Proteinase 3 on apoptotic cells disrupts immune silencing in autoimmune vasculitis
Arnaud Millet, … , Sylvain Perruche, Véronique Witko-Sarsat
Arnaud Millet, … , Sylvain Perruche, Véronique Witko-Sarsat
Published October 5, 2015
Citation Information: J Clin Invest. 2015;125(11):4107-4121. https://doi.org/10.1172/JCI78182.
View: Text | PDF
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

×

Figure 6

pDCs exposed in vivo to a PR3-induced microenvironment triggered the production of Th9 cells, and addition of anti-PR3 ANCA facilitated Th17 cell polarization.

Options: View larger image (or click on image) Download as PowerPoint
pDCs exposed in vivo to a PR3-induced microenvironment triggered the pro...
(A) pDCs isolated from mice following the i.v. injection of vehicle (white triangles), apoptotic controls (white circles), PR3- (black squares), or PR3/4H4A-expressing cells (black diamonds) were cultured with naive CD25–CD4+ T cells from OTII/Rag1–/– mice in the presence of OVA peptide. Flow cytometry analysis was performed 4 days later, and the percentages of Tregs (CD4+CD25+FOXP3+), Th9 (CD4+IL-9+), Th2 (CD4+IL-4+), Th1 (CD4+IFN-γ+), and Th17 (CD4+IL-17A+) cells determined. (B) The above experiments were also performed with the concomitant injection of anti-PR3 ANCA or control IgG in the presence of apoptotic control (white bars) or PR3-expressing cells (black bars). T cell polarization was analyzed using flow cytometry. Data are presented as mean ± SEM. n = 3 mice per group, each mice studied in triplicate. Significant differences between groups were determined by ANOVA with multicomparison analysis. *P < 0.05; **P < 0.01; ***P < 0.001. The experiment presented in A was reproduced twice with identical results.