Invited Review: From nose to gut - the role of the microbiome in neurological disease.

Inflammation and neurodegeneration are key features of many chronic neurological diseases, yet the causative mechanisms underlying these processes are poorly understood. There has been mounting interest in the role of the human microbiome in modulating the inflammatory milieu of the central nervous system (CNS) in health and disease. To date, most research has focussed on a gut-brain axis, with other mucosal surfaces being relatively neglected. We herein take the novel approach of comprehensively reviewing the roles of the microbiome across several key mucosal interfaces - the nose, mouth, lung and gut - in health and in Parkinson's disease (PD), Alzheimer's disease (AD) and multiple sclerosis (MS). This review systematically appraises the anatomical and microbiological landscape of each mucosal surface in health and disease before considering relevant mechanisms that may influence the initiation and progression of PD, AD and MS. The cumulative effects of dysbiosis from the nose to the gut may contribute significantly to neurological disease through a wide variety of mechanisms, including direct translocation of bacteria and their products, and modulation of systemic or CNS-specific immunity. This remains an understudied and exciting area for future research and may lead to the development of therapeutic targets for chronic neurological disease.

A protocol for a randomised double-blind placebo-controlled feasibility study to determine whether the daily consumption of flavonoid-rich pure cocoa has the potential to reduce fatigue in people with relapsing and remitting multiple sclerosis (RRMS).

BACKGROUND: Dietary interventions including consumption of flavonoids, plant compounds found in certain foods, may have the ability to improve fatigue. However, to date, no well-designed intervention studies assessing the role of flavonoid consumption for fatigue management in people with MS (pwMS) have been performed. The hypothesis is that the consumption of a flavonoid-rich pure cocoa beverage will reduce fatigue in pwMS. The aim of this study is to determine the feasibility and potential outcome of running a trial to evaluate this hypothesis. METHODS: Using a randomised (1:1) double-blind placebo-controlled feasibility study, 40 men and women (20 in each trial arm) with a recent diagnosis (< 10 years) of relapsing and remitting MS (RRMS) and who are over 18 years of age will be recruited from neurology clinics and throughout the Thames Valley community. During a 6-week nutrition intervention period, participants will consume the cocoa beverage, high flavonoid or low flavonoid content, at breakfast daily. At baseline, demographic factors and disease-related factors will be assessed. Fatigue, activity and quality of life, in addition to other measures, will be taken at three visits (baseline, week 3 and week 6) in a university setting by a researcher blinded to group membership. Feasibility and fidelity will be assessed through recruitment and retention, adherence and a quantitative process evaluation at the end of the trial.We will describe demographic factors (age, gender, level of education) as well as disease-related factors (disease burden scores, length of time diagnosed with MS) and cognitive assessment, depression and quality of life and general physical activity in order to characterise participants and determine possible mediators to identify the processes by which the intervention may bring about change. Feasibility (recruitment, safety, feasibility of implementation of the intervention and evaluation, protocol adherence and data completion) and potential for benefit (estimates of effect size and variability) will be determined to inform future planned studies. Results will be presented using point estimates, 95% confidence intervals and p values. Primary statistical analysis will be on an intention-to-treat basis and will use the complete case data set. DISCUSSION: We propose that a flavonoid-enriched cocoa beverage for the management of fatigue will be well received by participants. Further, if it is implemented early in the disease course of people diagnosed with RRMS, it will improve mobility and functioning by modifying fatigue. TRIAL REGISTRATION: Registered with ISRCTN Registry. Trial registration No: ISRCTN69897291; Date April 2016.

Review: the role of vitamin D in nervous system health and disease.

Vitamin D and its metabolites have pleomorphic roles in both nervous system health and disease. Animal models have been paramount in contributing to our knowledge and understanding of the consequences of vitamin D deficiency on brain development and its implications for adult psychiatric and neurological diseases. The conflation of in vitro, ex vivo, and animal model data provide compelling evidence that vitamin D has a crucial role in proliferation, differentiation, neurotrophism, neuroprotection, neurotransmission, and neuroplasticity. Vitamin D exerts its biological function not only by influencing cellular processes directly, but also by influencing gene expression through vitamin D response elements. This review highlights the epidemiological, neuropathological, experimental and molecular genetic evidence implicating vitamin D as a candidate in influencing susceptibility to a number of psychiatric and neurological diseases. The strength of evidence varies for schizophrenia, autism, Parkinson's disease, amyotrophic lateral sclerosis, Alzheimer's disease, and is especially strong for multiple sclerosis.

