Tau protein quantification in skin biopsies differentiates tauopathies from alpha-synucleinopathies.

Abnormal accumulation of microtubule-associated protein tau (τ) is a characteristic feature of atypical parkinsonisms with tauopathies, such as progressive supranuclear palsy and corticobasal degeneration. However, pathological τ has also been observed in α-synucleinopathies like Parkinson's disease and multiple system atrophy. Based on the involvement of the peripheral nervous system in several neurodegenerative diseases, we characterized and compared τ expression in skin biopsies of patients clinically diagnosed with Parkinson's disease, multiple system atrophy, progressive supranuclear palsy and corticobasal degeneration and in healthy control subjects. In all groups, τ protein was detected along both somatosensory and autonomic nerve fibres in the epidermis and dermis by immunofluorescence. We found by western blot the presence of mainly two different bands at 55 and 70 kDa, co-migrating with 0N4R/1N3R and 2N4R isoforms, respectively. At the RNA level, the main transcript variants were 2N and 4R, and both were more expressed in progressive supranuclear palsy/corticobasal degeneration by real-time PCR. Enzyme-linked immunosorbent assay demonstrated significantly higher levels of total τ protein in skin lysates of progressive supranuclear palsy/corticobasal degeneration compared to the other groups. Multivariate regression analysis and receiver operating characteristics curve analysis of τ amount at both sites showed a clinical association with tauopathies diagnosis and high diagnostic value for progressive supranuclear palsy/corticobasal degeneration versus Parkinson's disease (sensitivity 90%, specificity 69%) and progressive supranuclear palsy/corticobasal degeneration versus multiple system atrophy (sensitivity 90%, specificity 86%). τ protein increase correlated with cognitive impairment in progressive supranuclear palsy/corticobasal degeneration. This study is a comprehensive characterization of τ in the human cutaneous peripheral nervous system in physiological and pathological conditions. The differential expression of τ, both at transcript and protein levels, suggests that skin biopsy, an easily accessible and minimally invasive exam, can help in discriminating among different neurodegenerative diseases.

Alpha-synuclein oligomers and small nerve fiber pathology in skin are potential biomarkers of Parkinson's disease.

The proximity ligation assay (PLA) is a specific and sensitive technique for the detection of αSyn oligomers (αSyn-PLA), early and toxic species implicated in the pathogenesis of PD. We aimed to evaluate by skin biopsy the diagnostic and prognostic capacity of αSyn-PLA and small nerve fiber reduction in PD in a longitudinal study. αSyn-PLA was performed in the ankle and cervical skin biopsies of PD (n = 30), atypical parkinsonisms (AP, n = 23) including multiple system atrophy (MSA, n = 12) and tauopathies (AP-Tau, n = 11), and healthy controls (HC, n = 22). Skin biopsy was also analyzed for phosphorylated αSyn (P-αSyn) and 5G4 (αSyn-5G4), a conformation-specific antibody to aggregated αSyn. Intraepidermal nerve fiber density (IENFD) was assessed as a measure of small fiber neuropathy. αSyn-PLA signal was more expressed in PD and MSA compared to controls and AP-Tau. αSyn-PLA showed the highest diagnostic accuracy (PD vs. HC sensitivity 80%, specificity 77%; PD vs. AP-Tau sensitivity 80%, specificity 82%), however, P-αSyn and 5G4, possible markers of later phases, performed better when considering the ankle site alone. A small fiber neuropathy was detected in PD and MSA. A progression of denervation not of pathological αSyn was detected at follow-up and a lower IENFD at baseline was associated with a greater cognitive and motor decline in PD. A skin biopsy-derived compound marker, resulting from a linear discrimination analysis model of αSyn-PLA, P-αSyn, αSyn-5G4, and IENFD, stratified patients with accuracy (77.8%), including the discrimination between PD and MSA (84.6%). In conclusion, the choice of pathological αSyn marker and anatomical site influences the diagnostic performance of skin biopsy and can help in understanding the temporal dynamics of αSyn spreading in the peripheral nervous system during the disease. Skin denervation, not pathological αSyn is a potential progression marker for PD.

