An sLex-deficient variant of HL60 cells exhibits high levels of adhesion to vascular selectins: further evidence that HECA-452 and CSLEX1 monoclonal antibody epitopes are not essential for high avidity binding to vascular selectins.
Selectins are carbohydrate-binding cell adhesion molecules that play a key role in the initiation of inflammatory responses. Several studies have suggested that the sialylated, fucosylated tetrasaccharide sialyl Lewis X (sLex) is an important component of leukocyte ligands for E- and P-selectin. We have identified a stable variant of the HL60 cell line, HL60var, which displays a nearly complete absence of staining with several mAb directed against sLex and/or sLex-related structures. HL60var also exhibits a concomitant increase in reactivity with mAb directed against the unsialylated Lewis X (Lex/CD15) structure. Despite this sLex deficiency, HL60var binds well to both E- and P-selectin. No significant differences in expression of alpha1,3-fucosyltransferases, C2GnT (Core2 transferase), or P-selectin glycoprotein ligand-1 between HL60var and typical sLex(high) HL60 cells were detected. Although the precise molecular basis for the sLex(-/low) phenotype of HL60var remains uncertain, flow cytometric analysis with the sialic acid-specific Limax flavus lectin revealed a sharp reduction in HL60var surface sialylation. Thus, the loss in mAb reactivity may result from a loss of sialic acid residues from the mAb carbohydrate epitope. However, binding of HL60var to E- and P-selectin remains sensitive to neuraminidase treatment. Taken together, these data indicate that high levels of surface sLex and/or related epitopes are not essential for interactions with vascular selectins, implying that as yet unidentified sialylated, fucosylated structures serve as physiologically relevant ligands for E- and P-selectin.
Wagers
A J
AJ
Department of Microbiology-Immunology, Northwestern Medical School, Chicago, IL 60611, USA.
Stoolman
L M
LM
Craig
R
R
Knibbs
R N
RN
Kansas
G S
GS
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D019556:COS Cells; D019040:E-Selectin / Q000378:metabolism*; D000939:Epitopes; D016695:Glycosyltransferases / Q000378:metabolism; D018922:HL-60 Cells; D006801:Humans; D008562:Membrane Glycoproteins / Q000032:analysis; D019158:N-Acetylneuraminic Acid / Q000032:analysis; D009844:Oligosaccharides / Q000032:analysis*; D019007:P-Selectin / Q000378:metabolism*; D000080506:Sialyl Lewis X Antigen
D000911:Antibodies, Monoclonal; D019040:E-Selectin; D000939:Epitopes; D008562:Membrane Glycoproteins; D009844:Oligosaccharides; D019007:P-Selectin; C084882:P-selectin ligand protein; D000080506:Sialyl Lewis X Antigen; D016695:Glycosyltransferases; D019158:N-Acetylneuraminic Acid