Stimulated PBMC-produced IFN-γ and TNF-α are associated with altered relapse risk in multiple sclerosis: results from a prospective cohort study.

BACKGROUND: Altered reactivity of peripheral blood mononuclear cells (PBMC) and their production of cytokines may affect multiple sclerosis (MS) clinical course. We assessed the relationship of stimulated PBMC-produced IFN-γ, TNF-α, IL-4 and IL-10 in modulating relapse risk using a prospective cohort with established relapsing-remitting MS. METHODS: Cytokine production from PBMCs taken in summer and winter was measured by ELISA. Predictors of cytokines assessed by multilevel mixed-effects linear regression. Predictors of relapse assessed by survival analysis. RESULTS: Increasing IFN-γ was associated with increasing relapse risk, while increasing TNF-α reduced relapse risk after adjusting for IFN-γ. IL-10 and IL4 were not consistently associated with relapse risk. IFN-γ's effects on relapse were greatly attenuated by immunomodulatory therapies, by summer season and by higher serum vitamin D, whereas TNF-α's inverse association with relapse was only present in these circumstances. The TNF-α inverse association with relapse was only present among persons carrying the wild-type of the functional SNP rs1800693 in TNFRSF1A that has been previously associated with MS risk. CONCLUSIONS: We found strong effects of IFN-γ and TNF-α on relapse risk, these differing by immunomodulatory therapy, season, and serum vitamin D, as well as by genotype. These results indicate altered reactivity of immune cells modulate MS disease.

Novel modulating effects of PKC family genes on the relationship between serum vitamin D and relapse in multiple sclerosis.

BACKGROUND: The interplay between genes and environmental factors on multiple sclerosis (MS) clinical course has been little studied. METHODS: We conducted a prospective cohort study of 141 participants with relapsing-remitting MS (RRMS) and genotype data followed from 2002 to 2005 and examined genes in the vitamin D metabolism and vitamin D receptor (VDR)/retinoid X receptor (RXR) transcription factor formation pathway. Gene-vitamin D interactions and the genetic predictors of relapse were assessed using survival analysis. Genetic predictors of 25-hydroxyvitamin D (25(OH)D) were evaluated by multilevel mixed-effects linear regression. Significance threshold was adjusted by Bonferroni correction for the number of genes evaluated. RESULTS: The relationship between 25(OH)D and hazard of relapse was significantly different for different alleles of two intronic single nucleotide polymorphisms (SNPs) (rs908742 in PRKCZ and rs3783785 in PRKCH) in the protein kinase C (PKC) family genes (p(interaction)=0.001, p(adj)=0.021, respectively). Two other intronic SNPs (rs1993116 in CYP2R1and rs7404928 in PRKCB) were significantly associated with lower levels of 25(OH)D (p(interaction)=0.001, p(adj)=0.021, respectively). A cumulative effect of multiple 'risk' genotypes on 25(OH)D levels and hazard of relapse was observed for the significant SNPs (p(trend)=7.12×10(-6) for 25(OH)D levels, p(trend)=8.86×10(-6) for hazard of relapse). CONCLUSIONS: Our data support the hypothesis that gene-vitamin D interactions may influence MS clinical course and that the PKC family genes may play a role in the pathogenesis of MS relapse through modulating the association between 25(OH)D and relapse.

Association between multiple sclerosis risk-associated SNPs and relapse and disability--a prospective cohort study.

BACKGROUND: The modulating effects of the multiple sclerosis (MS) risk-associated single-nucleotide polymorphisms (SNPs) on MS clinical course are not well established. OBJECTIVES: The objective of this paper is to investigate whether known MS risk-associated SNPs were associated with clinical course, and whether these SNPs modified the 25(OH)D-relapse association. METHODS: Using a prospective cohort of 141 participants with relapsing-remitting MS and genotype data followed between 2002 and 2005, genotype-vitamin D interactions and the genetic predictors of relapse were assessed using survival analysis, and genetic predictors of 25(OH)D and disability progression were evaluated by multilevel mixed-effects linear regression. RESULTS: While no SNP reached statistical significance after multiple testing, five SNPs were associated with relapse, with significant cumulative genotype risk effects and two demonstrated significant allele dose-response. Two SNPs altered the 25(OH)D-relapse association with significant allele dose-response. Five SNPs modified levels of 25(OH)D, with significant cumulative genotype 'risk' effect, and three demonstrated significant allele dose-response. We found no consistent evidence for an association between any SNPs and disability. CONCLUSIONS: Our study provides evidence for an association between known MS risk-associated SNPs and relapse. Our findings indicate gene-environment interactions may be an important mechanism on MS clinical course, and provide support for the role of vitamin D in MS relapse.

