The presence of oligoclonal bands (OCB) in cerebrospinal fluid (CSF) is a typical finding in multiple sclerosis (MS). We applied data from Norwegian, Swedish and Danish (i.e. Scandinavian) MS patients from a genome-wide association study (GWAS) to search for genetic differences in MS relating to OCB status. GWAS data was compared in 1367 OCB positive and 161 OCB negative Scandinavian MS patients, and nine of the most associated SNPs were genotyped for replication in 3403 Scandinavian MS patients. HLA-DRB1 genotypes were analyzed in a subset of the OCB positive (n = 2781) and OCB negative (n = 292) MS patients and compared to 890 healthy controls. Results from the genome-wide analyses showed that single nucleotide polymorphisms (SNPs) from the HLA complex and six other loci were associated to OCB status. In SNPs selected for replication, combined analyses showed genome-wide significant association for two SNPs in the HLA complex; rs3129871 (p = 5.7×10−15) and rs3817963 (p = 5.7×10−10) correlating with the HLA-DRB1*15 and the HLA-DRB1*04 alleles, respectively. We also found suggestive association to one SNP in the Calsyntenin-2 gene (p = 8.83×10−7). In HLA-DRB1 analyses HLA-DRB1*15∶01 was a stronger risk factor for OCB positive than OCB negative MS, whereas HLA-DRB1*04∶04 was associated with increased risk of OCB negative MS and reduced risk of OCB positive MS. Protective effects of HLA-DRB1*01∶01 and HLA-DRB1*07∶01 were detected in both groups. The groups were different with regard to age at onset (AAO), MS outcome measures and gender. This study confirms both shared and distinct genetic risk for MS subtypes in the Scandinavian population defined by OCB status and indicates different clinical characteristics between the groups. This suggests differences in disease mechanisms between OCB negative and OCB positive MS with implications for patient management, which need to be further studied.
Published work assessing psychosocial stress (job strain) as a risk factor for coronary heart disease is inconsistent and subject to publication bias and reverse causation bias. We analysed the relation between job strain and coronary heart disease with a meta-analysis of published and unpublished studies.
We used individual records from 13 European cohort studies (1985–2006) of men and women without coronary heart disease who were employed at time of baseline assessment. We measured job strain with questions from validated job-content and demand-control questionnaires. We extracted data in two stages such that acquisition and harmonisation of job strain measure and covariables occurred before linkage to records for coronary heart disease. We defined incident coronary heart disease as the first non-fatal myocardial infarction or coronary death.
30 214 (15%) of 197 473 participants reported job strain. In 1·49 million person-years at risk (mean follow-up 7·5 years [SD 1·7]), we recorded 2358 events of incident coronary heart disease. After adjustment for sex and age, the hazard ratio for job strain versus no job strain was 1·23 (95% CI 1·10–1·37). This effect estimate was higher in published (1·43, 1·15–1·77) than unpublished (1·16, 1·02–1·32) studies. Hazard ratios were likewise raised in analyses addressing reverse causality by exclusion of events of coronary heart disease that occurred in the first 3 years (1·31, 1·15–1·48) and 5 years (1·30, 1·13–1·50) of follow-up. We noted an association between job strain and coronary heart disease for sex, age groups, socioeconomic strata, and region, and after adjustments for socioeconomic status, and lifestyle and conventional risk factors. The population attributable risk for job strain was 3·4%.
Our findings suggest that prevention of workplace stress might decrease disease incidence; however, this strategy would have a much smaller effect than would tackling of standard risk factors, such as smoking.
Finnish Work Environment Fund, the Academy of Finland, the Swedish Research Council for Working Life and Social Research, the German Social Accident Insurance, the Danish National Research Centre for the Working Environment, the BUPA Foundation, the Ministry of Social Affairs and Employment, the Medical Research Council, the Wellcome Trust, and the US National Institutes of Health.
The genetic association of the major histocompatibility complex (MHC) to rheumatoid arthritis risk has commonly been attributed to HLA-DRB1 alleles. Yet controversy persists about the causal variants in HLA-DRB1 and the presence of independent effects elsewhere in the MHC. Using existing genome-wide SNP data in 5,018 seropositive cases and 14,974 controls, we imputed and tested classical alleles and amino acid polymorphisms for HLA-A, B, C, DPA1, DPB1, DQA1, DQB1, and DRB1 along with 3,117 SNPs across the MHC. Conditional and haplotype analyses reveal that three amino acid positions (11, 71 and 74) in HLA-DRβ1, and single amino acid polymorphisms in HLA-B (position 9) and HLA-DPβ1 (position 9), all located in the peptide-binding grooves, almost completely explain the MHC association to disease risk. This study illustrates how imputation of functional variation from large reference panels can help fine-map association signals in the MHC.
