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Adolescent transplant recipients are at risk for nonadherence, development of de novo donor-specific antibody (dnDSA), and allograft loss. Belatacept, a selective T cell costimulatory blocker, is associated with reduced dnDSA, improved renal function, and prolonged allograft survival when compared to calcineurin inhibitor-based regimens in adults; however, its use in children is scant. Three adolescents were initiated on belatacept between August 2017 and September 2018 at the time of kidney transplantation. Selection criteria included age ≥ 14 and EBV IgG + serostatus. Intraoperative alemtuzumab and methylprednisolone were given as induction therapy. Tailored maintenance therapy included steroid-free belatacept and sirolimus for two patients. One patient was initially maintained steroid-free on belatacept and belimumab, an inhibitor of B cell activating factor to treat concurrent systemic lupus erythematous; steroids were added subsequently. Renal function, biopsy-proven rejection, dnDSA, allograft survival, infection, nonadherence, and proteinuria were monitored. Renal function was 86, 73, 52 mL/min/1.73 m2 at 20, 20, and 8 months, respectively. There was 100% adherence to therapy and no development of dnDSA. All patients had treatable infections. One developed steroid-responsive acute cellular rejection. Belatacept-based regimens can be tailored for adolescent recipients with good short-term clinical outcomes.

BACKGROUND AND OBJECTIVES: Steroid resistant nephrotic syndrome (SRNS) due to focal segmental glomerulosclerosis (FSGS) and minimal change disease (MCD) is a leading cause of end stage kidney disease in children. Recurrence of primary disease following transplantation is a major cause of allograft loss. The clinical determinants of disease recurrence are not completely known. Our objectives were to determine risk factors for recurrence of FSGS/MCD following kidney transplantation, and factors that predict response to immunosuppression following recurrence. STUDY DESIGN: Multicenter study of pediatric patients with kidney transplants performed for ESKD due to SRNS between 1/2006–12/2015. Demographics, clinical course, and biopsy data were collected. Patients with primary-SRNS (PSRNS) were defined as those initially resistant to corticosteroid therapy at diagnosis, and patients with late-SRNS (LSRNS) as those initially responsive to steroids who subsequently developed steroid resistance. We performed logistic regression to determine risk factors associated with nephrotic syndrome (NS) recurrence. RESULTS: We analyzed 158 patients; 64 (41%) had recurrence of NS in their renal allograft. Disease recurrence occurred in 78% of patients with LSRNS compared to 39% of those with PSRNS. Patients with MCD on initial native kidney biopsy had a 76% recurrence rate compared with a 40% recurrence rate in those with FSGS. Multivariable analysis showed that MCD histology (OR; 95% CI: 5.6; 1.3–23.7) compared to FSGS predicted disease recurrence. CONCLUSIONS: Pediatric patients with MCD and LSRNS are at higher risk of disease recurrence following kidney transplantation. These findings may be useful for designing studies to test strategies for preventing recurrence.

The number of genes associated with renal disease is increasing every day and this has led to a clearer understanding of the pathophysiology of renal disease in many disorders. It is also appreciated now that a genetic mutation(s) underlie many renal syndromes. Genetic testing may also offer the possibility to diagnose some renal diseases without the need for a renal biopsy. It also allows the prenatal diagnosis of certain renal diseases in at risk fetuses or identification of potential renal disease before it has become manifest. Finally, identification of a specific gene mutation holds the possibility of correction though gene therapy in the future. It is increasingly clear that many renal disorders in pediatrics are a consequence of genetic mutations. In the future, genetic testing will become as easy and as common as ordering a serum creatinine today.

