Title Anti-citrullinated protein antibody ( ACPA ) in rheumatoid arthritis : influence of an interaction between HLA-DRB 1 shared epitope and a deletion polymorphism in glutathione S-transferase in a cross-sectional study

Introduction: A deletion polymorphism in glutathione S-transferase Mu-1 (GSTM1-null) has previously been implicated to play a role in rheumatoid arthritis (RA) risk and progression, although no prior investigations have examined its associations with anticitrullinated protein antibody (ACPA) positivity. The purpose of this study was to examine the associations of GSTM1-null with ACPA positivity in RA and to assess for evidence of interaction between GSTM1 and HLA-DRB1 shared epitope (SE). Methods: Associations of GSTM1-null with ACPA positivity were examined separately in two RA cohorts, the Veterans Affairs Rheumatoid Arthritis (VARA) registry (n = 703) and the Study of New-Onset RA (SONORA; n = 610). Interactions were examined by calculating an attributable proportion (AP) due to interaction. Results: A majority of patients in the VARA registry (76%) and SONORA (69%) were positive for ACPA with a similar frequency of GSTM1-null (53% and 52%, respectively) and HLA-DRB1 SE positivity (76% and 71%, respectively). The parameter of patients who had ever smoked was more common in the VARA registry (80%) than in SONORA (65%). GSTM1-null was significantly associated with ACPA positivity in the VARA registry (odds ratio (OR), 1.45; 95% confidence interval (CI), 1.02 to 2.05), but not in SONORA (OR, 1.00; 95% CI, 0.71 to 1.42). There were significant additive interactions between GSTM1 and HLA-DRB1 SE in the VARA registry (AP, 0.49; 95% CI, 0.21 to 0.77; P < 0.001) in ACPA positivity, an interaction replicated in SONORA (AP, 0.38; 95% CI, 0.00 to 0.76; P = 0.050). Conclusions: This study is the first to show that the GSTM1-null genotype, a common genetic variant, exerts significant additive interaction with HLA-DRB1 SE on the risk of ACPA positivity in RA. Since GSTM1 has known antioxidant functions, these data suggest that oxidative stress may be important in the development of RA-specific autoimmunity in genetically susceptible individuals. * Correspondence: tmikuls@unmc.edu Omaha Veterans Affairs Medical Center and Nebraska Arthritis Outcomes Research Center, University of Nebraska Medical Center (UNMC), 986270 Nebraska Medical Center, Omaha, NE 68198-6270, USA Full list of author information is available at the end of the article Mikuls et al. Arthritis Research & Therapy 2010, 12:R213 http://arthritis-research.com/content/12/6/R213 © 2010 Mikuls et al.; licensee BioMed Central Ltd. This is an open access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Introduction The human leukocyte antigen (HLA) region accounts for approximately one half of the genetic risk of rheumatoid arthritis (RA). This risk is attributable to alleles encoding a conserved amino acid sequence in the third hypervariable region of the DRB1 chain (commonly referred to as the shared epitope [SE]) [1]. Recent efforts have examined the importance of interactions of SE with other genetic and environmental factors in RA risk and progression. Most notably, studies have yielded evidence of significant interactions between SE and cigarette smoking in the development of anticitrullinated protein antibody (ACPA)-positive RA [2,3], although the precise mechanisms underpinning this interaction are not understood. Genetic and environmental factors that mediate oxidative stress, including cigarette smoking, are postulated to play a central role in the pathogenesis of autoimmune disorders including RA. While oxidative stress represents a form of host defense, it can also result in tissue damage. Oxidative modification of proteins and other biologic molecules leads to the expression of neoantigens, a possible first step in the development of autoimmunity, which may herald the future onset of clinically relevant autoimmune disease [4]. Antioxidants, which mitigate tissue damage caused by reactive oxygen species, may serve important protective functions in RA. While not all studies have identified a similar protective effect [4,5], the dietary intake of small-molecule antioxidants has been reported to be inversely associated with RA risk [6-9]. Additionally, low circulating levels of antioxidants have been reported to portend the onset of RA [10]. In addition to the effects of exogenous antioxidants, oxidation is also regulated by several enzymes, including glutathione S-transferase (GST). A ubiquitous cytosolic protein, GST catalyzes the conjugation of glutathione to a variety of substrates, including reactive oxygen species and other toxins, facilitating their elimination. Four classes of GST have been identified: a, μ, π and θ. Approximately one half of all individuals of European ancestry are homozygous for a deletion at the GST Mu1 (GSTM1) locus (GSTM1-null) [11] located on chromosome 1 (1p13.1). The GSTM1-null genotype has been associated with an increased risk of RA and in most [12-14] but not all [15] case control studies. In addition to being implicated as a potential risk factor in RA, the GSTM1-null genotype is associated with higher levels of oxidative stress [16] and has been reported to be a risk factor for other smoking-related inflammatory diseases, including asthma, emphysema and atherosclerosis [17-21]. However, there have been no studies examining associations of GSTM1 genotypes with ACPA expression in patients with RA. This represents an important knowledge gap, since these antibodies are disease-specific, have significant prognostic and pathogenic significance and are increasingly recognized to characterize a unique subset of patients with RA [22,23]. In the present study, we have evaluated potential gene-gene interactions by exploring the GSTM1-null genotype as a risk factor for ACPA positivity in RA, providing evidence of an interaction with HLA-DRB1 shared epitope (SE)-containing alleles. Materials and methods