Gabriele Schumacher

Visualization of sialyl Lewisx glycosphingolipid microdomains in model membranes as selectin recognition motifs using a fluorescence label

Selectin-induced leukocyte rolling along the endothelial surface is an essential step in the cellular immune response. For efficient recognition, the relevant carbohydrate epitope sialyl Lewis(X) (sLe(X); alpha-Neup5Ac-(2-->3)-beta-Galp-(1-->4)-[alpha-Fucp-(1-->3)]GlcpNAc) has to be arranged in clusters. We describe the synthesis of the sLe(X)-glycosphingolipid (sLe(X)-GSL) with a NBD fluorescence label in the tail region, which allows the direct visualization of sLe(X)-GSL microdomains to very low concentrations (0.01mol%) in various planar phosphocholine matrices by fluorescence microscopy. Cell rolling experiments of E-selectin expressing cells along these membranes confirmed that the fluorescence analog behaves similar to the naturally occuring sLe(X)-GSL. This is direct evidence for recent hypotheses on multivalent sLe(X) binding as molecular basis for selectin-mediated cell rolling.