De-escalating rituximab dose results in stability of clinical, radiological, and serum neurofilament levels in multiple sclerosis.

BACKGROUND: Phase II and observational studies support the use of rituximab in multiple sclerosis. Standard protocols are lacking, but studies suggest comparable efficacy between low- and high-dose regimens. OBJECTIVE: To evaluate effectiveness and safety of de-escalating rituximab dose from 1000 to 500 mg/6 months in multiple sclerosis. METHODS: Patients were switched from rituximab 1000 to 500 mg/6 months and prospectively followed for 12 months. Relapses, disability, occurrence of brain/spinal magnetic resonance imaging (MRI) lesions, serum neurofilament light chain (NfL), CD19+ B cell, and IgG concentrations were analyzed. RESULTS: Fifty-nine patients were included (37 relapsing-remitting, 22 secondary progressive). No relapses occurred, with no difference in expanded disability status scale (EDSS) between baseline (4 (2.5-4.5) and 12 months (3.5 (2.5-5.5) p = 0.284). Overall, three new T2 lesions appeared during follow-up. NfL concentration was stable between baseline (7.9 (5.9-45.2) pg/mL) and 12 months (9.1 (5.9-21.3) pg/mL, p = 0.120). IgG concentrations decreased with greater rituximab load (coefficient = -0.439, p = 0.041). IgG deficient patients had greater risk of infections (OR = 6.27, 95% CI = 1.71-22.9, p = 0.005). CONCLUSION: De-escalating rituximab dose from 1000 to 500 mg/6 months is safe, results in clinical and radiological stability, and does not affect serum NfL over 12 months. Rituximab load negatively influences IgG concentrations, and IgG deficient patients are at higher risk of infections.

Increasing cancer risk over calendar year in people with multiple sclerosis: a case-control study.

BACKGROUND: Data on cancer prevalence and incidence in multiple sclerosis (MS) patients are controversial. This study is aimed at estimating cancer risk in MS patients. METHODS: Nested case-control study using data collected between 01/01/1987 and 28/02/2016 from the United Kingdom Clinical Practice Research Datalink. Cancer diagnoses after first MS code (index date) was counted in 10,204 MS patients and 39,448 controls matched by sex, age, general practitioner, and registration year. Cancer rates were compared using multivariable Cox regression models. Ethics approval was not required. RESULTS: Cancer was reported in 433 (4.41%) MS patients and 2014 (5.31%) controls after index date. Cancer risk was associated with gender (HR for female = 0.88, 95% CI = 0.81-0.96, p = 0.004), age at index date (HR = 1.06, 95% CI = 1.06-1.07, p < 0.001), and index year (HR = 1.01, 95% CI = 1.00-1.02, p = 0.016), but not with MS status (HR = 0.95, 95% CI = 0.86-1.05, p = 0.323). A significant interaction between MS status and index year was found (HR = 1.02, 95% CI = 1.00-1.04, p = 0.022). Cancer risk was positively associated with index year among MS patients (HR = 1.03, 95% CI = 1.01-1.05; p = 0.010), but not controls (HR = 1.01, 95% CI = 0.99-1.02; p = 0.144). MS patients compared to controls had no increased risk for any specific cancer type. CONCLUSIONS: Overall cancer risk was similar in multiple sclerosis patients and matched controls. The frequency of cancer diagnoses has increased over time among MS patients but not in controls.

Effectiveness and safety of Rituximab in multiple sclerosis: an observational study from Southern Switzerland.

