Anti-Proteinase 3 Anti-Neutrophil Cytoplasm Autoantibodies Recapitulate Systemic Vasculitis in Mice with a Humanized Immune System

Mark A Little,Caroline O Savage,Bahjat Al-Ani,Jeremy S Duffield,Shuyu Ren,Hamad Al-Nuaimi,Maurilo Leite,Charles E Alpers,Carol Feghali-Bostwick

doi:10.1371/journal.pone.0028626

Abstract

Evidence is lacking for direct pathogenicity of human anti-proteinase-3 (PR3) antibodies in development of systemic vasculitis and granulomatosis with polyangiitis (GPA, Wegener's granulomatosis). Progress in study of these antibodies in rodents has been hampered by lack of PR3 expression on murine neutrophils, and by different Fc-receptor affinities for IgG across species. Therefore, we tested whether human anti-PR3 antibodies can induce acute vasculitis in mice with a human immune system. Chimeric mice were generated by injecting human haematopoietic stem cells into irradiated NOD-scid-IL2Rγ−/− mice. Matched chimera mice were treated with human IgG from patients with: anti-PR3 positive renal and lung vasculitis; patients with non-vasculitic renal disease; or healthy controls. Six-days later, 39% of anti-PR3 treated mice had haematuria, compared with none of controls. There was punctate bleeding on the surface of lungs of anti-PR3 treated animals, with histological evidence of vasculitis and haemorrhage. Anti-PR3 treated mice had mild pauci-immune proliferative glomerulonephritis, with infiltration of human and mouse leukocytes. In 3 mice (17%) more severe glomerular injury was present. There were no glomerular changes in controls. Human IgG from patients with anti-PR3 autoantibodies is therefore pathogenic. This model of anti-PR3 antibody-mediated vasculitis may be useful in dissecting mechanisms of microvascular injury.

Highlights

Systemic vasculitis, comprising microscopic polyangiitis and granulomatosis with polyangiitis (GPA, Wegener’s granulomatosis), is a debilitating and frequently life threatening disease affecting the microvasculature, in its most severe form manifesting with acute kidney and lung injury or failure
We assessed the proportion of mCD452 leukocytes with high side scatter (SSC) by flow cytometry compared with total high SSC cells to give a sense of the proportion of circulating granulocytes that were of human origin (Fig. 1A, Table 1) Once the degree of chimerism was established, mice were divided into experimental groups, matched for the degree of chimerism
By comparison no leukocytes were recruited to the livers of these mice. These studies are the first to report a clear pathogenic role in vivo for IgG derived from patients with anti-proteinase 3 (PR3) antibody positive vasculitis

Summary

Introduction

Systemic vasculitis, comprising microscopic polyangiitis and granulomatosis with polyangiitis (GPA, Wegener’s granulomatosis), is a debilitating and frequently life threatening disease affecting the microvasculature, in its most severe form manifesting with acute kidney and lung injury or failure. A major breakthrough was made in the 1980’s when several groups discovered autoantibodies directed at cytoplasmic constituents of neutrophils (Anti-Neutrophil Cytoplasmic Antibodies [ANCA]) in patients affected by small vessel systemic vasculitides [1,2,3,4,5]. These antibodies, which have specificity for either neutrophil myeloperoxidase (MPO) or proteinase 3 (PR3), have been shown to activate primed neutrophils in vitro by binding to cell surface exposed antigens [6,7] and signalling via Fcc receptors, suggesting a direct pathogenic role for the circulating antibodies. Further in vitro studies lent support to a potential role for monocyte-ANCA interaction in vasculitis pathogenesis [8]

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