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Chemical measurement of calcium oxalate crystalluria: results in various causes of calcium urolithiasis.

Calcium oxalate and calcium phosphate crystalluria have been measured chemically in 1,173 urine samples whose chemical compositions were also analysed. The importance of urinary oxalate as a determinant for calcium oxalate crystalluria was confirmed. Significant concentrations of calcium oxalate crystals may be present in urine even though the crystals are too small for detection by light microscopy or by many particle-counting methods. Calcium phosphate crystals in urine always contain a small proportion of calcium oxalate. Results in various clinical situations are reviewed.

Calcium Oxalate↗

Idiopathic calcium oxalate urinary lithiasis: usefulness of Parks' and Tiselius' indices in the evaluation of the risk of stone formation.

Different indices of the risk of urinary calcium oxalate crystallization were compared to determine their usefulness in detecting the stone-formers particularly prone to recurrence. Urine volume and calcium, oxalate, citrate, magnesium or creatinine were determined in 55 patients presenting with an idiopathic calcium oxalate urolithiasis, as well as in 50 control subjects. On 24-hour urine samples, these elements allowed for the calculation and comparison of different indices of lithogenous risk as proposed by Parks and Tiselius. Both Parks' indices and the urinary citrate-calcium ratio varied significantly between the two groups, but conversely Tiselius' indices were statistically comparable. The three Tiselius' indices taking the 24-hour urine volume into account were also strongly correlated. Parks' index and the urinary citrate-calcium ratio are highly discriminating and potentially relevant to select the stone-formers with a high risk of relapse. Tiselius' indices basically reflect urinary calcium oxalate saturation, and can only be used clinically to control the treatment interfering with this. In this respect, the formula based simply on urine volume, calcium and oxalate over 24 h (Ca0.71.Ox.V-1.2) appears to be sufficient.

Adolescent↗

Histological observations of the adhesion and endocytosis of calcium oxalate crystals in MDCK cells and in rat and human kidney.

Adhesion and/or endocytosis of calcium oxalate crystals to the three kinds of tubular cells (Madin-Darby canine kidney (MDCK) cells, rat and human kidney) were demonstrated morphologically to presume the initial formation of kidney stone. After removal of the nonadhesion crystals, the cells were subsequently recultured in the vertical position. At various times thereafter, the interactions between COM crystals and MDCK cells were evaluated morphologically by SEM. COM crystals adhered to the surface of MDCK cells immediately, and the crystals were then endocytosed. The microvilli of the cells appeared to play an important role in these processes. At later times, some complexes that consist of aggregated calcium oxalate crystals and cell debris were observed sporadically. Kidney tissues were obtained from male Sprague-Dawley rats which were injected with sodium oxalate intraperitoneally. Experimentally induced calcium oxalate crystals were evaluated histologically using polarized light microscopy. Some crystals in the cortical portion were attached to the tubular epithelium or internalized into the luminal membrane. Whereas in the papilla, the aggregated crystals were observed lying free from the degenerated tubular lumen along with the cell debris. Human kidney tissues were obtained from 38 patients with calcium oxalate nephrolithiasis who underwent nephrolithotomy or partial nephrectomy before the era of ESWL. The specimens were examined for calcium crystals within the tubular lumen, attached to the tubular walls or internalized into the tubular cells, by polarized light microscopy. Approximately 50% of the specimens observed crystals attached to the tubular cell epithelium and some of them were seen inside the tubular cells. In conclusion, crystal-cell interaction resulted in movement of crystals from the lumen into the cells by an action of microvilli from the results of MDCK cells. However, it was not clear from the results in rats or human kidney tissue that crystal adhesion and/or endocytosis might be vital in the crystal growth in the kidney.

Animals↗

Absorption of oxalic acid in rats by means of a 14C method.

13 rats received a daily dose of 0.5 ml of a 1% solution of 14C-marked sodium oxalate (specific activity 29 nCi/ml) administered in one application with a throat probe. Feces and urine were collected every 24 h and the amount of 14C-oxalate was determined by liquid scintillation counting. The average distribution for 13 rats during 7 days showed 25.2% of 14C-oxalate in the urine and 73.4% in the feces. The rate of absorption is considerably higher than values given in literature (2.6--12% urine). These results indicate that in the pathogenesis of calcium oxalate stone formation some possible significance must be attributed to the exogenic oxalate.

