Conformational activation of CD11b without shedding of L-selectin on circulating human neutrophils

Abstract

Membrane-activated complex 1 (Mac-1; CD11b/CD18) is a β2 integrin implicated in the pathophysiology of neutrophil-mediated tissue injury whose functional capacity is determined by stimulus-induced conformational activation rather than upregulation. Mac-1 up-regulation and conformational activation, together with shedding of L-selectin, are reported after in vitro neutrophil activation. However, their regulation on circulating human neutrophils during acute inflammation is unclear. Using flow cytometry, we investigated neutrophil expression of Mac-1, its activation-reporter neo-epitope CBRM1/5, and L-selectin during the inflammatory stimulus of cardiac surgery. A subpopulation of circulating neutrophils expressed CBRM1/5 (CBRM1/ 5+) under basal conditions (6.28±2.59%) and was persistently expanded (9.95±4.0%-15.2±4.2%; P<0.0001) peri-operatively, whereas total CD11b expression increased only transiently, intra-operatively. L-selectin expression was unchanged on CBRM1/5+ neutrophils, and soluble L-selectin levels decreased intra-operatively (P<0.01), indicating that L-selectin was not shed. Increased CBRM1/5 expression without L-selectin loss or CD11b up-regulation was replicated in vitro by neutrophil stimulation with IL-8, C3a, and platelet-activating factor. Heparin, a known CD11b ligand, which is administered during cardiac surgery, markedly reduced neutrophil expression of conformationally active CD11b in vivo and in vitro, identifying a potential mechanism for its anti-inflammatory properties. We conclude that conformational activation of CD11b occurs on circulating neutrophils in vivo and can occur in the absence of CD11b up-regulation and L-selectin shedding.11 page(s