Abstract
Objective.To assess the potential association between the rs1343151 IL23R and the rs3790567 IL12RB2 polymorphisms and giant cell arteritis (GCA). We also studied whether these polymorphisms might influence the phenotypic expression of GCA.
Methods.In total, 357 Spanish patients with biopsy-proven GCA and 574 matched controls were assessed. DNA from patients and controls was obtained from peripheral blood. Samples were genotyped for the rs1343151 IL23R and the rs3790567 IL12RB2 polymorphisms using a predesigned TaqMan allele discrimination assay and by polymerase chain reaction amplification.
Results.Regarding the rs1343151 IL23R polymorphism, no significant differences in the genotype or allele frequencies between GCA patients and healthy controls were observed. The frequency of the minor allele A of the rs3790567 IL12RB2 variant was increased in GCA patients compared with controls (30.1% vs 25.7%, respectively; p = 0.039, OR 1.25, 95% CI 1.01–1.54). An increased frequency of subjects carrying the minor allele A (GA+AA genotypes) of the rs3790567 IL12RB2 polymorphism was found among GCA patients compared with controls (52.8% vs 44.4%; p = 0.013, OR 1.40, 95% CI 1.06–1.85). Although a higher frequency of the combination of minor alleles (A-A) in the subgroup of patients with visual ischemic complications compared with the combination of both major alleles (G-G; p = 0.029) or with the other allelic combinations (p = 0.035) was found, logistic regression analysis showed that this association was no longer significant after adjustment for potential confounding factors (A-A vs G-G: OR 2.10, 95% CI 0.88–5.04, p = 0.096).
Conclusion.Our results support a potential influence of the rs3790567 IL12RB2 polymorphism in the pathogenesis of GCA.
Giant cell arteritis (GCA) is the most common primary systemic vasculitis in the elderly in Western countries1. It is also a complex polygenic disease2. CD4 T cells are the dominant cell population in the vascular lesions3, with Th1 and Th17 cells playing a role in pathogenesis of GCA4. IL-12 and IL-23 drive the development of naïve T cells into either one of those Th cells. Both molecules share a receptor subunit (IL-12Rß1). For proper signaling, IL-12 also has to bind to IL-12Rß2 and IL-23 to IL-23R5,6, leading to different cellular responses. Both receptors are expressed in T cells; therefore, the responsiveness to either cytokine is determined by their respective expression6.
The coding genes for both these receptor subunits are located in chromosome 1, less than 50 kb apart. Both have been associated with different autoimmune and inflammatory diseases7,8,9.
The aim of our study was to analyze the potential influence of 2 polymorphisms, the rs1343151 variant in the IL23R gene and the rs3790567 variant in the IL12RB2 gene, isolated or in combination, in susceptibility to biopsy-proven GCA and in the risk of ischemic GCA-related manifestations.
MATERIALS AND METHODS
Patients
We recruited 357 Spanish patients with a positive temporal artery biopsy10 who fulfilled the 1990 American College of Rheumatology classification criteria for GCA11 from Departments of Rheumatology or Internal Medicine from 5 Spanish cities: Lugo, Madrid, L’Hospitalet de Llobregat, Sabadell, and Granada. A control population of 574 healthy controls from the corresponding cities matched by age and sex with GCA patients was also assessed. Approval from the local ethical committees and written informed consent from patients and controls were obtained.
Genotyping methods
DNA from patients and controls was obtained from peripheral blood, using standard methods. Samples were genotyped for the rs1343151 IL23R and the rs3790567 IL12RB2 polymorphisms using a TaqMan 5’ allele discrimination assay (Applied Biosystems, Foster City, CA, USA) following manufacturer’s instructions.
Duplicate samples and negative controls were included. The genotyping success rate in the healthy control group was 100% for both polymorphisms, in the GCA group, 98.6% (352/357) for the rs1343151 IL23R polymorphism and 97.5% (348/357) for the rs3790567 IL12RB2 polymorphism.
