P-selectin mediates neutrophil rolling on histamine-stimulated endothelial cells.
In postcapillary venules, marginating neutrophils (PMNs) are often seen rolling along the vessel wall prior to stopping and emigrating. There is substantial evidence in vitro and in vivo that the adhesion receptors E- and L-selectin participate in this phenomenon on cytokine-stimulated endothelium, and recent evidence has shown that a closely related adhesion receptor, P-selectin, is capable of mediating neutrophil rolling on an artificial membrane. Here we demonstrate and characterize PMN rolling on monolayers of human umbilical vein endothelial cells (HUVECs) stimulated with histamine to induce surface expression of P-selectin. Peak association of PMNs with the HUVECs occurs 10 min after histamine stimulation, and at a postcapillary venular wall shear stress of 2.0 dyn/cm2 the rolling velocity is 14 microns/s. Approximately 95% of the PMNs roll on the endothelial cells, 5% adhere firmly, and none migrate beneath the endothelial monolayer. Monoclonal antibody (MAb) G1, which binds P-selectin and blocks its adhesive function, completely prevents association of the PMNs with histamine-stimulated HUVEC, whereas the nonblocking anti-P-selectin MAb S12 does not. Treatment of PMNs with the anti-L-selectin MAb DREG56 reduces PMN adherence by approximately 50%. Anti-CD54 MAb R6.5 and anti-CD18 MAb R15.7 have little effect on the number of PMNs rolling on the HUVECs but completely prevent PMNs from stopping and significantly increase rolling velocity. Nonblocking control MAbs for R6.5 (CL203) and R15.7 (CL18/1D1) lack these effects. Rolling adhesion of PMNs on histamine-stimulated HUVECs therefore appears to be completely dependent on endothelial cell P-selectin, with a minor adhesion-stabilizing contribution from intercellular adhesion molecule 1 and beta 2 integrins. The partial inhibition of rolling with DREG56 suggests that L-selectin may also play a role in neutrophil interactions with histamine-stimulated endothelium. We further characterize these interactions by determining the effects of the various MAbs and wall shear stresses on adhesion patterns, rolling velocities, and distributions of rolling velocities.
Jones
D A
DA
Rice University, Houston, Texas 77251-1892.
Abbassi
O
O
McIntire
L V
LV
McEver
R P
RP
Smith
C W
CW
D000911:Antibodies, Monoclonal; D001667:Binding, Competitive; D001696:Biomechanical Phenomena; D055592:Biophysical Phenomena; D001703:Biophysics; D002448:Cell Adhesion / Q000502:physiology; D015815:Cell Adhesion Molecules / Q000276:immunology / Q000502:physiology*; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D006632:Histamine / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D009504:Neutrophils / Q000502:physiology*; D019007:P-Selectin; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D013314:Stress, Mechanical
D000911:Antibodies, Monoclonal; D015815:Cell Adhesion Molecules; D019007:P-Selectin; D010980:Platelet Membrane Glycoproteins; D006632:Histamine