Absorption↗

Evaluation of the risk of stone formation: study on crystalluria in patients with recurrent calcium oxalate urolithiasis.

OBJECTIVE: The aim of this study was to determine the usefulness of the morning calcium oxalate crystalluria in detecting stone formers particularly prone to recurrence. METHODS: Over a 24-hour period of urine collection, the morning calcium oxalate crystalluria was evaluated as well as the risk of stone formation, established with Tiselius and Parks indices, for 25 recurrent stone formers (group 1) and 25 normal controls (group 2). RESULTS: Morning crystalluria (type, size, number/ml and state of aggregate) and the Tiselius index were comparable in the two groups. Conversely, calciuria as well as the citrate/ calcium ratio and the Parks index varied significantly for stone formers and normal controls. No particular correlation appeared between crystalluria and indices of Tiselius and Parks, calciuria, calcium-oxalate product or calcium/ oxalate and citrate/calcium ratios. CONCLUSIONS: Morning calcium oxalate crystalluria does not enable an efficient characterization of recurrent stone formers. Its discordance with others potential indicators of the risk of stone formation poses the problem of their respective validity and evokes the prevalence of still unknown inhibiting agents in the phenomenon of crystallization.

Calcium Oxalate↗

Urine glycoprotein crystal growth inhibitors. Evidence for a molecular abnormality in calcium oxalate nephrolithiasis.

One reason that some people are prone to calcium oxalate nephrolithiasis is that they produce urine that is subnormal in its ability to inhibit the growth of calcium oxalate crystals. We have identified in human urine a glycoprotein (GCI) that inhibits calcium oxalate crystal growth strongly, and at low concentrations (10(-7) M); in this study, we have isolated GCI molecules from the urine of normal people and patients with calcium oxalate stones. GCI from stone formers is abnormal in three ways: it contains no detectable gamma-carboxyglutamic acid (Gla), whereas normal GCI contains 2-3 residues of Gla per mole; about half of the GCI in urine of patients inhibits crystal growth 4-20 times less than normal GCI as judged by its performance in a kinetic growth assay, in vitro; at the air-water interface, patient GCI has a film collapse pressure approximately half of normal. GCI molecules from the urine of patients with calcium oxalate nephrolithiasis are intrinsically abnormal, and these abnormalities could play a role in the genesis of stones.

1-Carboxyglutamic Acid↗

Effect of Tamm-Horsfall protein on calcium oxalate precipitation.

The effect of Tamm-Horsfall protein isolated from urine of healthy subjects on calcium oxalate precipitation was studied in model systems of precipitation. The study was performed using following conditions: concentrations of calcium chloride 10 mmol/l, sodium chloride 150 mmol/l, oxalic acid 300 mumol/l; pH 6.0, and temperature 310 K. The concentration of Tamm-Horsfall protein varied between 1-10 mg/l. The kinetics of calcium oxalate precipitation was observed by measuring the number and volume of particles in the suspension, and the precipitate composition by an optic microscope. In all the studied systems, the precipitate morphology corresponded to pure calcium oxalate monohydrate. Tamm-Horsfall protein was found to inhibit the growth of calcium oxalate monohydrate crystals and stimulate their aggregation in the given experimental conditions. Both effects were enhanced by increase in the concentrations of Tamm-Horsfall protein and were most pronounced at the concentration of Tamm-Horsfall protein of 10 mg/l.

Adult↗

The influence of freezer storage of urine samples on the BONN-Risk-Index for calcium oxalate crystallization.

This study was performed to quantify the effect of a 1-week freezer storage of urine on its calcium oxalate crystallization risk. Calcium oxalate is the most common urinary stone material observed in urolithiasis patients in western and affluent countries. The BONN-Risk-Index of calcium oxalate crystallization risk in human urine is determined from a crystallization experiment performed on untreated native urine samples. We tested the influence of a 1-week freezing on the BONN-Risk-Index value as well as the effect of the sample freezing on the urinary osmolality. In vitro crystallization experiments in 49 native urine samples from stone-forming and non-stone forming individuals were performed in order to determine their calcium oxalate crystallization risk according to the BONN-Risk-Index approach. Comparison of the results derived from original sample investigations with those obtained from the thawed aliquots by statistical evaluation shows that i) no significant deviation from linearity between both results exists and ii) both results are identical by statistical means. This is valid for both, the BONN-Risk-Index and the osmolality data. The differences in the BONN-Risk-Index results of both procedures of BONN-Risk-Index determination, however, exceed the clinically acceptable difference. Thus, determination of the urinary calcium oxalate crystallization risk from thawed urine samples cannot be recommended.

