Renal Tubular Brush Border Membrane Research Articles

FGF23 Regulation of Phosphorus Homeostasis Is Dependent on PTH

Fibroblast growth factor-23 (FGF23) is a key regulator of phosphorus homeostasis (1). FGF23 is a member of the FGF19 subfamily of fibroblast growth factors. The FGF23 gene is located on human chromosome 12p13 and mouse chromosome 6 and is composed of three coding exons with an open reading frame of 125 residues. FGF23 is most highly expressed in bone but also found in thymus, brain, heart, and other tissues. In bone, FGF23 mRNA is found in osteoblasts, osteocytes, bone-lining cells, and osteoprogenitor cells. FGF23 is secreted primarily by osteoblasts and osteocytes in response to increased serum phosphorus or 1,25-dihydroxyvitamin D. Circulating FGF23 binds to the canonical FGF receptor 1c coupled to the single transmembrane protein Klotho, a coreceptor required for FGF23 signaling, to form the specific receptor for FGF23 function (2). FGF23 targets the parathyroid gland to decrease PTH secretion and the kidney to increase urinary phosphorus excretion and to suppress synthesis of 1,25-dihydroxyvitamin D by decreasing renal tubular 1 -hydroxylase expression. These changes result in increased urinary loss of phosphorus as well as decreased proximal renal tubular production of 1,25dihydroxyvitamin D, both of which lead to a return to normal serum phosphorus levels (Fig. 1). In the kidney, circulating FGF23 reduces the expression of proximal renal tubular brush border membrane sodium-phosphate cotransporters NaPi2a and NaPi2c, which reduces the ability of the kidney to reabsorb phosphorus from proximal renal tubular glomerular filtrate, thereby causing increased urinary loss of phosphorus (3). FGF23 also reduces the activity of proximal renal tubular 1 -hydroxylase, the enzyme required for production of 1,25-dihydroxyvitamin D and increases the activity of 24-hydroxylase, the proximal renal tubular enzyme that metabolizes 1,25-dihydroxyvitamin D. Decreased serum 1,25-dihydroxyvitamin D leads to decreased intestinal calcium and phosphorus absorption, and low serum calcium and low serum 1,25-dihydroxyvitamin D both lead to increased production of PTH. The main physiological short-term role of circulating FGF23 appears to be to stimulate renal tubular phosphorus excretion when excessive phosphorus intake leads to hyperphosphatemia, whereas its main long-term role appears to be to limit intestinal absorption of phosphorus. Progressive increases in serum FGF23 play a major role in mineral metabolism abnormalities that develop in disorders characterized by hyperphosphatemia. Serum FGF23 levels increase in early-stage chronic kidney disease (CKD) and are associated with progressive hyperphosphatemia and hypocalcemia in late-stage CKD, secondary hyperparathyroidism, and low 1,25-dihydroxyvitamin D levels. Serum FGF23 levels increase beginning as early as stage 2 CKD because of decreased renal tubular clearance of phosphorus and result in increased fractional phosphorus excretion, reduced serum phosphorus levels, and reduced renal 1 -hydroxylase activity, which lead to normalization of serum phosphorus and reduced serum 1,25-dihydroxyvitamin D levels. Increased serum phosphorus levels in later-stage CKD, despite increased serum FGF23 levels and low serum 1,25dihydroxyvitamin D levels, lead to persistent stimulation of PTH secretion. As CKD progresses, serum FGF23 continues to increase, whereas responsiveness to FGF23 decreases as the number of intact nephrons declines. Decreased intact nephrons are associated with reduced expression of the FGF23 coreceptor Klotho. In end-stage

Intratubular crystallization of calcium oxalate in the presence of membrane vesicles: An in vitro study

