Melatonin Contributes to the Seasonality of Multiple Sclerosis Relapses.

Seasonal changes in disease activity have been observed in multiple sclerosis, an autoimmune disorder that affects the CNS. These epidemiological observations suggest that environmental factors influence the disease course. Here, we report that melatonin levels, whose production is modulated by seasonal variations in night length, negatively correlate with multiple sclerosis activity in humans. Treatment with melatonin ameliorates disease in an experimental model of multiple sclerosis and directly interferes with the differentiation of human and mouse T cells. Melatonin induces the expression of the repressor transcription factor Nfil3, blocking the differentiation of pathogenic Th17 cells and boosts the generation of protective Tr1 cells via Erk1/2 and the transactivation of the IL-10 promoter by ROR-α. These results suggest that melatonin is another example of how environmental-driven cues can impact T cell differentiation and have implications for autoimmune disorders such as multiple sclerosis.

Assessing robustness and generalization of a deep neural network for brain MS lesion segmentation on real-world data.

OBJECTIVES: Evaluate the performance of a deep learning (DL)-based model for multiple sclerosis (MS) lesion segmentation and compare it to other DL and non-DL algorithms. METHODS: This ambispective, multicenter study assessed the performance of a DL-based model for MS lesion segmentation and compared it to alternative DL- and non-DL-based methods. Models were tested on internal (n = 20) and external (n = 18) datasets from Latin America, and on an external dataset from Europe (n = 49). We also examined robustness by rescanning six patients (n = 6) from our MS clinical cohort. Moreover, we studied inter-human annotator agreement and discussed our findings in light of these results. Performance and robustness were assessed using intraclass correlation coefficient (ICC), Dice coefficient (DC), and coefficient of variation (CV). RESULTS: Inter-human ICC ranged from 0.89 to 0.95, while spatial agreement among annotators showed a median DC of 0.63. Using expert manual segmentations as ground truth, our DL model achieved a median DC of 0.73 on the internal, 0.66 on the external, and 0.70 on the challenge datasets. The performance of our DL model exceeded that of the alternative algorithms on all datasets. In the robustness experiment, our DL model also achieved higher DC (ranging from 0.82 to 0.90) and lower CV (ranging from 0.7 to 7.9%) when compared to the alternative methods. CONCLUSION: Our DL-based model outperformed alternative methods for brain MS lesion segmentation. The model also proved to generalize well on unseen data and has a robust performance and low processing times both on real-world and challenge-based data. CLINICAL RELEVANCE STATEMENT: Our DL-based model demonstrated superior performance in accurately segmenting brain MS lesions compared to alternative methods, indicating its potential for clinical application with improved accuracy, robustness, and efficiency. KEY POINTS: • Automated lesion load quantification in MS patients is valuable; however, more accurate methods are still necessary. • A novel deep learning model outperformed alternative MS lesion segmentation methods on multisite datasets. • Deep learning models are particularly suitable for MS lesion segmentation in clinical scenarios.

The frequency and characteristics of multiple sclerosis misdiagnosis in Latin America: A referral center study in Buenos Aires, Argentina.

OBJECTIVE: Most contemporary data concerning the frequency and causes of multiple sclerosis (MS) misdiagnosis are from North America and Europe with different healthcare system structure and resources than countries in Latin America. We sought to determine the frequency, and potential contributors to MS misdiagnosis in patients evaluated at an MS referral center in Argentina. METHODS: The study was a retrospective medical record review. We included patients evaluated at the MS Clinic at Fleni between April 2013 and March 2021. Diagnoses prior to consultation, final diagnoses after consultation, demographic, clinical and paraclinical data, and treatment were extracted and classified. RESULTS: Seven hundred thirty-six patients were identified. Five hundred seventy-two presented with an established diagnosis of MS and after evaluation, misdiagnosis was identified in 89 (16%). Women were at 83% greater risk of misdiagnosis (p = 0.034). The most frequent alternative diagnoses were cerebrovascular disease, radiological isolated syndrome (RIS), and headache. Seventy-four (83%) of misdiagnosed patients presented with a syndrome atypical for demyelination, 62 (70%) had an atypical brain magnetic resonance imaging (MRI), and 54 (61%) were prescribed disease-modifying therapy. CONCLUSION: Sixteen percent of patients with established MS were subsequently found to have been misdiagnosed. Women were at higher risk for misdiagnosis. Expert application of the McDonald criteria may prevent misdiagnosis and its associated morbidity and healthcare system cost.

