degeneration such as kidney fibrosis.

High dietary phosphate intake induces hypertension and augments exercise pressor reflex function in rats.

An increasing number of studies have linked high dietary phosphate (Pi) intake to hypertension. It is well established that the rise in sympathetic nerve activity (SNA) and blood pressure (BP) during physical exertion is exaggerated in many forms of hypertension, which are primarily mediated by an overactive skeletal muscle exercise pressor reflex (EPR). However, it remains unknown whether high dietary Pi intake potentiates the EPR-mediated SNA and BP response to exercise. Accordingly, we measured renal SNA (RSNA) and mean BP (MBP) in normotensive Sprague-Dawley rats fed a normal Pi diet (0.6%, n = 13) or high Pi diet (1.2%, n = 13) for 3 mo. As previously reported, we found that resting BP was significantly increased by 1.2% Pi diet in both conscious and anesthetized animals. Activation of the EPR by electrically induced hindlimb contraction triggered greater increases in ΔRSNA and ΔMBP in the 1.2% compared with 0.6% Pi group (126 ± 25 vs. 42 ± 9%; 44 ± 5 vs. 14 ± 2 mmHg, respectively, P < 0.01). Activation of the muscle mechanoreflex, a component of the EPR, by passively stretching hindlimb muscle also evoked greater increases in ΔRSNA and ΔMBP in the 1.2% compared with 0.6% Pi group (109 ± 27 vs. 24 ± 7%, 38 ± 7 vs. 8 ± 2 mmHg, respectively, P < 0.01). A similar response was produced by hindlimb intra-arterial capsaicin administration to stimulate the metaboreflex arm of the EPR. Thus, our data demonstrate a novel action of dietary Pi loading in augmenting EPR function through overactivation of both the muscle mechanoreflex and metaboreflex.

Effect of ischemia reperfusion on sodium-dependent phosphate transport in renal brush border membranes

The effect of ischemia induced acute renal failure (ARF) on the transport of phosphate (Pi) after early (15–30 min) and prolonged (60 min) ischemia in the brush border membrane vesicles (BBMV) from rat renal cortex was studied. Sodium-dependent transport of Pi declined significantly and progressively due to ischemia. Western blot analysis of BBM from ischemic rats showed decreased expression of NaPi-2. A compensatory increase was observed in Pi uptake in BBMV from contralateral kidneys. There was no significant difference in NaPi-2 expression between BBMV from sham and contralateral kidneys. Early blood reperfusion for 15 min after 30 min ischemia caused further decline in Pi uptake. Prolonged reperfusion for 120 min caused partial reversal of transport activities in 30-min ischemic rats. However, no improvement in the transport of Pi was observed in 60-min ischemic rats after 120 min of blood reperfusion. Kinetic studies showed that the effect of ischemia and blood reperfusion was dependent on the V max of the Na-Pi transporter. Western blot analysis showed increased expression of NaPi-2 in the BBMs from ischemia–reperfusion animals. Further, a shift in the association of Na ions to transport one molecule of Pi was observed under different extracellular Na concentrations [Na] o. Feeding rats with low Pi diet and/or treatment with thyroid hormone (T3) prior to ischemia resulted in increased basal Pi transport. Ischemia caused similar decline in Pi transport in BBM from LPD and/or T3 animals. However, recovery in these animals was faster than the normal Pi diet fed (NPD) animals. The study suggests a change in the intrinsic properties of the Na-Pi transporter in rat kidneys due to ischemia. The study also indicates that treatment with T3 and feeding LPD prior to ischemia caused faster recovery of phosphate uptake due to ischemia–reperfusion injury.

Internalization of renal type IIc Na-Pi cotransporter in response to a high-phosphate diet

