Effect of bolus and divided feeding on urine ions and supersaturation in genetic hypercalciuric stone-forming rats.

Because urine ion excretion varies throughout the day, clinicians monitor 24 h urine samples to measure ion excretion and supersaturation in kidney stone patients. However, these results are averages and may not reflect maximal supersaturation which drives stone formation. We measured ion excretion and saturation in genetic hypercalciuric stone-forming rats on both a normal or low calcium diet over 0-3, 3-6 and 6-24 h using two feeding protocols, where the daily food allotment was fed either as a bolus or divided into three portions. With a normal calcium diet, urine calcium, oxalate, volume, and calcium oxalate supersaturation were significantly greater on the bolus compared to the divided feeds in the prandial and postprandial periods. Bolus eaters also excreted more calcium and oxalate and had increased volume over 24 h. Maximal calcium oxalate supersaturation was greater during the initial time periods than during the entire 24 h, regardless of the feeding schedule. With the low calcium diet, the effect of bolus feeding was reduced. Thus, urine ion excretion and supersaturation vary with the type of feeding. If these results are confirmed in man, it suggests that eating as a bolus may result in greater prandial and postprandial calcium oxalate supersaturation. This may increase growth on Randall's plaques and promote stone disease.

Urine calcium excretion predicts bone loss in idiopathic hypercalciuria.

Although idiopathic hypercalciuria (IH) is associated with reduced bone mineral density (BMD), no studies to date have identified predictors of BMD change over an extended period of observation. We have studied change in femoral neck and spine BMD z-scores in men and women with IH and stone disease (IHSF) and their first-degree relatives in order to determine the predictive value of commonly made clinical measurements. Urine calcium excretion was inversely correlated with change in femoral neck z-score over 3 years, and marginally correlated with fall in spine z-score. Markers of bone turnover, serum calcitriol, and urine measurements of acid-base balance such as ammonium and sulfate had no predictive value, nor did calcium intake assessed using a well-established questionnaire. It would appear that IHSF with the highest 24-h urine calcium excretion rates are at highest risk for loss of femoral neck bone mineral over a 3-year period.

Effect of cinacalcet on urine calcium excretion and supersaturation in genetic hypercalciuric stone-forming rats.

Idiopathic hypercalciuria is the most common metabolic abnormality in patients with nephrolithiasis. Through successive inbreeding, we have developed a strain of rats whose urine calcium (UCa) excretion is approximately 8-10-fold greater than that of control rats and who spontaneously form kidney stones. We have termed these rats genetic hypercalciuric stone-forming (GHS) rats. The physiology of the hypercalciuria in the GHS rats closely parallels that of man. We have recently shown that the GHS rat kidneys have an increased number of receptors for calcium (CaR) compared to Sprague-Dawley rats, the strain of rats originally bred to develop the GHS rats. Calcimimetics, such as cinacalcet (Cin), increase the sensitivity of the CaR to Ca. The effects of Cin on UCa are complex and difficult to predict. We tested the hypothesis that Cin would alter urinary (U) Ca and supersaturation with respect to calcium hydrogen phosphate (CaHPO(4)) and calcium oxalate (CaOx). GHS or control rats were fed a normal Ca diet (0.6% Ca) for 28 days with Cin (30 mg/kg/24 h) added to the diet of half of each group for the last 14 days. The protocol was then repeated while the rats were fed a low Ca (0.02% Ca) diet. We found that Cin led to a marked reduction in circulating parathyroid hormone and a modest reduction in serum Ca. Cin did not alter UCa when the GHS rats were fed the normal Ca diet but lowered UCa when they were fed the low Ca diet. However, Cin did not alter U supersaturation with respect to either CaOx or CaHPO(4) on either diet. If these findings in GHS rats can be confirmed in man, it suggests that Cin would not be an effective agent in the treatment of human idiopathic hypercalciuria and resultant stone formation.

Urinary cystine excretion and capacity in patients with cystinuria.

