Mechanism of collagen network stabilization in human irreversible granulomatous liver fibrosis.

BACKGROUND & AIMS: Cross-linking participates in the increased stability of collagen towards proteolytic degradation. Liver collagen cross-linking by pyridinoline, from the lysyl oxidase pathway, and by pentosidine, issued from glycation, was investigated to determine their respective contribution to collagen stabilization in patients with an irreversible liver fibrosis caused by the parasitic granulomatous disease alveolar echinococcosis. METHODS: Liver pyridinoline and pentosidine were analyzed by high-performance liquid chromatography, and urinary pyridinoline was analyzed by immunoassay. Cross-linked type I collagen was localized by immunohistochemistry with an antibody against the C-terminal part of the molecule, involved in pyridinoline formation, that was measured in serum by radioimmunoassay. RESULTS: In contrast to pyridinoline, pentosidine decreased in fibrotic lesions. Cross-linked I collagen was located predominantly in collagen bundles in the periparasitic granuloma. Serum pentosidine and urinary pyridinoline levels did not differ significantly from controls, but the serum concentration of the C-terminal telopeptide of type I collagen increased significantly. CONCLUSIONS: Lysyl oxidase-mediated cross-linking is the major process contributing to the stabilization of collagen in granulomatous fibrosis, and glycation is not significantly involved in it. The changes induced by alveolar echinococcosis in liver collagen metabolism are associated with an increase in serum C-telopeptide of type I collagen.

Collaborative evaluation in seven laboratories of a standardized micromethod for yeast susceptibility testing.

The new micromethod for yeast susceptibility testing, MYCOTOTAL, was evaluated with 10 reference strains in seven laboratories. Ready-to-use microtitration plates and the same synthetic medium were used with two dilutions of imidazoles, flucytosine, and amphotericin B, permitting the categorization of each strain as susceptible, intermediate, or resistant. The results were compared with the MIC for each reference strain, and the repeatability and reproducibility were evaluated. The yeasts tested presenting different patterns of susceptibilities in reference MICs included six strains of Candida albicans, two strains of Candida tropicalis, one strain of Candida parapsilosis, and one strain of Torulopsis glabrata. For 4,200 antifungal agent-yeast results, the repeatability was 99.3% and the reproducibility was 96.3%. The correlation between the reference MICs and the category results was 91.5% for seven laboratories (and 92.7% for six laboratories excluding the laboratory which did not follow exactly the same protocol). We observed only 7.9% minor discrepancies, 0.5% (0.29% for six laboratories) major discrepancies, and 0.1% uninterpretable results. The percentages of concording results were similar for each strain and each antifungal agent tested. The overall results indicated that MYCOTOTAL was a reliable and reproducible method, well correlated with reference MICs. This ready-to-use micromethod with the same medium for all antifungal agents would be an important step in the necessary standardization of yeast susceptibility testing.

Highly sensitive biotin-avidin sandwich ELISA for the rapid detection of pneumococcal capsular polysaccharide antigens.

The immunological detection of soluble pneumococcal polysaccharide antigens in pathological products is of importance in the direct diagnosis of meningitis or pulmonary infections. We have developed a double antibody sandwich ELISA method using a biotin-avidin system using antibodies constituted with a mixture of IgGs from pooled and/or monospecific antipneumococcal sera provided by the Danish Statens Seruminstitut. The sensitivity of this rapid ELISA method was optimized with purified capsular polysaccharides of the 24 main pneumococcal serotypes. With incubation steps of 30 min at 37 degrees C for the antigens and the conjugates, the detection limit was close to 1 ng/ml for 75% of the purified polysaccharides. A retrospective study of 46 CSF samples established the validity of the assay. This type of modified ELISA system represents a specific, sensitive and rapid procedure for the potential detection of capsular soluble antigens of all pneumococcal serotypes.