title: Multivalent sialyl-LeX: potent inhibitors of E-selectin-mediated cell adhesion; reagent for staining activated endothelial cells.
abstract: Free, monovalent, SLeX (Neu5Ac alpha 2-3Gal beta 1-4(Fuc alpha 1-3)-GlcNAc), SLn (Neu5Ac alpha 2-3Gal beta 1-4GlcNAc) and corresponding BSA-conjugated forms--displaying different ratios of SLeX and SLn to protein--were tested for their ability to inhibit binding of HL-60 cells to immobilized E-selectin. Free SLeX and conjugated SLeX-BSA inhibited cell binding in a dose-dependent manner. SLn and SLn-BSA did not inhibit binding. SLeX16BSA (16 mol tetrasaccharide/mol BSA) and monovalent SLeX inhibited cell binding with measured inhibitory concentrations (IC50S) of 1 microM and 1 mM, respectively, demonstrating a three-order-of-magnitude enhancement of inhibitory activity with the multivalent form of SLeX. A SLex7BSA conjugate was 10-fold less potent than those with 11 or 16 mol SLeX/mol BSA. An assay which measured neutrophil rolling on interleukin (IL)-1 beta-activated human umbilical vein endothelial cells (HUVECs) showed 50% reduction in the number of rolling neutrophils in the presence of 1 microM SLeX16BSA, whereas the level of free, monovalent SLeX oligosaccharide required to produce the same effect was approximately 0.3 mM. SLeX-BSA was found to be an excellent reagent for staining endothelial cells expressing E-selectin. Biotinylated SLeX-BSA in conjunction with Texas red avidin-stained lipopolysaccharide (LPS)-activated HUVECs, and co-incubation of activated cells with anti-E-selectin, specifically blocked staining. The distribution of E-selectin, as determined by binding of SLeX-BSA, was virtually identical with that obtained by binding of anti-E-selectin antibody.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Monsanto Corporate Research, Department of Immunology, St Louis, MO 63167.
authors.forename: J K
authors.initials: JK
authors.lastname: Welply
authors.forename: S Z
authors.initials: SZ
authors.lastname: Abbas
authors.forename: P
authors.initials: P
authors.lastname: Scudder
authors.forename: J L
authors.initials: JL
authors.lastname: Keene
authors.forename: K
authors.initials: K
authors.lastname: Broschat
authors.forename: S
authors.initials: S
authors.lastname: Casnocha
authors.forename: C
authors.initials: C
authors.lastname: Gorka
authors.forename: C
authors.initials: C
authors.lastname: Steininger
authors.forename: S C
authors.initials: SC
authors.lastname: Howard
authors.forename: J J
authors.initials: JJ
authors.lastname: Schmuke
mesh_terms: D002448:Cell Adhesion / Q000187:drug effects*; D015815:Cell Adhesion Molecules / Q000502:physiology*; D019040:E-Selectin; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology*; D006801:Humans; D007202:Indicators and Reagents; D007375:Interleukin-1 / Q000494:pharmacology; D015473:Leukemia, Promyelocytic, Acute; D008070:Lipopolysaccharides / Q000494:pharmacology; D009504:Neutrophils / Q000502:physiology; D009844:Oligosaccharides / Q000494:pharmacology*; D000080506:Sialyl Lewis X Antigen; D013194:Staining and Labeling; D014407:Tumor Cells, Cultured; D014471:Umbilical Veins; D014966:Xanthenes
chemical_list: D015815:Cell Adhesion Molecules; D019040:E-Selectin; D007202:Indicators and Reagents; D007375:Interleukin-1; D008070:Lipopolysaccharides; D009844:Oligosaccharides; D000080506:Sialyl Lewis X Antigen; D014966:Xanthenes; C034657:Texas red