The effects of inhibiting l-selectin shedding by a metalloprotease inhibitor on leukocyte rolling behavior

The L-selectin adhesion molecule is one of a family of three similar glycoproteins (Selectins). L-selectin is expressed exclusively on leukocytes and is cleaved by an unusual proteolytic activity at a membrane-proximal site resulting in rapid shedding from the cell surface. While it has been demonstrated that L-selectin can mediate leukocyte rolling in vivo along vascular endothelium as well as in vitro on purified ligands, other leukocytes, and activated endothelial cells, the contribution of shedding to L-selectin function has remained unknown. We describe a hydroxamic-based peptide inhibitor of metalloproteases (KD-IX-73-4) that inhibits the shedding of L-selectin from activated leukocytes without affecting general cell activation. Leukocytes treated with KD-IX-73-4 roll at a significantly reduced velocity in vitro under hydrodynamic flow on Lselectin ligands, resulting in increased leukocyte accumulation. KD-IX73-4 does not affect leukocyte rolling on P-selectin. Our results suggest that L-selectin shedding is induced by ligand interaction during the course of leukocyte rolling, and that shedding of L-selectin contributes to the velocity of leukocyte rolling.