905 Enevold, et al: SNP screening in RA Personal non-commercial use only. The Journal of Rheumatology Copyright © 2010. All rights reserved. Multiplex Screening of 22 Single-Nucleotide Polymorphisms in 7 Toll-like Receptors: An Association Study in Rheumatoid Arthritis CHRISTIAN ENEVOLD, TIMOTHY R.D. RADSTAKE, MARIEKE J.H. COENEN, JAAP FRANSEN, ERIK J.M. TOONEN, KLAUS BENDTZEN, and PIET L.C.M. van RIEL ABSTRACT. Objective. Toll-like receptors (TLR) have been implicated in the pathogenesis of arthritis. We inves- tigated the role of functional variants of TLR in the disease phenotype and severity of rheumatoid arthritis (RA). Methods. All patients from a longterm observational inception cohort (n = 319) were genotyped for 22 single-nucleotide polymorphisms (SNP) in TLR2, 3, 4, 5, 7, 8, and 9 using multiplex assays. Clinical characteristics including sex, age at disease onset, rheumatoid factor (RF), and shared epi- tope positivity and disease activity score and radiological progression were taken into account. Genotypes were analyzed for association with Disease Activity Scores (DAS28) and joint damage (Rau scores) at 3 and 6 years. Results. After Bonferroni correction, there was a moderate association between RF positivity and TLR8-rs5741883. No other TLR variant was significantly associated with any RA clinical characteristics. Conclusion. Using a large inception cohort and strict statistical evaluation, we could not identify an association between functional TLR variants and RA phenotype and disease severity. This suggests the functional TLR variants do not play a major role in RA phenotype and disease severity. (First Release March 1 2010; J Rheumatol 2010;37:905–10; doi:10.3899/jrheum.090775) Key Indexing Terms: TOLL-LIKE RECEPTOR RHEUMATOIDARTHRITIS RHEUMATOID FACTOR RADIOLOGICAL JOINT DAMAGE DISEASE ACTIVITY SCORE From the Institute for Inflammation Research, National University Hospital, Copenhagen, Denmark; and the Department of Rheumatology and Department of Human Genetics, Radboud University Nijmegen Medical Centre, Nijmegen, The Netherlands. Supported by the Lundbeck Foundation and the Danish Biotechnology Program. Dr. Radstake is a VIDI Laureate of the Dutch Organization of Research (NWO). C. Enevold*, MSc, Institute for Inflammation Research, National University Hospital; T.R.D. Radstake*, MD, PhD, Program Leader, Translational Research, Department of Rheumatology; M.J.H. Coenen, PhD, Department of Human Genetics; J. Fransen, PhD, Department of Rheumatology; E.J.M. Toonen, MSc, Department of Human Genetics, Radboud University Nijmegen Medical Centre; K. Bendtzen, MD, PhD, Institute for Inflammation Research, National University Hospital; P.L.C.M. van Riel, MD, PhD, Department of Rheumatology, Radboud University Nijmegen Medical Centre. *Both authors contributed equally to this report. Address correspondence to Dr. T.R.D. Radstake, Department of Rheumatology, Radboud University Nijmegen Medical Centre, Geert Grooteplein 8, 6500 HB, Nijmegen, The Netherlands. E-mail: t.radstake@reuma.umcn.nl Accepted for publication December 3, 2009. Rheumatoid arthritis (RA) is an autoimmune disease affect- ing multiple synovial joints, leading to significant morbidi- ty and shortened life expectancy. Despite longstanding efforts, the precise mechanisms underlying the inflammato- ry processes remain to be elucidated. Toll-like receptors (TLR) were discovered to be crucial receptors triggering innate immune responses. Currently, 10 TLR subtypes have been described in humans, all thought to have their own spe- cific ligands and cellular localization (as reviewed 1 ). For example TLR1, 2, 4, 5, and 6 are located on the cell surface and scavenge the environment for ligands. In contrast, TLR3, 7, 8, and 9 have an intracellular localization and rec- ognize intracellular ligands, including ligands that are endo- cytosed 2,3 . Accumulating evidence suggests a pivotal role for TLR in the recognition of endogenous ligands and, as well, linking innate and adaptive immune responses. Recently, several groups have provided evidence for a role of TLR in arthritis in experimental disease conditions and in humans. It has been demonstrated that various TLR sub- types are expressed at higher levels in synovial tissues from patients with RA compared to those from healthy con- trols 4-6 . Ligands for TLR3 [host-derived (RNA) and TLR4 (HSPB8)] are also abundant in the circulation as well as in the synovial joints of patients with RA 4,5,7 . Additionally, TLR4-mediated stimulation of dendritic cells from patients with RA leads to significantly higher cytokine concentra- tions compared to similar cells from healthy controls, fur- ther supporting a deranged TLR response in RA. On these grounds, we hypothesized that TLR might be involved in the pathogenesis of RA. To test this, we investigated potential www.jrheum.org Downloaded on June 7, 2022 from