Altered conformational activity and cleavage of CD18 integrins on the human neutrophil surface exposed to fluid shear stress

Abstract

CD18 integrins are critical regulators of neutrophil adhesion and migration. Previously, we reported that shear stress inhibits neutrophil pseudopod formation while simultaneously reducing membrane expression of CD18 through cleavage by cathepsin B. The role of CD18 cleavage and bond detachment in shear‐induced pseudopod retraction, however, remains uncertain. To address this, we examined control of CD18 surface expression and shifts in affinity during shear‐induced pseudopod retraction. Using antibodies specific for activated CD18 and a FRET reporter for LFA‐1 (a CD18 integrin), we demonstrated that fluid shear induces shifts in the CD18 ectodomain to an open‐extended conformation associated with ligand binding. Western blots and immunofluorescence revealed cleavage of the extracellular domain of CD18 by cathepsin B in the presence, but not absence, of either shear stress or PMA. Furthermore, cathepsin B was necessary for neutrophil pseudopod retraction in response to shear. Together these results suggest a link between CD18 surface levels and shear‐induced pseudopod retraction involving conformational shifts in the CD18 ectodomain that increases its susceptibility to cathepsin B proteolysis. Moreover, these results point to an anti‐inflammatory role for fluid shear due to blood flow to maintain neutrophils in a rounded, non‐adhesive state under physiological conditions.