Primary vesicoureteral reflux (PVUR) is the most common malformation of the kidney and urinary tract, and reflux nephropathy is a major cause of chronic kidney disease in children. Recently, we reported mutations in the tenascin XB gene (TNXB) as a cause of PVUR with joint hypermobility. To define the role of rare variants in tenascin genes in the etiology of PVUR, we screened a cohort of patients with familial PVUR (FPVUR) and non-familial PVUR (NFPVUR) for rare missense variants inTNXB and the tenascin C gene (TNC) after excluding mutations in ROBO2 and SOX17. The screening procedure identified 134 individuals from 112 families with PVUR; two families with mutations in ROBO2 were excluded from further analysis. Rare missense variants in TNXB were found in the remaining 110 families, of which 5/55 (9 %) families had FPVUR and 2/55 (4 %) had NFPVUR. There were no differences in high-grade reflux or renal parenchymal scarring between patients with and without TNXB variants. All patients with TNXB rare variants who were tested exhibited joint hypermobility. Overall we were able to identify causes of FPVUR in 7/57 (12 %) families (9 % in TNXB and 3 % in ROBO2). In conclusion, the identification of a rare missense variant in TNXB in combination with a positive family history of VUR and joint hypermobility may represent a non-invasive method to diagnose PVUR and warrants further evaluation in other cohorts.

Background Mutations in podocin (NPHS2) are the most common cause of childhood onset autosomal recessive steroid-resistant nephrotic syndrome (SRNS). The disease is characterized by early-onset proteinuria, resistance to immunosuppressive therapy and rapid progression to end-stage renal disease. Compound heterozygous changes involving the podocin variant R229Q combined with another pathogenic mutation have been associated with a mild phenotype with disease onset often in adulthood. Methods We screened 19 families with early-onset SRNS for mutations in NPHS2 and WT1 and identified four disease-causing mutations (three in NPHS2 and one in WT1) prior to planned whole-exome sequencing. Results We describe two families with three individuals presenting in childhood who are compound heterozygous for R229Q and one other pathogenic NPHS2 mutation, either L327F or A297V. One child presented at age 4 years (A297V plus R229Q) and the other two at age 13 (L327F plus R229Q), one with steadily deteriorating renal function. Conclusions These cases highlight the phenotypic variability associated with the NPHS2 R229Q variant plus pathogenic mutation. Individuals may present with early aggressive disease.

Background: Mutations of the CYP24A1 gene, which encodes the 1,25-dihydroxyvitamin D-24-hydroxylase cytochrome P450, Cyp24A1, are predicted to result in elevated 1,25-dihydroxyvitamin D concentrations, hypercalcemia, hypercalciuria, nephrolithiasis, and bone disease. Treatment of hypercalcemia associated with CYP24A1 gene mutations has not been described. Methods: The genetic basis of a syndrome in a 44-yr-old Caucasian male characterized by intermittent hypercalcemia, hypercalciuria, elevated serum 1,25-dihydroxyvitamin D, undetectable serum 24,25-dihydroxyvitamin D, metabolically active nephrolithiasis, and reduced bone mineral density of the lumbar spine was examined. Sequencing of the CYP24A1 gene and biochemical and genetic analysis of seven family members in three generations was carried out. Because of hypercalcemia, hypercalciuria, and metabolically active nephrolithiasis, the patient was treated with a cytochrome 3A inhibitor, ketoconazole, 200 mg orally every 8 h, for 2 months. Results: The sequ...

Hypertension affects one billion people and is a principal reversible risk factor for cardiovascular disease. Pseudohypoaldosteronism type II (PHAII), a rare Mendelian syndrome featuring hypertension, hyperkalaemia and metabolic acidosis, has revealed previously unrecognized physiology orchestrating the balance between renal salt reabsorption and K(+) and H(+) excretion. Here we used exome sequencing to identify mutations in kelch-like 3 (KLHL3) or cullin 3 (CUL3) in PHAII patients from 41 unrelated families. KLHL3 mutations are either recessive or dominant, whereas CUL3 mutations are dominant and predominantly de novo. CUL3 and BTB-domain-containing kelch proteins such as KLHL3 are components of cullin-RING E3 ligase complexes that ubiquitinate substrates bound to kelch propeller domains. Dominant KLHL3 mutations are clustered in short segments within the kelch propeller and BTB domains implicated in substrate and cullin binding, respectively. Diverse CUL3 mutations all result in skipping of exon 9, producing an in-frame deletion. Because dominant KLHL3 and CUL3 mutations both phenocopy recessive loss-of-function KLHL3 mutations, they may abrogate ubiquitination of KLHL3 substrates. Disease features are reversed by thiazide diuretics, which inhibit the Na-Cl cotransporter in the distal nephron of the kidney; KLHL3 and CUL3 are expressed in this location, suggesting a mechanistic link between KLHL3 and CUL3 mutations, increased Na-Cl reabsorption, and disease pathogenesis. These findings demonstrate the utility of exome sequencing in disease gene identification despite the combined complexities of locus heterogeneity, mixed models of transmission and frequent de novo mutation, and establish a fundamental role for KLHL3 and CUL3 in blood pressure, K(+) and pH homeostasis.

