Detection of ligands for selectins in the oligosaccharide fraction of human milk

Summary

Background Human milk contains a large variety of oligosaccharides which show structural similarities with ligands for selectins, a family of cell adhesion molecules which are involved in many cell-cell interactions. Aim of the study Due to their structural similarity with selectin ligands, human milk oligosaccharides were labelled with phosphatidyl ethanolamine to be able to investigate specific effects of antibodies against carbohydrate epitopes. Methods Various monoclonal antibodies against physiological selectin ligands were used to determine whether epitopes within human milk oligosaccharides are recognized. Oligosaccharides were isolated from human milk, transferred into neoglycolipids and separated using high performance thin layer chromatography prior to incubation with monoclonal antibodies for the selectin ligands sialyl-Lewis a (sLe a), sialyl-Lewis x (sLe x), Lewis x (Le x) and Lewis y (Le y) after Western blotting. Fast atom bombardment-mass spectrometry was then used to identify antibody-binding compounds. Results In the immunoassays all ligand epitopes except for Le y were detected in the oligosaccharide fraction of human milk. Anti-sLe a showed the most distinct reaction with N-acetylneuraminic acid containing neoglycolipids of which two were identified as neuraminyl-fucosyl-lacto-N-hexaose and neuraminyl-lacto-N-tetraose. Such oligosaccharides as well as similar structures are present in relatively high concentrations in human milk. Conclusions The presence of sialyl-Lewis ligands on milk oligosaccharides together with their abundancy in human milk may suggest that they could be selectin ligands and they may be part of inflammatory processes.