STUDIES ON UROLITHIASIS

CRYSTAL AGGREGATION IN CALCIUM OXALATE STONE FORMERS

Abstract

The abnormalities in urine of calcium oxalate stone formers in comparison with control subjects were studied in order to clarify the mechanism of calcium stone formation.
The results were as follows:
1) The daily excretion of calcium, oxalic acid, uric acid and creatinine was determined in fifty-five normal men, fifty-two single stone formers and thirty-seven recurrent stone formers. The calcium excretion was higher in recurrent stone formers (190±21mg/day, mean±SEM) and single stone formers (160±11mg/day) than in normal controls (116±11mg/day). The calcium to creatinine ratio of these groups showed almost similar value. The oxalic acid excretion was higher in recurrent stone formers (34.3±4.0mg/day) than in single stone formers (24.0±2.3mg/day) and controls (24.8±1.6mg/day). The value of calcium (mg/day)×oxalic acid (mg/day) was 6815 in recurrent stone formers, 4689 in single stone formers and 3195 in controls. The value of calcium×oxalic acid/creatinine was also elevated in relation to the severity of the disease. There was no difference of uric acid excretion among the three groups.
2) The particle size distribution of calcium oxalate crystals were measured in fresh morning urine from ten recurrent stone formers and five controls using Coulter counter Model ZBl. The crystal volume excreted by the controls was 0.37-0.05×10⁶μm³/ml (mean±SEM), whereas the volume excreted by the stone formers was 2.21±0.21×10⁶μm³/ml. These values showed significant difference. It was concluded that patients with recurrent calcium oxalate stone disease excreted more crystals of calcium oxalate and the crystals were generally larger and more aggregated than those found in urine of normal subjects. These findings were also obtained with light microscopy. Using this technique calcium oxalate crystalluria may be expressed as either the number or the volume of crystals excreted per unit volume of urine.
3) A simple system was developed to study the effect of inhibitors of crystal aggregation in vitro under controlled conditions in which crystal growth could not be observed. A measure of the aggregation of added calcium oxalate crystals was obtained by calculating the percent increase in the fraction of particles greater than 9μm in diameter after 4hr incubation in metastable solution with respect to calcium oxalate. The aggregation of added crystals was determined by Coulter counter and compared with the control solution. The degree of aggregation was expressed as inhibitory activity.
In this system, urine of either controls or stone formers inhibited aggregation when added at concentration as low as 5%, where inhibitory activity was almost 100%. The inhibitory activity of the urine from ten recurrent stone formers was significantly decreased (38.4%) compared with fifteen controls (60.2%) and ten single stone formers (55.4%).
4) Effects of citrate, magnesium, urea, creatinine, methylene blue, sodium copper chlorophyllin, EDTA, pyrophosphate and glycosaminoglycans on calcium oxalate crystal aggregation were examined. Of the known inhibitors tested, citrate, magnesium, pyrophosphate, glycosaminoglycans and sodium copper chlorophyllin were effective, whereas methylene blue and EDTA had no effect on the aggregation of calcium oxalate crystals in vitro. Urea and creatinine had no effect. Potent inhibition was observed with glycosaminoglycans, especially with heparin.