Steroid-sensitive nephrotic syndrome (SSNS) accounts for >80% of cases of nephrotic syndrome in childhood. However, the etiology and pathogenesis of SSNS remain obscure. Hypothesizing that coding variation may underlie SSNS risk, we conducted an exome array association study of SSNS. We enrolled a discovery set of 363 persons (214 South Asian children with SSNS and 149 controls) and genotyped them using the Illumina HumanExome Beadchip. Four common single nucleotide polymorphisms (SNPs) in HLA-DQA1 and HLA-DQB1 (rs1129740, rs9273349, rs1071630, and rs1140343) were significantly associated with SSNS at or near the Bonferroni-adjusted P value for the number of single variants that were tested (odds ratio, 2.11; 95% confidence interval, 1.56 to 2.86; P=1.68×10(-6) (Fisher exact test). Two of these SNPs-the missense variants C34Y (rs1129740) and F41S (rs1071630) in HLA-DQA1-were replicated in an independent cohort of children of white European ancestry with SSNS (100 cases and ≤589 controls; P=1.42×10(-17)). In the rare variant gene set-based analysis, the best signal was found in PLCG2 (P=7.825×10(-5)). In conclusion, this exome array study identified HLA-DQA1 and PLCG2 missense coding variants as candidate loci for SSNS. The finding of a MHC class II locus underlying SSNS risk suggests a major role for immune response in the pathogenesis of SSNS.

Primary vesicoureteral reflux (VUR) is the most common congenital anomaly of the kidney and the urinary tract, and it is a major risk factor for pyelonephritic scarring and CKD in children. Although twin studies support the heritability of VUR, specific genetic causes remain elusive. We performed a sequential genome-wide linkage study and whole-exome sequencing in a family with hereditary VUR. We obtained a significant multipoint parametric logarithm of odds score of 3.3 on chromosome 6p, and whole-exome sequencing identified a deleterious heterozygous mutation (T3257I) in the gene encoding tenascin XB (TNXB in 6p21.3). This mutation segregated with disease in the affected family as well as with a pathogenic G1331R change in another family. Fibroblast cell lines carrying the T3257I mutation exhibited a reduction in both cell motility and phosphorylated focal adhesion kinase expression, suggesting a defect in the focal adhesions that link the cell cytoplasm to the extracellular matrix. Immunohistochemical studies revealed that the human uroepithelial lining of the ureterovesical junction expresses TNXB, suggesting that TNXB may be important for generating tensile forces that close the ureterovesical junction during voiding. Taken together, these results suggest that mutations in TNXB can cause hereditary VUR.

To determine the effects of low-dose dopamine on urine output (UOP) in very low birth weight premature neonates.Retrospective cohort study of all low-dose (3-5 μg kg(-1) per min) dopamine infusions24-h duration in neonates 1500 g and 32 weeks gestation from August 2009 through September 2011. Linear regression was used to estimate the impact of covariates on UOP.We identified 91 episodes of low-dose dopamine use in 65 neonates. Increased UOP occurred in 64% of episodes. Low-dose dopamine use was associated with a 0.6 ml kg(-1) h(-1) increase in UOP (P0.001) and a 1.3 ml kg(-1)h(-1) increase when baseline UOP was1.5 ml kg(-1) h(-1) (P0.001). The improvement remained statistically significant after controlling for medications (diuretics and hydrocortisone) and fluid intake.Low-dose dopamine use was associated with increased UOP in very low birth weight neonates.