Since urine spends only a few minutes in the renal tubules and has a low supersaturation with respect to calcium oxalate (CaOx), nucleation of CaOx crystals in the kidneys is most probably heterogeneous. We have proposed that membranes of cellular degradation products are the main substrate for crystal nucleation. The purpose of our study was to determine the site of membrane-mediated crystal nucleation within the renal tubules and the required lag time, factors that determine whether crystallization results in crystalluria or nephrolithiasis. Nucleation of CaOx was allowed to occur in five different artificial urine solutions with ionic concentrations simulating urine in proximal tubules (PTs), descending (DLH) and ascending (ALH) limbs of the loop of Henle, distal tubules (DTs), and collecting ducts (CDs). A constant composition crystallization system was used. Experiments were run for two hours with or without the renal tubular brush border membrane (BBM) vesicles. The addition of BBM significantly reduced the nucleation lag time and increased the rate of crystallization. The average nucleation lag time decreased from 84.6 +/- 43.4 minutes to 24.5 +/- 19 minutes in PTs, from 143.6 +/- 29 to 70.2 +/- 53.4 minutes in DLH, from 17.6 +/- 8.6 minutes to 0.625 +/- 0.65 minutes in DTs and from 9.54 +/- 3. 03 minutes to 0.625 +/- 0.65 minutes in CDs. There was no nucleation in the ALH solution without BBM for two hours. CaOx dihydrate (COD) was common in most solutions. Calcium phosphate (CaP) also nucleated in the DLH and CD solutions. In the absence of membrane vesicles, there was no crystallization in any of the solutions within the time urine spends in the renal tubules. As a result, homogeneous nucleation of crystals anywhere within the nephron appears unlikely. However, BBM-supported nucleation is possible in the DTs as well as CDs. A high crystallization rate in CDs would promote rapid crystal growth and aggregation, resulting in crystal retention within the kidneys and development of nephrolithiasis.

Cl− and Membrane Potential Dependence of Amino Acid Transport across the Rat Renal Brush Border Membrane

The relative roles of the anion present and the membrane potential in the operation of each of the seven amino acid transport systems in the renal tubular brush border membrane were explored by manipulating transmembrane potential and chemical gradients across the membrane. The effect of various external anions with different permeabilities of the membrane and of valinomycin-generated K+ diffusion potential on Na+-coupled amino acid accumulation by rat renal brush border membrane vesicles was examined. Accumulation of all amino acids examined, except for cystine, was membrane potential dependent. The highest voltage dependence was observed for taurine (equivalent to glucose) and l-methionine. Addition of taurine uptake values obtained under each electrical gradient (inside negative) and a chemical gradient (100 mM NaCl out) condition yielded markedly lower values than under conditions where there was a combined electrochemical gradient. Cl− gradient rather than merely imposing a voltage gradient was a specific mediator of Na+-coupled transport of l-proline, taurine, l-glutamic acid, and glycine across the brush border membrane. Cl− gradient alone under Na+-equilibrated conditions could energize an overshoot of taurine accumulation by vesicles providing evidence that taurine is energetically activated by and coupled to Cl− transport. These data suggest that Na+-linked transport of most amino acids across the tubular luminal membrane is an electrogenic positive process and for proline, taurine, glutamic acid, and glycine, a Cl−-requiring process. A negative intracellular potential combined with luminal chloride is required for optimal Na+-coupled transport of these amino acids across the luminal membrane of the proximal tubule. The coupling of Cl− to the transport of these osmoprotective amino acids may enhance their volume regulatory effect in kidney cells and other mammalian cells.

Effects of cyclosporin A on experimental nephritis in rats (2): Cyclosporin A suppresses the development of accelerated passive Heymann nephritis

We investigated the effects of cyclosporin A (CyA) on accelerated passive Heymann nephritis, an experimental model of membranous nephropathy, that is characterized by immune complex deposition on the glomerular basement membrane. The nephritis was induced in rats by injection of antiserum against the antigen located in the renal tubular brush border membrane and sensitization with rabbit gamma-globulin. CyA was administered p.o. at the dose of 2.5, 10 or 20 mg/kg/day for 40 days after the injection of the antiserum. The administration of CyA resulted in marked suppression of proteinuria and hypercholesterolemia in the nephritic rats. In light microscopy, nephritic control rats showed thickening of the glomerular basement membrane and spike formation in the glomeruli. CyA significantly reduced the appearance of the glomerular alteration. The production of antibody was dramatically attenuated by CyA administration. However, CyA did not decrease the number of circulating white blood cells and platelets below the normal level. In conclusion, CyA suppressed the progress of accelerated passive Heymann nephritis in a dose-dependent manner. The effect of CyA is likely attributable to the powerful depression of antibody production.