The treatment of cystinuria is hampered by methods used to measure urinary lithogenicity. Most cystine assays cannot reliably distinguish cystine from soluble thiol drug-cysteine complexes. We used a solid-phase assay of urinary cystine capacity in a large sample of patients with cystinuria. A known amount of solid-phase cystine is added to urine. In supersaturated urine, cystine precipitates onto added crystals, so the solid phase recovered after incubation will be greater than that added. We studied the effect of cystine-binding thiol drugs (CBTD) to solubilize cystine and determined correlates of cystine capacity in patients who were and were not taking CBTD. Increasing concentrations of D-penicillamine, tiopronin and captopril dissolved cystine in urine with similar efficacy. A general linear model in which 24 h cystine excretion was the dependent variable showed that creatinine, urea nitrogen, and sodium excretions were associated with cystine excretion (P<0.02, all three). Urine volume, pH, and cystine excretion strongly correlated with cystine capacity (P<0.001). Tiopronin had no effect on supersaturation in a cross-sectional analysis. A subset of supersaturated samples, with negative cystine capacity, occurred mainly among women not taking CBTD. For this subset, capacity differed significantly between CBTD users and non-users; use of CBTD avoided extremes of supersaturation. Female enrichment in the supersaturated group was accounted for in part by underprescription of CBTD to women. This assay of cystine capacity was reliable in the presence of CBTD. It should be useful in monitoring patients' response to dietary interventions and administration of fluid, citrate, and CBTD.

Patient adherence to long-term medical treatment of kidney stones.

PURPOSE: We determine patient adherence to and quality of outcome of medical kidney stone treatment during a 30-year duration at a single university based referral clinic. We also analyze time trends in adherence and timing of followup measurements, and supersaturation reduction during treatment. MATERIALS AND METHODS: Data on all patients who entered the University of Chicago Kidney Stone Prevention Program from 1970 to 2000 were analyzed. Fractions of new patients who had any followup and those remaining in followup at increasing intervals were analyzed. Timing of followup was measured. Changes in adherence during the 3 decades were also analyzed, as was reduction in supersaturation in regard to calcium oxalate, calcium phosphate and uric acid. RESULTS: A total of 70% to 80% of patients were retained at each successive followup cycle with 2 physicians, and a clinical protocol that always required 6-week followup with 24-hour urine collection and a yearly one thereafter for stone risk factors. Retention decreased during the last 5 years of the 1990s. Supersaturation reduction was present by the first followup and remained constant or improved with time. Timing of followup measurements was in accord with our protocol. CONCLUSIONS: At best, one can retain only 70% to 80% of patients in a followup program at each interval, and achieve supersaturation reductions that are constant and significant during the long term. Timing of followup measurements can be close to that of the protocol in use.

Proportional reduction of urine supersaturation during nephrolithiasis treatment.

PURPOSE: During metabolic stone therapy, urine supersaturation decreases in proportion to pretreatment levels. We gauge the quantitative contribution of regression to the mean for reducing urine supersaturation from high pretreatment to lower values during therapy. MATERIALS AND METHODS: The 24-hour urine supersaturations for calcium oxalate, calcium phosphate and uric acid were measured on 2 pretreatment and at least 1 treatment 24-hour collection for each of the 2,667 patients in 2 networks and at a university based specialty clinic. Changes in supersaturation between the first and second pretreatment collections were an estimate of random change and compared to therapeutic changes. RESULTS: Supersaturations decreased between the first and second pretreatment collections, proportional to the supersaturation in the first collection. However, the magnitude of this effect was minor compared to therapeutic changes. Also, mean change between pretreatment collections was 0, whereas mean change with therapy was greater than 0 for all 3 supersaturations. CONCLUSIONS: Although regression to the mean can be detected, it cannot be responsible for the decrease in urine supersaturation with therapy or the fact that the decrease is proportional to pretreatment mean supersaturation. The mechanisms responsible for proportional reduction remain to be clarified.

Effect of grapefruit juice on urinary lithogenicity.