Calcium Oxalate↗

Structure and transport mechanism of the bacterial oxalate transporter OxlT.

Membrane proteins that belong to the major facilitator superfamily (MFS) are found in organisms across the evolutionary spectrum and mediate the transport of a variety of substrates ranging from small metabolites to neurotransmitters. The oxalate transporter (OxlT) is a representative MFS protein, and exchanges formate for oxalate across the cytoplasmic membrane of the organism Oxalobacter formigenes. Here, we present a structural model for the protein conformational changes that occur during oxalate transport by combining a three-dimensional map of the oxalate-bound, "closed" state of OxlT at 6.5 A determined by cryo-electron microscopy with a model of the "open" state of OxlT based on the atomic structures of the related transporters, glycerol-3-phosphate transporter (GlpT) and lactose permease (LacY). We demonstrate that the principal structural change associated with substrate transport is a concerted rocking movement of the two structurally similar halves of the protein relative to each other. Our structural model places two positively charged residues, Arg-272 and Lys-355 in the central cavity, suggesting that electrostatic interactions between these residues and the oxalate anion is a key step in generating the conformational change between the open and closed states of the transporter.

Amino Acid Sequence↗

Effect of diuretics on urinary oxalate, calcium, and sodium excretion in very low birth weight infants.

OBJECTIVE: To study the effect of diuretic drugs on urinary oxalate excretion in premature infants, and to examine the relationship between urinary calcium and sodium excretion in premature infants. METHODOLOGY: We measured urinary oxalate, calcium, and sodium excretion in 32 premature infants at approximately 34 weeks gestational age. Seven infants were receiving furosemide, 5 infants were receiving thiazide, 8 infants were receiving furosemide plus thiazide, and 12 infants who were not receiving diuretics served as controls. RESULTS: Urinary oxalate to creatinine ratios in infants receiving furosemide (0.48 +/- .26), thiazide (0.54 +/- .20), furosemide plus thiazide (0.44 +/- .19), and control infants (0.51 +/- .43) were similar by analysis of variance (ANOVA). Data expressed as oxalate concentration gave similar results. Urinary calcium to creatinine ratios in infants receiving furosemide (0.81 +/- .30), thiazide (0.54 +/- .25), furosemide plus thiazide (0.75 +/- .49), and control infants (0.37 +/- .25) were similar by ANOVA. The urinary calcium concentration in infants receiving furosemide plus thiazide (0.085 +/- 0.042 mg/mL) was different from control infants (0.044 +/- .023) by ANOVA and Student-Newman-Keuls test. Urinary calcium to creatinine ratio was correlated with sodium to creatinine ratio (r = .751). CONCLUSION: Urinary oxalate excretion in premature infants is not affected by diuretic drugs. Urinary sodium and calcium excretion are closely linked in sodium supplemented premature infants receiving diuretics. The calciuric effect of furosemide is not decreased by the addition of thiazide in premature infants receiving sodium supplements.

Analysis of Variance↗

Kinetic spectrophotometric method for the determination of trace amounts of oxalate by an activation effect.

A new, simple and inexpensive kinetic catalytic spectrophotometric method for the determination of oxalate is described. The method is based on an activation effect of oxalate on a catalytic effect of iron(II) on the oxidation of iodide by bromate. The reaction is monitored by measuring the absorbance of triiodide ion at lambda max = 352 nm. A calibration graph was obtained from 0.10 - 7.0 microg cm(-3) of oxalate with a detection limit of 0.080 microg cm(-3). The standard deviations for ten replicate determinations of 0.50, 1.0 and 5.0 microg cm(-3) of oxalate were 4.0, 2.6 and 1.8%, respectively. The applicability of the method was demonstrated by the determination of oxalate ion in real samples.

Bromates↗

Effects of dietary potassium citrate supplementation on urine pH and urinary relative supersaturation of calcium oxalate and struvite in healthy dogs.

