Divi Cornec1, Alvise Berti2, Amber Hummel3, Tobias Peikert3, Jacques-Olivier Pers4, Ulrich Specks5. 1. Thoracic Disease Research Unit, Division of Pulmonary and Critical Care Medicine, Mayo Clinic, Rochester, MN, USA; INSERM UMR1227, Lymphocytes B et Autoimmunité, Université de Bretagne Occidentale, CHU de Brest, Brest, France. 2. Thoracic Disease Research Unit, Division of Pulmonary and Critical Care Medicine, Mayo Clinic, Rochester, MN, USA; Immunology, Rheumatology, Allergy and Rare Diseases Department, San Raffaele Scientific Institute, Milan, Italy. 3. Thoracic Disease Research Unit, Division of Pulmonary and Critical Care Medicine, Mayo Clinic, Rochester, MN, USA. 4. INSERM UMR1227, Lymphocytes B et Autoimmunité, Université de Bretagne Occidentale, CHU de Brest, Brest, France. 5. Thoracic Disease Research Unit, Division of Pulmonary and Critical Care Medicine, Mayo Clinic, Rochester, MN, USA. Electronic address: specks.ulrich@mayo.edu.
Abstract
OBJECTIVES: To develop a method to detect and phenotype circulating proteinase 3 (PR3)-specific B-cells in patients with PR3-ANCA associated vasculitis (AAV). METHODS: Recombinant human PR3 (rPR3) was tagged with FITC or biotin, and its binding characteristics were studied by flow cytometry using three hybridoma cell lines secreting antibodies (Ab) against human PR3, mouse PR3 (no cross-reactivity with human PR3), and human neutrophil elastase. We measured the proportion of PR3-specific B-cells and studied their surface phenotype in patients with PR3-AAV and healthy controls (HC). RESULTS: Labeled rPR3 efficiently and specifically bound to hybridoma cells producing anti-human-PR3-Ab but not anti-mouse-PR3-Ab or anti-human-elastase-Ab. The proportion of rPR3-stained B cells was higher in patients with PR3-AAV compared to HCs: median (IQR) 1.11% (0.81-2.43) vs 0.45% (0.26-0.62) respectively, p < 0.001. There was a trend towards a higher proportion of PR3-specific B cells among patients with active disease compared to patients in remission: 2.91% (1.18-6.52) vs 0.99% (0.72-1.58), p = 0.09. In HCs, the proportion of PR3-specific B cells was highest among the transitional B-cell subset, and decreased with the maturation of B cells. Conversely, in patients, the proportion of PR3-specific B cells progressively increased with the maturation of B cells (median 1.90% of naïve B cells, 2.30% of unswitched memory B cells, 2.37% of switched memory B cells, and 3.68% of plasmablasts). CONCLUSIONS: Circulating PR3-specific B cells can be detected in HC and patients with PR3-AAV. Their progressive enrichment during B-cell maturation suggests that they are actively selected and escape peripheral tolerance checkpoints in patients.
OBJECTIVES: To develop a method to detect and phenotype circulating proteinase 3 (PR3)-specific B-cells in patients with PR3-ANCA associated vasculitis (AAV). METHODS: Recombinant humanPR3 (rPR3) was tagged with FITC or biotin, and its binding characteristics were studied by flow cytometry using three hybridoma cell lines secreting antibodies (Ab) against humanPR3, mousePR3 (no cross-reactivity with humanPR3), and humanneutrophil elastase. We measured the proportion of PR3-specific B-cells and studied their surface phenotype in patients with PR3-AAV and healthy controls (HC). RESULTS: Labeled rPR3 efficiently and specifically bound to hybridoma cells producing anti-human-PR3-Ab but not anti-mouse-PR3-Ab or anti-human-elastase-Ab. The proportion of rPR3-stained B cells was higher in patients with PR3-AAV compared to HCs: median (IQR) 1.11% (0.81-2.43) vs 0.45% (0.26-0.62) respectively, p < 0.001. There was a trend towards a higher proportion of PR3-specific B cells among patients with active disease compared to patients in remission: 2.91% (1.18-6.52) vs 0.99% (0.72-1.58), p = 0.09. In HCs, the proportion of PR3-specific B cells was highest among the transitional B-cell subset, and decreased with the maturation of B cells. Conversely, in patients, the proportion of PR3-specific B cells progressively increased with the maturation of B cells (median 1.90% of naïve B cells, 2.30% of unswitched memory B cells, 2.37% of switched memory B cells, and 3.68% of plasmablasts). CONCLUSIONS: Circulating PR3-specific B cells can be detected in HC and patients with PR3-AAV. Their progressive enrichment during B-cell maturation suggests that they are actively selected and escape peripheral tolerance checkpoints in patients.