PURPOSE: An increased risk of nephrolithiasis has been associated with the ingestion of grapefruit juice in epidemiological studies. To our knowledge the basis of this effect of grapefruit juice has not been studied previously. We studied the effect of grapefruit juice consumption on urinary chemistry and measures of lithogenicity. MATERIALS AND METHODS: Ten healthy men and women between ages of 25 and 40 years participated. Each subject drank 240 ml. of tap water at least 3 times daily for 7 days during the control period. This period was followed by a second 7 days experimental period during which they drank 240 ml. of grapefruit juice 3 times daily. In each 7-day period urine was collected for 24 hours during the last 3 days. Urine chemical analysis was performed, supersaturations of calcium oxalate, calcium phosphate and uric acid were calculated and urinary lithogenicity was measured. RESULTS: Urine volume and creatinine excretion were the same during the control and experimental periods. Grapefruit juice ingestion was associated with an increase in mean oxalate excretion plus or minus standard deviation of 41.1 +/- 9.2 to 51.9 +/- 12.0 mg. per 24 hours (p = 0.001) and in mean citrate excretion of 504.8 +/- 226.5 to 591.4 +/- 220.0 mg. per 24 hours (p = 0.01). There was no net change in the supersaturation or upper limit of metastability of calcium oxalate, calcium phosphate or uric acid. Crystal aggregation and growth inhibition by urinary macromolecules was not affected by grapefruit juice ingestion. CONCLUSIONS: Offsetting changes in urine chemistry caused by the ingestion of grapefruit juice led to no net change in calculated supersaturation. No changes in lithogenicity were demonstrated. The results do not demonstrate an effect of grapefruit juice for increasing lithogenicity. The basis of the observations of epidemiological studies remain unexplained.

Differences in urine volume and supersaturation in 2 physician networks.

PURPOSE: We determined whether a network of practices devoted to a broad range of urological care would achieve a decrease in metabolic stone risk comparable to that achieved by a network of similar practices that emphasized kidney stone management as a distinct specialized interest, provided that each was given equivalent access to high level urine testing and software support. MATERIALS AND METHODS: Pretreatment and treatment 24-hour urine samples were obtained from patients in a large network of practices related by the shared use of lithotripsy facilities and instruments (group 2) and a contrasting network of practices that emphasize stone treatment over other concerns (group 1). All known urine risk factors, including supersaturation, were measured and calculated. RESULTS: Treatment supersaturation values in group 2 exceeded those in group 1. The reason was unpredicted and unexplained but highly consistent lower urine volume in group 2 patients that was present before and persisted during treatment. Group 2 physicians mostly achieved changes in urine volume and stone risk factors equivalent to those of group 1 physicians but began with higher supersaturation due to lower urine volume. CONCLUSIONS: A network of physicians not specialized for stone care may achieve a decreased risk equivalent to that of more specialized physicians. Initial patient characteristics may vary significantly in the groups for reasons that are unknown to date, greatly affecting treatment outcome.

Solid phase assay of urine cystine supersaturation in the presence of cystine binding drugs.

PURPOSE: We report a new kind of assay system for urine cystine supersaturation that is accurate in the presence of cystine binding thiol drugs. We measured the molar ratio of cystine dissolved per mole of drug. MATERIALS AND METHODS: Measured amounts of cystine crystals were incubated in buffer or urine for 48 hours with stirring. The solid phase remaining was pelleted by centrifugation, extracted into a high pH buffer and measured. D-penicillamine, tiopronin and captopril were added to determine their effect on solid phase dissolution. RESULTS: Total cystine calculated from urine and solid measurements closely matched the amounts of cystine weighed in, meaning that the assay system successfully recovered the total cystine from the 2 phases. Each drug dissolved solid cystine in a specific and fixed proportion to its molar concentration in the range of 0.2 to 0.4 mM. dissolution per mM. of drug. Solution measurements were not a reliable gauge to the actual amounts of cystine dissolved. CONCLUSIONS: Changes in solid phase cystine accurately reflect buffer or urine supersaturation when thiol drugs are present. The solid phase assay is a technically straightforward and reliable way of assessing cystine movement into and out of urine that avoids complexity of measurement and distortions of assay systems by drugs. This assay enables one to assess the level of drug effect and the need for a change in dosing.

Phosphorylated osteopontin peptides suppress crystallization by inhibiting the growth of calcium oxalate crystals.