Insulin-like growth factor-I gene expression in the tibial epiphyseal growth plate of growth hormone-treated uremic rats. To identify the molecular mechanisms involved in long bone growth of uremic animals, we evaluated the effects of recombinant human growth hormone (rhGH) supplementation on whole body growth, growth plate morphometries, and insulin-like growth factor-I (IGF-I) gene expression in the tibial epiphyseal growth plates of uremic rats. Uremia was induced by a two-stage subtotal nephrectomy (Nx) of 30-day-old rats, followed by rhGH (N = 6) or saline (N = 6) treatment from day 56 to day 70 of age. Controls (N = 4) were sham decapsulated. Treatment with rhGH on Nx animals caused: (1) a significant increase in weight, (2) longitudinal growth similar to controls, and (3) increased total growth plate width predominantly due to an increase in hypertrophic zone width. rhGH increased IGF-I mRNA abundance in both zones, but the increase was greater in the proliferative zone. These changes were accompanied by concomitant alterations in IGF-I immunoreactivity. In uremic animals, therefore, rhGH treatment induces local IGF-I gene expression in the growth plate and increases the hypertrophic zone width but not the proliferative zone width. The latter suggests resistance to IGF-I action in that zone.

Focal and segmental glomerulosclerosis (FSGS) is an important cause of steroid-resistant nephrotic syndrome in adults and children. It is responsible for 5–20% of all cases of end-stage kidney disease (ESKD) in the United States. The pathogenesis of FSGS has not been fully elucidated; however, data from molecular studies of familial cases in the last two decades suggest that FSGS is a defect of the podocyte. The therapeutic agents available for treatment of FSGS are not very effective and only a small percentage of affected individuals will achieve complete remission. Recent data from molecular biology and molecular genetics has provided insight into the mechanisms of action of old agents and also identification of other novel therapeutic targets. This review focuses on recent advances in the molecular pathogenesis of FSGS and currently available therapeutic agents as well as potential novel therapies.

The majority of human urinary stones are primarily composed of calcium salts. Although normal urine is frequently supersaturated with respect to calcium oxalate, most humans do not form stones. Inhibitors are among the multiple factors that may influence the complex process of urinary stone formation. We have isolated an inhibitor of calcium oxalate crystal growth from human urine by monoclonal antibody immunoaffinity chromatography. The N-terminal amino acid sequence and acidic amino acid content of this aspartic acid-rich protein, uropontin, are similar to those of other pontin proteins from bone, plasma, breast milk, and cells. The inhibitory effect of uropontin on calcium oxalate crystal growth in vitro supports the concept that pontins may have a regulatory role. This function would be analogous to that of other members of the aspartic acid-rich protein superfamily, which stereospecifically regulate the mineralization fronts of calcium-containing crystals.

The effect of loading renal tubule cells with cystine was studied by incubating them with cystine dimethylester. Proline uptake into brushborder membrane vesicles isolated from the cystine loaded cells was not different from that observed into brushborder vesicles isolated from tubules incubated in buffer alone. Incubating brushborder membranes with 2 mM cystine dimethylester for 10 minutes reduced the uptake of proline by 27% after 15 seconds of incubation and by 21% after 60 seconds of incubation. There was no effect after 20 minutes of incubation. Pre-incubating brushborder membrane vesicles with cystine dimethylester had no statistically significant effect on the affinity of priline for the carrier, but did reduce the maximal rate of proline uptake by 49%.

The pediatric ESRD patient is a member of a unique subpopulation of ESRD patients. The cause of ESRD in the pediatric patient differs markedly from the adult patient; treatment modality in the pediatric ESRD patient differs substantially from the adult patient; and outcomes such as growth

Nephronophthisis (NPHP), an autosomal recessive cystic kidney disease, leads to chronic renal failure in children. The genes mutated in NPHP1 and NPHP4 have been identified, and a gene locus associated with infantile nephronophthisis (NPHP2) was mapped. The kidney phenotype of NPHP2 combines clinical features of NPHP and polycystic kidney disease (PKD). Here, we identify inversin (INVS) as the gene mutated in NPHP2 with and without situs inversus. We show molecular interaction of inversin with nephrocystin, the product of the gene mutated in NPHP1 and interaction of nephrocystin with β-tubulin, a main component of primary cilia. We show that nephrocystin, inversin and β-tubulin colocalize to primary cilia of renal tubular cells. Furthermore, we produce a PKD-like renal cystic phenotype and randomization of heart looping by knockdown of invs expression in zebrafish. The interaction and colocalization in cilia of inversin, nephrocystin and β-tubulin connect pathogenetic aspects of NPHP to PKD, to primary cilia function and to left-right axis determination.