Microsomal PGE synthase 1 (mPGES-1) is the terminal enzyme in the induced state of prostaglandin E2 (PGE2) synthesis and constitutes a therapeutic target for rheumatoid arthritis (RA) treatment. We examined the role of the prostaglandin E synthase (PTGES) gene polymorphism in susceptibility to and severity of RA and related variations in the gene to its function. The PTGES gene polymorphism was analyzed in 3081 RA patients and 1900 controls from two study populations: Swedish Epidemiological Investigation of Rheumatoid Arthritis (EIRA) and the Leiden Early Arthritis Clinic (Leiden EAC). Baseline disease activity score (DAS28) was employed as a disease severity measure. mPGES-1 expression was analyzed in synovial tissue from RA patients with known genotypes using immunohistochemistry. In the Swedish study population, among women a significant association with risk for RA was observed for PTGES single-nucleotide polymorphisms (SNPs) in univariate analysis and for the distinct haplotype. These results were substantiated by meta-analysis of data from EIRA and Leiden EAC studies with overall OR 1.31 (95% confidence interval 1.11–1.56). Several PTGES SNPs were associated with earlier onset of disease or with higher DAS28 in women with RA. Patients with the genotype associated with higher DAS28 exhibited significantly higher mPGES-1 expression in synovial tissue. Our data reveal a possible influence of PTGES polymorphism on the pathogenesis of RA and on disease severity through upregulation of mPGES-1 at the sites of inflammation. Genetically predisposed individuals may develop earlier and more active disease owing to this mechanism.
mPGES-1; gene polymorphism; rheumatoid arthritis; gender
Tobacco smoking is a major contributor to the public health burden and healthcare costs worldwide, but the determinants of smoking behaviours are poorly understood. We conducted a large individual-participant meta-analysis to examine the extent to which work-related stress, operationalised as job strain, is associated with tobacco smoking in working adults.
Methodology and Principal Findings
We analysed cross-sectional data from 15 European studies comprising 166 130 participants. Longitudinal data from six studies were used. Job strain and smoking were self-reported. Smoking was harmonised into three categories never, ex- and current. We modelled the cross-sectional associations using logistic regression and the results pooled in random effects meta-analyses. Mixed effects logistic regression was used to examine longitudinal associations. Of the 166 130 participants, 17% reported job strain, 42% were never smokers, 33% ex-smokers and 25% current smokers. In the analyses of the cross-sectional data, current smokers had higher odds of job strain than never-smokers (age, sex and socioeconomic position-adjusted odds ratio: 1.11, 95% confidence interval: 1.03, 1.18). Current smokers with job strain smoked, on average, three cigarettes per week more than current smokers without job strain. In the analyses of longitudinal data (1 to 9 years of follow-up), there was no clear evidence for longitudinal associations between job strain and taking up or quitting smoking.
Our findings show that smokers are slightly more likely than non-smokers to report work-related stress. In addition, smokers who reported work stress smoked, on average, slightly more cigarettes than stress-free smokers.
The relationship between work-related stress and alcohol intake is uncertain. In order to add to the thus far inconsistent evidence from relatively small studies, we conducted individual-participant meta-analyses of the association between work-related stress (operationalised as self-reported job strain) and alcohol intake.
Methodology and Principal Findings
We analysed cross-sectional data from 12 European studies (n = 142 140) and longitudinal data from four studies (n = 48 646). Job strain and alcohol intake were self-reported. Job strain was analysed as a binary variable (strain vs. no strain). Alcohol intake was harmonised into the following categories: none, moderate (women: 1–14, men: 1–21 drinks/week), intermediate (women: 15–20, men: 22–27 drinks/week) and heavy (women: >20, men: >27 drinks/week). Cross-sectional associations were modelled using logistic regression and the results pooled in random effects meta-analyses. Longitudinal associations were examined using mixed effects logistic and modified Poisson regression. Compared to moderate drinkers, non-drinkers and (random effects odds ratio (OR): 1.10, 95% CI: 1.05, 1.14) and heavy drinkers (OR: 1.12, 95% CI: 1.00, 1.26) had higher odds of job strain. Intermediate drinkers, on the other hand, had lower odds of job strain (OR: 0.92, 95% CI: 0.86, 0.99). We found no clear evidence for longitudinal associations between job strain and alcohol intake.
Our findings suggest that compared to moderate drinkers, non-drinkers and heavy drinkers are more likely and intermediate drinkers less likely to report work-related stress.