BACKGROUND: Despite positive results from phase II and observational studies, Rituximab (RTX) is not currently approved for multiple sclerosis (MS) treatment and can only be used off-label. OBJECTIVE: To characterize MS patients treated with RTX and investigate its effectiveness and safety in a clinical practice setting. METHODS: Observational analysis of data collected from MS patients at the Neurocenter of Southern Switzerland. Relapses, EDSS worsening, MRI lesion accrual and "evidence of disease activity" (EDA) status were described by Cox regression. RTX and natalizumab treated patients were matched by propensity scores. RESULTS: Out of 453 MS patients, 82 were treated with RTX, 43 (52.4%) relapsing-remitting (RRMS) and 39 (47.6%) progressive MS (median age = 48 [40-54] years, females n = 60 [73.2%], EDSS = 4.0 [2.5-6.0], median follow-up = 1.5 [1.0-2.5] years). Three relapses occurred and 59 (75.6%) patients had not EDA at follow-up end. Time to EDA was similar in RTX and natalizumab treated RRMS patients (HR = 1.64, 95%CI = 0.46-5.85, p = 0.44). Twenty-four patients presented non infusion related adverse events (infections), requiring RTX discontinuation in 6 individuals. CONCLUSION: These results provide further evidence for RTX being effective in MS treatment, to a similar extent to natalizumab in RRMS. Clinicians must be vigilant for the potential occurrence of infections.

No evidence for higher risk of cancer in patients with multiple sclerosis taking cladribine.

OBJECTIVE: To compare the cancer risk of cladribine and other disease-modifying drugs (DMDs) in trials of people with relapsing multiple sclerosis (pwRMS). METHODS: Meta-analysis of phase III trials of licensed DMDs for pwRMS and a phase III trial of cladribine (CLARITY). Cancer rates were compared using Fisher exact test. RESULTS: Eleven trials were included. Investigated treatments included cladribine, dimethyl fumarate, fingolimod, teriflunomide, natalizumab, alemtuzumab, and glatiramer acetate. The cancer rate in the CLARITY treatment group (0.34%) was not increased compared to all other treatment groups, whether including placebo-controlled trials only (0.6%, p = 0.4631) or all trials, i.e., including those with an active comparator arm (0.67%, p = 0.3669). No cancer was reported in the CLARITY placebo group, whereas the combined cancer rate of all other placebo groups was 1.19% (p = 0.0159). The cancer rate of zero in the CLARITY placebo group was also lower than that in the phase III trial of cladribine in people with clinically isolated syndrome (ORACLE MS, 2.91%, p = 0.0012). In fact, no difference was detected between cancer rates in the treatment groups of CLARITY (0.34%) and ORACLE MS (0.49%) (p = 0.6546). CONCLUSIONS: Our study does not support an increased cancer risk from cladribine in the doses used in CLARITY and ORACLE MS, which previously contributed to refusal of market authorization of cladribine in Europe. Longer-term follow-up is required to assess the safety profile of cladribine, as well as currently licensed DMDs, to definitively assess cancer risk.

Impact of month of birth on the development of autoimmune thyroid disease in the United Kingdom and Europe.

CONTEXT: Viral/bacterial infection is proposed as a trigger for the autoimmune thyroid diseases (AITD): Graves' disease (GD) and Hashimoto's thyroiditis (HT). Previous studies in European Caucasian AITD subjects found higher birth rates in the autumn/winter, suggesting those born in the autumn/winter experience increased viral/bacterial exposure after birth, impacting upon immune system development and predisposing to AITD later in life. OBJECTIVE: Month of birth effects were investigated in three independent European Caucasian AITD datasets. DESIGN: Variation in GD and HT onset was compared across months and seasons, with fluctuations across all 12 months analyzed using a Walter-Elwood test. SETTING: The study was conducted at a research laboratory. PATIENTS: National UK Caucasian AITD Case Control Collection (2746 GD and 502 HT compared with 1 423 716 UK births), National UK Caucasian GD Family Collection (239 GD and 227 unaffected siblings), and OXAGEN AITD Caucasian Family Collection (885 GD, 717 HT, and 794 unaffected siblings of European Caucasian decent). MAIN OUTCOME MEASURES: Case-control and family-based association studies were measured. RESULTS: No consistent month of birth effects were detected in GD females or males across all three collections. In HT females from the OXAGEN AITD Caucasian Family Collection, slightly higher birth rates were detected in autumn (Walter's test statistic = 7.47, P = .024) however, this was not seen in the HT females from the case-control cohort. CONCLUSION: Our results suggest in UK/Northern European Caucasian GD subjects, month of birth does not impact on AITD development. Although some month of birth effects for HT females in one collection cannot be excluded, only further work in larger European Caucasian AITD collections can confirm these effects.

