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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.

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.

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.

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.

Purpose: We determined what metabolic features of the 24-hour urine predict calcium oxalate dihydrate in kidney stones. Prior studies have suggested that low urine magnesium, high urine calcium, high calcium-to-oxalate ratio and high urine supersaturation with respect to calcium oxalate monohydrate predict calcium oxalate dihydrate., Materials and Methods: Stone analyses and results from 2, 24-hour pretreatment urine collections from 96 patients with nephrolithiasis were drawn from 3 kidney stone prevention centers. Standard stone risk measurements were made on the urine, including supersaturation for calcium oxalate monohydrate, brushite and uric acid., Results: The main differences in metabolic urine findings were between patients with no calcium oxalate dihydrate and those with any calcium oxalate dihydrate in stones. Percent calcium oxalate dihydrate itself did not correlate with urine findings. Patients with no calcium oxalate dihydrate in stones showed a biphasic pattern of urine calcium oxalate monohydrate supersaturation, about half had values below almost any found among patients with calcium oxalate dihydrate in stones (less than 7) and the rest overlapped with the calcium oxalate dihydrate group. Except for higher calcium oxalate monohydrate supersaturation, patients with calcium oxalate dihydrate in stones had higher urine calcium excretion and lower urine citrate concentrations, even after calcium oxalate monohydrate supersaturation was considered., Conclusions: Patients with low calcium oxalate monohydrate supersaturation (less than 7) are unlikely to have calcium oxalate dihydrate in renal stones. However, many patients with no calcium oxalate dihydrate have higher calcium oxalate monohydrate supersaturation values, and so prediction of calcium oxalate dihydrate or its absence from urine findings is imperfect. Urine magnesium and the calcium-to-oxalate ratio are unrelated to calcium oxalate dihydrate.

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.

Formation of renal stones requires supersaturation (SS) high enough to induce crystallization; such a SS is referred to as the upper limit of metastability (ULM). The ULM for calcium oxalate (CaOx) or calcium phosphate can be measured by adding oxalate or calcium to urine, respectively, and noting the point at which overt crystallization occurs as evidenced by clouding. In principle, the urine should be more prone to form stone crystals as its SS approaches the ULM, and the SS ULM distance has been used as an index of stone forming potential. In addition, one would expect the ULM and initial SS to be unrelated, as the starting urine SS has no apparent link to the amount of calcium or oxalate that urine can dissolve without leading to crystal formation. However, in rats, we have found a surprising correlation between ULM and SS, such that ULM appears to rise with initial SS, for CaOx, and, to a lesser extent, for brushite (Br), a typical calcium phosphate initial phase. In this study, we measured CaOx and Br ULM, and SS, in urine of 50 patients and 11 normal people, to determine if ULM and SS were correlated, as in rats, and to explore the relationship between SS and ULM. We found the same dependence of ULM on SS as in rats, for both CaOx and Br, and found no differences between patients and normal people with respect to this dependency. However, for Br, patients showed a lower ULM than normals, but the same initial SS, meaning that patients were closer to their crystal formation threshold than normals. Treatments for stones had no apparent effect on the SS-ULM dependency. We conclude that in humans, as in rats, ULM is related to initial SS, and that this relationship is the same in patients as in normals for CaOx, but shifted in a stone forming direction for Br among patients. The ULM-SS interaction is unaffected by contemporary conventional stone treatments, and is more marked for CaOx than Br. The mechanisms of the dependence are unknown. The smaller difference between ULM and initial SS for Br in patients than normal supports prior evidence suggesting a defect in stone patients that could lead to calcium phosphate crystallization, subsequent nucleation of CaOx, and stone disease.

