Involvement of cell surface glycans in adhesion of human colon carcinoma cells to liver tissue in a frozen section assay: Role of endo-beta-galactosidase-sensitive structures

Adhesion of human colon carcinoma variant cell lines expressing different l evels of the cell surface sialyl Lewis X (sLe(x)) antigen to frozen section s of mouse liver was examined, KM12-HX cells that bound the monoclonal anti body (mAb) FH6 (anti-sLe(x)) and thus expressed a high level of sLe(x) demo nstrated a greater degree of adhesion to liver sections than their low-bind ing counterparts, KM12-LX cells, The adhesion of KM12-HX cells to liver sec tions was partially blacked by mAb FH6, but not by another anti-sLe(x) mAb, KM93. The adhesion was Ca2+ dependent but was not inhibited by anti-E-sele ctin. Endo-beta-galactosidase treatment significantly reduced adhesion and resulted in the loss of cell surface binding sites for mAb FH6. O-linked ol igosaccharides from KM12-HX cells incubated in the presence of p-nitropheny l-N-acetylgalactosaminide were fractionated by a combination of gel filtrat ion, anion exchange chromatography, and normal phase high-performance liqui d chromatography, The structure of a mAb FH6-reactive and endo-beta-galacto sidase-sensitive glycan was estimated by matrix-assisted laser desorption i onization time-of-flight mass spectrometry in a post source decay mode and by glycosidase digestions to be NeuAc alpha 2-3Gal beta 1-4GlcNAc beta 1-3G al beta 1-4Glc-NAc beta 1-3Gal beta 1-4(+/-Fuc alpha 1-3)GlcNAc beta 1-6(Ne uAc alpha 2-3Gal beta 1-3)GalNAc-pNP. Mild detergent lysates of mouse liver surface-labeled with sulfo-NHS biotin were incubated with glutaraldehyde-f ixed monolayers of I(KM12-HX cells, and hound components were isolated afte r EDTA treatment. A M-r 49,000 component that bound only to KM12-HX cells a nd not to KM12-LX cells was identified.