Regulatory genomic regions active in immune cell types explain a large proportion of the genetic risk of multiple sclerosis.

There is little understanding of how genetic variants discovered in recent genome-wide association studies are involved in the pathogenesis of multiple sclerosis (MS). We aimed to investigate which chromatin states and cell types explain genetic risk in MS. We used genotype data from 1854 MS patients and 5164 controls produced by the International Multiple Sclerosis Genetics Consortium and Wellcome Trust Case Control Consortium. We estimated the proportion of phenotypic variance between cases and controls explained by cell-specific chromatin state and DNase I hypersensitivity sites (DHSs) using the Genome-wide Complex Trait Analysis software. A large proportion of variance was explained by single-nucleotide polymorphisms (SNPs) in strong enhancer (SE) elements of immortalized B lymphocytes (5.39%). Three independent SNPs located within SE showed suggestive evidence of association with MS: rs12928822 (odds ratio (OR)=0.81, 95% confidence interval (CI)=0.73-0.89, P=2.48E-05), rs727263 (OR=0.75, 95% CI=0.66-0.85, P=3.26E-06) and rs4674923 (OR=0.85, 95% CI=0.79-0.92, P=1.63E-05). Genetic variants located within DHSs of CD19+ B cells explained the greatest proportion of variance. Genetic variants influencing the risk of MS are located within regulatory elements active in immune cells. This study also identifies a number of immune cell types likely to be involved in the causal cascade and that carry important implications for future studies of therapeutic design.

Hypovitaminosis-D and EBV: no interdependence between two MS risk factors in a healthy young UK autumn cohort.

Late Epstein-Barr virus infection and hypovitaminosis-D as environmental risk factors in the pathogenesis of multiple sclerosis are gaining great interest. We, therefore, tested for in-vivo interdependence between Epstein-Barr-virus (EBV)-status and 25-hydroxyvitamin D3 (25(OH)D3) -level in healthy young individuals from a United Kingdom (UK) autumn cohort. EBV-load was measured by quantitative polymerase chain reaction and 25(OH)D3 levels by isotope-dilution liquid chromatography-tandem mass spectrometry. This young, healthy UK autumn cohort showed surprisingly low levels of 25(OH)D3 (mean value: 40.5 nmol/L ± 5.02). Furthermore, we found that low 25(OH)D3 levels did not impact on EBV load and anti-EBV nuclear antigen-1 (EBNA-1) titers. However, we observed a correlation between EBV load and EBNA-1 titers. These observations should be of value in the study of the potential relationship between hypovitaminosis-D and EBV-status in the pathophysiology of multiple sclerosis.

DNase hypersensitive sites and association with multiple sclerosis.