Calcium oxalate (CaOx) and calcium phosphate (CaP) crystals do not precipitate in large amounts in normal urine despite considerable supersaturation (SS), partly because urine inhibits crystal nucleation, aggregation, and growth. In normal rats and rats bred for hypercalciuria (GHS), we varied SS by varying calcium intake to test the hypothesis that increased SS might deplete inhibitors and reduce inhibition of crystal formation. In normal rats when compared to a low calcium diet (0.02% Ca), a high calcium diet (1.2% Ca) raised the SS of CaOx from 0.8 to 8.2. The high calcium diet also raised the upper limit of metastability (ULM) of CaOx (the SS at which crystals form in urine) from 11.8 to 36. In GHS rats, diet change altered CaOx SS from 1.5 to 12, and ULM from 17 to 50 (all differences, P < 0.001). Because ULM rose with SS, the increased SS had little potential to increase CaOx stone risk. For CaP, however, SS rose from 0.6 to 2.4 and 1.1 to 8 in normal and GHS rats (P < 0.001 for both), respectively, whereas ULM for CaP did not increase significantly (8 vs. 7 and 7 vs. 11; P = NS, both changes). Therefore, CaP SS rose close to the ULM, posing a high stone risk. The stones formed by these rats are composed of CaP. Increasing CaOx SS by diet raises ULM for CaOx thereby offsetting the risk of CaOx stones in rats.

Uric acid is the end product of purine degradation in humans. It is a weak acid with a pK of 5.35 in urine. In acidic urine the undissociated form of uric acid predominates and is poorly soluble, leading to crystalluria and stone formation. Uric acid stones account for 5% to 10% of all stones in the United States. Acidic urine, hyperuricosuria, and low urine volume are the risk factors for uric acid stone formation. Uric acid stones may be idiopathic or secondary to a systemic disease such as gout. Treatment includes alkalinization of the urine, high fluid intake, and reduction of uric acid excretion if it is abnormal.

Urine from mammalian kidneys is regularly supersaturated with respect to calcium oxalate monohydrate, the most common solid phase in human nephrolithiasis, and also inhibits the nucleation, growth, and aggregation of calcium oxalate crystals. Nephrolithiasis is often associated with increased supersaturation, and it is assumed that this increase overbalances the inhibition effects, causing stones. However, some patients form stones in the absence of increased supersaturation, and in those patients, one might assume that reduced inhibition is the cause of their stones. This hypothesis was tested in 25 patients who formed at least ten stones each, yet lacked the usual metabolic abnormalities that increase supersaturation. Compared with 25 age- and sex-matched control subjects, urine supersaturation among the patients was not increased; this is an expected result of this study's selection criteria. Compared with the same age- and sex-matched control subjects, urine from the patients showed no evidence for reduced inhibition of calcium oxalate crystal growth, so low inhibition of growth did not contribute to pathogenesis of stones in our highly selected study population, despite their otherwise unexplained and active stone formation. These results do not support the hypothesis that growth inhibition defects are a cause of stone disease.

We have used published rat micropuncture data to construct a matrix of ion concentrations along the rat nephron. With an iterative computer model of known ion interactions, we calculated relative supersaturation ratios in all nephron segments. The collecting ducts and urine showed expected supersaturation with stone-forming salts. Fluid in the thin segment of the loop of Henle may be supersaturated with calcium carbonate and calcium phosphate under certain conditions. Because calculations cannot predict the actual course of crystallization, we made solutions to mimic, in vitro, presumed conditions in the loop of Henle. The solid phases that formed were analyzed by X-ray powder diffraction, electron microprobe, and infrared spectroscopy. All samples were identified as poorly crystallized or immature apatite. The descending limb of Henle's loop creates a unique condition as it extracts water but not sodium, bicarbonate, calcium, or phosphate, giving a calcium concentration at the bend of 3 mM, pH 7.4, and a phosphate concentration that varies from 0.8 to 48 mM, depending on parathyroid hormone and dietary phosphate. We conclude that conditions in the thin segment potentially could create a solid calcium phosphate phase, which may initiate nucleation of calcium oxalate salts in the collecting ducts, potentiating nephrolithiasis and nephrocalcinosis.