Selectin adhesion molecules are principally responsible for the initial rolling-adhesion interaction between circulating leukocytes and vascular endothelium. Rolling-adhesion is a necessary prelude to the leukocyte immobilization, transendothelia migration, and tissue accumulation that occurs as part of normal inflammation, as well as in pathological inflammation that causes tissue injury in autoimmune diseases. Endothelial E-, and P- Selectin, and leukocyte L-Selectin are a gene family of 3 closely related molecules, whose amino-terminal lectin domain is principally responsible for binding to fucosylaated, sialylated, and sulfated oligosacchardide counter-receptors expressed on leukocytes and endothelial cells respectively. Leukocyte L-Selectin was originally described as a lymphocyte homing receptor for lymph nodes. However L-Selectin is expressed on all leukocytes, including neutrophils, and it is clear that L-Selectin is required for expression of a normal inflammatory response in non-lymphoid tissues. This implies that endothelial cell ligands for L-Selectin are expressed in acute inflammation, however, none have been identified. In the first specific aims we propose to complete the characterization and identification of a novel L-Selectin ligand expressed by rabbit (non-lymphoid) aortic endothelial cells. Preliminary experiments have indicated a single molecular species of molecular weight 240,000 that is not functionally dependent on sialylation, but is however, dependent on sulfation. The molecule does not have the characteristics of a glycosaminoglycan, indicating that it is likely to be an O-linked mucin related to other L-Selectin ligands. The molecule is expressed on the apical surface of the endothelial cells, indicating its relevance to leukocyte adhesion. In the second specific aim we propose to utilize the mono-specific binding assay we have developed to assess L-Selectin binding to native ligands, to discover compounds, whose structure is based on the structure of known selectin ligands, that have potential to block L-Selectin-dependent leukocyte-endothelial adhesion.