Dietary phosphate levels regulate the renal brush-border type IIa Na-Pi cotransporter. Another Na-Pi cotransporter, type IIc, colocalizes with type IIa Na-Pi cotransporter in the apical membrane of renal proximal tubular cells. The goal of the present study was to determine whether dietary phosphate levels also rapidly regulate the type IIc Na-Pi cotransporter. Type IIa and type IIc transporter protein levels were increased in rats chronically fed a low-Pi diet compared with those fed a normal-Pi diet. Two hours after beginning a high-Pi diet, type IIa transporter levels were decreased, whereas type IIc protein levels remained unchanged. Western blot analysis of brush-border membrane prepared 4 h after beginning a high-Pi diet showed a significant reduction in type IIc transporter protein levels, and immunohistochemistry showed translocation of the type IIc-immunoreactive signal from the entire brush border to subapical membrane. Membrane fractionation studies revealed a decrease in apical membrane type IIc protein without changes in total cortical type IIc protein, which is compatible with redistribution of type IIc protein from the apical membrane to the dense membrane fraction. The microtubule-disrupting reagent colchicine prevented this reduction in apical type IIc transporter at the apical membrane but had no effect on type IIa transporter levels. These data suggest that the type IIc Na-Pi cotransporter level is rapidly regulated by rapid adaptation to dietary Pi in a microtubule-dependent manner. Furthermore, the mechanisms of the internalization of the type IIc transporter are distinct from those of the type IIa transporter.

Renal tubular sites of increased phosphate transport and NaPi-2 expression in the juvenile rat.

To determine the tubular sites and mechanisms involved in enhanced renal phosphate (P(i)) reabsorption seen in the juvenile animal, renal micropuncture experiments were performed in acutely thyroparathyroidectomized adult (>14 wk old) and juvenile (4 wk old) male Wistar rats fed either a normal P(i) diet (NPD, 0.6% P(i)) or low P(i) diet (0.07% P(i)) for 2 days, in the presence and absence of parathyroid hormone (PTH). P(i) reabsorption was greater in proximal convoluted (PCT) and straight tubules (PST) of the juvenile compared with adult rats fed NPD, whether or not PTH was present. These findings were consistent with a greater P(i) uptake in brush-border membrane (BBM) vesicles from both superficial (SC) and outer juxtamedullary (JMC) cortices of juvenile animals. Western blot analysis revealed a 2- and 1.8-fold higher amount of NaPi-2 protein in the SC and JMC, respectively, in juvenile rats. Immunofluorescence microscopy also indicated that NaPi-2 protein expression was present in the proximal tubule (PT) BBM to a greater extent in juvenile rats. Dietary P(i) restriction in juvenile rats resulted in a significant increase in P(i) reabsorption in the PCT and PST segments. NaPi-2 expression in the PT BBM was also increased, as was the expression of intracellular NaPi-2 protein. These studies indicate that P(i) reabsorption in both the PCT and PST segments of the renal tubule contributes to the attenuated response to PTH in the normal juvenile animal. In addition, dietary P(i) restriction in the juvenile rat upregulates BBM NaPi-2 expression, which is associated with a further increase in proximal tubular P(i) reabsorption.

Regulation of intestinal Na+-dependent phosphate co-transporters by a low-phosphate diet and 1,25-dihydroxyvitamin D3

In a study of the rat intestinal Pi transport system, an activator protein for rat Na/Pi co-transport system (PiUS) was isolated and characterized. We also investigated the effects of restriction of vitamin D and Pi (two of the most important physiological and pathophysiological regulators of Pi absorption in the small intestine) on intestinal Pi transport activity and the expression of Na/Pi co-transporters that are expressed in rat small intestine. Rat PiUS encodes a 424-residue protein with a calculated molecular mass of 51463 Da. The microinjection of rat PiUS into Xenopus oocytes markedly stimulated Na+-dependent Pi co-transport activity. In rats fed with a low-Pi diet, Na+-dependent Pi co-transport activity was increased approx. 2-fold compared with that of rats fed a normal Pi diet. Kinetic studies demonstrated that this increased activity was due to an elevation of Vmax but not Km. The PiUS mRNA levels showed an approximate doubling in the rats fed with the low-Pi diet compared with those fed with the normal Pi diet. In addition, after the administration of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] to vitamin D-deficient animals, the Pi uptake was significantly increased in the Na+-dependent component in the brush border membrane vesicle (BBMV) at 24 and 48 h. In addition, we found a further high-affinity Na/Pi co-transport system in the BBMV isolated from the vitamin D-replete animals. The levels of type III Na/Pi co-transporter PiT-2 mRNA were increased 24 and 48 h after 1,25-(OH)2D3 administration to vitamin D-deficent animals, whereas PiUS and the type IIb Na/Pi co-transporter mRNA levels were unchanged. In conclusion, we first cloned a rat activator protein, PiUS, and then studied its role along with that of other type III Na/Pi co-transporters. PiUS and PiT-2 might be important components in the regulation of the intestinal Pi transport system by Pi restriction and 1,25-(OH)2D3.