Genome-wide association studies (GWASs) have shown that approximately 60 genetic variants influence the risk of developing multiple sclerosis (MS). Our aim was to identify the cell types in which these variants are active. We used available data on MS-associated single nucleotide polymorphisms (SNPs) and deoxyribonuclease I hypersensitive sites (DHSs) from 112 different cell types. Genomic intervals were tested for overlap using the Genomic Hyperbrowser. The expression profile of the genes located nearby MS-associated SNPs was assessed using the software GRAIL (Gene Relationships Across Implicated Loci). Genomic regions associated with MS were significantly enriched for a number of immune DHSs and in particular T helper (Th) 1, Th17, CD8+ cytotoxic T cells, CD19+ B cells and CD56+ natural killer (NK) cells (enrichment = 2.34, 2.19, 2.27, 2.05 and 1.95, respectively; P < 0.0001 for all of them). Similar results were obtained when genomic regions with suggestive association with MS and additional immune-mediated traits were investigated. Several new candidate MS-associated genes located within regions of suggestive association were identified by GRAIL (CARD11, FCRL2, CHST12, SYK, TCF7, SOCS1, NFKBIZ and NPAS1). Genetic data indicate that Th1, Th17, cytotoxic T, B and NK cells play a prominent role in the etiology of MS. Regions with confirmed and suggestive association have a similar immunological profile, indicating that many SNPs truly influencing the risk of MS actually fail to reach genome-wide significance. Finally, similar cell types are involved in the etiology of other immune-mediated diseases.

Integrating multiple oestrogen receptor alpha ChIP studies: overlap with disease susceptibility regions, DNase I hypersensitivity peaks and gene expression.

BACKGROUND: A wealth of nuclear receptor binding data has been generated by the application of chromatin immunoprecipitation (ChIP) techniques. However, there have been relatively few attempts to apply these datasets to human complex disease or traits. METHODS: We integrated multiple oestrogen receptor alpha (ESR1) ChIP datasets in the Genomic Hyperbrowser. We analysed these datasets for overlap with DNase I hypersensitivity peaks, differentially expressed genes with estradiol treatment and regions near single nucleotide polymorphisms associated with sex-related diseases and traits. We used FIMO to scan ESR1 binding sites for classical ESR1 binding motifs drawn from the JASPAR database. RESULTS: We found that binding sites present in multiple datasets were enriched for classical ESR1 binding motifs, DNase I hypersensitivity peaks and differentially expressed genes after estradiol treatment compared with those present in only few datasets. There was significant enrichment of ESR1 binding present in multiple datasets near genomic regions associated with breast cancer (7.45-fold, p = 0.001), height (2.45-fold, p = 0.002), multiple sclerosis (5.97-fold, p < 0.0002) and prostate cancer (4.47-fold, p = 0.0008), and suggestive evidence of ESR1 enrichment for regions associated with coronary artery disease, ovarian cancer, Parkinson's disease, polycystic ovarian syndrome and testicular cancer. Integration of multiple cell line ESR1 ChIP datasets also increases overlap with ESR1 ChIP-seq peaks from primary cancer samples, further supporting this approach as helpful in identifying true positive ESR1 binding sites in cell line systems. CONCLUSIONS: Our study suggests that integration of multiple ChIP datasets can highlight binding sites likely to be of particular biological importance and can provide important insights into understanding human health and disease. However, it also highlights the high number of likely false positive binding sites in ChIP datasets drawn from cell lines and illustrates the importance of considering multiple independent experiments together.

Hospital admissions for vitamin D related conditions and subsequent immune-mediated disease: record-linkage studies.

BACKGROUND: Previous studies have suggested that there may be an association between vitamin D deficiency and the risk of developing immune-mediated diseases. METHODS: We analyzed a database of linked statistical records of hospital admissions and death registrations for the whole of England (from 1999 to 2011). Rate ratios for immune-mediated disease were determined, comparing vitamin D deficient cohorts (individuals admitted for vitamin D deficiency or markers of vitamin D deficiency) with comparison cohorts. RESULTS: After hospital admission for either vitamin D deficiency, osteomalacia or rickets, there were significantly elevated rates of Addison's disease, ankylosing spondylitis, autoimmune hemolytic anemia, chronic active hepatitis, celiac disease, Crohn's disease, diabetes mellitus, pemphigoid, pernicious anemia, primary biliary cirrhosis, rheumatoid arthritis, Sjogren's syndrome, systemic lupus erythematosus, thyrotoxicosis, and significantly reduced risks for asthma and myxoedema. CONCLUSIONS: This study shows that patients with vitamin D deficiency may have an increased risk of developing some immune-mediated diseases, although we cannot rule out reverse causality or confounding. Further study of these associations is warranted and these data may aid further public health studies.