The risk of infections for multiple sclerosis and neuromyelitis optica spectrum disorder disease-modifying treatments: Eighth European Committee for Treatment and Research in Multiple Sclerosis Focused Workshop Review. April 2021.

Over the recent years, the treatment of multiple sclerosis (MS) and neuromyelitis optica spectrum disorder (NMOSD) has evolved very rapidly and a large number of disease-modifying treatments (DMTs) are now available. However, most DMTs are associated with adverse events, the most frequent of which being infections. Consideration of all DMT-associated risks facilitates development of risk mitigation strategies. An international focused workshop with expert-led discussions was sponsored by the European Committee for Treatment and Research in Multiple Sclerosis (ECTRIMS) and was held in April 2021 to review our current knowledge about the risk of infections associated with the use of DMTs for people with MS and NMOSD and corresponding risk mitigation strategies. The workshop addressed DMT-associated infections in specific populations, such as children and pregnant women with MS, or people with MS who have other comorbidities or live in regions with an exceptionally high infection burden. Finally, we reviewed the topic of DMT-associated infectious risks in the context of the current SARS-CoV-2 pandemic. Herein, we summarize available evidence and identify gaps in knowledge which justify further research.

Toll-like receptor 2 and poly(ADP-ribose) polymerase 1 promote central nervous system neuroinflammation in progressive EAE.

Multiple sclerosis is an inflammatory disease of the central nervous system that begins as a relapsing-remitting disease (RRMS) and is followed by a progressive phase (SPMS). The progressive phase causes the greatest disability and has no effective therapy, but the processes that drive SPMS are mostly unknown. Here we found higher serum concentrations of 15alpha-hydroxicholestene (15-HC) in patients with SPMS and in mice with secondary progressive experimental autoimmune encephalomyelitis (EAE) but not in patients with RRMS. In mice, 15-HC activated microglia, macrophages and astrocytes through a pathway involving Toll-like receptor 2 (TLR2) and poly(ADP-ribose) polymerase 1 (PARP-1). PARP-1 activity was higher in monocytes of patients with SPMS, and PARP-1 inhibition suppressed the progression of EAE. Thus, the TLR2-PARP-1 pathway is a potential new therapeutic target in SPMS.

Brain volumes quantification from MRI in healthy controls: Assessing correlation, agreement and robustness of a convolutional neural network-based software against FreeSurfer, CAT12 and FSL.

BACKGROUND AND PURPOSE: There are instances in which an estimate of the brain volume should be obtained from MRI in clinical practice. Our objective is to calculate cross-sectional robustness of a convolutional neural network (CNN) based software (Entelai Pic) for brain volume estimation and compare it to traditional software such as FreeSurfer, CAT12 and FSL in healthy controls (HC). MATERIALS AND METHODS: Sixteen HC were scanned four times, two different days on two different MRI scanners (1.5 T and 3 T). Volumetric T1-weighted images were acquired and post-processed with FreeSurfer v6.0.0, Entelai Pic v2, CAT12 v12.5 and FSL v5.0.9. Whole-brain, grey matter (GM), white matter (WM) and cerebrospinal fluid (CSF) volumes were calculated. Correlation and agreement between methods was assessed using intraclass correlation coefficient (ICC) and Bland Altman plots. Robustness was assessed using the coefficient of variation (CV). RESULTS: Whole-brain volume estimation had better correlation between FreeSurfer and Entelai Pic (ICC (95% CI) 0.96 (0.94-0.97)) than FreeSurfer and CAT12 (0.92 (0.88-0.96)) and FSL (0.87 (0.79-0.91)). WM, GM and CSF showed a similar trend. Compared to FreeSurfer, Entelai Pic provided similarly robust segmentations of brain volumes both on same-scanner (mean CV 1.07, range 0.20-3.13% vs. mean CV 1.05, range 0.21-3.20%, p = 0.86) and on different-scanner variables (mean CV 3.84, range 2.49-5.91% vs. mean CV 3.84, range 2.62-5.13%, p = 0.96). Mean post-processing times were 480, 5, 40 and 5 min for FreeSurfer, Entelai Pic, CAT12 and FSL respectively. CONCLUSION: Based on robustness and processing times, our CNN-based model is suitable for cross-sectional volumetry on clinical practice.