Risk alleles for multiple sclerosis in multiplex families.

OBJECTIVE: We assessed the hypotheses that non-major histocompatibility complex multiple sclerosis (MS) susceptibility loci would be common to sporadic cases and multiplex families, that they would have larger effects in multiplex families, and that the aggregation of susceptibility loci contributes to the increased prevalence of MS in such families. METHODS: A set of 43 multiplex families comprising 732 individuals and 211 affected subjects was genotyped for 13 MS candidate genes identified by genome-wide association. A control data set of 182 healthy individuals was also genotyped to perform a case-control analysis alongside the family-based pedigree disequilibrium association test, although this may have been underpowered. RESULTS: An effect of the IL2RA and CD58 loci was shown in multiplex families as in sporadic MS. The aggregate of the IL2RA, IL7R, EVI5, KIAA0350, and CD58 risk genotypes in affected individuals from multiplex families was found to be notably different from controls (chi(2) = 112, p = 1 x 10(-22)). CONCLUSIONS: Although differences between individual families can only be suggested, the aggregate results in multiplex families demonstrate effect sizes that are increased as compared with those reported in previous studies for sporadic cases. In addition, they imply that concentrations of susceptibility alleles at IL2RA, IL7R, EVI5, KIAA0350, and CD58 are partly responsible for the heightened prevalence of multiple sclerosis within multiplex families.

Age of puberty and the risk of multiple sclerosis: a population based study.

BACKGROUND AND PURPOSE: Genetic and environmental factors have important roles in multiple sclerosis (MS) susceptibility. Given a potential role for sex hormones in MS, we have investigated whether or not the age of puberty influences the risk of developing MS in a population-based cohort. METHODS: We identified 5493 MS index cases and 1759 spousal controls with age of puberty information from the Canadian Collaborative Project on Genetic Susceptibility to MS. Age of puberty was compared between index cases and controls, and any effect of age of puberty on the age of onset of MS was also investigated. RESULTS: There were no significant differences between male index cases and controls with respect to age of puberty, P = 0.70. However, a significant difference was observed between female index cases and female controls, with average age of puberty being 12.4 and 12.6 years respectively, P = 0.00017, providing a relative risk decrease of 0.9 per year increase of age of puberty. There was no effect of the age of puberty on the age of MS onset in either sex. CONCLUSIONS: Earlier age at menarche increases the risk of MS in women. Whether this association is a surrogate for a disease causative factor or directly involved in MS disease aetiology needs to be uncovered.

An extremes of outcome strategy provides evidence that multiple sclerosis severity is determined by alleles at the HLA-DRB1 locus.

Multiple sclerosis (MS) is a common inflammatory disease of the central nervous system unsurpassed for variability in disease outcome. A cohort of sporadic MS cases (n = 163), taken from opposite extremes of the distribution of long-term outcome, was used to determine the role of the HLA-DRB1 locus on MS disease severity. Genotyping sets of benign and malignant MS patients showed that HLA-DRB1*01 was significantly underrepresented in malignant compared with benign cases. This allele appears to attenuate the progressive disability that characterizes MS in the long term. The observation was doubly replicated in (i) Sardinian benign and malignant patients and (ii) a cohort of affected sibling pairs discordant for HLA-DRB1*01. Among the latter, mean disability progression indices were significantly lower in those carrying the HLA-DRB1*01 allele compared with their disease-concordant siblings who did not. The findings were additionally supported by similar transmission distortion of HLA-DRB1*04 subtypes closely related to HLA-DRB1*01. The protective effect of HLA-DRB1*01 in sibling pairs may result from a specific epistatic interaction with the susceptibility allele HLA-DRB1*1501. A high-density (>700) SNP examination of the MHC region in the benign and malignant patients could not identify variants differing significantly between the two groups, suggesting that HLA-DRB1 may itself be the disease-modifying locus. We conclude that HLA-DRB1*01, previously implicated in disease resistance, acts as an independent modifier of disease progression. These results closely link susceptibility to long-term outcome in MS, suggesting that shared quantitative MHC-based mechanisms are common to both, emphasizing the central role of this region in pathogenesis.