BACKGROUND: Osteopontin isolated from human urine [uropontin (uOPN)] is a potent inhibitor of calcium oxalate (CaOx) monohydrate (COM) crystallization. However, specific structural features responsible for its effects on CaOx crystallization were not previously known. The present studies were designed to define molecular features responsible for interactions of uOPN with COM crystals and the inhibition of crystallization. METHODS: Peptides and phosphopeptides with sequences corresponding to potential crystal binding domains within the protein sequence of osteopontin were synthesized. Then the effects of these peptides on COM crystal growth and crystal aggregation were investigated and their secondary structures analyzed. RESULTS: Growth of COM crystals was inhibited by approximately 50% at 1000 nmol/L concentrations by the two unmodified peptides with the closest clustering of aspartic acid residues. Growth was not inhibited by the other two unmodified peptides, with aspartic residues more evenly distributed within their sequences. Phosphorylation markedly increased inhibition of COM crystal growth, so that each of the four phosphorylated peptides inhibited growth by at least 50% at concentrations of < or =200 nmol/L. Phosphorylation of these peptides did not cause changes in secondary structure that would favor interaction with COM crystal surfaces. CONCLUSIONS: These studies of synthetic peptides identify molecular features within the osteopontin molecule that contribute to the inhibition of one aspect of COM crystallization. The inhibition of crystal growth induced by phosphorylation appears to result from altered local patterns of charge density, since conformational changes favoring interaction with crystals were not caused by phosphorylation.

Effect of acidosis on urine supersaturation and stone formation in genetic hypercalciuric stone-forming rats.

BACKGROUND: We have successively inbred over 45 generations a strain of rats to maximize urine calcium excretion. The rats now consistently excrete 8 to 10 times as much calcium as controls and uniformly form poorly crystalline calcium phosphate kidney stones. In humans with calcium nephrolithiasis, consumption of a diet high in acid precursors is often cited as a risk factor for the development of calcium-based kidney stones; however, the effect of this diet on urinary supersaturation with respect to the common solid phases found in kidney stones has not been determined. METHODS: To determine the effect of the addition of an acid precursor on urine ion excretion, supersaturation, and stone formation, we fed these genetic hypercalciuric stone-forming (GHS) rats 13 g/day of a 1.2% calcium diet with 0.0, 0.5, 1.0, or 1.5% NH4Cl in the drinking water for 14 weeks (N = 8 for each). Urine was collected and analyzed every two weeks. RESULTS: As expected, the addition of dietary NH4Cl led to a progressive fall in urine pH and urine citrate, while urine ammonium increased. Urine calcium and phosphorus increased, while urine oxalate fell. Increasing dietary NH4Cl led to a fall in supersaturation with respect to CaHPO4 (brushite) and CaOx and a rise in supersaturation with respect to uric acid. In spite of differences in supersaturation, most rats in each group formed stones that contained calcium phosphate and not calcium oxalate. CONCLUSIONS: Thus, while the provision of additional dietary acids alters urinary ion excretion and lowers supersaturation with respect to CaHPO4 and CaOx, it does not change the character or rate of stone formation in the GHS rats.

Clinical use of cystine supersaturation measurements.

PURPOSE: We measured the concentration and solubility of cystine in urine from patients with cystinuria or calcium stones and from normal subjects to determine whether urine cystine supersaturation can be calculated from a standard nomogram of solubility versus pH or needs to be measured directly. We also evaluated whether increasing pH of the 24-hour collection recovered enough crystallized cystine to increase cystine supersaturation. MATERIALS AND METHODS: Cystine concentration, pH and usual stone risk factors were measured on 50 ml. aliquots of 24-hour collections from 24 patients with cystinuria, 22 calcium stone formers and 15 normal subjects. After 48 hours of incubation with sodium bicarbonate, a second aliquot was taken from the 24-hour collection for cystine concentration. The original urine at its ambient pH was incubated with an excess of cystine crystals for 24, 48, 72 or 96 hours at 37C to determine solubility and kinetics of equilibration. RESULTS: Cystine solubility varied so widely at any pH range that no predictive nomogram could be relied on for calculating supersaturation. Addition of sodium bicarbonate to the 24-hour urine significantly increased cystine concentration. Urine from stone formers had higher cystine solubility than urine from normal subjects. CONCLUSIONS: Clinical management of cystinuria can be improved by direct measurement of cystine solubility because it varies widely at any given pH. Increasing 24-hour collection pH with sodium bicarbonate additionally improves accuracy of supersaturation measurement by recovering crystallized cystine.