Although the mechanisms underlying the inhibitory effects of chronic renal insufficiency on growth are poorly understood, large doses of growth hormone (GH) have been used to improve growth. The present study examines the effects of rat GH and a reduced (8%) protein diet on 75% nephrectomized weanling rats by measuring changes in growth parameters, food utilization, serum albumin concentration, and muscle water content. Significantly greater improvement in growth was found in the GH-treated uremic rats compared with the uremic controls. The mean percent change in wt, length (nose to tail tip), and cranial biparietal diameter was significantly increased in the GH-treated uremic rats, compared with the uremic controls, but foot length and femur length showed only moderate improvement. Food utilization efficiency and serum albumin concentration were significantly higher in GH-treated uremic rats compared with uremic controls, achieving levels that were not different from sham-operated rats. Muscle water content was not significantly different between GH-treated uremic rats, uremic controls, and sham-operated rats. Thus, rat GH treatment administered at an early age in mild renal insufficiency significantly improved overall growth, food efficiency, and serum albumin concentrations, despite a low protein diet, suggesting that further evaluation of this form of therapy for growth failure of uremia is warranted.

Glutamate had no significant effect on the uptake of 0.025 mM cystine by isolated rat renal cortical tubules and brushborder membrane vesicles in contrast to lysine which significantly inhibits cystine transport. Glutamate, however, markedly inhibited cystine uptake by rat renal tubule cells grown in a serum-free, hormonally defined media for 5 days. Lysine also inhibited cystine transport in these cultured renal tubule cells.

Increased amino acid excretion is a characteristic of developing rat, dog and man. The mechanisms underlying this physiologic aminoaciduria are unclear. We have shown faster cystine entry into isolated renal tubule cells from newborn rats compared to adults, despite impaired in vivo reabsorption. To see if these observations extend to other animals, we examined the fractional reabsorption (FRc) of cystine in developing dogs and cystine uptake by isolated renal cortical tubule fragments from newborn (NB) and adult dogs. FRc was 67% of the filtered load in five day old dogs but reached the adult value of 99% by 21 days of life. Isolated renal cortical tubules from NB dogs progressively accumulated label when incubated for 60 min. with a physiologic concentration of cystine (0.025 mM), although this uptake was lower than that observed in adults. Nearly complete conversion of transported labelled cystine to cysteine occurred in the NB and adult as previously noted for rat tubules. Kinetic analysis of uptake indicated two systems for cystine entry in both adult and NB. The affinity constants (Km) for the newborn systems were Km1 = 0.08 ± 0.01 and Km2 = 0.33 ± 0.03 mM. The corresponding values for adult were significantly higher - Km1 = 0.14 ± 0.02 and Km2 = 0.66 ± 0.09 mM. The maximal uptake rate for each NB system was only 1/3 that of the adult. These data suggest that, in contrast to the rat, the impaired cystine reabsorption by NB dog kidney in vivo is related in part to a slower flux of cystine into the renal tubule cells.

Intracellular levels of cysteine (CSH) and reduced glutathione (GSH) in the kidney and liver of rats from the newborn to the adult period have been determined using a sensitive high performance liquid chromatography method. In the kidney, the intracellular level of free CSH increased 4-fold from 1 to 4 nmol/mg protein with GSH levels which ranged from 20 to 25 nmol/mg protein from the 10th to 21st postnatal day, respectively. In contrast, intracellular free hepatic CSH showed a biphasic pattern with development. Intracellular free hepatic GSH, on the other hand, increased 2-fold over the 3- to 21-day postnatal period. In adult tissues, intracellular levels of free CSH and GSH decreased as compared with levels in 21-day postnatal animals. When ratios of CSH to GSH were compared between tissues from the 3-day-old postnatal and adult rat, CSH:GSH increased approximately 4-fold in the kidney and decreased 2- to 3-fold in the liver.