Multiple sclerosis susceptibility and the X chromosome.

Multiple sclerosis (MS) is a chronic autoimmune complex trait with strong evidence for a genetic component. A female gender bias is clear but unexplained and a maternal parent-of-origin effect has been described. X-linked transmission of susceptibility has been previously proposed, based on pedigree, association and linkage studies. We genotyped 726 relative pairs including 552 affected sib-pairs for 22 X-chromosome microsatellite markers and a novel dataset of 195 aunt-uncle/niece-nephew (AUNN) affected pairs for 18 markers. Parent-of-origin effects were explored by dividing AUNN families into likely maternal and paternal trait transmission. For the sib-pair dataset we were able to establish exclusion at a lambda s = 1.9 for all markers using an exclusion threshold of LOD < or = -2. Similarly for the AUNN dataset, we established exclusion at lambdaAV = 1.9. For the combined dataset we estimate exclusion of lambda = 1.6. We did not identify significant linkage in either the sib-pairs or the AUNN dataset nor when datasets were stratified for the presence/absence of the HLA-DRB1*15 allele or for paternal or maternal transmission. This comprehensive scrutiny of the X-chromosome suggests that it is unlikely to harbour an independent susceptibility locus or one which interacts with the HLA. Complex interactions including epigenetic ones, and masking by balanced polymorphisms are mechanisms not excluded by the approach taken.

The role of hereditary spastic paraplegia related genes in multiple sclerosis. A study of disease susceptibility and clinical outcome.

Multiple sclerosis (MS) is a common inflammatory disease of the central nervous system unsurpassed for its variability in disease outcome. It has been observed that axonal loss in MS is significant and that irreversible clinical disability relates to such axonal loss. The clinical similarities between Hereditary Spastic Paraplegia (HSP) and progressive MS, along with their analogous profiles of axonal loss in the long tracts, make the genes known to cause HSP biologically relevant candidates for the study of clinical outcome in MS. A cohort of sporadic MS cases and a set of unaffected controls were used to determine the role of HSP genes on MS susceptibility and disease severity. The MS cases were taken from opposite extremes of the putative distribution of long-term outcome using the most stringent clinical criteria to date. Genotyping the two sets of MS patients and controls could not provide any evidence to suggest that genes involved in the pathogenesis of HSP (Paraplegin, NIPA1, KIF5A, HSPD1, Atlastin, Spartin, Spastin, PLP1, L1CAM, Maspardin and BSCL2) play a role in susceptibility to, or modifying the course of, MS, although small effects of these genes cannot be ruled out.

No effect of APOE and PVRL2 on the clinical outcome of multiple sclerosis.

Multiple sclerosis (MS) is a common inflammatory disease of the central nervous system unsurpassed for its variability in disease outcome. Apolipoprotein E (APOE) is involved in neuronal remodelling and several studies have attempted to examine the effect of APOE on MS disease severity, but its function in modifying the course of MS is controversial. It has been suggested recently that PVRL2, not APOE, is the locus on chromosome 19 which influences clinical outcome of MS. A cohort of sporadic MS cases, taken from opposite extremes of the putative distribution of long-term outcome using the most stringent clinical criteria to date, was used to determine the role of APOE and PVRL2 on MS disease severity. The MS cases selected represent the prognostic best 5% (benign MS) and worst 5% (malignant MS) of cases in terms of clinical outcome assessed by the EDSS. Genotyping the two sets of MS patients (112 benign and 51 malignant) and a replication cohort from Sardinia provided no evidence to suggest that APOE or PVRL2 have any outcome modifying activity. We conclude that APOE and PVRL2 have little or no effect on the clinical outcome of MS.

The contribution of demyelination to axonal loss in multiple sclerosis.