Multiple sclerosis (MS) is a complex disease of the central nervous system of unknown etiology. The human leukocyte antigen (HLA) locus on chromosome 6 confers a considerable part of the susceptibility to MS, and the most important factor is the class II allele HLA-DRB1*15:01. In addition, we and others have previously established a protective effect of HLA-A*02. Here, we genotyped 1,784 patients and 1,660 healthy controls from Scandinavia for the HLA-A, HLA-B, HLA-C and HLA-DRB1 genes and investigated their effects on MS risk by logistic regression. Several allele groups were found to exert effects independently of DRB1*15 and A*02, in particular DRB1*01 (OR = 0.82, p = 0.034) and B*12 (including B*44/45, OR = 0.76, p = 0.0028), confirming previous reports. Furthermore, we observed interaction between allele groups: DRB1*15 and DRB1*01 (multiplicative: OR = 0.54, p = 0.0041; additive: AP = 0.47, p = 4×10−06), DRB1*15 and C*12 (multiplicative: OR = 0.37, p = 0.00035; additive: AP = 0.58, p = 2.6×10−05), indicating that the effect size of these allele groups varies when taking DRB1*15 into account. Analysis of inferred haplotypes showed that almost all DRB1*15 bearing haplotypes were risk haplotypes, and that all A*02 bearing haplotypes were protective as long as they did not carry DRB1*15. In contrast, we found one class I haplotype, carrying A*02-C*05-B*12, which abolished the risk of DRB1*15. In conclusion, these results confirms a complex role of HLA class I and II genes that goes beyond DRB1*15 and A*02, in particular by including all three classical HLA class I genes as well as functional interactions between DRB1*15 and several alleles of DRB1 and class I genes.
Rheumatoid arthritis (RA) is a multifactorial autoimmune disease in which genetic and environmental factors interact in the etiology. In this study, we investigated whether smoking and HLA-DRB1 shared-epitope (SE) alleles interact differently in the development of the two major subgroups of rheumatoid arthritis (RA), anti-citrullinated proteins antibody (ACPA)-positive and ACPA-negative disease, in a multiethnic population of Asian descent.
A case-control study comprising early diagnosed RA cases was carried out in Malaysia between 2005 and 2009. In total, 1,076 cases and 1,612 matched controls participated in the study. High-resolution HLA-DRB1 genotyping was performed for shared-epitope (SE) alleles. All participants answered a questionnaire on a broad range of issues, including smoking habits. The odds ratio (OR) of developing ACPA-positive and ACPA-negative disease was calculated for smoking and the presence of any SE alleles separately. Potential interaction between smoking history (defined as "ever" and "never" smoking) and HLA-DRB1 SE alleles also was calculated.
In our multiethnic study, both the SE alleles and smoking were associated with an increased risk of developing ACPA-positive RA (OR SE alleles, 4.7; 95% confidence interval (CI), 3.6 to 6.2; OR smoking, 4.1; 95% CI, 1.9 to 9.2). SE-positive smokers had an odds ratio of ACPA-positive RA of 25.6 (95% CI, 10.4 to 63.4), compared with SE-negative never-smokers. The interaction between smoking and SE alleles was significant (attributable proportion due to interaction (AP) was 0.7 (95% CI, 0.5 to 1.0)). The HLA-DRB1*04:05 SE allele, which is common in Asian populations, but not among Caucasians, was associated with an increased risk of ACPA-positive RA, and this allele also showed signs of interaction with smoking (AP, 0.4; 95% CI, -0.1 to 0.9). Neither smoking nor SE alleles nor their combination was associated with an increased risk of ACPA-negative RA.
The risk of developing ACPA-positive RA is associated with a strong gene-environment interaction between smoking and HLA-DRB1 SE alleles in a Malaysian multiethnic population of Asian descent. This interaction seems to apply also between smoking and the specific HLA-DRB1*04:05 SE allele, which is common in Asian populations but not in Caucasians.
HLA-DRB1 shared epitope (SE) alleles are the strongest genetic determinants for autoantibody positive rheumatoid arthritis (RA). One of the key regulators in expression of HLA class II receptors is MHC class II transactivator (CIITA). A variant of the CIITA gene has been found to associate with inflammatory diseases.
We wanted to explore whether the risk variant rs3087456 in the CIITA gene interacts with the HLA-DRB1 SE alleles regarding the risk of developing RA. We tested this hypothesis in a case-control study with 11767 individuals from four European Caucasian populations (6649 RA cases and 5118 controls).
We found no significant additive interaction for risk alleles among Swedish Caucasians with RA (n = 3869, attributable proportion due to interaction (AP) = 0.2, 95%CI: −0.2–0.5) or when stratifying for anti-citrullinated protein antibodies (ACPA) presence (ACPA positive disease: n = 2945, AP = 0.3, 95%CI: −0.05–0.6, ACPA negative: n = 2268, AP = −0.2, 95%CI: −1.0–0.6). We further found no significant interaction between the main subgroups of SE alleles (DRB1*01, DRB1*04 or DRB1*10) and CIITA. Similar analysis of three independent RA cohorts from British, Dutch and Norwegian populations also indicated an absence of significant interaction between genetic variants in CIITA and SE alleles with regard to RA risk.
