Neutrophil rolling altered by inhibition of L-selectin shedding in vitro

THE L-selectin adhesion molecule is involved in guiding leuko-cytes to sites of inflammation 1 . L-selectin is cleaved by an unusual proteolytic activity at a membrane-proximal site resulting in rapid shedding from the cell surface 2–7 . Although it has been demonstrated that L-selectin mediates, in part, the early event of leukocyte rolling under hydrodynamic flow 8–10 , the contribution of shedding to L-selectin function has remained unknown. Here we show that hydroxamic acid-based metalloprotease inhibitors block L-selectin downregulation from the cell surface of stimu-lated neutrophils, without affecting Mac-1 mobilization or gen-eral neutrophil activation, and inhibit cleavage of L-selectin in a cell-free system. Unexpectedly, the hydroxamic acid-based inhi-bitors reduced neutrophil rolling velocity under hydrodynamic flow, resulting in increased neutrophil accumulation. These results suggest that L-selectin is cleaved in seconds—much faster than previously suspected—during the process of rolling under hydrodynamic flow, and that shedding of L-selectin may contribute significantly to the velocity of leukocyte rolling. L-selectin shedding during rolling interactions may be physio-logically important for limiting leukocyte aggregation and accumulation at sites of inflammation.