The selectins and the beta 2-integrins (CD11/CD18) mediate distinct adhesive interactions between neutrophils and endothelial cells. Selectins are believed to initiate binding by mediating neutrophil rolling, whereas beta 2-integrins are required for subsequent activation-induced firm sticking and emigration. In vitro evidence suggests that two endothelial cell selectins, P- and E-selectin, can mediate rolling by binding to the carbohydrate ligand sialyl-Lewisx (sLex) on neutrophil surface glycoconjugates. To test the relative contribution of selectins and beta 2-integrins in vivo we used intravital microscopy to study the behavior of neutrophils from two patients with distinct inherited leukocyte adhesion deficiency syndromes. Neutrophils from a patient suffering from CD18 deficiency showed normal rolling behavior but were incapable of sticking or emigrating upon chemotactic stimulation. Neutrophils from a second patient with a newly described adhesion deficiency had normal CD18 but did not express sLex. These neutrophils rolled poorly and also failed to stick in venules under shear force. Under static conditions, however, chemoattractant-induced sticking and emigration could be observed. This demonstrates that both selectin-carbohydrate-mediated initiation of adhesion and subsequent activation-induced beta 2-integrin engagement are essential for the normal function of human neutrophils in vivo.

The glycoprotein (GP) L-selectin initiates adhesive interactions between leukocytes and endothelial cells (EC). It functions as a lymphocyte-lectin homing receptor recognizing carbohydrate determinants of the peripheral lymph node addressing on high endothelial venules. It also mediates neutrophil rolling, the earliest interaction of neutrophils with acutely inflamed venules. Neutrophil L-selectin presents sialyl-LewisX (sLe(X)) as a ligand to P- and E-selectin in vitro, and we have proposed that this is a major mechanism of L-selectin-mediated rolling in vivo. In contrast, the contribution of neutrophil L-selectin as a receptor protein recognizing one (or more) ligand(s) on inflamed EC is unclear. To address this question, an sLe(X)-negative murine pre-B cell line, L1-2, that can neither bind vascular selectins nor roll in inflamed rabbit venules, was transfected with human L-selectin cDNA. L-selectin expression in stable transfectants was sufficient to confer significant rolling in vivo. Rolling was unaffected by neuraminidase treatment but completely blocked by anti-L-selectin monoclonal antibody (MoAb) DREG-56. Thus, L-selectin can initiate leukocyte interactions with EC determinants potentially through recognition of endothelial carbohydrates. In contrast, when human neutrophils were tested, rolling was reduced, but not abolished, by MoAb DREG-56. Likewise, treatment with neuraminidase or anti-sLe(X) MoAbs decreased, but did not abrogate, neutrophil rolling, consistent with residual EC recognition via L-selectin. Combination of MoAb DREG-56 and neuraminidase resulted in almost complete loss of rolling, as did removal of glycosylated L-selectin by chymotrypsin. Together with the demonstrable rolling of L-selectin transfectants, our results support the concept of a bidirectional interaction between L-selectin bearing sLe(X) on neutrophils and activated EC in vivo. These findings also suggest that L-selectin may mediate rolling of lymphocytes that lack carbohydrate ligands for E- or P-selectin, although probably less efficiently than through bidirectional recognition.

Several sequential steps characterized by distinct molecular mechanisms are required for intravascular neutrophil adhesion in acute inflammation under the unique physical conditions found in the microvasculature in vivo. Adhesion is initiated by L-selectin, a constitutively functional glycoprotein on circulating neutrophils which may interact with several ligands on inflamed endothelial cells. L-selectin binding allows margination and rolling of neutrophils in venules. Subsequently, rolling cells may encounter activating or chemotactic stimuli which trigger functional upregulation of beta 2 integrins (CD11/CD18); the integrins mediate firm attachment and are required for diapedesis. This paper will discuss the physiologic importance of L-selectin-mediated rolling and activation-induced beta 2 integrin engagement for neutrophil function in vivo.