Transloation of slexon the azurophilic granule membrane to the plasma membrane in activated human neutrophils

: Using immunogold electron microscopy, we found that human neutrophilic sialyl Lewis x (sLe(x)), an adhesive ligand for selectins, detectable by a monoclonal antibody, KM-93, is present in the sacculi of the Golgi apparatus as well as on the membranes of large electron-lucent azurophilic granules and the plasma membrane, including surface projections and microvilli. Neutrophilic sLe(x), however, was not detected on the membranes of specific granules. In comparison with the distribution of sLe(x), CD18 was localized on the plasma membrane and specific granule membrane but not on the azurophilic granule membrane. We also found by immunogold electron microscopy and flow cytometry that treatment of neutrophils with sialidase resulted in a loss of sLex on the plasma membrane. In contrast, intracellular sLex on the azurophilic granule membrane was not destroyed by sialidase. When sialidase-treated neutrophils were stimulated with N-formyl-methionyl-leucyl-phenylalanine (fMLP), an inflammatory mediator peptide, in the presence of cytochalasin B, we observed by immunogold electron microscopy and flow cytometry that sLe(x) again appeared on the plasma membrane. These results indicate that stimulation by fMLP induces the up-regulation of sLe(x) on the cell surface by promoting translocation of sLe(x) from the azurophilic granule membrane to the plasma membrane in human neutrophils.