Increased urinary hyaluronic acid and interstitial cystitis.

PURPOSE: We compared urinary levels of hyaluronic acid in patients who met the National Institute for Diabetes, and Digestive and Kidney Diseases criteria for interstitial cystitis and in age matched healthy female controls. MATERIALS AND METHODS: Urinary hyaluronic acid was measured by solid phase radiometric assay using hyaluronic acid binding protein. Hyaluronic acid and symptom scores were compared in interstitial cystitis patients who gave multiple urine samples during treatment. Since hyaluronic acid changed with treatment in some patients, 17 samples from untreated interstitial cystitis patients were selected and compared with 17 control samples. RESULTS: Mean plus or minus standard deviation urinary hyaluronic acid concentrations were similar in the 2 groups (interstitial cystitis group 574 +/- 496, controls 512 +/- 324 ng./ml., p = 0.77). When normalized to creatinine urinary hyaluronic acid was significantly higher in interstitial cystitis patients (interstitial cystitis group 674 +/- 220, controls 446 +/- 220 ng./mg. creatinine, p = 0.0019). Urinary creatinine concentrations did not differ significantly (interstitial cystitis group 842 +/- 715, controls 1,162 +/- 516 mg./l., p = 0.12). CONCLUSIONS: Urinary hyaluronic acid was higher in interstitial cystitis patients than healthy controls. Since bladder hyaluronic acid is below the epithelium, this finding may indicate leakage across the epithelium into the urine in interstitial cystitis patients.

Differential binding of Pseudomonas aeruginosa to normal and cystic fibrosis tracheobronchial mucins.

Pseudomonas aeruginosa infection is a leading cause of deterioration of pulmonary function in patients with cystic fibrosis (CF). The interaction of the bacterium with CF and non-CF tracheobronchial mucins was examined to understand the biochemical basis for the high susceptibility of the lungs of CF patients to infection by P.aeruginosa. The binding of radiolabelled bacteria to pure mucins in solid-phase assays was not significantly above non-specific binding to various blocking agents, such as bovine serum albumin, Tween 20, milk powder and polyvinyl pyrrolidine. Further, there was a tendency for the bacteria to be excluded from plastic wells and membranes coated with mucin. Therefore, an indirect approach involving the binding of radiolabelled P.aeruginosa to asialo GM1 ganglioside, the putative receptor for the bacteria on tracheal cells, was used to compare the interaction of CF and non-CF mucins with the bacteria. Highly purified preparations of CF mucin were consistently better inhibitors of the binding of the bacteria to asialo GM1 ganglioside than non-CF mucin preparations. In the case of the binding of a stable mucoid strain, the difference was statistically significant (P < 0.001) at all concentrations of mucin tested. For the non-mucoid strain, the difference was significant only at the higher concentrations. Of the saccharides tested similarly, sialyl lactose and the oligosaccharide portion of asialo GM1 were found to be good inhibitors. The increased binding of the bacteria to CF mucin was further confirmed by a solution binding assay in which the binding of 125I-labelled mucin to unlabelled bacteria was determined.(ABSTRACT TRUNCATED AT 250 WORDS)

Purification and characterization of monkey (Macaca nemestrina) tracheobronchial mucin.

A major mucin glycoprotein was purified from monkey (Macaca nemestrina) bronchoalveolar lavages by gel filtration, delipidation, and a series of density gradient centrifugations in cesium trifluoroacetate/guanidinium chloride. Lipids noncovalently associated with the mucin amounted to 24-36% by weight and consisted primarily of phospholipids and glycolipids. The mucin preparation was free of low-molecular-weight protein/glycoprotein contaminants, glycosaminoglycans/proteoglycans, and nucleic acids. The weight-average molecular weight and radius of gyration of the mucin in buffer containing 6 M guanidinium chloride was estimated to be approximately 1.56 x 10(6) and 100 nm, respectively, by laser light scattering technique. When the mucin was dissolved in 0.15 M NaCl, a considerably higher molecular weight of approximately 5.05 x 10(6) and a larger radius of gyration of approximately 127 nm were observed suggesting aggregation of the mucin molecules. Amino acid composition of the glycoprotein was characteristic of mucins with threonine, serine, glutamic acid, proline, glycine, and alanine comprising 63%. The total carbohydrate content was 71.5% and consisted of GalNAc, GlcNAc, Gal, sialic acids, and fucose in the molar ratio of 1.0:2.2:2.4:1.4:1.2 with no detectable mannose. Alkaline borohydride treatment indicated that 65% of the threonine and 27% of the serine are substituted by saccharides via GalNAc residues. An antisera produced against the purified mucin was found to react well with the native and weakly with the deglycosylated mucins and will be useful for immunoassays. A second, minor, mucin glycoprotein obtained during the purification was also partially characterized.

Colorimetric determination of N-acetylhexosamine-terminating O-glycosidically linked saccharides in mucins and glycoproteins.

A sensitive colorimetric assay for detecting mucins and glycoproteins rich in O-glycosidically linked saccharides is reported. The method combines the susceptibility of N-acetylgalactosamine terminating O-glycosidically linked saccharides to beta-elimination with the Morgan-Elson reaction for N-acetylhexosamines with free reducing ends. All mucin and mucin-type glycoproteins but none of the serum-type glycoproteins tested resulted in characteristic color production. All mucins tested gave linear responses in the range 5 to 200 micrograms and the assay was also adapted to the microscale involving the use of 96-well microtiter plates. The microassay in which the volumes of samples and reagents are scaled down 2.5-fold was particularly useful in monitoring of mucins, in the presence of other glycoconjugates, in large numbers of samples obtained during fractionation procedures. Cesium chloride, cesium bromide, potassium thiocyanate, and various detergents do not interfere with the colorimetric determination. Guanidine hydrochloride, cesium trifluoroacetate, and beta-mercaptoethanol decreased color by 30 to 45%; however, the interference was not serious to prevent the use of the method for detection of mucins in their presence. The use of the method for the specific detection of mucin during fractionation by gel filtration and density gradient centrifugation of cystic fibrosis sputum samples is demonstrated.