The risk of developing multiple sclerosis in individuals seronegative for Epstein-Barr virus: a meta-analysis.

BACKGROUND: Epstein-Barr virus (EBV) infection is widely considered to be a risk factor for multiple sclerosis (MS). A previous meta-analysis estimated an odds ratio (OR) for MS in individuals seronegative for EBV of 0.06. Given the potential importance of this finding, we aimed to establish a more precise OR for adult and paediatric onset MS in EBV seronegative individuals. METHODS: PubMed and EMBASE searches were undertaken to identify studies investigating the association between MS and EBV. Twenty-two adult and three paediatric studies were included. ORs were calculated using a fixed effects model. A sub-group analysis based on the method of EBV detection was performed. RESULTS: The OR for developing adult MS in EBV seronegatives was 0.18 (95% confidence interval (CI) 0.13-0.26)) and for paediatric MS was 0.18 (95% CI 0.11-0.30). Sub-group analysis on EBV detection method showed that studies which used immunofluoresence generated an OR=0.07 (95% CI 0.03-0.16); for those that used enzyme-linked immunosorbent assay (ELISA) OR=0.33 (95% CI 0.22-0.50) and for studies which used ELISA and immunofluoresence OR=0.00 (95% CI 0-0.43). CONCLUSION: The sensitivity and specificity of the assay used to measure EBV antibody titres have an influence on the association between MS and EBV. Looking at studies where two independent methods are used and therefore are likely to be the most robust, EBV appears to be present in 100% of MS patients. This has implications for future studies of EBV in MS. MS patients without EBV infection, if they truly exist, should be studied in more detail.

Epstein-Barr virus, latitude and multiple sclerosis.

Several lines of evidence support a role for Epstein-Barr virus (EBV) in the aetiology of multiple sclerosis (MS). This includes the observation that nearly all MS patients show serological markers of past EBV infection. Given the well-known association between MS prevalence and latitude, we investigated whether EBV seropositivity also increases with distance from the equator. We found that the proportion of EBV positive individuals is positively associated with latitude independently of MS status (odds ratio = 1.06, 95% CI = 1.02-1.09, p = 0.002). Latitude-related factors may be implicated in the immune response to EBV and its role in MS aetiology.

Month of birth, vitamin D and risk of immune-mediated disease: a case control study.

BACKGROUND: A season of birth effect in immune-mediated diseases (ID) such as multiple sclerosis and type 1 diabetes has been consistently reported. We aimed to investigate whether season of birth influences the risk of rheumatoid arthritis, Crohn's disease, ulcerative colitis and systemic lupus erythematosus in addition to multiple sclerosis, and to explore the correlation between the risk of ID and predicted ultraviolet B (UVB) light exposure and vitamin D status during gestation. METHODS: The monthly distribution of births of patients with ID from the UK (n = 115,172) was compared to that of the general population using the Cosinor test. Predicted UVB radiation and vitamin D status in different time windows during pregnancy were calculated for each month of birth and correlated with risk of ID using the Spearman's correlation coefficient. RESULTS: The distributions of ID births significantly differed from that of the general population (P = 5e-12) with a peak in April (odds ratio = 1.045, 95% confidence interval = 1.024, 1.067, P < 0.0001) and a trough in October (odds ratio = 0.945, 95% confidence interval = 0.925, 0.966, P < 0.0001). Stratification by disease subtype showed seasonality in all ID but Crohn's disease. The risk of ID was inversely correlated with predicted second trimester UVB exposure (Spearman's rho = -0.49, P = 0.00005) and third trimester vitamin D status (Spearman's rho = -0.44, P = 0.0003). CONCLUSIONS: The risk of different ID in the UK is significantly influenced by the season of birth, suggesting the presence of a shared seasonal risk factor or factors predisposing to ID. Gestational UVB and vitamin D exposure may be implicated in the aetiology of ID.