Our data suggest that risk from the CIITA locus is independent of the major risk for RA from HLA-DRB1 SE alleles, given that no significant interaction between rs3087456 and SE alleles was observed. Since a biological link between products of these genes is evident, the genetic contribution from CIITA and class II antigens in the autoimmune process may involve additional unidentified factors.
Earlier studies have demonstrated that smoking and genetic risk factors interact in providing an increased risk for Rheumatoid Arthritis (RA). Less is known on how smoking contributes to RA in the context of genetic variability, and what proportion of RA that may be caused by smoking.
To determine the association between amount of smoking and risk of RA in the context of different HLA-DRB1 shared epitope (SE) alleles, and to estimate proportions of RA cases attributed to smoking.
Design, Setting, and Participants
Data from the Swedish Epidemiological Investigation of Rheumatoid Arthritis (EIRA) case-control study encompassing 1204 cases and 871 controls were analysed.
Main Outcome Measure
Estimated odds ratio to develop RA and excess fraction of cases attributable to smoking according to amount of smoking and genotype.
Smoking was estimated to be responsible for 35 % of the ACPA+ cases. For each HLA-DRB1 SE genotype, smoking was dose-dependently associated with increased risk of ACPA+ RA (p-trend<0.001). In individuals carrying two copies of the HLA-DRB1 shared epitope, 55 % of ACPA-positive RA were attributable to smoking.
Smoking is a preventable risk factor for RA. The increased risk due to smoking is dependent on amount of smoking and genotype.
Rheumatoid Arthritis; Smoking; HLA-DRB1; Case-control studies; Epidemiology
The aim of this study was to investigate the association between exposure to cosmetics, often containing mineral oil, and the risk of developing rheumatoid arthritis (RA). The study was performed against the background that occupational exposure to mineral oil has recently been shown to be associated with an increased risk for RA in man, and that injection of or percutaneous exposure to mineral-oil-containing cosmetics can induce arthritis in certain rat strains.
A population-based case-control study of incident cases of RA was performed among the population aged 18 to 70 years in a defined area of Sweden during May 1996 to December 2003. A case was defined as an individual from the study base, who received for the first time a diagnosis of RA according to the 1987 American College of Rheumatology criteria. Controls were randomly selected from the study base with consideration taken for age, gender and residential area. Cases (n = 1,419) and controls (n = 1,674) answered an extensive questionnaire regarding environmental and lifestyle factors including habits of cosmetic usage. The relative risk of developing RA was calculated for subjects with different cosmetic usage compared with subjects with low or no usage. Analysis was also performed stratifying the cases for presence/absence of rheumatoid factor and antibodies to citrulline-containing peptides.
The relative risks of developing RA associated with use of cosmetics were all close to one, both for women and men, for different exposure categories, and in relation to different subgroups of RA.
This study does not support the hypothesis that ordinary usage of common cosmetics as body lotions, skin creams, and ointments, often containing mineral oil, increase the risk for RA in the population in general. We cannot exclude, however, that these cosmetics can contribute to arthritis in individuals carrying certain genotypes or simultaneously being exposed to other arthritis-inducing environmental agents.
Job strain (i.e., high job demands combined with low job control) is a frequently used indicator of harmful work stress, but studies have often used partial versions of the complete multi-item job demands and control scales. Understanding whether the different instruments assess the same underlying concepts has crucial implications for the interpretation of findings across studies, harmonisation of multi-cohort data for pooled analyses, and design of future studies. As part of the 'IPD-Work' (Individual-participant-data meta-analysis in working populations) consortium, we compared different versions of the demands and control scales available in 17 European cohort studies.
Six of the 17 studies had information on the complete scales and 11 on partial scales. Here, we analyse individual level data from 70 751 participants of the studies which had complete scales (5 demand items, 6 job control items).
We found high Pearson correlation coefficients between complete scales of job demands and control relative to scales with at least three items (r > 0.90) and for partial scales with two items only (r = 0.76-0.88). In comparison with scores from the complete scales, the agreement between job strain definitions was very good when only one item was missing in either the demands or the control scale (kappa > 0.80); good for job strain assessed with three demand items and all six control items (kappa > 0.68) and moderate to good when items were missing from both scales (kappa = 0.54-0.76). The sensitivity was > 0.80 when only one item was missing from either scale, decreasing when several items were missing in one or both job strain subscales.
Partial job demand and job control scales with at least half of the items of the complete scales, and job strain indices based on one complete and one partial scale, seemed to assess the same underlying concepts as the complete survey instruments.