Does a high concentration of calcium in the urine cause an important renal concentrating defect in human subjects?

The objective of this study was to evaluate the hypothesis that a high concentration of ionized calcium in the lumen of the medullary collecting duct causes an osmole-free water diuresis. The urine flow rate and osmolality were measured in normal human subjects, as well as in patients with a history of nephrolithiasis who excreted more than 5 mmol of calcium per 24 h. There was an inverse relationship between the concentration of calcium in the urine and the 24 h urine volume both in normal subjects and in patients with a history of nephrolithiasis. When the concentration of calcium in the urine was greater than 5 mmol/l, the urine volume was less than 1 litre per day in the majority of subjects. After 16 h of water deprivation, when the concentration of calcium in the urine was as high as 17 mmol/l (ionized calcium 7.4 mmol/l), urine osmolality was 1258 mOsm/kg of water and the urine flow rate was 0.30 ml/min. We conclude that, although a calcium receptor may be present in the lumen of the medullary collecting duct in human subjects, an extremely high concentration of urinary total and ionized calcium does not cause a clinically important defect in the renal concentrating process.

Isolated hypercalciuria with mutation in CLCN5: relevance to idiopathic hypercalciuria.

UNLABELLED: Isolated hypercalciuria with mutation in CLCN5: Relevance to idiopathic hypercalciuria. BACKGROUND: Idiopathic hypercalciuria (IH) is the most common risk factor for kidney stones and often has a genetic component. Dent's disease (X-linked nephrolithiasis) is associated with mutations in the CLCN5 chloride channel gene, and low molecular weight (LMW) proteinuria was universally observed in affected males. We sought to identify mutations in CLCN5 or abnormalities in LMW protein excretion in a large group of patients with IH and in a rat model of genetic hypercalciuria. METHODS: One hundred and seven patients with IH (82 adults and 25 children) and one asymptomatic hypercalciuric man with a known inactivating mutation in CLCN5 were studied. Secondary causes of hypercalciuria were excluded in all. The excretion of retinol-binding protein and beta2-microglobulin was measured by immunoassay in 101 patients with IH. Mutation analysis of the CLCN5 gene was performed in 32 patients with IH and in the genetic hypercalciuric stone-forming (GHS) rat strain. RESULTS: LMW protein excretion was normal in 92 patients with IH, and only slight abnormalities were found in the other nine, none of whom had a mutation in CLCN5. One 27-year-old man who had a CLCN5 mutation was found to have isolated hypercalciuria without LMW proteinuria, renal failure, or other evidence of renal disease. Mutation analysis was normal in 32 patients with IH. The CLCN5 sequence was normal in the GHS rat. CONCLUSIONS: Inactivation of CLCN5 can be found in the setting of hypercalciuria without other features of X-linked nephrolithiasis. However, mutations in CLCN5 do not represent a common cause of IH.

Calcium phosphate supersaturation regulates stone formation in genetic hypercalciuric stone-forming rats.

