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Sporadic amyotrophic lateral sclerosis (ALS), a devastating neurodegenerative disease, most likely results from complex genetic and environmental interactions. Although a number of association studies have been performed in an effort to find genetic components of sporadic ALS, most of them resulted in inconsistent findings due to a small number of genes investigated in relatively small sample sizes, while the replication of results was rarely attempted. Defects in retrograde axonal transport, vesicle trafficking and xenobiotic metabolism have been implicated in neurodegeneration and motor neuron death both in human disease and animal models. To assess the role of common genetic variation in these pathways in susceptibility to sporadic ALS, we performed a pathway-based candidate gene case-control association study with replication. Furthermore, we determined reliability of whole genome amplified DNA in a large-scale association study. In the first stage of the study, 1277 putative functional and tagging SNPs in 134 genes spanning 8.7 Mb were genotyped in 822 British sporadic ALS patients and 872 controls using whole genome amplified DNA. To detect variants with modest effect size and discriminate among false positive findings 19 SNPs showing a trend of association in the initial screen were genotyped in a replication sample of 580 German sporadic ALS patients and 361 controls. We did not detect strong evidence of association with any of the genes investigated in the discovery sample (lowest uncorrected P-value 0.00037, lowest permutation corrected P-value 0.353). None of the suggestive associations was replicated in a second sample, further excluding variants with moderate effect size. We conclude that common variation in the investigated pathways is unlikely to have a major effect on susceptibility to sporadic ALS. The genotyping efficiency was only slightly decreased ( approximately 1%) and genotyping quality was not affected using whole genome amplified DNA. It is reliable for large scale genotyping studies of diseases such as ALS, where DNA sample collections are limited because of low disease prevalence and short survival time. [ABSTRACT FROM AUTHOR]

Functional and structural age-associated changes in the blood-brain barrier (BBB) may affect the neurovascular unit and contribute to the onset and progression of age-associated neurodegenerative pathologies, including Alzheimer's disease. The current study interrogated the RNA profile of the BBB in an ageing human autopsy brain cohort and an ageing mouse model using combined laser capture microdissection and expression profiling. Only 12 overlapping genes were altered in the same direction in the BBB of both ageing human and mouse cohorts. These included genes with roles in regulating vascular tone, tight junction protein expression and cell adhesion, all processes prone to dysregulation with advancing age. Integrated mRNA and miRNA network and pathway enrichment analysis of the datasets identified 15 overlapping miRNAs that showed altered expression. In addition to targeting genes related to DNA binding and/or autophagy, many of the miRNAs identified play a role in age-relevant processes, including BBB dysfunction and regulating the neuroinflammatory response. Future studies have the potential to develop targeted therapeutic approaches against these candidates to prevent vascular dysfunction in the ageing brain.

Aims: While vascular pathology is a common feature of a range of neurodegenerative diseases, we hypothesized that vascular changes occur in association with normal ageing. Therefore, we aimed to characterize age-associated changes in the blood-brain barrier (BBB) in human and mouse cohorts., Methods: Immunohistochemistry and Evans blue assays were used to characterize BBB dysfunction (tight junction protein expression and serum plasma protein accumulation), vascular pathology (pericyte loss and vascular density) and glial pathology (astrocyte and microglial density) in ageing neurological control human prefrontal cortex (a total of 23 cases from 5 age groups representing the spectrum of young adult to old age: 20-30 years, 31-45 years, 46-60 years, 61-75 years and 75+) and C57BL/6 mice (3 months, 12 months, 18 months and 24 months, n = 5/6 per group)., Results: Quantification of the tight junction protein ZO-1 within the cortex and cerebellum of the mouse cohort showed a significant trend to both increased number (cortex P < 0.001, cerebellum P < 0.001) and length (cortex P < 0.001, cerebellum P < 0.001) of junctional breaks associated with increasing age. GFAP expression significantly correlated with ageing in the mice (P = 0.037). In the human cohort, assessment of human protein accumulation (albumin, fibrinogen and human IgG) demonstrated cells morphologically resembling clasmatodendritic astrocytes, indicative of BBB dysfunction. Semiquantitative assessment of astrogliosis in the cortex expression revealed an association with age (P = 0.003), while no age-associated changes in microglial pathology, microvascular density or pericyte coverage were detected., Conclusions: This study demonstrates BBB dysfunction in normal brain ageing, both in human and mouse cohorts., (© 2017 The Authors. Neuropathology and Applied Neurobiology published by John Wiley & Sons Ltd on behalf of British Neuropathological Society.)