Anti-HHV-6 IgG titer significantly predicts subsequent relapse risk in multiple sclerosis.

BACKGROUND: Some of the strongest associations with MS onset are for human herpesviruses, particularly Epstein-Barr virus (EBV) and human herpesvirus 6 (HHV-6). Their role in MS clinical course is less clear, however. METHODS: Prospective cohort of 198 persons with clinically definite MS, followed 2002-5, and serum samples obtained from all subjects at study entry to measure anti-HHV-6 and anti-EBV (Epstein-Barr nuclear antigen [EBNA] and viral capsid antigen [VCA]) IgG titers. Association with relapse evaluated using survival analysis; association with disability/progression evaluated using linear regression or multilevel mixed-effects linear regression. RESULTS: For the 145 persons with relapsing-remitting MS followed beyond one review, anti-HHV-6 IgG titer was positively associated with the hazard of relapse with a dose-dependent trend (p = 0.003), not affected by adjustment for anti-EBV IgG titers, neither of which were independently associated with relapse. There was no significant association between anti-human herpesvirus IgG titers and baseline-measured disability scores, or change in disability scores; however, anti-HHV-6 IgG titers were 2.8 times higher among progressive-course females than progressive-course males. DISCUSSION: These findings suggest that, in addition to a potential etiological role in MS, HHV-6 infection or the immune response to HHV-6 antigens may have an effect on the risk of MS relapses and possibly on progressive courses of MS. The observed effect was directly related to anti-HHV-6 IgG titers and may indicate that either HHV-6 infection or factors associated with an altered humoral immune response to HHV-6 may have an effect on MS clinical course. Anti-HHV-6 IgG titer may be a useful prognostic factor in relapsing-remitting MS clinical course.

Higher 25-hydroxyvitamin D is associated with lower relapse risk in multiple sclerosis.

OBJECTIVE: A protective association between higher vitamin D levels and the onset of multiple sclerosis (MS) has been demonstrated; however, its role in modulating MS clinical course has been little studied. We investigated whether higher levels of serum 25-hydroxyvitamin D (25-OH-D) were associated with a lower risk of relapses in people with MS. METHODS: We conducted a prospective cohort study of 145 participants with relapsing-remitting MS from 2002 to 2005. Serum 25-OH-D levels were measured biannually, and the hazard of relapse was assessed using survival analysis. RESULTS: There was an inverse linear relationship between 25-OH-D levels and the hazard of relapse over the subsequent 6 months, with hazard ratio (HR) 0.91 (95% confidence interval [CI]: 0.85-0.97) per 10 nmol/l increase in 25-OH-D level (p = 0.006). When variation due to timing of blood collection was removed by estimating 25-OH-D at the start of each season, this association persisted, with HR 0.90 (95% CI, 0.83-0.98) per 10 nmol/l increase (p = 0.016). Taking into account the biological half-life of 25-OH-D, we estimated 25-OH-D at monthly intervals, resulting in a slightly enhanced association, with HR 0.88 (95% CI, 0.82-0.95) per 10 nmol/l increase (p = 0.001). Adjusting for potential confounders did not alter these findings. INTERPRETATION: In this prospective population-based cohort study, in a cohort largely on immunomodulatory therapy, higher 25-OH-D levels were associated with a reduced hazard of relapse. This occurred in a dose-dependent linear fashion, with each 10 nmol/l increase in 25-OH-D resulting in up to a 12% reduction in risk of relapse. Clinically, raising 25-OH-D levels by 50 nmol/l could halve the hazard of a relapse.

