Emilie Sundqvist

Smoking and two human leukocyte antigen genes interact to increase the risk for multiple sclerosis

Both genetic and environmental factors display low or modest associations with multiple sclerosis. Hypothetically, gene-environment interactions may exert much stronger effects. In this study, we investigated potential interactions between genetic risk factors and smoking in relation to risk of developing multiple sclerosis. A population-based case-control study involving incident cases of multiple sclerosis (843 cases, 1209 controls) was performed in Sweden. Cases and controls were classified according to their smoking status and human leukocyte antigen DRB1 as well as human leukocyte antigen A genotypes. Subjects with different genotypes and smoking habits were compared with regard to incidence of multiple sclerosis, by calculating odds ratios with 95% confidence intervals employing logistic regression. The potential interaction between different genotypes, as well as between genotype and smoking, was evaluated by calculating attributable proportion due to interaction. A significant interaction between two genetic risk factors, carriage of human leukocyte antigen DRB1*15 and absence of human leukocyte antigen A*02, was observed among smokers whereas such an interaction was absent among non-smokers. There were considerable differences in odds ratios between the various groups. Compared with non-smokers with neither of the genetic risk factors, the odds ratio was 13.5 (8.1-22.6) for smokers with both genetic risk factors. The odds ratio for smokers without genetic risk was 1.4 (0.9-2.1) and the odds ratio for non-smokers with both genetic risk factors was 4.9 (3.6-6.6). Among those with both genetic risk factors, smoking increased the risk by a factor of 2.8 in comparison with a factor of 1.4 among those without the genetic risk factors. The risk of developing multiple sclerosis associated with human leukocyte antigen genotypes may be strongly influenced by smoking status. The findings are consistent with our hypothesis that priming of the immune response in the lungs may subsequently lead to multiple sclerosis in genetically susceptible people.

Epstein-Barr virus and multiple sclerosis: interaction with HLA.

Epstein-Barr virus (EBV) infection, history of infectious mononucleosis (IM) and HLA-A and DRB1 have all been proposed as risk factors for multiple sclerosis (MS). Our aim was to analyse possible interactions between antibodies against Epstein-Barr virus nuclear antigen 1 (EBNA1) or EBNA1 fragments, presence of DRB1*15 and absence of A*02. The study population includes newly diagnosed cases and matched controls. Interaction on the additive scale was calculated using attributable proportion due to interaction (AP), which is the proportion of the incidence among individuals exposed to two interacting factors that is attributable to the interaction per se. IM showed association with MS, odds ratio (OR)=1.89 (1.45–2.48% confidence interval (CI)), as did raised EBNA1 IgG OR=1.74 (1.38–2.18 95%CI). All EBNA1 fragment IgGs were associated with MS risk. However, EBNA1 fragment 385–420 IgG levels were more strongly associated to MS than total EBNA1 IgG, OR=3.60 (2.75–4.72 95%CI), and also interacted with both DRB1*15 and absence of A*02, AP 0.60 (0.45–0.76 95%CI) and AP 0.39 (0.18–0.61 95%CI), respectively. The observed interaction between HLA class I and II genotype and reactivity to EBV-related epitopes suggest that the mechanism through which HLA genes influence the risk of MS may, at least in part, involve the immune control of EBV infection.

Sunlight is associated with decreased multiple sclerosis risk: no interaction with human leukocyte antigen-DRB1*15.

Background:  Both insufficient exposure to sunlight and vitamin D deficiency have been associated with an increased risk for multiple sclerosis (MS). An interaction between human leukocyte antigen HLA‐DRB1*15 and vitamin D in MS was recently proposed. We investigated the association between previous exposure to ultraviolet radiation (UVR), vitamin D status at inclusion in the study, and MS risk including the interaction of these factors with HLA‐DRB1*15.

Confirmation of association between multiple sclerosis and CYP27B1

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.