Heterogeneous nucleation of calcium oxalate crystals in the presence of membrane vesicles

Membrane-assisted crystallization of calcium oxalate was studied in vitro, using constant composition methodology. Rat renal tubular brush border membrane vesicles were incubated in supersaturated solution of calcium oxalate. Calcium and oxalate depletion started much earlier in the presence of the vesicles than in their absence; within 8, 32, or 258 min of the incubation of vesicles in calcium oxalate solutions of relative supersaturation of 12, 10 or 6 respectively. Thin plate-like crystals with jagged edges formed in association with the membrane vesicles. Since crystal nucleation in the presence of membrane vesicles started within 8 min at a relative supersaturation as low as 12, it will start significantly earlier in the urine of stone formers which is known to have higher relative supersaturation with respect to calcium oxalate. These results demonstrate that cellular membranes can efficiently induce nucleation of calcium oxalate crystals from a metastable solution in an vitro system. Similar membrane induced heterogeneous nucleation of calcium oxalate in vivo within the renal tubules is a distinct possibility.

The biochemical modifications of the brush border membrane induced by vitamin D and parathyroid hormone in their actions on phosphate transport.

Since the development of techniques for the preparation and isolation of brush border membrane vesicles (BBMV),1,2 from the luminal cell membrane of renal tubular epthelial cells considerable progress has been made in our understanding of renal tubular cell phosphate transport. A sodium dependent co-transport system capable of moving phosphate uphill against an electrochemical gradient has been characterized in BBMV. The stoichiometry of the carrier mechanism remains controversial. According to one report, it is either 2Na+:HPO 4 −2 or Na+:H2PO4−, dependent upon the charge of the phosphate species present in the bathing fluid, and phosphate transfer is electroneutral.3 However, others have reported that at pH 6 the stoichiometry is 2Na:H2PO4− and the carrier may be electrogenic at low pH.4,5 Carrier activity increases as the pH of the medium is increased in the range of 6.0–8.5.3–5 Thus, a mechanism of secondary active phosphate transport in the renal tubular brush border membrane (BBM) has been partially characterized.

A search for nephrotoxic factors within the uremic milieu.

The purpose of this study was to ascertain whether selected components of the uremic milieu adversely affected glomerular filtration rate (GFR), the glomerular protein filtration barrier, or the integrity of the proximal renal tubular brush border membrane. To achieve these goals, GFR and the excretion rates of albumin and of brush border derived-renal tubular epithelial antigens (RTE) were measured in normal rats and in rats with experimental nephropathies before and after the intravenous infusion of concentrated urine. This experimental protocol uniformly produced severe biochemical manifestations of uremia (for example 10-50-fold increases in BUN and creatinine, hyperphosphatemia, hyperkalemia, metabolic acidosis). However, despite these perturbations, GFR, albuminuria, and RTE excretion remained constant. To assess the influence of uremic hormonal derangements on renal function, GFR, albuminuria, and RTE excretion were measured in normal rats before and after inducing acute serum elevations of seven hormones whose concentrations are known to be increased in uremia (parathyroid hormone, growth hormone, insulin, glucagon, gastrin, prolactin, gastric inhibitory peptide). Again, GFR, albuminuria, and RTE excretion were not adversely affected. These results suggest that glomerular capillary function and proximal tubular brush border membranes are acutely resistant to many of the solute and hormonal derangements which are characteristic of uremia.