Trends in the epidemiology of multiple sclerosis in Greater Hobart, Tasmania: 1951 to 2009.

BACKGROUND: Hobart, Tasmania, has been the site of two major studies of multiple sclerosis (MS) frequency, in 1951-1961 and 1971-1981. Since then, there have been no studies of MS frequency in Hobart. METHODS: Using a prevalent cohort of 226 cases in 2001 and 265 in 2009, the authors undertook a two-stage survey of MS frequency in Hobart. Combined with the published data from the two preceding studies, the authors conducted a time-trend analysis of MS epidemiology over 1951-2009. RESULTS: The age-standardised prevalence in 2001 was 96.6/100 000, and 99.6/100 000 in 2009, a significant increase from the 1961 prevalence of 32.5/100 000 (p<0.001). Female prevalence increased over each time point; male prevalence increased between 1961 and 2001 but was unchanged thereafter. Incidence over 2001-2009 was 3.7/100 000, significantly increased from the 1951-1961 incidence of 2.2/100 000 (p=0.004), though the majority of this was between 1951-1961 and 1971-1981. Mortality fell by half from 2.4/100 000 in 1951-1959 to 1.0/100 000 in 2001-2009-this decreased mortality and an older cohort contribute to the increase in prevalence. Neither prevalence (p=0.48) nor incidence (p=0.18) sex ratios changed significantly between 1951 and 2009. CONCLUSIONS: Between 1951 and 2009, the age-standardised prevalence of MS in Hobart increased threefold, and the incidence nearly doubled. Part of the increase in prevalence was due to an increased longevity, decreased mortality and increased incidence. Differences in patterns by birthplace may be explained by the Australian assisted-migration programme of 1945-1981. These data do not demonstrate the strong and significant changes in sex ratio observed elsewhere.

Monthly ambient sunlight, infections and relapse rates in multiple sclerosis.

BACKGROUND: Monthly variation in multiple sclerosis (MS) relapses has been found. The relationship between seasonal environmental factors, infections, serum vitamin D [25(OH)D] and MS relapses is undetermined. METHODS: We prospectively followed a population-based cohort of relapsing-remitting (RR) MS patients in Southern Tasmania for a mean 2.3 years (January 2002-April 2005). Associations between monthly ambient environmental factors, estimated serum 25(OH)D, upper respiratory tract (URT) infections and relapse rates were examined using weighted Pearson's correlation and linear regression. RESULTS: Of 199 definite MS patients, 142 had RRMS. The lowest relapse rate of 0.5 per 1,000 days (95% CI: 0.2-1.3) occurred in February (mid-late summer) versus the March-January RR of 1.1 per 1,000 days (95% CI: 0.9-1.3; p = 0.018, weighted regression). Monthly relapse rates correlated with: (1) prior erythemal ultraviolet radiation (EUV): lagged 1.5 months, r = -0.32, p = 0.046; (2) URT infection rate: no lag, r = 0.39, p = 0.014; (3) 25(OH)D: no lag, r = -0.31, p = 0.057. The association between URT infections and relapses was reduced after adjustment for monthly EUV. CONCLUSIONS: Relapse rates were inversely associated with EUV and serum 25(OH)D levels and positively associated with URT infections. The demonstrated lag between EUV but not 25(OH)D and relapse rates is consistent with a role for EUV-generated 25(OH)D in the alteration of relapse rates. Future work on the association between URT infections and relapses should be considered in the context of ultraviolet radiation and vitamin D.

Adherence to the immunomodulatory drugs for multiple sclerosis: contrasting factors affect stopping drug and missing doses.