HLA-DRB1 and month of birth in multiple sclerosis

Background: Multiple sclerosis (MS) displays a month-of-birth effect, with an excess of individuals being born in the spring and a deficit in the winter. This effect was shown to be more pronounced in familial cases of MS. In the present study, we investigated whether this month-of-birth association has any relation to the principal MS susceptibility gene, HLA-DRB1. Methods: A total of 4,834 patients with MS, 4,056 controls, and 659 unaffected siblings from Canada, Sweden, and Norway were genotyped for the HLA-DRB1 gene. Month of birth was compared for patients, controls, and unaffected siblings with and without the MS risk allele HLA-DRB1*15. Results: Significantly fewer patients with MS carrying the HLA-DRB1*15 risk allele were born in November compared with patients not carrying this allele (p = 0.02). Additionally, patients with MS carrying HLA-DRB1*15 had a higher number of April births compared with patients with MS not carrying HLA-DRB1*15 (p = 0.004). These differences were not present in controls or unaffected siblings. Conclusions: Month of birth, HLA-DRB1 genotype, and risk of multiple sclerosis are associated. The interaction of a seasonal risk factor with loci at or near HLA-DRB1 during gestation or shortly after birth is implicated.

RGMA and IL21R show association with experimental inflammation and multiple sclerosis

Rat chromosome 1 harbors overlapping quantitative trait loci (QTL) for cytokine production and experimental models of inflammatory diseases. We fine-dissected this region that regulated cytokine production, myelin oligodendrocyte glycoprotein (MOG)-induced experimental autoimmune encephalomyelitis (EAE), anti-MOG antibodies and pristane-induced arthritis (PIA) in advanced intercross lines (AILs). Analysis in the tenth and twelfth generation of AILs resolved the region in two narrow QTL, Eae30 and Eae31. Eae30 showed linkage to MOG-EAE, anti-MOG antibodies and levels of interleukin-6 (IL-6). Eae31 showed linkage to EAE, PIA, anti-MOG antibodies and levels of tumor necrosis factor (TNF) and IL-6. Confidence intervals defined a limited set of potential candidate genes, with the most interesting being RGMA, IL21R and IL4R. We tested the association with multiple sclerosis (MS) in a Nordic case–control material. A single nucleotide polymorphism in RGMA associated with MS in males (odds ratio (OR)=1.33). Polymorphisms of RGMA also correlated with changes in the expression of interferon-γ (IFN-γ) and TNF in cerebrospinal fluid of MS patients. In IL21R, there was one positively associated (OR=1.14) and two protective (OR=0.87 and 0.68) haplotypes. One of the protective haplotypes correlated to lower IFN-γ expression in peripheral blood mononuclear cells of MS patients. We conclude that RGMA and IL21R and their pathways are crucial in MS pathogenesis and warrant further studies as potential biomarkers and therapeutic targets.

Cytomegalovirus seropositivity is negatively associated with multiple sclerosis.

Background: Epidemiological data suggest a role for common viruses in the pathogenesis of multiple sclerosis (MS), and recent data showed a negative association of past cytomegalovirus (CMV) infection on pediatric MS risk. Objective: Our aim was to analyze the association of CMV infection with MS risk in an adult case-control material. A meta-analysis was performed to validate our findings. Methods: Epidemiological Investigation in MS (EIMS) is a case-control study with incident cases and population-based controls. Anti-CMV antibody titers were measured with ELISA, and HLA-A and DRB1 genotyping was performed with SSP-PCR, in 658 MS cases, who all fulfilled the McDonald criteria for MS, and 786 controls. Results: CMV seropositivity was associated with a decreased MS risk, OR = 0.73 (0.58–0.92 95% CI), p = 0.005, adjusted for index age, gender, smoking, sun exposure, EBNA1 IgG titer and HLA-A*02 and DRB1*15. When we removed all cases and controls younger than 18 years at index, the protective effect was still apparent. Conclusions: CMV is negatively associated with adult-onset MS pathology, consistent with results from a study on pediatric MS cases. It remains to be shown whether this negative association is due to a true protective effect of CMV infection on MS risk.

