Regulation of CD18 expression on neutrophils in response to fluid shear stress

When leukocytes adhere to endothelial cells and are exposed to fluid shear stresses, they often retract pseudopods and reduce their attachment. Leukocytes use CD18 for membrane adhesion, but the kinetics of such integrin adhesion molecules under fluid shear is unknown. We examine on neutrophils with confocal microscopy of single adherent cells and flow cytometry of cell suspensions the CD18 expression under fluid shear after labeling with fluorescent antibodies. Fluid shear causes reduction of CD18-associated immunofluorescence of extracellular epitopes, especially in areas of the membrane exposed to elevated levels of shear (1.5 dyne/cm2 maximum shear stress; 1 dyne = 10 mN). CD18 was also translocated over the leukocyte surface from regions of higher shear to lower shear and into the membrane contact areas with the substrate. We obtained no evidence for cytoplasmic internalization of CD18. Fluid shear (5 dyne/cm2) in a suspension of human leukocytes resulted in cleavage of the extracellular domain but not against a cytoplasmic domain of CD18. Chelation of extracellular Ca2+ abolished the down-regulation of CD18. Cysteine protease inhibitors and a selective inhibitor for cathepsin B, but no blockade of other cysteine proteases such as cathepsin L and calpain, aminopeptidases, elastase, or metalloproteinases, suppressed shear-induced CD18 down-regulation. The evidence suggests that physiological levels of fluid shear cause release of cysteine protease(s) including cathepsin B, leading to cleavage of the extracellular domain of CD18 molecules and possible membrane detachment.