Authors: Alvise Berti; Roscoe Warner; Kent Johnson; Divi Cornec; Darrell Schroeder; Brian Kabat; Carol A Langford; Gary S Hoffman; Fernanado C Fervenza; Cees G M Kallenberg; Philip Seo; Robert Spiera; E William St Clair; Paul Brunetta; John H Stone; Peter A Merkel; Ulrich Specks; Paul A Monach Journal: Arthritis Rheumatol Date: 2018-05-07 Impact factor: 10.995
Authors: Peter Lamprecht; Anja Kerstein; Sebastian Klapa; Susanne Schinke; Christian M Karsten; Xinhua Yu; Marc Ehlers; Jörg T Epplen; Konstanze Holl-Ulrich; Thorsten Wiech; Kathrin Kalies; Tanja Lange; Martin Laudien; Tamas Laskay; Timo Gemoll; Udo Schumacher; Sebastian Ullrich; Hauke Busch; Saleh Ibrahim; Nicole Fischer; Katrin Hasselbacher; Ralph Pries; Frank Petersen; Gesche Weppner; Rudolf Manz; Jens Y Humrich; Relana Nieberding; Gabriela Riemekasten; Antje Müller Journal: Front Immunol Date: 2018-04-09 Impact factor: 7.561
Authors: Anouk von Borstel; Judith Land; Wayel H Abdulahad; Abraham Rutgers; Coen A Stegeman; Arjan Diepstra; Peter Heeringa; Jan Stephan Sanders Journal: Front Immunol Date: 2019-09-24 Impact factor: 7.561
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Alicia Martínez-Romero; Glòria Martrus; Ivana Marventano; Wlodzimierz Maslinski; Giuseppe Matarese; Anna Vittoria Mattioli; Christian Maueröder; Alessio Mazzoni; James McCluskey; Mairi McGrath; Helen M McGuire; Iain B McInnes; Henrik E Mei; Fritz Melchers; Susanne Melzer; Dirk Mielenz; Stephen D Miller; Kingston H G Mills; Hans Minderman; Jenny Mjösberg; Jonni Moore; Barry Moran; Lorenzo Moretta; Tim R Mosmann; Susann Müller; Gabriele Multhoff; Luis Enrique Muñoz; Christian Münz; Toshinori Nakayama; Milena Nasi; Katrin Neumann; Lai Guan Ng; Antonia Niedobitek; Sussan Nourshargh; Gabriel Núñez; José-Enrique O'Connor; Aaron Ochel; Anna Oja; Diana Ordonez; Alberto Orfao; Eva Orlowski-Oliver; Wenjun Ouyang; Annette Oxenius; Raghavendra Palankar; Isabel Panse; Kovit Pattanapanyasat; Malte Paulsen; Dinko Pavlinic; Livius Penter; Pärt Peterson; Christian Peth; Jordi Petriz; Federica Piancone; Winfried F Pickl; Silvia Piconese; Marcello Pinti; A Graham Pockley; Malgorzata Justyna Podolska; Zhiyong Poon; Katharina Pracht; Immo Prinz; Carlo E M Pucillo; Sally A Quataert; Linda Quatrini; Kylie M Quinn; Helena Radbruch; Tim R D J Radstake; Susann Rahmig; Hans-Peter Rahn; Bartek Rajwa; Gevitha Ravichandran; Yotam Raz; Jonathan A Rebhahn; Diether Recktenwald; Dorothea Reimer; Caetano Reis e Sousa; Ester B M Remmerswaal; Lisa Richter; Laura G Rico; Andy Riddell; Aja M Rieger; J Paul Robinson; Chiara Romagnani; Anna Rubartelli; Jürgen Ruland; Armin Saalmüller; Yvan Saeys; Takashi Saito; Shimon Sakaguchi; Francisco Sala-de-Oyanguren; Yvonne Samstag; Sharon Sanderson; Inga Sandrock; Angela Santoni; Ramon Bellmàs Sanz; Marina Saresella; Catherine Sautes-Fridman; Birgit Sawitzki; Linda Schadt; Alexander Scheffold; Hans U Scherer; Matthias Schiemann; Frank A Schildberg; Esther Schimisky; Andreas Schlitzer; Josephine Schlosser; Stephan Schmid; Steffen Schmitt; Kilian Schober; Daniel Schraivogel; Wolfgang Schuh; Thomas Schüler; Reiner Schulte; Axel Ronald Schulz; Sebastian R Schulz; Cristiano Scottá; 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