Proteinase 3 on apoptotic cells disrupts immune silencing in autoimmune vasculitis
Arnaud Millet, Katherine R. Martin, Francis Bonnefoy, Philippe Saas, Julie Mocek, Manal Alkan, Benjamin Terrier, Anja Kerstein, Nicola Tamassia, Senthil Kumaran Satyanarayanan, Amiram Ariel, Jean-Antoine Ribeil, Loïc Guillevin, Marco A. Cassatella, Antje Mueller, Nathalie Thieblemont, Peter Lamprecht, Luc Mouthon, Sylvain Perruche, Véronique Witko-Sarsat
Arnaud Millet, Katherine R. Martin, Francis Bonnefoy, Philippe Saas, Julie Mocek, Manal Alkan, Benjamin Terrier, Anja Kerstein, Nicola Tamassia, Senthil Kumaran Satyanarayanan, Amiram Ariel, Jean-Antoine Ribeil, Loïc Guillevin, Marco A. Cassatella, Antje Mueller, Nathalie Thieblemont, Peter Lamprecht, Luc Mouthon, Sylvain Perruche, Véronique Witko-Sarsat
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Research Article Immunology

Proteinase 3 on apoptotic cells disrupts immune silencing in autoimmune vasculitis

Abstract

Granulomatosis with polyangiitis (GPA) is a systemic necrotizing vasculitis that is associated with granulomatous inflammation and the presence of anti-neutrophil cytoplasmic antibodies (ANCAs) directed against proteinase 3 (PR3). We previously determined that PR3 on the surface of apoptotic neutrophils interferes with induction of antiinflammatory mechanisms following phagocytosis of these cells by macrophages. Here, we demonstrate that enzymatically active membrane-associated PR3 on apoptotic cells triggered secretion of inflammatory cytokines, including granulocyte CSF (G-CSF) and chemokines. This response required the IL-1R1/MyD88 signaling pathway and was dependent on the synthesis of NO, as macrophages from animals lacking these pathways did not exhibit a PR3-associated proinflammatory response. The PR3-induced microenvironment facilitated recruitment of inflammatory cells, such as macrophages, plasmacytoid DCs (pDCs), and neutrophils, which were observed in close proximity within granulomatous lesions in the lungs of GPA patients. In different murine models of apoptotic cell injection, the PR3-induced microenvironment instructed pDC-driven Th9/Th2 cell generation. Concomitant injection of anti-PR3 ANCAs with PR3-expressing apoptotic cells induced a Th17 response, revealing a GPA-specific mechanism of immune polarization. Accordingly, circulating CD4+ T cells from GPA patients had a skewed distribution of Th9/Th2/Th17. These results reveal that PR3 disrupts immune silencing associated with clearance of apoptotic neutrophils and provide insight into how PR3 and PR3-targeting ANCAs promote GPA pathophysiology.

Authors

Arnaud Millet, Katherine R. Martin, Francis Bonnefoy, Philippe Saas, Julie Mocek, Manal Alkan, Benjamin Terrier, Anja Kerstein, Nicola Tamassia, Senthil Kumaran Satyanarayanan, Amiram Ariel, Jean-Antoine Ribeil, Loïc Guillevin, Marco A. Cassatella, Antje Mueller, Nathalie Thieblemont, Peter Lamprecht, Luc Mouthon, Sylvain Perruche, Véronique Witko-Sarsat

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Figure 10

Apoptotic cells expressing phosphatidylserine-associated PR3 induce proinflammatory responses and favor Th9 polarization: in vivo switch to Th17 in the presence of PR3 ANCA.

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Apoptotic cells expressing phosphatidylserine-associated PR3 induce proi...
Phagocytosis of apoptotic cells expressing phosphatidylserine-associated PR3 stimulates the production of inflammatory chemokines by macrophages (depicted in red) through the IL-1R1/MyD88 signaling pathway and NO production, thus promoting the recruitment of inflammatory cells expressing PR3, including neutrophils and monocytes. This process acts as an amplification loop and perpetuates inflammation. The presence of phosphatidylserine-associated PR3 on apoptotic cells generates a proinflammatory microenvironment, which facilitates the production of Th2/Th9- and Th1-activated CD4+ T cells through the interaction between pDCs and naive T cells. Importantly, the presence of anti-PR3 ANCA induces Th17 cell production. Increased G-CSF production by macrophages and by T cells may result in an increased PR3 biosynthesis and mobilization of neutrophils from BM, suggesting that PR3 participates in an autoamplification loop, which in turn sustains inflammation. Expression of phosphatidylserine-associated PR3 on apoptotic neutrophils, which prevents the resolution of inflammation, appears to be a key element in the pathogenesis of GPA. PMN, neutrophils.