Diagnosis of Rapidly Progressive Dementia in a Referral Center in Argentina.

INTRODUCTION: Rapidly progressive dementia (RPD) is a broadly defined clinical syndrome. Our aim was to describe clinical and ancillary study findings in patients with RPD and evaluate their diagnostic performance for the identification of nonchronic neurodegenerative rapidly progressive dementia (ncnRPD). METHODS: We reviewed clinical records and ancillary methods of patients evaluated for RPD at our institution in Buenos Aires, Argentina from 2011 to 2017. We compared findings between chronic neurodegenerative RPD and ncnRPD and evaluated the diagnostic metrics using receiver operating characteristic curves. RESULTS: We included 104 patients with RPD, 29 of whom were chronic neurodegenerative RPD and 75 of whom were ncnRPD. The 6-month time to dementia cutpoint had a sensitivity of 89% and specificity of 100% for ncnRPD, with an area under the receiver operating characteristic curve of 0.965 (95% confidence interval=0.935-0.99; P<0.001). A decision tree that included time to dementia, brain magnetic resonance imaging, and cerebrospinal fluid analysis identified ncnRPD patients with a sensitivity of 100%, specificity of 79%, positive predictive value of 93%, and negative predictive value of 100% overall. DISCUSSION: RPD is a clinical syndrome that comprises different diagnoses, many of them for treatable diseases. Using the time to dementia, brain magnetic resonance imaging, and cerebrospinal fluid analysis when triaging these patients could help identify those diseases that need to be studied more aggressively.

Intravenous Chlorpromazine as Potentially Useful Treatment for Chronic Headache Disorders.

OBJECTIVE: The aim of this study was to describe a group of patients with chronic headache disorders (CH) and medication overuse headache (MOH) treated with intravenous chlorpromazine (IVC). We hypothesized that IVC is an effective and safe addition to well-known treatment strategies for CH and MOH management. INTRODUCTION: Up to 4% of the general population could experience CH. Most cases occur in women, in association with MOH. To date, evidence to support different treatment strategies is lacking. Although IVC is frequently used in the emergency room (ER), documentation on its use as supportive treatment for CH and for withdrawal management of MOH is poor. METHODS: A retrospective cohort of patients hospitalized to receive treatment for CH in a specialized neurological center in Argentina was analyzed. RESULTS: A total of 35 CH patients were included. Of the 35 patients, 33 (94%) patients also presented MOH. Patients reported only minor side effects to IVC administration (mainly drowsiness and symptomatic hypotension). Three months after inpatient treatment, the number of ER visits made by these patients decreased from an average of 2.8 in the 3 months prior to hospitalization to 0.7 after it (72%, P = .009). Headache frequency decreased in 20/34 (59%) patients during the same time period. Pain levels had dropped from a mean of 8 points at admission (in the scale of 1-10) to 2 points at discharge. In the first 3 months of follow-up, the average number of days per month in which patients experienced headache decreased from 28.9 to 15.4 days (53.3%, P < .0001). CONCLUSION: In this particular group of inpatients, there were no significant safety issues with IVC administration and the study might suggest that the efficacy of IVC as an add-on treatment for CH and MOH.

The role of infections in multiple sclerosis.

Several lines of evidence suggest that multiple sclerosis (MS), like other autoimmune diseases, may be triggered by microbial infections. Pathogens associated with development or exacerbation of MS include bacteria, such as Chlamydia pneumoniae, Staphylococcus aureus-produced enterotoxins that function as superantigens, and viruses of the Herpesviridae (Epstein-Barr virus and human herpes virus 6) and human endogenous retrovirus families. However, to date, no single pathogen has been accepted as causal agent. In addition, common upper respiratory, gastrointestinal, and urogenital tract infections have also been associated with MS exacerbations. Although evidence of an infectious etiology as cause of MS in humans remains inconclusive, microbial agents may modulate the neuroimmunological system of genetically susceptible individuals. Decoding the epidemiological contribution of different microorganisms to MS, along with their pathogenic mechanisms, may help develop new treatment strategies and prevent relapses.

A Population-Based Survey of Stroke Knowledge in Argentina: The SIFHON Study.