OBJECTIVE: To assess the effect of dietary potassium citrate supplementation on the urinary pH, relative supersaturation of calcium oxalate and struvite (defined as the activity product/solubility product of the substance), and concentrations of magnesium, ammonium, phosphate, citrate, calcium, and oxalate in dogs. ANIMALS: 12 healthy adult dogs. PROCEDURE: Canned dog food was fed to dogs for 37 days. Dogs were randomly allocated to 3 groups and fed test diets for a period of 8 days. Study periods were separated by 6-day intervals. During each study period the dogs were fed either standard diet solus (control) or standard diet plus 1 of 2 types of potassium citrate supplements (150 mg potassium citrate/kg of body weight/d) twice daily. Urinary pH, volume and specific gravity, relative supersaturation of calcium oxalate and struvite, and concentrations of magnesium, ammonium, phosphate, calcium, oxalate, and citrate were assessed for each treatment. RESULTS: Mean urine pH was not significantly affected by dietary potassium citrate supplementation, although urine pH did increase by 0.2 pH units with supplementation. Diets containing potassium citrate maintained a higher urine pH for a longer part of the day than control diet. Three Miniature Schnauzers had a significantly lower urinary relative calcium oxalate supersaturation when fed a diet supplemented with potassium citrate, compared with control diet. CONCLUSIONS AND CLINICAL RELEVANCE: Dietary potassium citrate supplementation has limited effects on urinary variables in most healthy dogs, although supplementation results in maintenance of a higher urine pH later in the day. Consequently, if supplementation is introduced, dogs should be fed twice daily and potassium citrate should be given with both meals or with the evening meal only.

Animals↗

How elevated oxalate can promote kidney stone disease: changes at the surface and in the cytosol of renal cells that promote crystal adherence and growth.

The present review assesses the mechanisms by which oxalate-induced alterations in renal cell function may promote stone disease focusing on 1) changes in membrane surface properties that promote the attachment of nascent crystals and 2) changes in the expression/secretion of urinary macromolecules that alter the kinetics of crystal nucleation, agglomeration and growth. The general role of renal cellular injury in promoting these responses and the specific role of urinary oxalate in producing injury is emphasized, and the signaling pathways that lead to the observed changes in cell surface properties and in the viability and growth of renal cells are discussed. Particular attention is paid to evidence linking oxalate-induced activation of cytosolic phospholipase A2 to changes in gene expression and to the activation of a second signaling pathway involving ceramide. The effects of the lipid signals, arachidonic acid, lysophosphatidylcholine and ceramide, on mitochondrial function are considered in some detail since many of the actions of oxalate appear to be secondary to increased production of reactive oxygen molecules within these organelles. Data from these studies and from a variety of other studies in vitro and in vivo were used to construct a model that illustrates possible mechanisms by which an increase in urinary oxalate levels leads to an increase in kidney stone formation. Further studies will be required to assess the validity of various aspects of this proposed model and to determine effective strategies for countering these responses in stone-forming individuals.

Animals↗

N-acetyl-beta-D-glucosaminidase excretion in calcium oxalate stone patients and its relation to the risk of stone formation.

A tubular renal damage induced by crystals in the renal tubuli can be the triggering primary but also secondary cause of stone formation. In 72 calcium oxalate stone patients (45 men, 27 women) N-acetyl-beta-D-glucosaminidase [NAG] excretion in 24h-urine was investigated. 48 healthy test persons (27 men, 21 women) served as a control group and helped to establish reference values. In order to find out a possible relation between NAG reference values and the risks of urolithiasis, lithogenous and inhibitory substances were determined in both groups. In the following, relative calcium oxalate supersaturation, representing a risk of urinary stone formation, was analyzed and compared to NAG excretion values. The threshold value of pathologic NAG excretion was determined in the group of healthy test subjects (mean value +/- 2 standard deviation) and was fixed at 4.2 U/d (women 3.06 U/d, men 4.24 U/d). In our investigations on NAG excretion significantly (p < 0.05) increased values were found in stone patients (4.43 U/d +/- 4.27) as compared to healthy test persons (2.13 U/d +/- 1.02). In the case of increased NAG excretion in stone patients there was a positive correlation between NAG excretion and increased phosphate, sulphate, uric acid, oxalate, and creatinine excretion. Only in female patients there was a relation between relative calcium oxalate supersaturation and NAG excretion values. As a result, NAG determination is considered to be an appropriate means of tracing a certain risk group within calcium oxalate stone patients.

