HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY Antineutrophil cytoplasm autoantibodies from patients with systemic vasculitis activate neutrophils through distinct signaling cascades: comparison with conventional Fcg receptor ligation

In systemic vasculitis, interactions between antineutrophil cytoplasm autoantibodies (ANCAs) and neutrophils initiate endothelial and vascular injury. ANCAs directed against either myeloperoxidase (MPO) or proteinase 3 (PR3) can activate cytokine-primed neutrophils by binding cell surface–expressed MPO or PR3, with the concurrent engagement of Fcg receptors (FcgR). Because roles for phospholipase D (PLD) and phosphatidylinositol 3 kinase (PI3K) have been demonstrated in FcgR activation of neutrophils, this study investigated the hypothesis that ANCA stimulation of neutrophils involved a similar engagement of FcgR and activation of PLD and PI3K. Pretreatment of tumor necrosis factor (TNF) a-primed neutrophils with antibodies against FcgRII and FcgRIII inhibited MPO-ANCA and PR3ANCA induced superoxide generation, confirming that FcgR ligation is involved in ANCA-mediated neutrophil activation. However, although stimulation of TNF-a– primed neutrophils by conventional FcgR ligation, either using antibody-mediated cross-linking of FcgR or aggregated IgG, induced PLD activation, ANCA stimulation did not. Moreover, although ANCAinduced neutrophil activation results in significant PI3K activation—as assessed by phosphatidylinositol 3,4,5-triphosphate generation—conventional FcgR ligation, but not ANCA, activates the p85/p110 PI3K subtype. Inhibition of ANCA-induced superoxide generation with pertussis toxin suggests that ANCAs activate the p101/ p110g PI3K isoform. In addition, the kinetics of activation of protein kinase B differs between conventional FcgR ligation and ANCA stimulation of neutrophils. These results demonstrate that though ligation of FcgRIIa and FcgRIIIb may be necessary, it is likely that ANCAs require other membrane cofactors for neutrophil activation. (Blood. 2001;98:1448-1455)