BACKGROUND AND OBJECTIVES: Information about stroke awareness in Latin America is scant. We conducted a large population survey in Argentina to assess stroke knowledge. METHODS: We distributed 110,000 multiple-choice anonymous questionnaires using the house distribution system of a bottled water dispensing company. The survey assessed demographic characteristics and stroke knowledge. RESULTS: A total of 12,710 surveys were returned (12%). Even though 95% of the respondents reported some prior information about stroke, only 37% had adequate knowledge based on prespecified criteria. The Spanish acronym for accidente cerebrovascular, was the most frequently identified name for stroke. Sixty nine percent of respondents were able to identify stroke main risk factors and only 29% knew about transient ischemic attacks. If a hypothetical scenario of stroke was presented, 63% knew the existence of a time-dependent treatment, 25% would call an ambulance, and 50% would go to an emergency room by own means. A lower degree of knowledge was present in young, single, and nonuniversity men. CONCLUSIONS: This study represents the largest stroke awareness survey in a Spanish-speaking population. There was good recognition of some basic facts of stroke. However, the population had poor knowledge of prevalence and severity of the disease, transient ischemic attacks, and treatment availability.

Differences between acute-onset chronic inflammatory demyelinating polyneuropathy and acute inflammatory demyelinating polyneuropathy in adult patients.

Acute inflammatory demyelinating polyneuropathy (AIDP) and acute-onset chronic inflammatory demyelinating polyneuropathy (A-CIDP) are conditions presenting overlapping clinical features during early stages (first 4 weeks), although the latter may progress after 8 weeks. The aim of this study was to identify predictive factors contributing to their differential diagnosis. Clinical records of adult patients with AIDP or A-CIDP diagnosed at our institution between January 2006 and July 2017 were retrospectively reviewed. Demographic characteristics, clinical manifestations, cerebrospinal-fluid (CSF) findings, treatment and clinical evolution were analyzed. Nerve conduction studies were performed in all patients with at least 12 months follow-up. A total of 91 patients were included (AIDP, n = 77; A-CIDP, n = 14). The median age was 55.5 years in patients with A-CIDP vs 43 years in AIDP (P = .07). The history of diabetes mellitus was more frequent in A-CIDP (29% vs 8%, P = .04). No significant differences between groups were observed with respect to: human immunodeficiency virus (HIV) status, presence of auto-immune disorder or oncologic disease. Cranial, motor and autonomic nerve involvement rates were similar in both groups. Patients in the A-CIDP group showed higher frequency of proprioceptive disturbances (83% vs 28%; P < .001), sensory ataxia (46% vs 16%; P = .01), and the use of combined immunotherapy with corticoids (29% vs 3%; P = .005). There were no significant differences in CSF findings, intensive care unit (ICU) admission, or mortality rates. During the first 8 weeks both entities are practically indistinguishable. Alterations in proprioception could suggest A-CIDP. Searching for markers that allow early differentiation could favor the onset of corticotherapy without delay.

Anti-inflammatory effects of melatonin in multiple sclerosis.

Melatonin is a hormone with complex roles in the pathogenesis of autoimmune disorders. Over the years, it has become clear that melatonin may exacerbate some autoimmune conditions, whereas it alleviates others such as multiple sclerosis. Multiple sclerosis is an autoimmune disorder characterized by a dysregulated immune response directed against the central nervous system. Indeed, the balance between pathogenic CD4(+) T cells secreting IFN-γ (TH 1) or IL-17 (TH 17); and FoxP3(+) regulatory T cells and IL-10(+) type 1 regulatory T cells (Tr1 cells) is thought to play an important role in disease activity. Recent evidence suggests that melatonin ameliorates multiple sclerosis by controlling the balance between effector and regulatory cells, suggesting that melatonin-triggered signaling pathways are potential targets for therapeutic intervention. Here, we review the available data on the effects of melatonin on immune processes relevant for MS and discuss its therapeutic potential.

Age may contribute to the increased frequency of axonal Guillain-Barré syndrome.

INTRODUCTION: The frequency of axonal Guillain-Barré syndrome (GBS) varies among countries. Previous studies supporting the high frequency of axonal GBS in South America have been carried out with pediatric populations. We seek to determine the frequency of axonal GBS in both children and adults in South America. METHODS: This is a retrospective cohort analysis of patients who were diagnosed with GBS between January 2006 and December 2013 in a neurological center in Buenos Aires, Argentina. Adults and children with a diagnosis of GBS were included and classified by applying Ho and colleagues' criteria1 for axonal GBS. RESULTS: The study included 105 patients with GBS. Among 58 adults, only 5 individuals were classified as axonal GBS compared with 16 of 47 children. The frequency of axonal GBS was significantly higher in children than in adults (34% vs. 8.6%, P = 0.0001). DISCUSSION: As shown in a cohort of South American patients, age may impact the frequency of axonal GBS. Muscle Nerve 56: 1311-1313, 2017.