Job demands; Job control; Job strain; Work stress; Agreement
Multiple sclerosis, MS (OMIM No. 126200), is a complex inflammatory disease that is characterized by lesions in the central nervous system. Both genes and other environmental factors influence disease susceptibility. One of the environmental factors that has been implicated in MS and autoimmune disease, such as type 1 diabetes, is vitamin D deficiency, in which patients have lower levels of 25-hydroxyvitamin D3 (25-OHD3) in blood than do controls. Previtamin D3 is produced in the skin, and turned into 25-OHD3 in the liver. In the kidney, skin and immune cells, 25-OHD3 is turned into bioactive 1,25(OH)2D3 by the enzyme coded by CYP27B1 (cytochrome P450 family 27 subfamily B peptide 1) on chromosome 12q13.1–3. 1,25(OH)2D3 binds to the vitamin D receptor, expressed in T cells and antigen-presenting cells. 1,25(OH)2D3 has a suppressive role in the adaptive immune system, decreasing T-cell and dendritic cell maturation, proliferation and differentiation, shifting the balance between T-helper 1 (Th1) and Th2 cells in favor of Th2 cells and increasing the suppressive function of regulatory T cells. Rs703842 in the 12q13–14 region was associated with MS in a recent study by the Australian and New Zealand Multiple Sclerosis Genetics Consortium (ANZgene). We show associations with three SNPs in this region in our Swedish materials (2158 cases, 1759 controls) rs4646536, rs10877012 and rs10877015 (P=0.01, 0.01 and 3.5 × 10−3, respectively). We imputed rs703842 SNP and performed a joint analysis with the ANZgene results, reaching a significant association for rs703842 (P=5.1 × 10−11; odds ratio 0.83; 95% confidence interval 0.79–0.88). Owing to its close association with 25-OHD3, our results lend further support to the role of vitamin D in MS pathology.
multiple sclerosis; CYP27B1; association study; vitamin D; case–control study; meta-analysis
Glutathione S-transferase (GST) and heme oxygenase-1 (HMOX1) genes encode enzymes that detoxify carcinogens and protect against oxidative stress. We studied gene-smoking interactions on rheumatoid arthritis (RA) susceptibility.
549 matched Caucasian RA cases and controls were selected from the Nurses' Health Study. Genotyping by TaqMan and BioTrove identified GSTM1, GSTT1 homozygous deletions (null) and GSTP1 (rs1695), HMOX1 (rs2071746) alleles, respectively. We studied gene-smoking interactions on the risk of all RA and serologic RA phenotypes in separate logistic models adjusted for covariates. We assessed multiplicative interactions with product terms in the logistic models and additive interactions with the attributable proportion due to interaction (AP). For replication, we repeated significant analyses in an independent case-control sample from the Epidemiologic Investigation of RA.
For all RA risk, we observed: multiplicative (p=0.05) and additive (AP=0.53, p=0.0005) interactions between GSTT1-null and smoking and multiplicative interaction (p=0.05) between HMOX1 and smoking. For seropositive RA risk, we found multiplicative (p=0.01) and additive (AP=0.63, p<0.0001) interactions between GSTT1-null and smoking and additive interaction (AP=0.41, p=0.03) between HMOX1 and smoking. After correction for multiple comparisons, additive interactions for GSTT1-null and smoking remained significant. GSTM1-null and GSTP1 did not show significant interactions. No associations were seen with seronegative RA. In replication analyses, we observed significant multiplicative (p=0.04) and additive (AP=0.32, p=0.02) interactions between GSTT1-null and smoking for ACPA-positive RA risk.
We observed significant gene-environment interactions between GSTT1-null and heavy smoking on RA risk. Future studies are needed to assess the impact of these interactions on RA prediction.
To discover novel RA risk loci, we systematically examined 370 SNPs from 179 independent loci with p<0.001 in a published meta-analysis of RA GWAS of 3,393 cases and 12,462 controls1. We used GRAIL2, a computational method that applies statistical text mining to PubMed abstracts, to score these 179 loci for functional relationships to genes in 16 established RA disease loci1,3-11. We identified 22 loci with a significant degree of functional connectivity. We genotyped 22 representative SNPs in an independent set of 7,957 cases and 11,958 matched controls. Three validate convincingly: CD2/CD58 (rs11586238, p=1×10−6 replication, p=1×10−9 overall), and CD28 (rs1980422, p=5×10−6 replication, p=1×10−9 overall), PRDM1 (rs548234, p=1×10−5 replication, p=2×10−8 overall). An additional four replicate (p<0.0023): TAGAP (rs394581, p=0.0002 replication, p=4×10−7 overall), PTPRC (rs10919563, p=0.0003 replication, p=7×10−7 overall), TRAF6/RAG1 (rs540386, p=0.0008 replication, p=4×10−6 overall), and FCGR2A (rs12746613, p=0.0022 replication, p=2×10−5 overall). Many of these loci are also associated to other immunologic diseases.