BACKGROUND: Long-term immunomodulatory drug (IMD) treatment is now common in multiple sclerosis (MS). However, predictors of adherence are not well understood; past studies lacked lifestyle factors such as alcohol use and predictors of missed doses have not been evaluated. We examined both levels of non-adherence-stopping IMD and missing doses. METHODS: This longitudinal prospective study followed a population-based cohort (n = 199) of definite MS patients in Southern Tasmania (January 2002 to April 2005, source population 226 559) every 6 months. Baseline factors (demographic, clinical, psychological and cognitive) affecting adherence were examined by logistic regression and a longitudinal analysis (generalized estimating equation (GEE)). RESULTS: Of the 97 patients taking an IMD (mean follow-up = 2.4 years), 73% (71/97) missed doses, with 1 in 10 missing > 10 doses in any 6-month period. Missed doses were positively associated with alcohol amount consumed per session (p = 0.008). A history of missed doses predicted future missed doses (p < 0.0005). Over one-quarter (27/97) stopped their current IMD, which was associated with lower education levels (p = 0.032) and previous relapses (p = 0.05). No cognitive or psychological test predicted adherence. CONCLUSIONS: There were few strong predictors of missed doses, although people with MS consuming more alcoholic drinks per session are at a higher risk of missing doses. Divergent factors influenced the two levels of non-adherence indicating the need for a multifaceted approach to improving IMD adherence. In addition, missed doses should be assessed and incorporated into clinical trial design and clinical practice as poor adherers could impact on clinical outcomes.

The effect of season on cytokine expression in multiple sclerosis and healthy subjects.

We measured the levels of IFNgamma, TNFalpha, Il-4 and Il-10 produced by mitogen-stimulated peripheral blood mononuclear cells (PBMC) from healthy people, and those with the relapse/remitting form of multiple sclerosis. Blood was taken in summer and winter. Healthy people had a summer excess of Il-4, Il-10 and TNFalpha, and a winter excess of IFNgamma. Untreated MS cases had a summer excess of Il-10, whereas those treated with Interferon-beta had lower levels of all cytokines, and displayed no seasonal effect.

The cost of multiple sclerosis in Australia.

Multiple sclerosis (MS) represents a significant economic burden both to the patient and to society. This study aims to provide information about direct and indirect costs of MS in Australia. Detailed questionnaires were completed for 100 patients over a 6-month period (12 months for hospitalization costs). Overall, the average annual direct and indirect costs per patient were AU$20 396 and AU$15 085, respectively. The greatest uses of resources were for immunomodulating drugs, consultations and district nursing. Various factors significantly increased overall direct costs, including secondary progressive MS, severe MS symptoms and higher Expanded Disability Status Scale scores. This study confirms that MS is a costly disease with a high economic burden on society. In order to minimize MS costs and improve quality of life, the ideal aim of MS treatment should be to stabilize patients on a low disability (low cost) level at an early stage of the disease utilising a cost-effective therapy.

Genetic variation in PBMC-produced IFN-γ and TNF-α associations with relapse in multiple sclerosis.

BACKGROUND: Alterations in peripheral blood mononuclear cell (PBMC) cytokine production have been found in multiple sclerosis (MS) compared to healthy controls. We have previously found that stimulated PBMC-produced TNF-α and IFN-γ modulated MS relapse risk, such that raised TNF-α was protective, while raised IFN-γ increased relapse risk. OBJECTIVE: To assess whether SNPs within genes for relevant cytokines and their receptors modulate the associations of TNF-α and IFN-γ with relapse, thus providing additional information about these cytokine effects and the roles of these genes in MS. METHODS: Prospective cohort of 91 participants with relapsing-remitting MS and cytokine and genotype data. SNPs (N=361) within a window of 10 kb around each cytokine/cytokine receptor gene (N=84) were selected for analysis. Predictors of PBMC cytokines were evaluated by multilevel mixed-effects linear regression. Predictors of relapse were evaluated by Cox proportional hazards regression. Bonferroni correction was used to adjust for multiple testing; thus p<1.39 × 10(-4) was defined as significant. RESULTS: Individuals of GG genotype of rs3218295 (within the gene IL2RB) demonstrated a significant protective effect of TNF-α on relapse while those of GA/AA genotype showed a significant positive association (pinteraction=5.04 × 10(-5)). Carriers of CC genotype of rs522807 (3' region of TNFRSF1B) and the AA genotype of rs25879 (5' region of IL3) showed a strong association between IFN-γ and increased relapse risk (pinteraction=8.21 × 10(-5) and 1.70 × 10(-5), respectively). CONCLUSIONS: Our results show novel modulation of TNF-α and IFN-γ associations with relapse by SNPs in major cytokines. These findings suggest the potential for these genes and/or their products as potential therapeutic targets in MS.

