title: Blockade of Toll-like receptor 2 prevents spontaneous cytokine release from rheumatoid arthritis ex vivo synovial explant cultures.
abstract: BACKGROUND
  The aim of this study was to examine the effect of blocking Toll-like receptor 2 (TLR2) in rheumatoid arthritis (RA) synovial cells.
  
  
  METHODS
  RA synovial tissue biopsies, obtained under direct visualization at arthroscopy, were established as synovial explant cultures ex vivo or snap frozen for immunohistology. Mononuclear cell cultures were isolated from peripheral blood and synovial fluid of RA patients. Cultures were incubated with the TLR1/2 ligand, Pam3CSK4 (200 ng, 1 and 10 μg/ml), an anti-TLR2 antibody (OPN301, 1 μg/ml) or an immunoglobulin G (IgG) (1 μg/ml) matched control. The comparative effect of OPN301 and adalimumab (anti-tumour necrosis factor alpha) on spontaneous release of proinflammatory cytokines from RA synovial explants was determined using quantitative cytokine MSD multiplex assays or ELISA. OPN301 penetration into RA synovial tissue explants cultures was assessed by immunohistology.
  
  
  RESULTS
  Pam3CSK4 significantly upregulated interleukin (IL)-6 and IL-8 in RA peripheral blood mononuclear cells (PBMCs), RA synovial fluid mononuclear cells (SFMCs) and RA synovial explant cultures (P < 0.05). OPN301 significantly decreased Pam3CSK4-induced cytokine production of tumour necrosis factor alpha (TNF-α), IL-1β, IL-6, interferon (IFN)-γ and IL-8 compared to IgG control in RA PBMCs and SFMCs cultures (all P < 0.05). OPN301 penetration of RA synovial tissue cultures was detected in the lining layer and perivascular regions. OPN301 significantly decreased spontaneous cytokine production of TNF-α, IL-1β, IFN-γ and IL-8 from RA synovial tissue explant cultures (all P < 0.05). Importantly, the inhibitory effect of OPN on spontaneous cytokine secretion was comparable to inhibition by anti-TNFα monoclonal antibody adalimumab.
  
  
  CONCLUSIONS
  These findings further support targeting TLR2 as a potential therapeutic agent for the treatment of RA.
authors.affiliation: Department of Rheumatology, St Vincent's University Hospital, Dublin Academic Medical Centre, Elm Park, Dublin 4, Ireland. douglas.veale@ucd.ie
authors.forename: Sinéad Nic An
authors.initials: SN
authors.lastname: Ultaigh
authors.forename: Tajvur P
authors.initials: TP
authors.lastname: Saber
authors.forename: Jennifer
authors.initials: J
authors.lastname: McCormick
authors.forename: Mary
authors.initials: M
authors.lastname: Connolly
authors.forename: Jerome
authors.initials: J
authors.lastname: Dellacasagrande
authors.forename: Brian
authors.initials: B
authors.lastname: Keogh
authors.forename: William
authors.initials: W
authors.lastname: McCormack
authors.forename: Mary
authors.initials: M
authors.lastname: Reilly
authors.forename: Luke A
authors.initials: LA
authors.lastname: O'Neill
authors.forename: Peter
authors.initials: P
authors.lastname: McGuirk
authors.forename: Ursula
authors.initials: U
authors.lastname: Fearon
authors.forename: Douglas J
authors.initials: DJ
authors.lastname: Veale
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D018501:Antirheumatic Agents / Q000494:pharmacology; D001172:Arthritis, Rheumatoid / Q000276:immunology* / Q000378:metabolism; D018929:Cell Culture Techniques; D016207:Cytokines / Q000096:biosynthesis; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D008297:Male; D008875:Middle Aged; D013583:Synovial Membrane / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D051195:Toll-Like Receptor 2 / Q000037:antagonists & inhibitors*
chemical_list: D000911:Antibodies, Monoclonal; D018501:Antirheumatic Agents; D016207:Cytokines; C495342:TLR2 protein, human; D051195:Toll-Like Receptor 2