α-1 Antitrypsin regulates human neutrophil chemotaxis induced by soluble immune complexes and IL-8
David A. Bergin, … , Shane J. O’Neill, Noel G. McElvaney
David A. Bergin, … , Shane J. O’Neill, Noel G. McElvaney
Published November 8, 2010
Citation Information: J Clin Invest. 2010;120(12):4236-4250. https://doi.org/10.1172/JCI41196.
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α-1 Antitrypsin regulates human neutrophil chemotaxis induced by soluble immune complexes and IL-8

Abstract

Hereditary deficiency of the protein α-1 antitrypsin (AAT) causes a chronic lung disease in humans that is characterized by excessive mobilization of neutrophils into the lung. However, the reason for the increased neutrophil burden has not been fully elucidated. In this study we have demonstrated using human neutrophils that serum AAT coordinates both CXCR1- and soluble immune complex (sIC) receptor–mediated chemotaxis by divergent pathways. We demonstrated that glycosylated AAT can bind to IL-8 (a ligand for CXCR1) and that AAT–IL-8 complex formation prevented IL-8 interaction with CXCR1. Second, AAT modulated neutrophil chemotaxis in response to sIC by controlling membrane expression of the glycosylphosphatidylinositol-anchored (GPI-anchored) Fc receptor FcγRIIIb. This process was mediated through inhibition of ADAM-17 enzymatic activity. Neutrophils isolated from clinically stable AAT-deficient patients were characterized by low membrane expression of FcγRIIIb and increased chemotaxis in response to IL-8 and sIC. Treatment of AAT-deficient individuals with AAT augmentation therapy resulted in increased AAT binding to IL-8, increased AAT binding to the neutrophil membrane, decreased FcγRIIIb release from the neutrophil membrane, and normalization of chemotaxis. These results provide new insight into the mechanism underlying the effect of AAT augmentation therapy in the pulmonary disease associated with AAT deficiency.

Authors

David A. Bergin, Emer P. Reeves, Paula Meleady, Michael Henry, Oliver J. McElvaney, Tomás P. Carroll, Claire Condron, Sanjay H. Chotirmall, Martin Clynes, Shane J. O’Neill, Noel G. McElvaney

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Figure 4

Release of biologically active AAT from TNF-α– or IL-8–treated MM neutrophils.

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Release of biologically active AAT from TNF-α– or IL-8–treated MM neutro...
(A) Membrane expression of L-selectin or AAT on neutrophils at baseline (green) or in response to TNF-α (red). (B) Western blot showing extracellular released AAT (52 kDa) following treatment with TNF-α or IL-8 in a dose- (B) and time-dependent manner (C). Serum AAT (52 kDa) was loaded as a positive antibody control, and untreated cells are in lanes labeled as control (Con). (D) Time course analysis of the release of AAT (*P < 0.05). (E) Gel filtration chromatography of neutrophil-released AAT. The start material (St) was released AAT from IL-8–treated neutrophils (10 ng). Inset: Western blot analysis illustrating altered migration of purified serum AAT (arrow) compared with neutrophil-released AAT (arrowheads). By Western blot analysis, fraction 28 contained neutrophil-released AAT and chromatographed at a molecular mass of 100–110 kDa. Fractions 30–33 were run on separate gels. (F) Formation of AAT-protease complexes. Samples were immunoblotted employing a rabbit anti-human AAT antibody. Lanes 1 and 3 show serum purified AAT and AAT released from neutrophils, respectively. Lanes 2 and 4 illustrate serum and neutrophil-released AAT incubated with NE. FACS analysis illustrated in A is a representative result of 3 independent experiments. B, C, and F are each representative Western blots of 3 separate experiments. Results in D represent experiments performed in triplicate. Each measurement is the mean ± SEM. FPLC analyses (E) were performed in triplicate on 2 consecutive days.