It has repeatedly been suggested that the development of complex diseases can be elucidated by gene–gene interactions. Recently, we found that HTR2A, a member of the serotonin receptor family, is associated with rheumatoid arthritis (RA). This study was aimed to investigate the possibility of a gene–gene interaction between HTR2A and the major genetic risk factor for RA, HLA-DRB1 shared epitope (SE) alleles. We studied 4095 RA cases and 3223 controls from three different populations – from Sweden, the United States and the Netherlands – to test for interaction between the protective HTR2A haplotype and HLA-DRB1 SE alleles. Further, we analyzed mRNA and/or protein expression of HTR2A and HLA-DR in biopsy samples and in synovial fibroblasts from RA patients. The interaction was defined as departure from additivity of effects using attributable proportion due to interaction. First, we could demonstrate and further replicate an interaction between a protective haplotype in HTR2A and HLA-DRB1 SE alleles regarding risk of developing autoantibody-positive RA. Second, we could show that both genes are constitutively expressed in fibroblasts from synovial tissue of RA patients, and, by double immunofluorescence staining, we demonstrated that these two proteins are colocalized in these cells. In conclusion, our data demonstrate a statistical interaction between HTR2A and HLA-DRB1 SE alleles and colocalization of the product of these two genes in inflamed synovial tissue, which suggest a possible biological relationship between these two proteins. This finding may lead to the development of treatment based on enhancing the protective features of 5-HT2A in individuals with a certain HLA genotype.
serotonin receptor; MHC class II; RA; gene–gene interaction
To investigate the associations between HLA-DRB1 shared epitope (SE) alleles and rheumatoid arthritis in subsets of rheumatoid arthritis defined by autoantibodies in three Asian populations from Malaysia.
1,079 rheumatoid arthritis patients and 1,470 healthy controls were included in the study. Levels of antibodies to citrullinated proteins (ACPA) and rheumatoid factors were assessed and the PCR-SSO method was used for HLA-DRB1 genotyping.
The proportion of ACPA positivity among Malay, Chinese and Indian rheumatoid arthritis patients were 62.9%, 65.2% and 68.6%, respectively. An increased frequency of SE alleles was observed in ACPA-positive rheumatoid arthritis among the three Asian ethnic groups. HLA-DRB1*10 was highly associated with rheumatoid arthritis susceptibility in these Asian populations. HLA-DRB1*0405 was significantly associated with susceptibility to rheumatoid arthritis in Malays and Chinese, but not in Indians. HLA-DRB1*01 did not show any independent effect as a risk factor for rheumatoid arthritis in this study and HLA-DRB1*1202 was protective in Malays and Chinese. There was no association between SE alleles and ACPA- negative rheumatoid arthritis in any of the three Asian ethnic groups.
The HLA-DRB1 SE alleles increase the risk of ACPA-positive rheumatoid arthritis in all three Asian populations from Malaysia.
Recent discoveries of risk alleles have made it possible to define genetic risk profiles for patients with rheumatoid arthritis (RA). We examined whether a cumulative score based on 22 validated genetic risk alleles for seropositive RA would identify high-risk, asymptomatic individuals who might benefit from preventive interventions.
We genotyped 14 single nucleotide polymorphisms (SNPs) at 13 validated RA risk loci and 8 HLA alleles among (1) 289 Caucasian seropositive cases and 481 controls from the US Nurses' Health Studies (NHS), and (2) 629 Caucasian CCP antibody positive cases and 623 controls from the Swedish Epidemiologic Investigation of RA (EIRA). We created a weighted genetic risk score (GRS), where the weight for each risk allele is the log of the published odds ratio. We used logistic regression to study associations with incident RA. We compared AUCs from a clinical-only model and clinical + genetic model in each cohort.
Patients with GRS > 1.25 standard deviations of the mean had a significantly higher OR of seropositive RA in both NHS (OR=2.9, 95%CI 1.8–4.6) and EIRA (OR=3.4, 95% CI 2.3–5.0) referent to the population average. In NHS, the AUC for a clinical model was 0.57 and for a clinical + genetic model was 0.66, and in EIRA was 0.63 and 0.75, respectively.
The combination of 22 risk alleles into a weighted genetic risk score significantly stratifies individuals for RA risk beyond clinical risk factors alone. However, given the low incidence of RA, the clinical utility of a weighted genetic risk score is limited in the general population.
rheumatoid arthritis; polymorphism; autoantibodies; anti-CCP; smoking
The GEIRA (Gene-Environment and Gene–Gene Interaction Research Application) algorithm and subsequent program is dedicated to genome-wide gene-environment and gene–gene interaction analysis. It implements concepts of both additive and multiplicative interaction as well as calculations based on dominant, recessive and co-dominant genetic models, respectively. Estimates of interactions are incorporated in a single table to make the output easily read. The algorithm is coded in both SAS and R. GEIRA is freely available to non-commercial users at http://www.epinet.se. Additional information, including user’s manual and example datasets is available online at http://www.epinet.se.