The traditional notion that multiple sclerosis is a primary demyelinating disease has led to a plaque-centred view of both aetiology and the pathogenesis of disease progression. The presence of axonal loss has received increasing recognition. However, the relative roles of demyelination and axonal loss have not been fully clarified in multiple sclerosis nor have their possible interrelationships been elucidated. Post-mortem material from the cerebrum, brainstem and spinal cord of 55 multiple sclerosis patients (29 males) with an age range of 25-83 years (mean = 57.5 years) and length of disease history ranging from 2 to 43 years (mean = 17.1 years) was stained for myelin. Plaque load was calculated by summing the relative proportion of plaque area compared with total white matter area of the corticospinal and sensory tracts at each level. This was related to estimates of axonal density and of total axon number in these tracts in the spinal cord. Our results indicate that plaque load did not correlate with brain weight. Unexpectedly, after adjusting for sex, age and duration of disease, correlations between total plaque load and axonal loss in both the corticospinal tract and sensory tracts were weak or absent at each level investigated. Since there was little correlation between plaque load and axonal loss, the possibility that demyelination is not the primary determinant of spinal cord axonal loss warrants consideration.

Pathological study of spinal cord atrophy in multiple sclerosis suggests limited role of local lesions.

Imaging studies in multiple sclerosis have shown that spinal cord atrophy correlates with clinical disability. The pathological substrate of atrophy has not as yet been investigated adequately. In order to determine the cause of spinal cord atrophy in multiple sclerosis, five different sections of the spinal cord were examined histopathologically in 33 controls and 55 multiple sclerosis cases. In the multiple sclerosis cases in each section the total lesion load and the cross-sectional area of the cord were measured. Multiple regression models were estimated, controlling for sex, age, duration of the disease and location of the cord sections. The multiple sclerosis cords were found to be significantly smaller than the controls. The duration of the disease played the most important role in determining cord atrophy. The degree of atrophy varied in different parts of the cord. Individual lesions played a minor role in local atrophy. Our findings suggest that axonal degeneration, possibly caused by the cumulative number of lesions in the brain and cord, or an alternative atrophic process, is responsible for spinal cord atrophy in multiple sclerosis, rather than tissue loss within individual lesions.

The extent of axonal loss in the long tracts in hereditary spastic paraplegia.

Hereditary spastic paraplegia (HSP) comprises a group of inherited neurodegenerative disorders with the shared characteristics of progressive weakness and spasticity predominantly affecting the lower limbs. Limited pathological accounts have described a 'dying back' axonal degeneration in this disease. However, the distribution and extent of axonal loss has not been elucidated in a quantitative way. We have studied post-mortem material from six HSP patients and 32 controls in detail. The population of axons was examined quantitatively in the corticospinal tracts from the medulla to the lumbar spinal cord and the sensory tracts from the lumbar to upper cervical spinal cord. Myelin and axon-stained sections were employed to estimate the notional area and axonal density, respectively, of both tracts. Our results indicate that in the corticospinal tracts there is a significant reduction in area and axonal density at all levels investigated in HSP compared to controls. In the corticospinal tracts, the ratio of medulla and lumbar total axonal number was significantly greater in HSP cases compared to controls suggesting more pronounced axonal loss in the distal neuraxis in HSP than in controls. The sensory tracts in HSP, in contrast, showed a significant reduction in area and axonal density only in the upper regions of the spinal cord. Similar to the corticospinal tracts, the ratio of lumbar and upper cervical cord total axonal number in the sensory tracts was increased in HSP cases compared to controls. These findings are consistent with a length-dependent 'dying back' axonopathy. Nerve fibre loss was not size-selective with both small and large diameter fibres affected. In HSP, axonal loss is widespread and symmetrical and its extent tract-specific. The characterization of the nature of axonal loss in HSP, where this is a primary phenomenon, may help the interpretation of axonal loss in conditions such as multiple sclerosis where the sequence of events is less clear.

Evidence of axonal damage in human acute demyelinating diseases.

UNLABELLED: Substantial axon damage, detected by immunostaining for beta amyloid precursor protein (betaAPP) has been demonstrated in acute demyelinating lesions in multiple sclerosis. AIMS: The present study aimed to determine if this was also the case in the other human acute demyelinating diseases, acute hemorrhagic leucoencephalitis (AHLE), acute disseminated encephalomyelitis (ADEM) and central pontine myelinolysis (CPM). METHODS: BetaAPP immunostaining was used as a marker of axonal damage in autopsy material from these conditions. RESULTS: Axonal damage was detected in all these conditions. Its extent varied within and between them. Axonal damage was largely confined to tissue adjacent to veins and venules in AHLE and ADEM but was unrelated to proximity to these vessels in CPM. CONCLUSION: Substantial axon damage occurs in fatal cases of AHLE, ADEM and CPM.