JC Polyomavirus Infection Is Strongly Controlled by Human Leucocyte Antigen Class II Variants

JC polyomavirus (JCV) carriers with a compromised immune system, such as in HIV, or subjects on immune-modulating therapies, such as anti VLA-4 therapy may develop progressive multifocal leukoencephalopathy (PML) which is a lytic infection of oligodendrocytes in the brain. Serum antibodies to JCV mark infection occur only in 50–60% of infected individuals, and high JCV-antibody titers seem to increase the risk of developing PML. We here investigated the role of human leukocyte antigen (HLA), instrumental in immune defense in JCV antibody response. Anti-JCV antibody status, as a surrogate for JCV infection, were compared to HLA class I and II alleles in 1621 Scandinavian persons with MS and 1064 population-based Swedish controls and associations were replicated in 718 German persons with MS. HLA-alleles were determined by SNP imputation, sequence specific (SSP) kits and a reverse PCR sequence-specific oligonucleotide (PCR-SSO) method. An initial GWAS screen displayed a strong HLA class II region signal. The HLA-DRB1*15 haplotype was strongly negatively associated to JCV sero-status in Scandinavian MS cases (OR = 0.42, p = 7×10−15) and controls (OR = 0.53, p = 2×10−5). In contrast, the DQB1*06:03 haplotype was positively associated with JCV sero-status, in Scandinavian MS cases (OR = 1.63, p = 0.006), and controls (OR = 2.69, p = 1×10−5). The German dataset confirmed these findings (OR = 0.54, p = 1×10−4 and OR = 1.58, p = 0.03 respectively for these haplotypes). HLA class II restricted immune responses, and hence CD4+ T cell immunity is pivotal for JCV infection control. Alleles within the HLA-DR1*15 haplotype are associated with a protective effect on JCV infection. Alleles within the DQB1*06:03 haplotype show an opposite association. These associations between JC virus antibody response and human leucocyte antigens supports the notion that CD4+ T cells are crucial in the immune defence to JCV and lays the ground for risk stratification for PML and development of therapy and prevention.

Lack of replication of interaction between EBNA1 IgG and smoking in risk for multiple sclerosis

Background: Epstein-Barr virus infection, smoking, HLA-A*02, and DRB1*15 have all been proposed as risk factors for multiple sclerosis (MS). In 2010, Simon et al. described an interaction on the multiplicative scale between EBNA1 immunoglobulin G (IgG) and smoking regarding risk of MS, a finding that we attempted to replicate. Methods: This Swedish case-control study consisted of patients with newly diagnosed MS and matched controls. Using logistic regression, we analyzed association to MS risk and interactions between EBNA1 IgG and smoking, HLA-DRB1*15, and A*02, respectively, on the multiplicative scale. In addition, we analyzed interactions on the additive scale using attributable proportion due to interaction (AP). Results: We did not observe any interaction on the multiplicative scale between EBNA1 IgG and any of the 3 risk factors, smoking, DRB1*15, or absence of A*02, although in a conditional analysis the interaction with absence of A*02 becomes significant. However, we observed interactions on the additive scale between EBNA1 IgG and DRB1*15 (AP = 0.34, 95% confidence interval 0.11–0.57, p = 5 × 10−3) and between EBNA1 IgG and absence of A*02 (AP = 0.36, 0.13–0.59, p = 2 × 10−3) but not between smoking and DRB1*15 and EBNA1 IgG. The interaction between EBNA1 IgG and DRB1*15 was not significant in the conditional analysis. Conclusion: We did not observe any interaction between EBNA1 IgG and smoking, regardless of scale used, and thus did not replicate the observations from Simon et al.

No evidence of IL21 association with multiple sclerosis in a Swedish population

Multiple sclerosis (MS) patients, with a second autoimmune disease after lymphocyte depletion, had elevated serum IL-21 before and after treatment which correlated to IL21 genotypes. In addition, the IL21 gene has been associated to several other autoimmune diseases. However, in a Spanish population there was no association to MS. Here, in a Swedish cohort (2090 MS cases and 1732 controls) 12 single nucleotide polymorphisms (SNPs) tagging IL21 were not associated to disease. There was no interaction with risk alleles of IL21R and HLA-DRB1*15. Lack of genetic association was confirmed in a meta-analysis with pooled data from the present study and the Spanish study. In conclusion, IL21 has not been shown to be a major risk gene for MS.