Statistical analysis
Genotype data were checked for deviation from Hardy-Weinberg equilibrium using the program http://ihg.gsf.de/cgi-bin/hw/hwa1.pl (Institute of Human Genetics, Newcastle University, Newcastle upon Tyne, UK). Linkage disequilibrium (LD) values (D’, r2) and allelic combinations were generated using UNPHASED software15. Only those allelic combinations with a frequency in healthy controls higher than 5% were analyzed. The chi-square or Fisher tests were used to compare allelic, genotypic, or allelic combination distributions. Odds ratios (OR) and 95% confidence intervals (95% CI) were calculated according to Woolf’s method. Multiple logistic regression was used to adjust the association between GCA-related ischemic manifestations and alleles, genotypes, or allelic combinations of both polymorphisms, by sex, age at GCA diagnosis, and traditional cardiovascular risk factors as potential confounders.
RESULTS
Clinical features of patients with GCA are summarized in Table 1. No evidence of departure from Hardy-Weinberg equilibrium was observed in controls or GCA patients. The power of this study for finding a difference between GCA patients and controls with an estimated OR between 1.5 and 2.0, a type I error rate of 0.05, a dominant inheritance mode, and 0.0001% of population risk, was 84.5% to 99.9% for the rs3790567 IL12RB2 and 81.6% to 99.7% for the rs1343151 IL23R variant. No differences in genotype or allele distribution, ethnicity, or age at diagnosis were observed among the GCA patients and controls of the different study centers. However, a higher percentage of females with biopsy-proven GCA were recruited in Madrid compared with the other centers (83% vs 61%, respectively; p < 0.01). The 2 polymorphisms were not in LD (D’ = 0.21, r2 = 0.033).
Influence of rs1343151 IL23R and rs3790567 IL12RB2 polymorphisms in susceptibility to GCA
Regarding the rs3790567 IL12RB2 polymorphism, a higher frequency of the minor allele A (30.1% vs 25.7%; p = 0.039, OR 1.25, 95% CI 1.01–1.54) and carriers of the minor allele (GA+AA: 52.8% vs 44.4%, p = 0.013, OR 1.40, 95% CI 1.06–1.85) was observed in GCA patients compared with controls, respectively (Table 2). No significant differences were observed in the distribution of the rs1343151 IL23R.
Influence of rs1343151 IL23R and rs3790567 IL12RB2 polymorphisms in the risk of GCA-related ischemic manifestations
As shown in Table 3, a higher frequency of the combination of minor alleles (A-A) in the subgroup of patients with visual ischemic complications compared with the combination of both major alleles (G-G; p = 0.029, OR 2.10, 95% CI 1.01–4.35) or with the other allelic combinations (p = 0.035, OR 1.99, 95% CI 0.99–3.96) was found. However, no significant differences were observed in the adjusted analysis. In this regard, the association between the allelic combination and visual ischemic manifestations remained out of the range of significance (A-A vs G-G: OR 2.10, 95% CI 0.88–5.04, p = 0.096).
DISCUSSION
In this study we assessed for the first time the contribution of the rs1343151 IL23R and rs3790567 IL12RB2 polymorphisms, isolated and in combination, in susceptibility to biopsy-proven GCA. Our results support a potential role of the rs3790567 IL12RB2 polymorphism, but not the rs1343151 IL23R variant, in susceptibility to this condition. Although both Th1 and Th17 cells seem to play a role in GCA4, our results may indicate a more important role of Th1 cells in initiation of this condition.
The potential association between allelic combination and visual ischemic complications observed in this study might suggest a potential collaborative effect of both Th1 and Th17 cells in the development of these manifestations. However, this association was lost after adjustment for demographic and classic cardiovascular risk factors.
Although our cohort encompassed the largest series of patients with biopsy-proven GCA assessed in a genetic study on GCA and this vasculitis is a relatively uncommon condition, we are aware of the potential limitation of the study related to the sample size. Therefore, further studies are required to fully characterize the clinical significance of our findings and the contribution of these polymorphisms to the pathogenesis of GCA.
Acknowledgment
We thank Sofia Vargas, Gema Robledo, and Sonia García Ruíz for collection, isolation, and storage of the DNA samples. We also thank Sara Abel Liz, Maria Soledad Folgosa Rodriguez, and Ana Maria Ramos Gandoy, nurses from the Rheumatology Division, Hospital Xeral-Calde, Lugo, Spain, for help in collection of the samples. We thank all the patients and controls for their selfless collaboration.
Footnotes
-
Supported by grants PI06-0024 and PS09/00748 from Fondo de Investigaciones Sanitarias (Spain); and by RETICS Program, RD08/0075 (RIER) from Instituto de Salud Carlos III (ISCIII).
- Accepted for publication December 6, 2010.