TCR beta polymorphisms and multiple sclerosis.

A total of 267 families with two or more siblings with multiple sclerosis (MS) were genotyped with 14 restriction fragment length polymorphisms at the TCR beta locus. A nonparametric linkage analysis of the data showed no evidence for linkage to this locus (mlod=0.11). No significant allelic or haplotype transmissions were observed in the total sample of 565 patients. After stratification for the presence of HLA DRB1*15, an association was observed between the BV25S1*1-BV26S1*1-BV2S1*1 haplotype and MS (P=0.00089). This was not significant upon correction for multiple comparisons. It was also not significant when the haplotype frequency in affected individuals was compared to a normal control sample (P=0.77). Furthermore, the associated haplotype was followed-up in an independent sample of 97 nuclear families with a single DRB1*15-positive child with MS. The BV25S1*1-BV26S1*1-BV2S1*1 haplotype did not show significant evidence for transmission distortion but the same trend was seen (P=0.21). There were no significant associations observed in the DRB1*15-negative patients and no detectable difference was seen in the DRB1*15-positive BV25S1*1-BV26S1*1-BV2S1*1 association when comparing different subgroups based on clinical course of MS. These results show no evidence for linkage and fail to establish an association between MS susceptibility and the TCR beta locus.

Axonal loss in multiple sclerosis: a pathological survey of the corticospinal and sensory tracts.

Clinical, imaging, and pathological studies in multiple sclerosis have generally emphasized the relative preservation of axons in comparison with myelin. Recent evidence, however, demonstrates that axonal loss is also significant, affects long tracts such as the corticospinal and sensory tracts and relates closely to functional disability. Accordingly, the distribution and extent of this axonal loss is the focus of the current investigation. Post-mortem material of 55 multiple sclerosis patients and 32 matched controls was used to examine quantitatively the population of axons in the corticospinal tracts from the medulla to the lumbar spinal cord and the sensory tracts from the lumbar to the upper cervical spinal cord. Myelin- and axon-stained sections have been prepared to estimate the notional area and axon density, respectively of both tracts. Our results indicate that in the corticospinal tracts there is a significant reduction of the area and axon density at all levels investigated in multiple sclerosis cases when compared with controls. In contrast, the sensory tracts in multiple sclerosis cases showed a significant reduction in area and axon density only in the upper regions of the spinal cord. As has been found with MRI plaque load and T2 burden, correlations of axonal loss with duration of disease were not strong. Of the fibres lost in multiple sclerosis, we have found that small fibres (<3 microm2) seem to be particularly affected, with large fibres remaining relatively preserved in both the corticospinal and sensory tracts. In multiple sclerosis, axonal loss is widespread, and its extent is tract specific and size selective.

Evidence for a role for apoptosis in central pontine myelinolysis.

We have performed an immunocytochemical study of autopsy material from five cases of central pontine myelinolysis (CPM) and five age and sex-matched control subjects with the aim of exploring the possible involvement of apoptotic mechanisms in oligodendrocytes in this condition. We searched for immunoreactivity of glial cells for the apoptotic-related markers death receptor (DR) 3, Bax and Bak, the anti-apoptotic marker Bcl-2 and the cell cycle marker Ki-67. The latter marker was studied because it is known that entry of cells into the cell division cycle can trigger apoptosis if it does not result in mitosis. In CPM there was marked up-regulation of expression of the myelination-related enzyme carbonic anhydrase isoenzyme II and also markedly increased expression of Ki-67 in nuclei of glial cells of all types but particularly those of cells morphologically resembling oligodendrocytes. Despite this, there was no clear increase in density of such cells in CPM. The pro-apoptotic markers Bax, Bak and DR3 were all modestly increased in glial cell cytoplasm in CPM, while there was no change in expression of Bcl-2, which was only sparsely detected in glial cells both in controls and cases of CPM. The ratio of pro- to anti-apoptotic factors would appear from this evidence to have altered in favour of apoptosis in glial cells, most of which had the appearance of oligodendrocytes and many of which expressed Ki-67. We interpret this preliminary study as favouring apoptosis as a mechanism of oligodendrocytic death in CPM. Further study of this mechanism in CPM may lead to identification of factors that could reduce or prevent this rare but serious and often fatal disease.