Interferon-ß and serum 25-hydroxyvitamin D interact to modulate relapse risk in MS.

OBJECTIVE: To determine whether interferon-ß (IFN-ß) medication use is associated with vitamin D levels and whether the two interact in exerting effects on relapse risk. METHODS: In a prospective cohort of 178 persons with clinically definite multiple sclerosis (MS) living in southern Tasmania in 2002-2005, serum 25-hydroxyvitamin D [25(OH)D] was measured biannually, with assessment by questionnaire for relevant factors, including IFN-ß treatment. RESULTS: Subjects reporting IFN-ß use had significantly higher mean 25(OH)D than persons who did not (p < 0.001). This was mediated by an interaction between personal sun exposure and IFN-ß, with treated persons realizing nearly three times 25(OH)D per hour of sun exposure of persons not on therapy. The association between 25(OH)D and 1,25-dihydroxyvitamin D did not differ by IFN-ß therapy (p = 0.82). 25(OH)D was associated with a reduced relapse risk only among persons on IFN-ß (p < 0.001). Importantly, IFN-ß was only protective against relapse among persons with higher 25(OH)D (hazard ratio [HR] 0.58 [95% confidence interval (CI) 0.35-0.98]), while among 25(OH)D-insufficient persons, IFN-ß increased relapse risk (HR 2.01 [95% CI 1.22-3.32]). CONCLUSION: In this study, we found that IFN-ß therapy is associated with greater production of vitamin D from sun exposure, suggesting part of the therapeutic effects of IFN-ß on relapse in MS may be through modulation of vitamin D metabolism. These findings suggest persons being treated with IFN-ß should have vitamin D status monitored and maintained in the sufficiency range. CLASSIFICATION OF EVIDENCE: This study provided Class III evidence that IFN-ß is associated with reduced risk of relapse, and this effect may be modified by a positive effect of IFN-ß on serum 25(OH)D levels.

Smoking is associated with progressive disease course and increased progression in clinical disability in a prospective cohort of people with multiple sclerosis.

BACKGROUND: Multiple sclerosis has a variable disease course. The contribution of modifiable lifestyle factors to disease course has not been well studied, although one cohort has reported that smoking is associated with conversion to secondary progressive MS course and another that smoking is not. METHODS: We conducted a prospective cohort study of people with MS in Southern Tasmania from 2002 to 2004 with 78% (203/259) of eligible participating and 198 with one or more reviews and confirmed MS. The cohort had a high retention rate (90% (183/203)). The median follow-up time was 909 days. Smoking data were collected at baseline and six-monthly reviews. Clinical disability assessments were conducted annually in conjunction with a real time clinical notification system for relapses. A repeated measures analysis and other statistical methods were used. RESULTS: Cumulative pack-years (p-y) smoked after cohort entry was associated with an increase in longitudinal MSSS (p < 0.001). Relative to the 0 pack years (p-y) category (in the year prior to the MSSS measure) those in the 0 to 1 p-y category had an adjusted mean difference in MSSS of 0.34 (95% CI 0.28, 0.66); those in the 1 to 2 p-y category had a 0.41 (95% CI -0.03, 0.85) increase; and those in the 2 or more p-y category had a 0.99 (95% CI 0.41, 1.58) increase in MSSS. Similar results were found using a variety of statistical approaches or EDSS as a clinical outcome. Smoking during the cohort period was not associated with relapse (cumulative pack years smoked after cohort entry, HR 0.94 (0.69, 1.26) per pack year). CONCLUSION: A better understanding of the mechanisms underlying smoking and multiple sclerosis, particularly progressive forms of the disease, may provide new insights for the eventual goal of better treatment and prevention of multiple sclerosis.