Idiopathic calcium nephrolithiasis. 1. Differences in urine crystalloids,urine saturation with brushite and urine inhibitors of calcification between persons with and persons without recurrent kidney stone formation.

The propensity of urine to promote calcium stone formation was compared in 64 patients with recurrent idiopathic calcium nephrolithiasis and 30 healthy individuals without such a history. The rates of excretion of urine crystalloids, the urine saturation with brushite (CaHPO4-2H2O), the ability of the urine to calcify collagen in vitro, and the concentration of urine inhibitors of collagen calcification were measured. The patients had a reduced urine citrate excretion rate in addition to an increased urine calcium excretion rate, while the rates for urine magnesium, phosphate, uric acid and oxalate were not significantly different in the two groups of subjects. The urine concentration of magnesium, phosphate and uric acid was decreased in the patients because of the higher urine volume. The urine creatinine excretion rate correlated with the rates of excretion of urine calcium, magnesium, phosphate, uric acid and oxalate in both groups, which suggested that increased lean body mass, possibly associated with greater food intake, may be an important determinant of crystalloid excretion. The urine of the patients was significantly more saturated with brushite than the urine of the control subjects and resulted in greater collagen calcification when incubated in vitro. The urine concentration of inhibitors of collagen calcification, however, was not significantly different in the two groups. Thus, the urine of patients with recurrent idiopathic calcium nephrolithiasis is more highly saturated with brushite, largely as a result of an increased urine calcium excretion rate, and contains a lower concentration of magnesium and citrate, substances that tend to prevent the precipitation and growth of crystals in urine.     

The effect of seed crystals of hydroxyapatite and brushite on the crystallization of calcium oxalate in undiluted human urine in vitro: implications for urinary stone pathogenesis

BACKGROUND: The aim of this study was to determine whether crystals of hydroxyapatite (HA) or brushite (BR) formed in urine promote the epitaxial deposition of calcium oxalate (CaOx) from undiluted human urine in vitro and thereby explain the occurrence of phosphate in the core of urinary stones consisting predominantly of CaOx. MATERIALS AND METHODS: Crystals of HA, BR, and CaOx were generated from human urine and their identity confirmed by X-ray analysis. Standard quantities of each crystal were then added to separate aliquots of pooled undiluted human urine and CaOx crystallization was induced by the addition of identical loads of sodium oxalate. Crystallization was monitored by Coulter Counter and (14) C-oxalate analysis and the precipitated crystals were examined by scanning electron microscopy. RESULTS: In comparison with the control to which no seeds were added, addition of CaOx crystals increased the deposition of (14) C-oxalate by 23%. On the other hand, seeds of HA and BR had no effect. These findings were supported by Coulter Counter analysis, which showed that the average modal sizes of crystal particles precipitated in the presence of HA and BR seeds were indistinguishable from those in the control, whereas those deposited in the presence of CaOx were significantly larger. Scanning electron microscopy confirmed these results, demonstrating that large aggregates of CaOx dihydrates were formed in the presence of CaOx seeds, whereas BR and to a lesser extent HA seeds were scattered free on the filtration membrane and attached like barnacles on the surface of the freshly precipitated CaOx crystals. CONCLUSION: Seed crystals of HA or BR do not promote CaOx deposition in urine in vitro and are therefore unlikely to influence CaOx crystal formation under physiologic conditions. However, binding of HA and BR crystals to, and their subsequent enclosure within, actively growing CaOx crystals might occur in vivo, thereby explaining the occurrence of mixed oxalate/phosphate stones.     

Excretion of inhibitors of calcification in urine Part II. Findings in patients with chronic renal failure.

By means of a semiquantitative method incorporating the rachitic rat cartilage technique, the total urinary inhibitory activity with respect to calcification was compared in 11 control subjects and 20 patients with renal failure. The patients had significantly lower mean values of inhibiting units per day than did the control subjects. Both groups showed a significant positive correlation between the number of inhibiting units per day and urine volume. When urine volume was taken into account in the comparison, the numbers of inhibiting units for patients continued to be lower than the numbers for controls. These findings are consistent with the hypothesis that the increase of inhibitory activity observed in uremic serum is secondary to a decrease in excretion of the responsible factor (or factors) in the urine, and that the factor (or factors) in serum responsible for the inhibition are identical to those in the urine.     

Interference with thyroid histogenesis by inhibitors of collagen synthesis

Histogenesis of thyroid follicles in the chick embryo begins with a penetration by cells of the mesenchymal capsule into a solid epithelial primordium. Before penetration occurs, slits containing fibrillar material form between the epithelial cells. The fibrillar material is an epithelial cell product as shown by its formation within channels that form in cultures of isolated epithelial primordia. The drugs L- azetidine-2-carboxylic acid (LACA) and alpha, alpha'-dipyridyl, which interfere with collagen synthesis, prevent the formation of fibrils in cultured epithelial primordia and in cultures of whole thyroids. Furthermore, mesenchymal cells do not invade when whole thyroid primordia are cultured in the presence of either drug. The effects of alpha, alpha'-dipyridyl are reversed by washing out the drug; the effects of LACA are reversed by incubation with equimolar or greater amounts of L-proline added to the medium along with the drug. The results are interpreted to mean that the fibrillar material is collagen of epithelial origin, that the collagen in some way plays a role in mesenchymal penetration of the epithelial primordium, and that the epithelium is responsible for the pattern of lobulation within the developing gland.     

Excretion of inhibitors of calcification in urine. Part I. Findings in control subjects and patients with renal stones.