Multiple sclerosis: risk factors and their interactions.

Multiple sclerosis (MS) is a highly debilitating immune mediated disorder of the central nervous system and represents a substantial burden to the developed world. Despite the recent advances in MS research, which risk factors are implicated and how they contribute to MS pathogenesis is largely unknown. However, in line with older studies investigating the genetic and geographical epidemiology of this complex disease, more recent studies have highlighted how MS arises from a combination of genetic susceptibility and environmental exposures acting from gestation to early adulthood. Vitamin D deficiency, season of birth, Epstein Barr virus infection, and smoking behaviour are strongly implicated and able to influence genetic predisposition to MS. Furthermore, these factors appear to act synergistically and the risk of MS in individuals exposed to more than one factor combines multiplicatively. Current evidence suggests that a large part of MS could be prevented and understanding how and when during life risk factors act will ultimately aid the development of prevention strategies.

Genomic regions associated with multiple sclerosis are active in B cells.

More than 50 genomic regions have now been shown to influence the risk of multiple sclerosis (MS). However, the mechanisms of action, and the cell types in which these associated variants act at the molecular level remain largely unknown. This is especially true for associated regions containing no known genes. Given the evidence for a role for B cells in MS, we hypothesized that MS associated genomic regions co-localized with regions which are functionally active in B cells. We used publicly available data on 1) MS associated regions and single nucleotide polymorphisms (SNPs) and 2) chromatin profiling in B cells as well as three additional cell types thought to be unrelated to MS (hepatocytes, fibroblasts and keratinocytes). Genomic intervals and SNPs were tested for overlap using the Genomic Hyperbrowser. We found that MS associated regions are significantly enriched in strong enhancer, active promoter and strong transcribed regions (p = 0.00005) and that this overlap is significantly higher in B cells than control cells. In addition, MS associated SNPs also land in active promoter (p = 0.00005) and enhancer regions more than expected by chance (strong enhancer p = 0.0006; weak enhancer p = 0.00005). These results confirm the important role of the immune system and specifically B cells in MS and suggest that MS risk variants exert a gene regulatory role. Previous studies assessing MS risk variants in T cells may be missing important effects in B cells. Similar analyses in other immunological cell types relevant to MS and functional studies are necessary to fully elucidate how genes contribute to MS pathogenesis.

Smoking and multiple sclerosis: an updated meta-analysis.

BACKGROUND: Multiple sclerosis (MS) is a leading cause of disability in young adults. Susceptibility to MS is determined by environmental exposure on the background of genetic risk factors. A previous meta-analysis suggested that smoking was an important risk factor for MS but many other studies have been published since then. METHODS/PRINCIPAL FINDINGS: We performed a Medline search to identify articles published that investigated MS risk following cigarette smoking. A total of 14 articles were included in this study. This represented data on 3,052 cases and 457,619 controls. We analysed these studies in both a conservative (limiting our analysis to only those where smoking behaviour was described prior to disease onset) and non-conservative manner. Our results show that smoking is associated with MS susceptibility (conservative: risk ratio (RR) 1.48, 95% confidence interval (CI) 1.35-1.63, p < 10⁻¹5; non-conservative: RR 1.52, 95% CI 1.39-1.66, p < 10⁻¹9). We also analysed 4 studies reporting risk of secondary progression in MS and found that this fell just short of statistical significance with considerable heterogeneity (RR 1.88, 95% CI 0.98-3.61, p = 0.06). DISCUSSION: Our results demonstrate that cigarette smoking is important in determining MS susceptibility but the effect on the progression of disease is less certain. Further work is needed to understand the mechanism behind this association and how smoking integrates with other established risk factors.