Abstract
We studied the inhibitory effect of urinary glycosaminoglycans (GAGs), in the natural form, on calcium oxalate crystals.
Control urine was collected from five healthy subjects and filtered by 5μm and 0.22μm membranes. The urine was cut off by a membrane of 30, 000 M. W. and the remaining urine was fractionated into three groups by anion-exchange chromatography on DEAE-cellulose: peak A contained only protein, peak B both protein and GAGs and peak C only GAGs. Peak A and peak B were fractionated again through a Sephacryl S-200 gel column.
Each fractionated sample was examined for the inhibitory effect on calcium oxalate crystal grwoth using the 14C-oxalate seeded crystal growth assay in a metastable solution developed by Koide et al. GAGs were determined by two-dimensional electrophoresis on cellulose acetate membrane. SDS-polyacrylamide electrophoresis was used to determine the molecular weights of proteins.
Peak A was found to have many kinds of proteins which had low inhibitory effects. Peak B had three kinds of proteins such as those with molecular weights of 130, 000, 45, 000 and 35, 000 as well as uronic acid. These proteins and uronic acid had high inhibitory effect and the highest inhibitory effect was found in the fraction of around 50, 000 M. W. where protein and uronic acid were contained. The molecular weight of this protein was 42, 000 by SDS electrophoresis. Uronic acid was a keratan sulphate by a two-dimensional electrophoresis.
In peak C there were mainly chondroitin sulphate and a small amount of heparan sulfate which had high inhibitory effect.
This study showed that urinary GAGs in the natural form have high inhibitory effects on calcium oxalate crystal growth.
