PL
Przygotowano lipopolisacharydy jednego nieenterotoksynotwórczego (NTBF) i trzech enterotoksynotwórczych (ETBF) szczepów B. fragilis oraz enterotoksynę B. fragilis. Badano wpływ polimyksyny B na stymulację ekspresji cząsteczek adhezyjnych: selektyny E, ICAM-1 i VCAM-1 na powierzchni komórek ludzkiego śródbłonka naczyniowego przez przygotowane preparaty bakteryjne. Do oznaczenia cząsteczek adhezyjnych zastosowano test ELISA z przeciwciałami monoklonalnymi. Stwierdzono, że polimyksyna B zmienia poziom ekspresji selektyny E, ICAM-1 i VCAM-1 podczas jednoczesnego działania z lipopolisacharydami lub enterotoksyną B. fragilis.
EN
The aim of this study was to examine the influence of polymyxin B on the level of expression of adhesion molecules E-selectin, ICAM-1, and VCAM-1 on human vascular endothelium activated with B. fragilis endotoxins or enterotoxin. Lipopolysaccharides were extracted by phenol-water method from one nonenterotoxigenic (NTBF) and three enterotoxigenic (ETBF) B. fragilis strains. LPS preparations were purified with nucleolytic enzymes and ultracentrifugation. Enterotoxin (BFT) was prepared from the supernatant of reference B. fragilis ATCC 43858 culture by precipitation with ammonium sulphate. BFT preparations were purified with the application of ion-exchange chromatography and hydrophobic chromatography. Adhesion molecule expression on the surface of human vascular endothelial cells (HMEC-1 cell line) was determined after simultaneous stimulation with bacterial compounds at the concetration of 10 μg/ml and polymyxin B at the concentration of 20 μg/ml. Endothelial cells were activated for 4 hours (E-selectin expression) or for 24 hours (ICAM-1 and VCAM-1 expression). Adhesion molecules were detected in immunoenzymatic test (ELISA) with the use of mouse, monoclonal antibodies against human ICAM-1, VCAM-1, and E-selectin. The results of performed experiments suggest, that polymyxin B changes the level of adhesion molecule expression on human vascular endothelium. This antibiotic causes changes in the expression of endothelial ICAM-1, VCAM-1, and E-selectin during simultaneous stimulationof endothelium with B. fragilis endotoxins or enterotoxin. In the majority of cases the addition of polymyxin B leads to the up-regulation of examined adhesion molecules.