thromboplastin and Anti-Neutrophil-Cytoplasmic-Antibody-Associated-Vasculitis

Patients with antineutrophil cytoplasmic antibody-associated vasculitis (AAV) have a high frequency of venous thromboembolic events and a hypercoagulable state. As C5a-primed neutrophils play an important role in the development of AAV, we investigated whether C5a-induced neutrophil tissue factor (TF)-expressing microparticles (MPs) and neutrophil extracellular traps (NETs) might promote hypercoagulability in AAV.. TF-expressing MPs were measured by flow cytometry. TF-expressing NETs were assessed by confocal microscopy. Levels of thrombin-antithrombin complexes were determined by enzyme-linked immunosorbent assay. The effect of C5a in sera from AAV patients was evaluated by treating neutrophils with C5a receptor antagonist before incubation with sera from AAV patients with active disease.. Treatment of C5a-primed neutrophils with antineutrophil cytoplasmic antibody (ANCA)-positive IgG resulted in the release of TF-bearing MPs and NETs. Neutrophils from healthy donors treated with sera from patients with active AAV released TF-bearing MPs and NETs, which were abolished by treatment with C5a receptor antagonist. Involvement of TF in MP- or NET-dependent thrombin generation was indicated by the findings of antibody neutralization studies. NETs with thrombin-generating capacity were demonstrated by DNase I treatment.. C5a-primed neutrophils produce TF-expressing MPs and NETs after stimulation with ANCAs, indicating a mechanism for hypercoagulability in AAV that was not previously recognized.

Topics: Adult; Aged; Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis; Antibodies, Antineutrophil Cytoplasmic; Case-Control Studies; Cell-Derived Microparticles; Cells, Cultured; Complement C5a; Extracellular Traps; Female; Humans; Male; Microscopy, Confocal; Middle Aged; Neutrophils; Nucleosomes; Thrombin; Thrombophilia; Thromboplastin

Antineutrophil cytoplasmic antibody (ANCA) associated vasculitis (AAV) is characterised by neutrophil activation. An elevated prevalence of venous thromboembolic events has been reported in AAV. Because of the critical role of neutrophils in inflammation associated thrombosis, we asked whether neutrophil tissue factor (TF) may be implicated in the thrombotic diathesis in AAV.. Neutrophils from four patients and sera from 17 patients with ANCA associated vasculitis with active disease and remission were studied. TF expression was assessed by immunoblotting and confocal microscopy. Circulating DNA levels were evaluated. TF expressing microparticles (MPs) were measured by flow cytometry and thrombin-antithrombin complex levels by ELISA.. Peripheral blood neutrophils from four patients with active disease expressed elevated TF levels and released TF expressing neutrophil extracellular traps (NETs) and MPs. TF positive NETs were released by neutrophils isolated from the bronchoalveolar lavage and were detected in nasal and renal biopsy specimens. Elevated levels of circulating DNA and TF expressing neutrophil derived MPs were further observed in sera from patients with active disease. Induction of remission attenuated the aforementioned effects. Control neutrophils treated with sera from patients with active disease released TF bearing NETs and MPs which were abolished after IgG depletion. Treatment of control neutrophils with isolated IgG from sera from patients with active disease also resulted in the release of TF bearing NETs. TF implication in MP dependent thrombin generation was demonstrated by antibody neutralisation studies.. Expression of TF in NETs and neutrophil derived MPs proposes a novel mechanism for the induction of thrombosis and inflammation in active AAV.

Topics: Adult; Aged; Anti-Neutrophil Cytoplasmic Antibody-Associated Vasculitis; Antibodies, Antineutrophil Cytoplasmic; Bronchoalveolar Lavage Fluid; Case-Control Studies; Cell-Derived Microparticles; Extracellular Traps; Female; Humans; Immunoglobulin G; Male; Middle Aged; Neutrophil Activation; Neutrophils; Remission Induction; Thrombophilia; Thromboplastin; Tumor Necrosis Factor-alpha