Regulation of intestinal Na+-dependent phosphate co-transporters by a low-phosphate diet and 1,25-dihydroxyvitamin D3

In a study of the rat intestinal P(i) transport system, an activator protein for rat Na/P(i) co-transport system (PiUS) was isolated and characterized. We also investigated the effects of restriction of vitamin D and P(i) (two of the most important physiological and pathophysiological regulators of P(i) absorption in the small intestine) on intestinal P(i) transport activity and the expression of Na/P(i) co-transporters that are expressed in rat small intestine. Rat PiUS encodes a 424-residue protein with a calculated molecular mass of 51463 Da. The microinjection of rat PiUS into Xenopus oocytes markedly stimulated Na(+)-dependent P(i) co-transport activity. In rats fed with a low-P(i) diet, Na(+)-dependent P(i) co-transport activity was increased approx. 2-fold compared with that of rats fed a normal P(i) diet. Kinetic studies demonstrated that this increased activity was due to an elevation of V(max) but not K(m). The PiUS mRNA levels showed an approximate doubling in the rats fed with the low-P(i) diet compared with those fed with the normal P(i) diet. In addition, after the administration of 1, 25-dihydroxyvitamin D(3) [1,25-(OH)(2)D(3)] to vitamin D-deficient animals, the P(i) uptake was significantly increased in the Na(+)-dependent component in the brush border membrane vesicle (BBMV) at 24 and 48 h. In addition, we found a further high-affinity Na/P(i) co-transport system in the BBMV isolated from the vitamin D-replete animals. The levels of type III Na/P(i) co-transporter PiT-2 mRNA were increased 24 and 48 h after 1,25-(OH)(2)D(3) administration to vitamin D-deficient animals, whereas PiUS and the type IIb Na/P(i) co-transporter mRNA levels were unchanged. In conclusion, we first cloned a rat activator protein, PiUS, and then studied its role along with that of other type III Na/P(i) co-transporters. PiUS and PiT-2 might be important components in the regulation of the intestinal P(i) transport system by P(i) restriction and 1,25-(OH)(2)D(3).

Evidence for induction of a phosphate appetite in juvenile rats.

This study examined whether dietary phosphate (Pi) restriction stimulates an appetite for Pi in the juvenile rat, which normally has a high metabolic Pi demand for growth. Juvenile Wistar rats were placed in individual cages with unrestricted access to tap water and a low (LPD, 0.02% Pi) or normal Pi diet (NPD, 0.6% Pi) for 7 days. On day 8, both groups of rats were given unlimited access to a solution of 0.3 M potassium phosphate water (PiH2O) for 8 additional days. Rats fed LPD consumed 70-100% more PiH2O then those rats fed NPD (P < 0.001). The increase in PiH2O intake resulted in a marked rise in the growth rate of rats fed LPD during days 8-15. A similar Pi intake was inducible after only 2 days of LPD and was associated with significant reductions in both plasma and cerebrospinal fluid (CSF) Pi levels; these levels remained low throughout Pi restriction, despite a significant PiH2O intake. Furthermore, the renal adaptation to enhance Pi reabsorption (TmPi) during Pi deprivation remained elevated despite enhanced PiH2O intake. Replenishment with a high-Pi diet rapidly quenched the PiH2O appetite and was associated with restoration of both plasma and CSF Pi levels. These findings suggest that an appetite for Pi can be induced in juvenile rats, perhaps through lowered plasma and CSF Pi levels. This behavioral response may serve as an additional mechanism to maintain an adequate supply of Pi necessary for growth and development of the animal.

Regulation of renal phosphate transport by acute and chronic metabolic acidosis in the rat