BACKGROUND: Hypercalciuria is the most common metabolic abnormality observed in patients with nephrolithiasis. Hypercalciuria raises urine supersaturation with respect to the solid phases of calcium oxalate and calcium phosphate, leading to an enhanced probability for nucleation and growth of crystals into clinically significant stones. However, there is little direct proof that supersaturation itself regulates stone formation. Through successive inbreeding of the most hypercalciuric progeny of hypercalciuric Sprague-Dawley rats, we have established a strain of rats, each of which excrete abnormally large amounts of urinary calcium and each of which forms calcium phosphate kidney stones. We used these hypercalciuric (GHS) rats to test the hypothesis that an isolated reduction in urine supersaturation, achieved by decreasing urine phosphorus excretion, would decrease stone formation in these rats. METHODS: Thirty 44th-generation female GHS rats were randomly divided into three groups. Ten rats received a high-phosphorus diet (0.565% phosphorus), 10 a medium-phosphorus diet (0.395% phosphorus), and 10 a low-phosphorus diet (0.225% phosphorus) for a total of 18 weeks. The lowered dietary phosphorus would be expected to result in a decrease in urine phosphorus excretion and a decrease in urinary supersaturation with respect to the calcium phosphate solid phase. Every two weeks, 24-hour urine collections were obtained. All relevant ions were measured, and supersaturation with respect to calcium oxalate and calcium hydrogen phosphate were determined. At the conclusion of the experiment, each rat was killed, and the kidneys, ureters, and bladder were dissected en block and x-rayed to determine whether any stones formed. A decrease in stone formation with a reduction in urinary supersaturation would support the hypothesis that supersaturation alone can regulate stone formation. RESULTS: Decreasing the dietary phosphorus intake led to a progressive decrease in urine phosphorus excretion and an increase in urine calcium excretion, the latter presumably caused by decreased intestinal calcium phosphate binding and increased calcium absorption. With decreasing dietary phosphorus intake, there was a progressive decrease in saturation with respect to the calcium phosphate solid phase. Fifteen of the 20 kidneys from the 10 rats fed the high-phosphorus diet had radiographic evidence of kidney stone formation, whereas no kidneys from the rats fed either the medium- or low-phosphorus diet developed kidney stones. CONCLUSIONS: A decrease in urine phosphorus excretion not only led to a decrease in urine supersaturation with respect to the calcium phosphate solid phase but to an elimination of renal stone formation. The results of this study support the hypothesis that variation in supersaturation alone can regulate renal stone formation. Whether a reduction of dietary phosphorus will alter stone formation in humans with calcium phosphate nephrolithiasis remains to be determined.

Reduced crystallization inhibition by urine from men with nephrolithiasis.

BACKGROUND: Human urine is known to inhibit growth, aggregation, nucleation, and cell adhesion of calcium oxalate monohydrate (COM) crystals, the main solid phase of human kidney stones. This study tests the hypothesis that low levels of inhibition are present in men with calcium oxalate stones and could therefore promote stone production. METHODS: In 17 stone-forming men and 17 normal men that were matched in age to within five years, we studied the inhibition by dialyzed urine proteins of COM growth, aggregation, and binding to cultured BSC-1 renal cells, as well as whole urine upper limits of metastability (ULM) for COM and calcium phosphate (CaP) in relationship to the corresponding supersaturation (SS). RESULTS: Compared with normals, patient urine showed reduced COM growth inhibition and reduced ULM in relationship to SS. When individual defects were considered, 15 of the 17 patients were abnormal in one or more inhibition measurements. ULM and growth inhibition defects frequently coexisted. CONCLUSIONS: Reduced COM growth and CaP and CaOx ULM values in relationship to SS are a characteristic of male stone formers. Both defects could promote stones by facilitating crystal nucleation and growth. Abnormal inhibition may be a very important cause of human nephrolithiasis.

Divergence between stone composition and urine supersaturation: clinical and laboratory implications.

PURPOSE: In general high urine supersaturation with respect to calcium oxalate, calcium phosphate or uric acid is associated with that phase in stones. We explore the exceptions when supersaturation is high and a corresponding solid phase is absent (type 1), and when the solid phase is present but supersaturation is absent or low (type 2). MATERIALS AND METHODS: Urine supersaturation values for calcium oxalate, calcium phosphate and uric acid, and other accepted stone risk factors were measured in 538 patients at a research clinic and 178 at stone prevention sites in a network served by a single laboratory. RESULTS: Of the patients 14% lacked high supersaturation for the main stone constituent (type 2 structural divergence) because of high urine volume and low calcium excretion, perhaps from changes in diet and fluid intake prompted by stones. Higher calcium excretion and low urine volume caused type 1 divergences, which posed no clinical concern. CONCLUSIONS: Type 1 divergence appears to represent a condition of low urine volume which raises supersaturation in general. Almost all of these patients are calcium oxalate stone formers with the expected high supersaturation with calcium oxalate as well as high uric acid and calcium phosphate supersaturations without either phase in stones. Type 2 divergence appears to represent an increase in urine volume and decrease in urine calcium excretion between stone formation and urine testing.

