Antibodies Against Granule Proteins Activate Neutrophils In Vitro

Abstract

Polymorphonuclear leukocyte (PMN) respiratory burst was stimulated by heterologous antibodies against PMN granule proteins but not by control antibodies. Fluorescence-activated cell sorter (FACS) analysis of activated PMN demonstrated the presence of two primary granule proteins, proteinase 3 (PR-3) and cationic protein 57 (CAP-57) at the membrane surface. The presence of myeloperoxidase (MPO) at the cell surface of primed and unprimed PMN was confirmed by immunoelectron microscopy. Priming doses of recombinant tumor necrosis alpha (rTNF alpha) enhanced the rate of superoxide (O2-) production by these antibodies and increased the amount of surface protein accessible to these antibodies. Anti-neutrophil cytoplasmic autoantibodies (ANCA) with specificities for PMN granule proteins are present in patients with Wegener's granulomatosis, polyarteritis nodosa, and idiopathic and crescentic glomerulonephritis. The demonstration that antibodies against granule proteins activate PMN supports the hypothesis that the vasculitis seen in these diseases is due in part to PMN mediated oxidative injury following PMN stimulation by ANCA.

Anti-neutrophil cytoplasmic antibody (ANCA) in malaria is directed against cathepsin G

“…This may be mediated through ANCA binding to PR3 expressed on the surface of activated endothelial cells, or through impaired clearance of PR3 by its natural inhibitor when bound to its specific autoantibody. ANCA also activate primed neutrophils to produce reactive oxygen species, and to release lysosomal enzymes, both important in the inflammatory process [25].…”

The cell biology of leukocyte-mediated proteolysis

“…Perhaps most importantly, PR3 is the antigen recognized by cytoplasmic type anti-neutrophil cytoplasmic autoantibodies (c-ANCA) in patients with Wegener's granulomatosis, which is characterized by a neutrophilic vasculitis [19,20]. PR3 may contribute to the pathogenesis of this disease by its presence on the surface of neutrophils that are activated in response to the binding of ANCA to membrane-bound PR3 [21,22].…”

Activation of the Gi Heterotrimeric G Protein by ANCA IgG F(ab′)2 Fragments Is Necessary but not Sufficient to Stimulate the Recruitment of Those Downstream Mediators Used by Intact ANCA IgG

“…ANCA IgG activation of TNF-␣-primed neutrophils, unlike conventional Fc␥R cross-linking, does not activate phospholipase D (PLD) or generate phosphatidate or diacylglycerol (10). Unlike conventional Fc␥R cross-linking, stimulation with ANCA IgG did not activate the phosphotyrosine-associated p85/p110 isoform of phosphatidylinositol (PI) 3-kinase, although there was significant production of PIP 3 after ANCA IgG stimulation (10). Both ANCA IgG and conventional Fc␥R cross-linking also activated protein kinase B (PKB/Akt), although the kinetics of the response were different (10).…”

“…Priming of neutrophils with cytokines such as TNF-␣, as is likely to occur in vivo during episodes of infection or inflammatory disease, induces translocation of target antigens (PR3 and MPO) from cytoplasmic granules to the extracellular surface, where they are accessible to autoantibody binding (3)(4).…”