Metabolic acidosis results in impaired renal tubular phosphate reabsorption and proximal tubular apical brush border membrane (BBM) sodium gradient-dependent phosphate transport (Na/Pi cotransport) activity. In the present study we investigated the cellular mechanisms responsible for decreased Na/Pi cotransport activity following six hours to 10 days of metabolic acidosis induced by ingestion of NH4Cl. Urinary Pi excretion was significantly increased and BBM Na/Pi cotransport activity was progressively and significantly decreased by 18% at six hours, 24% at 12 hours, 32% at 24 hours, and 61% after 10 days of metabolic acidosis. The progressive and time-dependent decreases in BBM cotransport activity were associated with progressive decreases in BBM NaPi-2 protein (43% at 12 hr, 54% at 24 hr and 66% at 10 days) and cortical NaPi-2 mRNA (22% at 12 hr, 54% at 24 hr and 56% at 10 days) abundance. Interestingly, following six hours of metabolic acidosis, there was a significant 29% decrease in BBM NaPi-2 protein abundance that was not associated with decreases in either cortical homogenate NaPi-2 protein or cortical NaPi-2 mRNA abundance. In additional studies we found that the effects of chronic metabolic acidosis on Na/Pi cotransport activity were independent of endogenous parathyroid hormone activity, but were somewhat dependent on dietary Pi intake. In rats fed a high or a normal Pi diet metabolic acidosis caused significant decreases in Na/Pi cotransport activity, NaPi-2 protein and NaPi-2 mRNA abundance, however, in rats fed a low Pi diet the inhibitory effect of metabolic acidosis on Na/Pi cotransport were minimal and not significant. These results indicate that in chronic (> or = 12 hr) metabolic acidosis the progressive decrease in BBM Na/Pi cotransport activity is most likely mediated by decrease in BBM NaPi-2 protein and cortical mRNA abundance. In contrast, in acute (< or = 6 hr) metabolic acidosis the decrease in BBM Na/Pi cotransport activity is likely mediated by changes in the trafficking of the NaPi-2 protein that is, enhanced internalization from and/or impaired delivery of the NaPi-2 protein to the apical BBM.

Renal adaptation to low-phosphate diet in diabetic rats.

Insulin stimulates the Na(+)-Pi cotransport system in the brush-border membrane (BBM) of the renal proximal tubule, and an acute decrease in plasma insulin leads to a decrease in renal reabsorption of Pi. It has been proposed that insulin may play a role in the rapid renal adaptation to dietary deprivation of Pi. This hypothesis was tested using rats with low plasma insulin due to streptozotocin-induced diabetes. Both control and diabetic rats were housed in metabolic cages and fed either a normal Pi diet or a low Pi diet for 3 days. At the end of the third day, BBM vesicles were prepared from renal cortex and Na(+)-Pi cotransport was measured. At the whole kidney level, diabetic rats showed a normal adaptive response. There was a prompt and marked decrease in urinary Pi excretion when the rats ate a low Pi diet. At the BBM level, however, the adaptive response was absent. There was no increase in Na(+)-Pi cotransport in diabetic rats fed low Pi diet. Treatment of diabetic rats with exogenous insulin before feeding low Pi diet restored the adaptive increase in Pi transport by BBM. Insulin appears to be required for the adaptation of proximal tubule Pi transport to low Pi diet. In the absence of this adaptation in proximal tubule BBM, a compensatory response in the kidney may produce an increase in Pi reabsorption in later segments of the nephron.

Restoration of a phosphaturic response to parathyroid hormone in the immature rat.

Recent studies have shown that immature rats display a diminished sensitivity to the phosphaturic effects of parathyroid hormone (PTH), and that the responsiveness to PTH increases with age. The attenuated phosphaturia may reflect an inability of the neonate to respond to the hormone because of functional immaturity of the developing kidney. Alternatively, PTH may actually inhibit tubular phosphate reabsorption in the neonate but, due to other phosphate conservation mechanisms, no phosphaturia occurs. Our objective was to determine whether a phosphaturic response to PTH would be elicited in immature rats during infusion of moderate amounts of phosphate (Pi). Clearance experiments were performed on 26 acutely thyroparathyroidectomized immature Wistar rats (3-5 wk of age) fed a normal Pi diet (0.63%). In response to infusion of either Pi (1 mumol/min.100 g) (group I) or PTH (8.3 ng/min.100 g) (group II) alone, the fractional excretion of phosphate rose minimally (from 0.01 +/- 0.01% to 4.9 +/- 1.9% and from 0.12 +/- 0.12% to 2.9 +/- 1.4% for groups I and II, respectively). However, when Pi and PTH were combined either Pi first followed by PTH (group III) or PTH first followed by Pi (group IV), the fractional excretion of Pi rose dramatically (from 0.01 +/- 0.01 to 21.8 +/- 3.5% and from 0.04 +/- 0.04 to 27.7 +/- 3.3% for groups III and IV, respectively). A significant increase in urinary cAMP excretion occurred during infusion of PTH even when Pi excretion was minimal, but there was no further increase in urinary cAMP during the combined infusion of Pi and PTH.(ABSTRACT TRUNCATED AT 250 WORDS)

Dynamics of NAD in cortical nephron segments: effect of nicotinamide and of dietary phosphate intake.