The uptake of l -lysine was examined in isolated renal cortical tubules. Lysine was actively taken up by the renal tubule cells isolated from 7-week-old rats. No metabolism of the transported lysine was found. There was no evidence for sodium-dependence of lysine uptake. Concentration dependence studies revealed that the lysine was taken up by one saturable transport system with a K m of 1.66 mmol/l and V max of 7 mmol/l intracellular fluid per 10 min. Lysine also entered by a non-saturable pathway. Arginine and ornithine inhibited the initial uptake of lysine. Cystine increased the efflux of lysine from preloaded renal cells via hetero-exchange, indicating that a common system exists for these two amino acids.

A B S T RA C T Isolated renal tubule fragments prepared from adult Sprague-Dawley rats were used to study the cellular uptake of hypoxanthine. This uptake was rapid, reaching a steady state after 30 min of incubation. Analysis ofthe intracellular pool during the initial uptake and at the steady state revealed a concentration gradient ofhypoxanthine consistent with active transport, although only one-third of the transported hypoxanthine remained unmetabolized. The remainder of the transported hypoxanthine was converted to inosine and inosinic acid, but detectable conversion to uric acid was not noted. A kinetic analysis of uptake revealed that two systems for cellular entry of hypoxanthine existed with Km, = 0.005 and Km2 = 0.80 mM. Hypoxanthine uptake at physiologic concentrations was oxygen, sodium, and temperature dependent, but the addition ofmetabolic fuels and alteration of the medium pH over the range of from 6.1 to 7.4 had no effect. Adenine, guanine, and inosine inhibited the uptake of hypoxanthine via the low-Km system which mediates the majority of uptake at physiologic levels. Xanthine, uric acid, and probenecid inhibited uptake via the high-Km system, but did not affect uptake via the low-Km system. The data indicate that hypoxanthine at physiologic levels is transported into the renal tubule cell via a system different from that for other oxypurines.

Summary: α-Methyl-D-glucoside (AMG) uptake was examined in isolated renal cortical tubules from newborn, 3-month-old, and adult dogs. All three age groups demonstrated active sugar transport. The initial rate of AMG uptake was similar in the 3-month-old and adult tubules which was twice that of the newborn. At steadystate, the adult and newborn tubules had achieved a similar intracellular AMG concentration which was 45% greater than that of the 3-month-old. Determination of the flux constants of these uptake patterns revealed that there was an age-dependent increase in both the net flux and the fractional influx rate constant. However, the 3-month-old had the highest fractional efflux rate constant and the newborn the lowest value with the adult in between. Kinetic analysis of AMG uptake showed a single saturable transport system for each age group. The newborn and adult had similar Km values but the 3-month-old had a value that was 60% higher. The 3-month-old tubules had the highest Vmax and the newborn tubules the lowest with the adult value in between. AMG uptake by tubules from each age group demonstrated a similar pattern of inhibition in a low sodium buffer and by glucose and phlorizin. This indicated that, aside from kinetic changes with maturation, the saturable transport system for AMG is similar in each age group.

Within the past several decades disorders with deranged transport functions in renal proximal tubule cells have become clinically prominent. Every textbook of medicine or pediatrics contains a discussion of inherited or acquired diseases involving defects in the absorption of specific amino acids, sugars, or other solutes. The inherited disorders may involve specific amino acids, such as in classic cystinuria with dibasic aminoaciduria, iminoglycinuria, and Hartnup disease with neutral aminoaciduria, sugars as in renal glycosuria and glucose-galactose malabsorption, or phosphate as in hypophosphatemic rickets or uric acid. Although the exact defect in cell function in these disorders has not been delineated, the likely explanation has been that they result from a malfunctioning or absent specific carrier mechanism for the transport of the particular substances involved. The renal Fanconi syndrome (De Toni-Debre-Fan-coni syndrome), which has been thoroughly discussed by Morris et al [1] and by Brodehl [2], on the other hand, is characterized by a generalized disorder in proximal renal tubule transport affecting amino acids, glucose, and phosphate, as well as uric acid, bicarbonate, and other substances. The clinical consequences are acidosis, rickets, impaired growth, polyuria, dehydration, and hypokalemia.

