Standardised assessment of membrane proteinase 3 expression. Analysis in ANCA-associated vasculitis and controls.
OBJECTIVE
Increased numbers of neutrophils expressing proteinase 3 on their membrane (mPR3) have been reported in anti-neutrophil cytoplasm antibody (ANCA)-associated vasculitis (AAV) and are suggested to be involved in AAV immunopathogenesis. In most studies, neutrophils were analysed for mPR3 expression without priming with TNFalpha, suggesting that mPR3 expression on neutrophils is dependent on other priming events, such as isolation procedures . These priming events can be variable. Therefore, we analysed mPR3 expression on neutrophils before and after priming with TNFalpha to assess whether standardised assessment of mPR3 expression requires priming. Using neutrophils before and after priming with TNFalpha, we assessed percentages of mPR3(+) neutrophils in patients with AAV and in disease and healthy controls.


METHODS
Neutrophils from patients with PR3-AAV and MPO-AAV, systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), and from healthy controls were analysed before and after priming with TNFalpha for mPR3 expression.


RESULTS
42% of all individuals analysed showed minimal expression for mPR3 on all neutrophils before priming with TNFalpha, whereas after priming a clear mPR3(+) subset was observed next to mPR3(-) neutrophils, corresponding to bimodal mPR3 expression. In patients with PR3-AAV or MPO-AAV, the percentage of mPR3(+) neutrophils after priming with TNFalpha was significantly increased (p<0.01 and p<0.05, respectively) compared with healthy controls. Percentages of mPR3(+) PMN were also increased in patients with SLE (p<0.01) but not in RA.


CONCLUSIONS
Standardised assessment of proteinase 3 on the membrane of neutrophils requires priming with TNFalpha. Percentages of mPR3(+) PMN are increased in AAV and SLE, but not in RA.
van Rossum
André P
AP
Department of Rheumatology and Clinical Immunology, Groningen University Medical Centre, University of Groningen, The Netherlands. c.g.m.kallenberg@int.umcg.nl
Huitema
Minke G
MG
Stegeman
Coen A
CA
Bijl
Marc
M
de Leeuw
Karina
K
Van Leeuwen
Miek A
MA
Limburg
Pieter C
PC
Kallenberg
Cees G M
CG
D000328:Adult; D019268:Antibodies, Antineutrophil Cytoplasmic / Q000276:immunology*; D001172:Arthritis, Rheumatoid / Q000201:enzymology / Q000276:immunology; D015415:Biomarkers / Q000032:analysis; D002462:Cell Membrane / Q000201:enzymology / Q000276:immunology; D005260:Female; D006801:Humans; D008180:Lupus Erythematosus, Systemic / Q000201:enzymology / Q000276:immunology; D008297:Male; D008875:Middle Aged; D053146:Myeloblastin / Q000032:analysis*; D018375:Neutrophil Activation / Q000276:immunology; D009504:Neutrophils / Q000201:enzymology / Q000276:immunology; D009195:Peroxidase / Q000276:immunology; D017463:Receptors, Complement 3b / Q000032:analysis; D018124:Receptors, Tumor Necrosis Factor / Q000032:analysis; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology; D014657:Vasculitis / Q000201:enzymology / Q000276:immunology*
D019268:Antibodies, Antineutrophil Cytoplasmic; D015415:Biomarkers; D017463:Receptors, Complement 3b; D018124:Receptors, Tumor Necrosis Factor; D014409:Tumor Necrosis Factor-alpha; D009195:Peroxidase; D053146:Myeloblastin