NAD content and the rate of NAD hydrolysis were determined in proximal convoluted tubules (PCT), proximal straight tubules (PST), and adjacent cortical nephron segments microdissected from kidneys of thyroparathyroidectomized (TPTX) rats. In the basal state, rats fed a normal phosphate diet had an NAD content higher in PCT, PST, and in cortical ascending limb (CAL) than in glomeruli. After intraperitoneal injection of nicotinamide, the NAD content increased significantly in all nephron segments except CAL; the greatest (delta + 277%) increase was found in PCT, with less (delta + 82%) in PST. In experiments conducted on TPTX rats stabilized on a low-phosphorus diet, NAD content increased in response to a nicotinamide injection in PCT (delta + 197%), but did not change significantly in PST. The catabolism of NAD was determined by generation of [3H]adenosine, a major metabolite of [adenine-2,8-3H]NAD. The rate of [3H]adenosine generation from [3H]NAD was significantly (P less than 0.001) higher in PST than in PCT. We conclude that, in response to nicotinamide administration in vivo, the NAD content increases more in PCT than in PST and that this difference may be, at least partly, due to a lower rate of NAD breakdown in PCT. In a state of dietary phosphate deprivation, NAD also increases significantly in response to intraperitoneal nicotinamide in PCT, but it does not increase significantly in PST. The nicotinamide-elicited increase of NAD content in proximal tubules, mainly in PCT, may be related to inhibition of Na+-gradient-dependent inorganic phosphate (Pi) reabsorption across the brush-border membrane of proximal tubules and to the phosphaturic effect of nicotinamide in rats fed normal-Pi diet.

Possible role of nicotinamide adenine dinucleotide as an intracellular regulator of renal transport of phosphate in the rat.

In these experiments we investigated whether NAD could serve as an intracellular modulator of the brush border membrane (BBM) transport of inorganic phosphate (Pi). NAD, both oxidized (NAD+) and reduced (NADH) form, inhibited the Na+-dependent uptake of 32Pi in the concentration range of 10-300 microM NAD when added in vitro to BBM vesicles isolated from rat kidney cortex, but did not inhibit BBM uptake of D-[3H]glucose or BBM uptake of 22Na+. Neither nicotinamide (NiAm) nor adenosine alone influenced BBM uptake of 32Pi. NAD had a similar relative effect (percent inhibition) in BBM from rats stabilized on low Pi diet (0.07% Pi), high Pi diet (1.2% Pi), or normal Pi diet (0.7% Pi). Subsequently, we examined the renal effects of changing the tissue NAD level in vivo. Rats stabilized on low Pi diet were injected intraperitoneally with NiAm (0.25-1.0 g/kg body wt); urinary excretions of Pi (UPiV), of fluid, and of other solutes were measured before and after NiAm injection, then renal cortical tissue nucleotide content was determined, and a BBM fraction was isolated for transport measurements. In BBM from NiAm-treated rats, the Na+-dependent uptake of 32Pi was decreased, but BBM uptake of D-[3H]glucose and BBM uptake of 22Na+ were not changed. NiAm injection elicited an increase in NAD+ (maximum change, 290%), a lesser increase in NADH (maximum change, +45%), but no change in the content of ATP or cyclic AMP in the renal cortex. Na+-dependent BBM uptake of 32Pi ws inversely correlated with NAD+ content in renal cortex (r = -0.77 +/- 0.1; P less than 0.001) and with UPiV (r = -0.67 +/- 0.13; P less than 0.01). NAD+ in renal cortex was positively correlated with UPiV (r = 0.88 +/- 0.05; P less than 0.001). Injection of NiAm elicited a marked increase in UPiV, but no change in excretions of creatinine or K+, or in urine flow; excretion of Na+ and Ca declined. NiAm injection caused similar renal responses, in normal and in thyroparathyroidectomized rats, as well as in rats on normal Pi diet and low Pi diet. We conclude that NAD can serve as an intracellular modulator (inhibitor) of Na+-dependent transport of Pi across the renal luminal BBM and across the proximal tubular wall by its direct interaction with BBM. We propose that at least some hormonal and/or metabolic stimuli elicit phosphaturia by increasing NAD+ in cytoplasm of proximal tubular cells.