Glutamine uptake was examined in isolated renal brush-border and basolateral-membrane vesicles from control and acidotic rats. In brush-border vesicles from acidotic animals, there was a significant increase in the initial rate of glutamine uptake compared with that in controls. Lowering the pH of the medium increased the initial rate of glutamine uptake in brush-border vesicles from acidotic, but not from control, rats. In brush-border vesicles from both groups of animals, two saturable transport systems mediated glutamine uptake. There was a 2-fold increase in the Vmax. of the low-affinity high-capacity system in the brush-border vesicles from the acidotic animals compared with that from control animals, with no alteration in the other kinetic parameters. There was no difference in glutamine uptake by the two saturable transport systems in basolateral vesicles from control and acidotic animals. Lowering the incubation-medium pH increased the uptake of glutamine by basolateral vesicles from both control and acidotic rats to a similar extent. The data indicate that during acidosis there are alterations in glutamine transport by both the basolateral and brush-border membrane which could enhance its uptake by the renal-tubule cell for use in ammoniagenesis.

EFFECT OF CYSTINE DIMETHYLESTER (CDM) ON RENAL TUBULAR FUNCTION, A MODEL OF FANCONI SYNDROME

Thrombosis and renovascular hypertension are complications of umbilical artery catheters (UAC). There are no reported cases of resolution of bilateral renal artery thromobosis leading to complete renal failure. We report an infant with complete aortic occlusion. A 1.3 kg 30 week gestation male infant was delivered via cesearean section for pre-eclampsia. UAC and umbilical venous catheter (UVC) were placed. Respiratory distress was managed with CPAP and oxygen therapy. UAC was removed on day 5. Complete renal failure (max: BUN 40, Creat 4.2) and hypertension (systolic BP > 110 mmHg) presented on day 8; UVC was removed. Renal scan showed no perfusion; abdominal ultrasound demonstrated complete aortic thrombosis from the superior mesenteric artery to the iliac arteries and including both renal arteries. Renal failure was managed with peritoneal dialysis without complications. Hypertension was controlled with hydralazine (max. 4.6 mg/kg/24 hr), propranolol (max. 0.5 mg/kg/24 hr), alphamethyldopa (35 mg/kg/24 hr), and intermittent diazoxide (2.5 mg/kg/dose). Urine production resumed (0.6 cc/kg/hr) on day 22 and increased progressively without surgical or thrombolytic therapy. Renal function was markedly improved (BUN 3, Creat 1.4) at three months. Previous reports of management of renovascular hypertension have suggested revascularization, thrombectomy or thrombolysis. Management with invasive therapy must be carefully considered as supportive management of this infant resulted in resolution of arterial obstruction and return of reasonable renal function.

In vitro studies of human renal transport have been limited in part because of the difficulty in obtaining adequate tissue. Cell culturing techniques afford the opportunity to perform such studies with limited amounts of tissue. Starting with small samples of normal human cortex obtained at nephrectomy for cancer, we were able to culture renal cortex epithelial cells that were free of fibroblast contamination by using a hormonally defined, serum-free medium. These cells proliferated and formed a monolayer, adhering to the surface of the culture flask. The cells could be passed for up to 5-6 times. The cell membrane facing the media had microvilli. There were connections between the cells resembling the tight junctions observed in vivo between renal tubule cells. When they were nearly confluent, "dome" formation, the lifting of the monolayer off the supporting structure, occurred suggesting active sodium and water movement. Dome formation became more evident with the addition of fetal calf serum to the medium. The cultured cells had alkaline phosphatase activity on the plasma membrane suggesting a proximal tubule origin. Because of this proximal tubule characteristic, we examined the ability of these cultured cells to transport the amino acid cystine. The monolayer progressively transported 35S-cystine for up to 60 min. of incubation. Lysine inhibited cystine uptake by these cells. This is the first in vitro evidence for the interaction between cystine transport and lysine which has been observed in vivo in the human kidney. Cultured renal cortex cells offer a new approach to the study of renal transport in humans.