Acetylglucosaminidase↗

Renal injury mediated calcium oxalate nephrolithiasis: role of lipid peroxidation.

The role of lipid peroxidation (LPO) in renal tubular damage mediated calcium oxalate retention was investigated in a rat model. Hyperoxaluria, without deposition of oxalate in kidney, was induced by administration of ethylene glycol (EG), a precursor of oxalate. Oxidative stress condition was produced by administration of buthionine sulfoximine (BSO), an inhibitor of glutathione biosynthesis. BSO-treated rats showed a significant (p < 0.001) increase in LPO over EG-treated rats and it was almost doubled in BSO + EG treated rats. LPO was accompanied by significant urinary excretion of renal damage marker enzymes such as gamma-glutamyl transpeptidase (gamma-GT), alkaline phosphatase (ALP) and cathepsin D, mucoproteins, and glycosaminoglycans (GAGs) in the BSO and BSO + EG groups but not in the EG group. Urinary excretion of gamma-GT (r = +0.90) (p < 0.001) and deposition of oxalate (r = +0.78) (p < 0.001) in kidney positively correlated with LPO. These results suggest that LPO initiates renal damage, thereby leading to calcium oxalate retention and stone formation.

Animals↗

FKBP-12 exhibits an inhibitory activity on calcium oxalate crystal growth in vitro.

Urolithiasis and calcium oxalate crystal deposition diseases are still significant medical problems. In the course of nephrocalcin cDNA cloning, we have identified FKBP-12 as an inhibitory molecule of calcium oxalate crystal growth. lambdagt 11 cDNA libraries were constructed from renal carcinoma tissues and screened for nephrocalcin cDNA clones using anti-nephrocalcin antibody as a probe. Clones expressing recombinant proteins, which appeared to be antigenically cross-reactive to nephrocalcin, were isolated and their DNA sequences and inhibitory activities on the calcium oxalate crystal growth were determined. One of the clone lambda gt 11 #31-1 had a partial fragment (80 bp) of FKBP-12 cDNA as an insert. Therefore, a full-length FKBP-12 cDNA was PCR-cloned from the lambda gt 11 renal carcinoma cDNA library and was subcloned into an expression vector. The resultant recombinant FKBP-12 exhibited an inhibitory activity on the calcium oxalate crystal growth (Kd=10(-7) M). Physiological effect of the extracellular FKBP-12 was investigated in terms of macrophage activation and proinflammatory cytokine gene induction. Extracellular FKBP-12 failed to activate macrophages even at high concentrations. FKBP-12 seems an anti-stone molecule for the oxalate crystal deposition disease and recurrent stone diseases.

Animals↗

[Reduction of urinary oxalate excretion by administration of calcium and citrate].

Oxalic acid seems to be more important for the formation of calcium oxalate stone than calcium. Three grams of calcium lactate and 3 g of uraly U were administered to 35 urolithiasis patients, whose stones were mainly composed of calcium oxalate. Urinary oxalate level was reduced significantly without raising urinary calcium level by the administration of the two drugs for two weeks. The reduction of oxalic acid was particularly remarkable in patients without hypercalciuria. The mechanism of action of these drugs and the relation to dietary management were discussed.

Adult↗

[The effect of glycosaminoglycans on calcium oxalate crystal formation].

The effect of glycosaminoglycans on calcium oxalate crystal formation in the supersaturated solution was studied by examining the size and shape of calcium oxalate crystals generated under an optical microscope. It was found that heparan sulfate and heparin were more effective growth inhibitors than chondroitin sulfate and hyaluronic acid at concentrations within their respective urinary range. With increasing calcium and/or glycosaminoglycans concentration in the solution, the degree of growth inhibition caused by glycosaminoglycans was enhanced. Calcium oxalate crystal shapes generated with various glycosaminoglycans varied with glycosaminoglycan species. One of the causes of those differences in the shape and degree of growth inhibition might be the structural differences between them, that is, the number of sulfate residue and O- or N-form they contain. Calcium oxalate crystal shapes in the presence of heparin or heparan sulfate at higher concentrations were similar to those of calcium oxalate monohydrate crystals in the urinary sediments of hyperoxaluric patients. These facts might suggest the possibility that heparin and/or heparan sulfate were present in the crystal forming region.

Calcium Oxalate↗