The total excretion of inhibitors of in vitro calcification was measured (in inhibiting units per day) in 24-hour urine samples of 11 control subjects and 20 patients with renal calculi. A semiquantitative method incorporating the rachitic rat cartilage technique was used. In both groups there was a significant positive correlation between the number of inhibiting units per day and the daily urine volume. The mean number of inhibiting units per day was significantly (P smaller than 0.05) higher in the stone patients than in the controls. However, the stone-formers had significantly larger (P smaller than 0.01) 24-hour urine volumes. When corrections were made for urine volume there was no significant difference between the two groups. These data suggest that the underlying abnormality responsible for renal stone formation is not a persistent decrease in the total concentration of urinary inhibitors of calcification.     

Proteolipid and human aorta calcification, in vitro

The objective of this study was to determine whether in vitro calcification of human aorta is proteolipid dependent. Homogenates were prepared from tissue with no gross pathologic manifestations. Samples were examined for calcifiability in a metastable calcium phosphate solution before and after lipid extraction. Fractions of the extracted lipid were similarly examined. The tissue calcified before but not after lipid extraction. Calcifiability was restricted to the proteolipid portion of the lipid extract. Under the conditions employed, therefore, proteolipid is required for calcification of human aorta, in vitro.     

Effect of nifedipine on kidney output of calcium, oxalic acid, and saturation of urinary calcium oxalate]

The study involved 30 patients treated with nifedipine in daily dose of 30 mg for 7 days. Calcium, magnesium, phosphate, oxalate, and uric acid levels in the urine were measured. It was found that nifedipine significantly decreased oxaluria urinary excretion of calcium, magnesium, phosphate, and uric acid remained unchanged following nifedipine therapy. Results may suggest that nifedipine may exert an influence on renal stone formation.     

Effect of some alcohols on in vivo and in vitro aged collagen

1. The effects of mixtures of ethanol or n-propanol with 0.15 M NaCl or 0.1 N HCl on the isometric melting and load/extension behaviour of rat tail tendon were examined. 2. Both intra and intermolecular effects were observed but aging of the sample had little effect upon these. 3. The extension modulus in 0.15 M NaCl begins to decrease rapidly at temperatures above the molecular melting temperature (TD). This reduction in modulus is accelerated by alcohol, although the alcohol is not effective below TD.     

Prostaglandin (PG) analysis in urine of humans and rats by different radioimmunoassays: Effect on PG-excretion by PG-synthetase inhibitors, laparotomy and furosemide

Thw radioimmunological (RIA) determination of prostaglandin (PG) E₂ and of PGF_2α in urine humans and rats is described in detail. After extraction and chromatography PGE₂ was determined by using a PGE specific antibody or by using either PGB or PGF_2α specific antibodies after the respective conversion procedures. The three different RIA procedures were compared to each other. PGF_2α was determined by a specific antibody to PGF_2α. Basal excretion of PGE₂ and of PGF_2α in healthy women on free diet was 9.3 ng/hour ± 0.96 and 18.3 ng/hour ± 2.5 respectively. Furosemide increased the excretion of PGE₂ and of PGF_2α in humans significantly, while PG-excretion rates decreased on indomethacin. In rat urine PGE₂ and PGE_2α increased markedly from 46.2 pg/min ± 9.3 and 27 ± 3.4 to 253.8 ± 43.3 and 108 ± 12.6 pg/min (per one kidney) in the anesthetized-laparotomized animal. This increase was abolished after giving two different PG synthetase inhibitors.     

Immunological evaluation of urinary trypsin inhibitors in blood and urine: Role of N- & O-linked glycoproteins

Urinary trypsin inhibitors (uTi) suppress serine proteases during inflammation. After liberation from proinhibitors (P-alpha-I and I-alpha-I) by the white blood cell (WBC) response, uTi readily pass through the kidneys into urine. A key uTi, bikunin, is attached to O-linked and N-linked glycoconjugates. Recently, uTi inhibitors, called uristatins, were found to lack the O-linked glycoconjugates. Monoclonal antibodies were produced using purified uristatin and screened for binding differences to uristatin, bikunin, P-α-I, and I-α-I. Antibody-binding patterns were characterized using immunoaffinity binding onto protein-chip surfaces and analysis by Surface Enhanced Laser Desorption/Ionization mass spectrometry (SELDI), using specimens from patients and from purified uTi standards. Antibodies were developed and used in an enzyme-linked immunosorbent assay (ELISA) method for uTi measurement in urine and plasma specimens. ELISA was performed on specimens from normal, presumed healthy, controls and from patients who had been screened for inflammation using a high sensitivity C-reactive protein (CRP) test and a complete blood count (CBC). Polyclonal antibody against uTi showed cross-reactivity with the Tamm–Horsfall protein (THP) and with proinhibitors. Screening of anti-uTi monoclonal antibodies (Mab) revealed antibodies that did not cross-react with either of the above, thus providing a tool to measure both uristatin and bikunin in urine with Mab 3G5 and in plasma with Mab 5D11. The monoclonal antibody 5D11 cross-reacts with specific N-linked glycoconjugates of uristatin present in plasma. In ca 96% of healthy adults, uTi were present at <12 mg/l in urine and <4 mg/l in plasma. We also found that patients with an inflammation and a CRP of >2.0 mg/l had higher urinary concentrations of uTi than the control population in every subject. Free uristatin and bikunin pass readily into urine and are primarily bound to heavy chains that constitute the proinhibitor form in plasma.