GWAS; Interaction; Program
Previous studies have provided inconsistent results on whether variants in the MBL2 gene, coding for the complement-activating mannan-binding lectin (MBL) protein, associate with rheumatoid arthritis (RA). We re-evaluated this in context of the main environmental and genetic risk factors (smoking, HLA-DRB1 'shared epitope' (SE), PTPN22*620W), which predispose to rheumatoid factor (RF) and/or anti-citrullinated-protein antibody (ACPA)-positive RA.
In this population-based EIRA study, rheumatoid factor (RF), ACPA, smoking, SE and PTPN22*620W status was determined in incident RA cases and matched controls. MBL-high (n = 1330) and MBL-low (n = 1257) genotypes predicting MBL levels were constructed from four promoter and exon-1 polymorphisms in the MBL2 gene. Odds ratios with 95% confidence interval (OR, 95% CI) were calculated by logistic regression. In extended families (n = 316), previously reported data were re-analyzed, considering RF and smoking.
MBL-high genotypes tended to be associated with RF-negative (OR = 1.20, 95% CI 0.96-1.51) but not RF-positive (OR = 1.00, 95% CI 0.83-1.20) RA. Results divided by ACPA status did not differ. When stratified for smoking, MBL-high genotype was strongly associated with RF-negative RA in never smokers (OR = 1.82, 95% CI 1.24-2.69) but not in ever smokers (OR = 0.96, 95% CI 0.73-1.30). In never smokers, the association was observed in both the RF-negative/ACPA-negative (OR = 1.67, 95% CI 1.10-2.55) and RF-negative/ACPA-positive subgroups (OR = 3.07, 95% CI 1.37-6.89), and remained on an SE/PTPN22*620W negative background. In the extended families, the reported association between high MBL and RA was in fact confined to never smokers.
High MBL may predispose to RF-negative RA but only in individuals who have never smoked. This illustrates the importance of phenotypic subgrouping in genetic studies.
Glutathione-S-transferases (GSTs) play an important role in tobacco smoke detoxification, interestingly approximately 50% of individuals in most human populations lack the gene GSTM1 due to copy number variation (CNV). We aimed to investigate GSTM1 CNV in Rheumatoid Arthritis (RA) in relation to smoking and HLA-DRB1 shared epitope; the two best known risk factors for RA and in addition, to perform subanalyses in patients where relations between variations in GSTM1 and RA have previously been described.
qPCR was performed using TaqMan Copy Number assays (Applied Biosystems) for 2426 incident RA cases and 1257 controls from the Swedish EIRA. Odds ratio (OR) together with 95% confidence intervals (CI) was calculated and used as a measure of the relative risk of developing RA.
No association between RA and GSTM1 CNV was observed when analyzing whole EIRA. However, ≥1 copy of GSTM1 appears to be a significant risk factor for autoantibody positive RA in non-smoking females ≥60 years (OR: 2.00 95% CI: 1.07–3.74), a population where such relationships have previously been described. Our data further suggest a protective effect of GSTM1 in ACPA-negative smoking men (OR: 0.56 95% CI: 0.35–0.90).
We assessed the exact number of GSTM1 gene copies in relation to development and severity of RA. Our data provide support for the notion that variations in copy numbers of GSTM1 may influence risk in certain subsets of RA, but do not support a role for GSTM1 CNV as a factor that more generally modifies the influence of smoking on RA.
Rheumatoid arthritis (RA) can be divided into two major subsets based on the presence or absence of antibodies to citrullinated peptide antigens (ACPA). Until now, data from genome-wide association studies (GWAS) have only been published from ACPA-positive subsets of RA or from studies that have not separated the two subsets. The aim of the current study is to provide and compare GWAS data for both subsets.
and results GWAS using the Illumina 300K chip was performed for 774 ACPA-negative patients with RA, 1147 ACPA-positive patients with RA and 1079 controls from the Swedish population-based case–control study EIRA. Imputation was performed which allowed comparisons using 1 723 056 single nucleotide polymorphisms (SNPs). No SNP achieved genome-wide significance (2.9 × 10–8) in the comparison between ACPA-negative RA and controls. A case–case association study was then performed between ACPA-negative and ACPA-positive RA groups. The major difference in this analysis was in the HLA region where 768 HLA SNPs passed the threshold for genome-wide significance whereas additional contrasting SNPs did not reach genome-wide significance. However, one SNP close to the RPS12P4 locus in chromosome 2 reached a p value of 2 × 106 and this locus can thus be considered as a tentative candidate locus for ACPA-negative RA.
ACPA-positive and ACPA-negative RA display significant risk allele frequency differences which are mainly confined to the HLA region. The data provide further support for distinct genetic aetiologies of RA subsets and emphasise the need to consider them separately in genetic as well as functional studies of this disease.