Functional immaturity of the proximal tubule as reflected in a decreased maximal tubular reabsorptive capacity for many substrates, such as glucose (TmG), has been demonstrated in infants when compared to older children and adults. But when these measurements are factored by GFR, values comparable to those found in adults are obtained suggesting that the lower TmG in infants is related to the diminished GFR. To explore the nature of sugar handling by the developing kidney independent of GFR,we examined the uptake of the model sugar, α-methylglucoside (αMG), by isolated renal cortical tubule fragments from dogs over the age range of 5 days to adulthood. In tubules from pups less than 1 week old,concentrative uptake of 2mM 14C-αMG occurred by 1 min of incubation and reached a steady-state by 60 min with a distribution ratio of concentration (D.R.) of 8.4. With dogs 3 months old, the initial uptake was more rapid than the newborn pups reaching steady-state by 15 min, but the steady-state D.R. was lower(6.3). The kinetic parameters of initial uptake were for the 1 week old dog Km=4.87,Vmax=22.68mmoles/1 intracellular fluid/5 min; the 3 month old Km=8.05, Vmax=54.17; and the adult Km=7.42, Vmax=44.19. A sugar transport system is present in immature animals which resembles the adult system. With maturation there is a marked increase in the Vmax but only a small change in the Km. In the dog, the decreased TmG in immature animals appears to be related, in part, to a decreased number of carriers mediating sugar uptake into the renal tubule cell.

Because children with myelomeningocele (MM) are at increased risk of renal insufficiency, a simple method of estimating GFR from the plasma creatinine (PCr) and height would be useful. However, this estimate is predicated on a normal muscle mass and creatinine production for weight. Muscle mass may be diminished in children with paralysis. We therefore compared the estimate of GFR/1.73M2 from the formula 0.55 height (cm)÷ PCr with that determined with inulin in 14 children with MM ranging in age from 4-17 years. Seven had severe paralysis requiring wheelchair (WC) mobility. IVPs were abnormal in 9/14; 7/14 had inulin GFR < 80 ml/min/1.73M2. In 8 patients (57%) the estimated GFR averaged 85% higher than the inulin GFR. Of these 8 patients, 6 were WC-bound. In 5 patients (36%), there was close correlation between the two GFR methods, with an average difference of 11%. Only 1 of these was WC-bound. In 1 child, the estimated GFR was 64% lower than inulin GFR. All but 1 of the patients had PCr < 1 mg/dl. Our data suggest that an estimate of GFR determined from the serum creatinine and height in children with MM and significant paralysis is not reliable. This probably results from a diminished muscle mass and creatinine production per unit of body weight. Further studies are necessary to establish whether or not there is a reliable method for estimating GFR in this population.

GGT, which is concentrated on the renal proximal tubule brush-border membrane, is postulated to be important for the renal reabsorption of amino acids. Acivicin (AT-125) is a potent inhibitor of GGT allowing examination of this hypothesis. AT-125 inhibition of GGT activity in brushborder membrane vesicles (BBMV) from adult rats was both temperature and time dependent. After 20 min. of incubation at 37°C, 0.25 mM AT-125 inhibited 98% of GGT activity in BBMV. This inhibition was not reversed with repeated washing consistent with an affinity site inhibitor. AT-125 had no effect on another BBMV enzyme, alkaline phosphatase. AT-125 (20 mg/kg body weight) given to adult rats prior to BBMV isolation inhibited GGT activity by 90%. Cystine and glutamine uptake by AT-125 treated BBMV was unaffected, despite the fact that these amino acids are excellent substrates for GGT. AT-125 also had no effect on the BBMV uptake of proline, glycine, methionine, leucine and lysine. Cystine uptake by isolated renal cortical tubules prepared from rats treated with AT-125 was also unaffected, indicating that GGT inhibition had no effect on whole cell amino acid transport. These data suggest that GGT plays little or no role in renal amino acid transport.