Coexpression of CD177 and membrane proteinase 3 on neutrophils in antineutrophil cytoplasmic autoantibody–associated systemic vasculitis: Anti–proteinase 3–mediated neutrophil activation is independent of the role of CD177‐expressing neutrophils

Abstract

Objective. Wegener's granulomatosis (WG) is strongly associated with antineutrophil cytoplasmic autoantibodies (ANCAs) directed against proteinase 3 (PR3). Recent studies have shown that membranebound PR3 (mPR3) is differentially expressed and colocalizes with CD177/NB1 on circulating neutrophils. We undertook this study to assess the differential expression of CD177 on neutrophils from patients with ANCA-associated systemic vasculitis (ASV) in comparison with patients with systemic lupus erythematosus (SLE), patients with rheumatoid arthritis (RA), and healthy individuals, and to investigate whether colocalization of mPR3 and CD177 affects anti-PR3-mediated neutrophil activation.Methods. Expression of CD177 and mPR3 was analyzed by flow cytometry on isolated neutrophils from patients with ASV (n ‫؍‬ 53), those with SLE (n ‫؍‬ 30), those with RA (n ‫؍‬ 26), and healthy controls (n ‫؍‬ 31). Neutrophil activation mediated by anti-PR3 antibodies was assessed by measuring the oxidative burst with a dihydrorhodamine assay.Results. Percentages of CD177-expressing neutrophils were significantly higher in patients with ASV and those with SLE than in healthy controls. In 3 healthy donors, CD177 expression was not detected. After priming with tumor necrosis factor ␣, neutrophils remained negative for CD177 while mPR3 expression was induced. Neutrophils from CD177-negative donors or CD177؊ neutrophils sorted from donors with bimodal expression were susceptible to anti-PR3-mediated oxidative burst. Variation in the extent of anti-PR3-mediated neutrophil activation among different donors occurred independent of the percentage of CD177-expressing neutrophils.Conclusion. Membrane expression of CD177 on circulating neutrophils is increased in patients with ASV and in those with SLE, but not in RA patients. However, primed neutrophils from CD177-negative individuals also express mPR3 and are susceptible to anti-PR3-mediated oxidative burst, suggesting that recruitment of CD177-independent mPR3 is involved in anti-PR3-induced neutrophil activation.

Complement Receptor Mac-1 Is an Adaptor for NB1 (CD177)-mediated PR3-ANCA Neutrophil Activation

“…In a recent report, the percentage of NB1-expressing neutrophils was higher in ANCA vasculitis patients, compared with healthy controls. Furthermore, within the ANCA group, the percentage was higher in those patients who had relapsing disease (20). Together, these data indicate that the mNB1 pos /PR3 high phenotype is clinically relevant in ANCA vasculitis.…”

“…PR3, but not MPO shows a bimodal membrane expression pattern on neutrophils. High mPR3 expression is only found on those neutrophils that co-express the GPI-linked NB1 receptor (17,18,20). We performed previously ectopic expression studies showing that NB1 was a sufficient receptor for PR3 (18,35).…”

IgG Glycan Hydrolysis Attenuates ANCA-Mediated Glomerulonephritis

“…37 Freshly isolated healthy donor neutrophils (final concentration 2.5 ϫ 10 6 /ml) were gradually warmed to 37°C and incubated with 5 g/ml cytochalasin B (SigmaAldrich, Zwijndrecht, Netherlands) for 5 minutes at 37°C to enhance oxygen radical production. Then cells were loaded with 0.05 mM DHR (D632; Molecular Probes, Eugene, OR) for 10 minutes at 37°C.…”

Endothelium-Neutrophil Interactions in ANCA-Associated Diseases

“…52 -56 Positive correlations are also reported between a high percentage of mPR3 + neutrophils and abnormal renal function parameters in patients with GPA. 57 Priming with TNF-a increases the mPR3 expression in both CD177 2 and CD177 + subpopulations 55 and the low level of mPR3 then present on CD177 2 neutrophils appears to be sufficient for some of the effects promoted by anti-PR3 antibodies, such as actin polymerization and oxidative bursts. 55,58 More recently, anti-PR3 ANCA were shown phox , and PR3 from secretory vesicles to the plasma membrane, resulting in a co-localization of gp91 phox , b2-integrins, and FcgRII.…”