Objective To assess the risk of attempted suicide before, during, and after treatment with isotretinoin for severe acne.
Design Retrospective cohort study linking a named patient register of isotretinoin users (1980-9) to hospital discharge and cause of death registers (1980-2001).
Setting Sweden, 1980-2001.
Population 5756 patients aged 15 to 49 years prescribed isotretinoin for severe acne observed for 17 197 person years before, 2905 person years during, and 87 120 person years after treatment.
Main outcome measures Standardised incidence ratio (observed number divided by expected number of suicide attempts standardised by sex, age, and calendar year), calculated up to three years before, during, and up to 15 years after end of treatment.
Results 128 patients were admitted to hospital for attempted suicide. During the year before treatment, the standardised incidence ratio for attempted suicide was raised: 1.57 (95% confidence interval 0.86 to 2.63) for all (including repeat) attempts and 1.36 (0.65 to 2.50) counting only first attempts. The standardised incidence ratio during and up to six months after treatment was 1.78 (1.04 to 2.85) for all attempts and 1.93 (1.08 to 3.18) for first attempts. Three years after treatment stopped, the observed number of attempts was close to the expected number and remained so during the 15 years of follow-up: standardised incidence ratio 1.04 (0.74 to 1.43) for all attempts and 0.97 (0.64 to 1.40) for first attempts. Twelve (38%) of 32 patients who made their first suicide attempt before treatment made a new attempt or committed suicide thereafter. In contrast, 10 (71%) of the 14 who made their first suicide attempt within six months after treatment stopped made a new attempt or committed suicide during follow-up (two sample test of proportions, P=0.034). The number needed to harm was 2300 new six month treatments per year for one additional first suicide attempt to occur and 5000 per year for one additional repeat attempt.
Conclusions An increased risk of attempted suicide was apparent up to six months after the end of treatment with isotretinoin, which motivates a close monitoring of patients for suicidal behaviour for up to a year after treatment has ended. However, the risk of attempted suicide was already rising before treatment, so an additional risk due to the isotretinoin treatment cannot be established. As patients with a history of suicide attempts before treatment made new attempts to a lesser extent than did patients who started such behaviour in connection with treatment, patients with severe acne should not automatically have isotretinoin treatment withheld because of a history of attempted suicide.
Breast cancer is a heterogeneous disease and may be characterized on the basis of whether estrogen receptors (ER) are expressed in the tumour cells. ER status of breast cancer is important clinically, and is used both as a prognostic indicator and treatment predictor. In this study, we focused on identifying genetic markers associated with ER-negative breast cancer risk.
We conducted a genome-wide association analysis of 285,984 single nucleotide polymorphisms (SNPs) genotyped in 617 ER-negative breast cancer cases and 4,583 controls. We also conducted a genome-wide pathway analysis on the discovery dataset using permutation-based tests on pre-defined pathways. The extent of shared polygenic variation between ER-negative and ER-positive breast cancers was assessed by relating risk scores, derived using ER-positive breast cancer samples, to disease state in independent, ER-negative breast cancer cases.
Association with ER-negative breast cancer was not validated for any of the five most strongly associated SNPs followed up in independent studies (1,011 ER-negative breast cancer cases, 7,604 controls). However, an excess of small P-values for SNPs with known regulatory functions in cancer-related pathways was found (global P = 0.052). We found no evidence to suggest that ER-negative breast cancer shares a polygenic basis to disease with ER-positive breast cancer.
ER-negative breast cancer is a distinct breast cancer subtype that merits independent analyses. Given the clinical importance of this phenotype and the likelihood that genetic effect sizes are small, greater sample sizes and further studies are required to understand the etiology of ER-negative breast cancers.
Prior evidence demonstrates that the PRL -1149 T (minor) allele decreases prolactin expression and may be associated with autoimmune diseases. Our goal was to determine the role of the PRL -1149 G/T polymorphism (rs1341239) in rheumatoid arthritis (RA) susceptibility.
We examined the association between PRL -1149 G/T and RA risk in four separate collections consisting of 3,405 RA cases and 4,111 controls of self-described “white” European ancestry. Samples were genotyped using one of three genotyping platforms, and strict quality control metrics were applied. We tested for association with a two-tailed Cochran Mantel-Haenszel additive, fixed effects model.
In the individual collections, odds ratios for an association between PRL -1149 T and RA risk ranged from 0.80 to 0.97. In a joint meta-analysis across all four collections, the odds ratio for an association between PRL -1149 T and RA risk was 0.90 (95% CI 0.84-0.96) with p = 0.001.
Our results suggest an association between the PRL -1149 T allele and decreased RA risk. The effect size is small but similar to odds ratios for other genetic polymorphisms associated with complex traits, including RA. Further studies will be necessary to confirm association unequivocally.