Recombinant Protein to Analyze Autoantibodies to Proteinase 3 in Systemic Vasculitis

Rarok, Agnieszka A.; Huitema, Minke G.; Leij, Marcel J. van der; Geld, Ymke M. van der; Berthold, Heike; Schmitt, Jacky; Stegeman, Coen A.; Limburg, Pieter C.; Kallenberg, Cees G. M.

doi:10.1309/ytu2fuhbexjlwmld

Cited by 9 publications

(5 citation statements)

References 38 publications

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“…Also recombinant PR3 has not been successfully introduced in clinical practice due to problems with expression and purification procedures 20. A direct ELISA using recombinant, proteolytically inactive PR3 produced in the baculovirus expression system, appeared less sensitive than a direct ELISA using native PR3 21. Recombinant PR3, however, is preferentially expressed in human cell lines that constitutively secrete the wild-type precursor, but are unable to generate active PR3 in vivo 8 22.…”

Section: Discussionmentioning

confidence: 99%

A novel enzyme-linked immunosorbent assay using a mixture of human native and recombinant proteinase-3 significantly improves the diagnostic potential for antineutrophil cytoplasmic antibody-associated vasculitis

et al. 2008

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Owing to the very high sensitivity of the novel anti-PR3-hn-hr ELISA for the detection of PR3-ANCA in C-ANCA-positive samples of patients with AAV this assay has an excellent diagnostic performance. This feature is combined with a good predictability of clinical relapses in patients with PR3-AAV. These characteristics challenge the dogma that, for detection of PR3-ANCA, capture ELISAs are superior for diagnosis and follow-up.

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Section: Discussionmentioning

confidence: 99%

A novel enzyme-linked immunosorbent assay using a mixture of human native and recombinant proteinase-3 significantly improves the diagnostic potential for antineutrophil cytoplasmic antibody-associated vasculitis

et al. 2008

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“…This suggests that the PR3 N-terminus is an element of the PR3-NB1 binding site. Studies using recombinant PR3 demonstrated that nearly all ANCA sera from patients with vasculitis recognize PR3 irrespective of its N-terminus [30,31,[47][48][49]. In fact, clinical data show that ANCA binding not only to mature PR3 but also to proPR3 have the same or even better predictive power for disease progression in patients with ANCA vasculitis [8,50].…”

Section: Discussionmentioning

confidence: 99%

Neutrophil surface presentation of the anti-neutrophil cytoplasmic antibody-antigen proteinase 3 depends on N-terminal processing

Vietinghoff

Eulenberg

Wellner

et al. 2008

Clinical and Experimental Immunology

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SummaryThe neutrophil serine protease proteinase 3 (PR3) is a main autoantigen in anti-neutrophil cytoplasmic antibody-associated vasculitis. PR3 surface presentation on neutrophilic granulocytes, the main effector cells, is pathogenically important. PR3 is presented by the NB1 (CD177) glycoprotein, but how the presentation develops during neutrophil differentiation is not known. An N-terminally unprocessed PR3 (proPR3) is produced early during neutrophil development and promotes myeloid cell differentiation. We therefore investigated if PR3 presentation depended on NB1 during neutrophil differentiation and if PR3 and proPR3 could both be presented by NB1. In contrast to mature neutrophils, differentiating neutrophils showed an early NB1-independent PR3 surface display that was recognized by only two of four monoclonal anti-PR3 antibodies and occurred in parallel with proPR3, but not PR3 secretion, suggesting that the NB1-independent surface PR3 was proPR3. PR3 gene expression preceeded NB1. When the NB1 receptor was detected on the surface, a mode of PR3 surface display similar to mature neutrophils developed together with the degranulation system. Ectopic expression studies showed that NB1 was a sufficient receptor for PR3 but not proPR3. ProPR3 display on the plasma membrane may influence the bone marrow microenvironment. NB1-mediated PR3 presentation depended on PR3 N-terminal processing implicating the PR3-N-terminus as NB1-binding site.

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“…Here, for the first time, we used a flow cytometric approach to measure early detectable (actin polymerization) and late detectable (oxidative burst) responses of neutrophils [36] to anti-PR3 antibody at the single-cell level in relation to PR3 expression on the neutrophil surface. In our system, we only used the monoclonal anti-PR3G-3 as a model antibody for activation of neutrophils because of its adequate binding characteristics in capture ELISA in comparison with other PR3-specific mAb such as mAb 12.8 [37]. We also measured F-actin responses with PR3-ANCA-positive IgG fractions from patients with active WG (Fig.…”

Section: Discussionmentioning

confidence: 99%

Constitutive membrane expression of proteinase 3 (PR3) and neutrophil activation by anti-PR3 antibodies

Rossum

Rarok

Huitema

et al. 2004

Journal of Leukocyte Biology

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Antineutrophil cytoplasm autoantibodies with specificity for proteinase 3 (PR3) are thought to play a major role in the pathogenesis of Wegener's granulomatosis (WG), presumably by their potential to activate neutrophils. In patients with WG, high expression of PR3 on the surface of nonprimed neutrophils is associated with an increased incidence and rate of relapse. In this study, we analyzed the functional significance of constitutive PR3 expression for neutrophil activation as induced by anti-PR3 antibody. Therefore, primed and nonprimed neutrophils were stimulated with the monoclonal anti-PR3 antibody PR3G-3. Activation was measured as actin polymerization by the phalloidin assay as an early, detectable activation event and oxidative burst by the dihydrorhodamine assay, as a late, detectable activation event. In contrast to the oxidative burst, we found that anti-PR3 antibody-induced actin polymerization could be triggered in neutrophils without priming with tumor necrosis factor alpha (TNF-alpha). In addition, a correlation was found between the level of PR3 expression on the surface of these nonprimed neutrophils and the degree of actin polymerization. However, after priming with TNF-alpha, no correlation was found between membrane expression of PR3 and the level of actin polymerization or respiratory burst as induced by anti-PR3 antibody. These data suggest that the presence of PR3 on the surface of nonprimed neutrophils has consequences for their susceptibility to the initial activation step by anti-PR3 antibodies. These data may be relevant in view of the observed relation between membrane expression of PR3 on nonprimed neutrophils of patients with WG and their susceptibility for relapses.

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Recombinant Protein to Analyze Autoantibodies to Proteinase 3 in Systemic Vasculitis

Cited by 9 publications

References 38 publications

A novel enzyme-linked immunosorbent assay using a mixture of human native and recombinant proteinase-3 significantly improves the diagnostic potential for antineutrophil cytoplasmic antibody-associated vasculitis

A novel enzyme-linked immunosorbent assay using a mixture of human native and recombinant proteinase-3 significantly improves the diagnostic potential for antineutrophil cytoplasmic antibody-associated vasculitis

Neutrophil surface presentation of the anti-neutrophil cytoplasmic antibody-antigen proteinase 3 depends on N-terminal processing

Constitutive membrane expression of proteinase 3 (PR3) and neutrophil activation by anti-PR3 antibodies

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