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Show DETERMINATION OF SIALIC ACID LOCALIZATION AND LINKAGE ON THE ENDOTHELIAL SURFACE Kristina Giantsos (Randal Dull) Department of Anesthesiology, University of Utah Podocalyxin and endomucin are two sialomucin found on the en- dothelial cell surface of vascular endothelial cells. It is believed that these mucins may play a role in regulating capillary perme-ability via intercellular interactions. The opening and closing of junctions between endothelial cells may be modulated, in part, by charge repulsions induced by the heavy sialic acid content of these mucins. Immunostaining using lectins was used to deter-mine approximate sialic acid content and its distribution on the endothelial cell surface. The lectins Maakia amurensis (MAA) and Sambucus nigra (SNA) were chosen because of their selectiv-ity with respect to sialic acid linkage, as MAA binds sialic acids with 2,3 linkages and SNA binds to those with 2,6 linkage. Bo-vine endothelial cells were previously cultured then stained with both lectins and examined by immunoflurorescence microscopy. Photos obtained after staining show a significant amount of 2,3-linked sialic acids on the surface of endothelial cells with little or no 2,6 linkages. The areas of heaviest sialation were found to be along the periphery of the cells coinciding with the location of the cell junctions. We are developing the methodology to selectively quantify the amount of sialic acids on the endothelial surface us-ing selective enzyme digestion followed by HPLC analysis. In pre-liminary studies, sialomucins were cleaved from the endothelial surface with O-sialoglycoprotein endopeptidase (OSGE) and then subjected to a second digestion with sialidase. Liberated sialic acids were then quantified by cation-exchange HPLC. Total cell surface sialic acid content will be evaluated following complete digestion of the endothelial surface with sialidase alone. These data, when combined with functional assays of endothelial per-meability will aid in developing a structure-function model of si-alomucins and junctional permeability. Kristina Giantsos Biochemistry Faculty Sponsor Randal Dull |