SUBCELLULAR-LOCALIZATION OF GLYCOSPHINGOLIPIDS IN HUMAN NEUTROPHILS

The subcellular distribution of five major glycosphingolipids (GSLs) i n human neutrophils was analyzed, The neutrophils were isolated from t he blood of six donors and subdivided in three fractions containing th e cell membranes, and the primary and the secondary granules, respecti vely. The separation was confirmed with antibodies detecting establish ed subcellular fraction-specific molecules, The two main neutral GSLs Gal beta 1-4Glc beta 1-1'Cer (lactosylceramide, LacCer) said nLc(4)Cer (paragloboside, PG) and the three gangliosides IV(3)NeuAcnLc(4)Cer (2 -3SPG), IV(6)NeuAcnLc(4)Cer (2-6SPG), and VI(3)NeuAcnLc(6)Cer (2-3SnHC ) were quantitated using the immunochemical digoxigenin (DIG) staining procedure. Secondary granules contained the highest amount of these G SLs, They are followed by the primary granules and the cell membranes. Based on this quantitation, we conclude that the majority of the GSLs of neutrophils occur intracellularly. These findings are in striking contrast to the general assumption of GSLs being mainly concentrated i n the cell membrane.