Standardised assessment of membrane proteinase 3 expression. Analysis in ANCA-associated vasculitis and controls

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Abstract
Objectives: 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 TNFα, 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 TNFα to assess whether standardised assessment of mPR3 expression requires priming. Using neutrophils before and after priming with TNFα, 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 TNFα for mPR3 expression. Results: 42% of all individuals analysed showed minimal expression for mPR3 on all neutrophils before priming with TNFα, 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 TNFα was significantly increased (p+ PMN were also increased in patients with SLE (pConclusion: Standardised assessment of proteinase 3 on the membrane of neutrophils requires priming with TNFα. Percentages of mPR3+ PMN are increased in AAV and SLE, but not in RA.