Headache and Treatment of Unruptured Intracranial Aneurysms.

BACKGROUND AND PURPOSE: The relationship between unruptured intracranial aneurysms (UIAs) and chronic headache and the impact of aneurysm treatment on headache outcome are controversial. The aim of this study was to determine clinical features of a supposedly primary headache in patients with UIA. We also assessed changes in headache characteristics after UIA treatment. METHODS: We examined clinical and imaging data of patients in whom a UIA was diagnosed during diagnostic workup of a suspected primary headache. Medical records were reviewed and personal telephone follow-ups were performed after UIA treatment to assess changes in the frequency and intensity of the headache. RESULTS: Forty-two patients (76%) reported a substantial improvement in headache frequency and intensity after UIA treatment. Forty-five patients (81%) reported a decrease in headache frequency from a median of 8 days/month before treatment to 1 day/month after treatment (95% confidence interval [CI] 81-83, P < .001). The average intensity in an analog pain scale was 7.7 ± 1.6 before treatment and 5.6 ± 2.4 after treatment (P < .001). Higher headache frequency was associated with a greater odd of improvement after treatment (odds ratio 1.12, 95% CI 1.0-1.26, P = .03). No associations were found between the type of headache, type of treatment (endovascular versus surgical), number, size, or localization of the aneurysms and the response to treatment. CONCLUSIONS: The treatment of UIA had a robust beneficial effect on previous headache. Although a "placebo" effect of aneurysm treatment cannot be ruled out, these results suggest a potential association between UIA and certain chronic headaches usually considered to be primary.

Clinical and cerebrospinal fluid findings contribute to the early differentiation between infectious and noninfectious encephalitis.

Early recognition and prompt specific treatment are crucial factors influencing the outcome of patients with acute encephalitis. The aim of this study was to determine the main causes of acute encephalitis in our population and to find predictors that may lead to specific diagnosis. Adult patients admitted to our hospital with suspected diagnosis of encephalitis in the period 2006-2013 were included. One hundred and five medical records were analyzed. Eighty-two patients with infectious encephalitis were identified (78% of total cases), 53 (65%) men and 29 (35%) women, mean age 47.8 years. The most common microorganisms identified were: HSV-1 (11%), VZV (10%), HSV-2 (5%) and EBV (5%). Twenty-three patients (22% of the series) had non-infectious encephalitis. Headache (p < 0.0001) and fever (p = 0.008) were more frequent in encephalitis of infectious origin. Protein levels and white blood cell counts in the cerebrospinal fluid were significantly higher in patients affected by infectious encephalitis than in those affected by noninfectious encephalitis (OR 95% CI 12.3 [2.9-51.7] and OR 95% CI 7.4 [2-27], respectively). Identifying specific causal agents of acute encephalitis remains a major challenge. Cerebrospinal fluid markers, as well as specific clinical findings, may however contribute to initial differentiation between infectious and noninfectious causes.

Sodium intake is associated with increased disease activity in multiple sclerosis.

BACKGROUND: Recently, salt has been shown to modulate the differentiation of human and mouse Th17 cells and mice that were fed a high-sodium diet were described to develop more aggressive courses of experimental autoimmune encephalomyelitis. However, the role of sodium intake in multiple sclerosis (MS) has not been addressed. We aimed to investigate the relationship between salt consumption and clinical and radiological disease activity in MS. METHODS: We conducted an observational study in which sodium intake was estimated from sodium excretion in urine samples from a cohort of 70 relapsing-remitting patients with MS who were followed for 2 years. The effect of sodium intake in MS disease activity was estimated using regression analysis. We then replicated our findings in a separate group of 52 patients with MS. RESULTS: We found a positive correlation between exacerbation rates and sodium intake in a multivariate model adjusted for age, gender, disease duration, smoking status, vitamin D levels, body mass index and treatment. We found an exacerbation rate that was 2.75-fold (95% CI 1.3 to 5.8) or 3.95-fold (95% CI 1.4 to 11.2) higher in patients with medium or high sodium intakes compared with the low-intake group. Additionally, individuals with high-sodium intake had a 3.4-fold greater chance of developing a new lesion on the MRI and on average had eight more T2 lesions on MRI. A similar relationship was found in the independent replication group. CONCLUSIONS: Our results suggest that a higher sodium intake is associated with increased clinical and radiological disease activity in patients with MS.