Medical reduction of stone risk in a network of treatment centers compared to a research clinic.

PURPOSE: We determined whether a network of 7 comprehensive kidney stone treatment centers supported by specialized stone management software and laboratory resources could achieve reductions in urine supersaturation comparable to those in a single research clinic devoted to metabolic stone prevention. MATERIALS AND METHODS: Supersaturation values for calcium oxalate, calcium phosphate and uric acid in 24-hour urine samples were calculated from a set of kidney stone risk factor measurements made at a central laboratory site for the network and research laboratory for the clinic. Individual results and group outcomes were presented to each center in time sequential table graphics. The decrease in supersaturation with treatment was compared in the network and clinic using analysis of variance. RESULTS: Supersaturation was effectively reduced in the network and clinic, and the reduction was proportional to the initial supersaturation value and increase in urine volume. The clinic achieved a greater supersaturation reduction, higher fraction of patient followup and greater increase in urine volume but the treatment effects in the network were, nevertheless, substantial and significant. CONCLUSIONS: Given proper software and laboratory support, a network of treatment centers can rival but not quite match results in a dedicated metabolic stone research and prevention clinic. Therefore, large scale stone prevention in a network system appears feasible and effective.

Evidence that calgranulin is produced by kidney cells and is an inhibitor of calcium oxalate crystallization.

Urine produced by normal human kidneys is almost always supersaturated with respect to calcium oxalate (CaOx), the most common constituent of human kidney stones. Crystallization, with risk of renal damage and kidney stones, appears to be affected by molecules in urine that retard nucleation, growth, aggregation, and renal cell adherence of CaOx. The repertoire of such molecules is incompletely known. We have purified a 28-kDa protein from urine using salt precipitation, preparative isoelectric focusing, and sizing chromatography. Amino acid composition and NH2-terminal amino sequence analysis showed complete homology to calgranulin. Calgranulin was found to be a potent inhibitor of CaOx crystal growth (44% of control) and aggregation (50% of control) in the nanomolar range. Calgranulin cDNA was cloned from a human kidney expression library. Western analysis of human and rat kidney homogenates and mRNA temporal expression from two independent renal epithelial cell lines showed that calgranulin is produced in the kidney. Given its urinary abundance and potency, calgranulin may contribute importantly to the normal urinary inhibition of crystal growth and aggregation and therefore to the renal defense against clinical stone disease.

Contribution of human uropontin to inhibition of calcium oxalate crystallization.

Uropontin (UP) is known to inhibit the growth and nucleation of calcium oxalate monohydrate (COM) crystals, and it also impedes attachment of calcium oxalate crystals to cultured renal epithelial cells. However, its role in normal defense against renal crystallization, and in pathogenesis of nephrolithiasis is unclear. In this study we determined the effect of UP on aggregation of COM crystals as well as the inhibitory activity of UP on COM crystal growth and nucleation in a series of normal subjects, in order to assess the potential of UP as an important urinary inhibitor. The mean urinary excretion of UP measured by ELISA was 185 +/- 12 nmol/24 hr (mean +/- SEM) with a mean urine UP concentration of 131 +/- 13 nM. Uropontin isolated by immunoaffinity chromatography was a very potent inhibitor of COM crystal aggregation, with a mean UP concentration of 28 +/- 4 nM required for a 50% reduction in aggregation. The kDa for COM crystal growth inhibition determined from Langmuir type isotherms was 21 +/- 3 nM and the concentration required for 50% reduction in COM crystal growth rate was 16 +/- 2 nM. Inhibition of secondary nucleation was measured at a single concentration of 200 nM, which reduced the nucleation rate to 42 +/- 3% of control. Using a theoretical model of growth and aggregation inhibition at varying urine flow rates, we showed that inhibitory activity of UP would be significant for all subjects over a wide range of urine flow rates. Overall, UP is a potent inhibitor of COM aggregation as well as growth and nucleation. The urinary concentration of UP is in the range in which its contribution to inhibition of growth and aggregation are likely to be substantial. Thus, UP appears to be an important natural defense against renal crystallizations and nephrolithiasis.