title: Toll-like receptor 9 activation enhances B cell activating factor and interleukin-21 induced anti-proteinase 3 autoantibody production in vitro.
abstract: OBJECTIVE
  Granulomatosis with polyangiitis (GPA) is a relapsing small-vessel vasculitis characterized by circulating ANCA against PR3. The mechanisms that trigger PR3-ANCA production are unknown. The aim of this study was to determine whether endogenous factors [B cell activating factor (BAFF) and IL-21] and exogenous factors [oligodeoxynucleotides containing CpG motifs (CpG-ODN)] synergize in stimulating PR3-ANCA production in GPA patients.
  
  
  METHODS
  Peripheral blood mononuclear cells from GPA patients and healthy controls (HCs) were cultured in the presence of BAFF and IL-21, with or without CpG-ODN, for 12 days. PR3-ANCA production in culture supernatants was quantified by Phadia EliA. Phenotypic characterization and the influence of CpG-ODN treatment on IL-21 receptor (IL-21R), transmembrane activator and calcium-modulator and cyclophilin ligand interactor (TACI) and BAFF receptor (BAFF-R) expression on B cells was analysed by flow cytometry.
  
  
  RESULTS
  Stimulation with BAFF and IL-21 significantly increased ANCA production in patient samples, which could be augmented further by addition of CpG-ODN. Stimulation with CpG-ODN increased the percentage of IL-21R(+) and TACI(+) B cells but did not affect BAFF-R expression. GPA patients had an increased percentage of circulating IL-21R(+) and a decreased percentage of TACI(+) circulating memory B cells when compared with HCs. Additionally, patients had decreased expression of BAFF-R on B cells, which was inversely correlated with BAFF concentrations in plasma.
  
  
  CONCLUSIONS
  Our data demonstrate that endogenous and exogenous factors can synergize to promote PR3-ANCA production. Mechanistically, CpG-ODN up-regulated IL-21R and TACI expression on B cells, possibly sensitizing these cells for IL-21- and BAFF-mediated signals. Agents inhibiting Toll-like receptor 9, BAFF and IL-21 signalling pathways may serve as potential therapeutics for intervention in GPA patients.
authors: Department of Pathology and Medical Biology, p.heeringa@umcg.nl.
authors: Nikola
authors: N
authors: Lepse
authors: Department of Rheumatology and Clinical Immunology and.
authors: Judith
authors: J
authors: Land
authors: Department of Rheumatology and Clinical Immunology and.
authors: Abraham
authors: A
authors: Rutgers
authors: Department of Rheumatology and Clinical Immunology and.
authors: Cees G M
authors: CG
authors: Kallenberg
authors: Department of Internal Medicine, Division of Nephrology, University of Groningen, University Medical Center Groningen, Groningen, The Netherlands.
authors: Coen A
authors: CA
authors: Stegeman
authors: Department of Rheumatology and Clinical Immunology and.
authors: Wayel H
authors: WH
authors: Abdulahad
authors: Department of Pathology and Medical Biology, p.heeringa@umcg.nl.
authors: Peter
authors: P
authors: Heeringa
mesh_terms: D000328:Adult; D000368:Aged; D001323:Autoantibodies / Q000276:immunology*; D053264:B-Cell Activating Factor / Q000378:metabolism*; D001402:B-Lymphocytes / Q000276:immunology / Q000378:metabolism / Q000473:pathology; D049109:Cell Proliferation; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005434:Flow Cytometry; D014890:Granulomatosis with Polyangiitis / Q000276:immunology* / Q000378:metabolism / Q000473:pathology; D006801:Humans; D007378:Interleukins / Q000378:metabolism*; D008213:Lymphocyte Activation / Q000276:immunology*; D008297:Male; D008875:Middle Aged; D053146:Myeloblastin / Q000276:immunology* / Q000378:metabolism; D051217:Toll-Like Receptor 9 / Q000276:immunology / Q000378:metabolism*; D000098546:Interleukin-21
chemical_list: D001323:Autoantibodies; D053264:B-Cell Activating Factor; D007378:Interleukins; C495371:TLR9 protein, human; D051217:Toll-Like Receptor 9; D053146:Myeloblastin; D000098546:Interleukin-21
keywords: ANCA; B cells; BAFF; BAFF-R; GPA; IL-21; IL-21R; TACI; TLR9; vasculitis