Parasite infections in multiple sclerosis modulate immune responses through a retinoic acid-dependent pathway.

We recently demonstrated better outcomes in helminth-infected multiple sclerosis (MS) patients, compared with uninfected ones. The present study evaluates the role of TLR2 and retinoic acid (RA) in parasite-driven protection in MS patients. RA serum levels were significantly higher in helminth-infected MS patients than in uninfected MS subjects or healthy controls. Genes involved in RA biosynthesis and metabolism, such as Adh1 and Raldh2, as well as RA receptors and IL-10, were induced in dendritic cells (DCs) via TLR2-dependent ERK signaling. This programmed DCs to induce FOXP3(+) T regulatory cells and suppressed production of proinflammatory cytokines (IL-6, IL-12, IL-23, and TNF-α) via induction of suppressor of cytokine signaling 3 (SOCS3), an effect mediated by soluble egg Ag (SEA) obtained from Schistosoma mansoni, and by RA. SEA-activated DCs also inhibited IL-17 and IFN-γ production through autoreactive T cells. These inhibitory effects were abrogated when SOCS3 gene expression was silenced, indicating that SEA-mediated signaling inhibited production of these cytokines by T cells, through a SOCS3-dependent pathway. Overall, helminth-related immunomodulation observed in MS patients was mediated by TLR2- and RA-dependent pathways, through two different mechanisms, as follows: 1) induction of IL-10 and FOXP3(+) T regulatory cells, and 2) suppression of proinflammatory cytokine production mediated by SOCS3.

Control of T(reg) and T(H)17 cell differentiation by the aryl hydrocarbon receptor.

Regulatory T cells (T(reg)) expressing the transcription factor Foxp3 control the autoreactive components of the immune system. The development of T(reg) cells is reciprocally related to that of pro-inflammatory T cells producing interleukin-17 (T(H)17). Although T(reg) cell dysfunction and/or T(H)17 cell dysregulation are thought to contribute to the development of autoimmune disorders, little is known about the physiological pathways that control the generation of these cell lineages. Here we report the identification of the ligand-activated transcription factor aryl hydrocarbon receptor (AHR) as a regulator of T(reg) and T(H)17 cell differentiation in mice. AHR activation by its ligand 2,3,7,8-tetrachlorodibenzo-p-dioxin induced functional T(reg) cells that suppressed experimental autoimmune encephalomyelitis. On the other hand, AHR activation by 6-formylindolo[3,2-b]carbazole interfered with T(reg) cell development, boosted T(H)17 cell differentiation and increased the severity of experimental autoimmune encephalomyelitis in mice. Thus, AHR regulates both T(reg) and T(H)17 cell differentiation in a ligand-specific fashion, constituting a unique target for therapeutic immunomodulation.

[Immunopathology of multiple sclerosis]. / Inmunopatología de la esclerosis múltiple.

Multiple sclerosis is an inflammatory demyelinating disease affecting the central nervous system and considered one of the leading causes of disability in young adults. The precise cause of multiple sclerosis is unknown, although the current evidence points towards a combination of genetic and environmental factors leading to an autoimmune response that promotes neuronal degeneration. In this review, we will describe the association between the immune response and neurodegeneration in multiple sclerosis.

Sex-specific environmental influences affecting MS development.

Vitamin D status, smoking, and Epstein-Barr virus infection (EBV) may all contribute to explain differences in disease prevalence and incidence of Multiple Sclerosis (MS). MS affects women more often than men, and recent cross-sectional study assessments provide evidence of increased female to male prevalence in relapsing remitting MS patients, suggesting that sex hormones may exert an active role in disease pathogenesis. Studies in both humans and animal disease models demonstrate a functional synergy for the immunomodulatory effects of Vitamin D3 and 17-ß estradiol. Both smoking and EBV infection clearly increase MS risk, and smoking history has also been associated with poorer disease prognosis. However, neither factor can explain the recent trend indicating greater female prevalence. Therefore, large population-based case-control studies from well defined geographic areas with homogeneous populations should be performed, in order to define environmental factor effects, and sex hormone influences, to better understand prevalence and incidence gender differences observed.