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OP0210 TLR-2 Induces Pro-Inflammatory/Angiogenic Mechanisms in GCA Temporal Artery Explant Cultures Ex Vivo
  1. A. Maher1,
  2. D. Molloy1,
  3. J. McCormick1,
  4. L. O’Neill1,
  5. D. Veale1,
  6. C. Murphy2,
  7. U. Fearon1,
  8. E. Molloy1
  1. 1Rheumatology Department, Dublin Academic Medical Centre, St. Vincent’s University Hospital, Dublin, Ireland
  2. 2Ophthalmology, Royal Victoria Eye and Ear Hospital, Dublin, Ireland

Abstract

Background Giant cell arteritis (GCA) is a common form of primary vasculitis characterised by dysfunctional vessels and inflammatory infiltration leading to luminal occlusion. Toll-Like receptor-2 (TLR2) has been implicated in the pathogenesis of GCA; however, the mechanisms involved have yet to be elucidated.

Objectives This study examines the mechanistic role of TLR2 activation in regulating the pro-inflammatory mechanisms in GCA.

Methods Temporal artery (TA) sections from patients positive and negative for GCA were assessed for TLR2 expression by immunohistology. Ex-vivo TA explant cultures were stimulated with Pam3CSK4 (TLR2 agonist) (1ug/ml) for 24 hrs. VEGF, Ang2, IL-6, IL-8 and MMP2/9 expression was quantified in cultured supernatants by ELISA and gelatin zymography. The effect of Pam3CSK4-induced GCA TA explant conditioned media on endothelial cell (dHMVEC) function was assessed by angiogenic assays. GCA TA explant myofibroblast outgrowth was assessed using matrigel assays (1-20 days)1. Finally the effect of GCA TA explant conditioned media on Human embryonic kidney (HEK)-TLR2 cells was quantified by NFκB luciferase reporter assays.

Results TLR2 expression was higher in positive versus negative TA sections. Pam3CSK4 significantly increased VEGF, IL-6 and IL-8 expression (all p<.0.05), with differential effects observed for Ang2 and MMP2/9 in TA explant cultures. Pam3CSK4-induced GCA TA conditioned media promoted dHMVEC tube formation (p<0.05). Pam3CSK4 induced myofibroblast outgrowths from GCA TA explants embedded in matrigel. Finally GCA TA conditioned media induced TLR2 activation through induction of NF-κB activation in HEK-TLR2 cells (p<0.05), confirming the presence of endogenous ligands for TLR2 at the site of inflammation in GCA.

Conclusions Using ex-vivo TA explant cultures, TLR2 activation induced angiogenic and invasive functional mechanisms in GCA. Furthermore activation of TLR2 by GCA TA conditioned media, suggests that TLR2 may represent a potential therapeutic target for GCA.

References

  1. Lozano E, Segarra M, García-Matínez A, Hernández-Rodríguez J, Cid MC. (2008) Imatinib mesylate inhibits in vitro and ex vivo biological responses related to vascular occlusion in giant cell arteritis. Ann Rheum Dis., 67(11): 1581-8.

Disclosure of Interest A. Maher: None Declared, D. Molloy: None Declared, J. McCormick: None Declared, L. O’Neill: None Declared, D. Veale Grant/research support from: Abbott, Opsona, Pfizer, Roche, Consultant for: MSD, Pfizer, Roche, Speakers bureau: Janssen, MSD, Pfizer, UCB, C. Murphy: None Declared, U. Fearon: None Declared, E. Molloy: None Declared

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