Amyotrophic lateral sclerosis (ALS) is a fatal, neurodegenerative condition characterized by loss of motor neurones and progressive muscle wasting. There is no diagnostic test for ALS therefore robust biomarkers would not only be valuable for diagnosis, but also for the classification of disease subtypes, monitoring responses to drugs and tracking disease progression. As regulators of gene expression, microRNAs (miRNAs) are increasingly used for diagnostic and prognostic purposes in various disease states with increasing exploration in neurodegenerative disorders. We hypothesize that circulating blood-based miRNAs will serve as biomarkers and use miRNA profiling to determine miRNA signatures from the serum of sporadic ALS patients compared to healthy controls and patients with diseases that mimic ALS. A number of differentially expressed miRNAs were identified in each set of patient comparisons. Validation in an additional patient cohort showed that miR-206 and miR-143-3p were increased and miR-374b-5p was decreased compared to controls. A continued change in miRNA expression persisted during disease progression indicating the potential use of these particular miRNAs as longitudinal biomarkers in ALS., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

Amyotrophic lateral sclerosis (ALS) is characterized by motor neurone loss resulting in muscle weakness, spasticity and ultimately death. 5-10% are caused by inherited mutations, most commonly C9ORF72, SOD1, TARDBP and FUS. Rarer genetic causes of ALS include mutation of optineurin (mt OPTN). Furthermore, optineurin protein has been localized to the ubiquitylated aggregates in several neurodegenerative diseases, including ALS. This study: (i) investigated the frequency of mt OPTN in ALS patients in England; (ii) characterized the clinical and neuropathological features of ALS associated with a mt OPTN; and (iii) investigated optineurin neuropathology in C9ORF72-related ALS (C9ORF72-ALS). We identified a heterozygous p.E322K missense mutation in exon 10 of OPTN in one familial ALS patient who additionally had a C9ORF72 mutation. This patient had bulbar, limb and respiratory disease without cognitive problems. Neuropathology revealed motor neurone loss, trans-activation response DNA protein 43 (TDP-43)-positive neuronal and glial cytoplasmic inclusions together with TDP-43-negative neuronal cytoplasmic inclusions in extra motor regions that are characteristic of C9ORF72-ALS. We have demonstrated that both TDP-43-positive and negative inclusion types had positive staining for optineurin by immunohistochemistry. We went on to show that optineurin was present in TDP-43-negative cytoplasmic extra motor inclusions in C9ORF72-ALS cases that do not carry mt OPTN. We conclude that: (i) OPTN mutations are associated with ALS; (ii) optineurin protein is present in a subset of the extramotor inclusions of C9ORF72-ALS; (iii) It is not uncommon for multiple ALS-causing mutations to occur in the same patient; and (iv) studies of optineurin are likely to provide useful dataregarding the pathophysiology of ALS and neurodegeneration., (© 2015 Japanese Society of Neuropathology.)

Aims: Amyotrophic lateral sclerosis (ALS) and primary lateral sclerosis (PLS) are two syndromic variants within the motor neurone disease spectrum. As PLS and most ALS cases are sporadic (SALS), this limits the availability of cellular models for investigating pathogenic mechanisms and therapeutic targets. The aim of this study was to use gene expression profiling to evaluate fibroblasts as cellular models for SALS and PLS, to establish whether dysregulated biological processes recapitulate those seen in the central nervous system and to elucidate pathways that distinguish the clinically defined variants of SALS and PLS., Methods: Microarray analysis was performed on fibroblast RNA and differentially expressed genes identified. Genes in enriched biological pathways were validated by quantitative PCR and functional assays performed to establish the effect of altered RNA levels on the cellular processes., Results: Gene expression profiling demonstrated that whilst there were many differentially expressed genes in common between SALS and PLS fibroblasts, there were many more expressed specifically in the SALS fibroblasts, including those involved in RNA processing and the stress response. Functional analysis of the fibroblasts confirmed a significant decrease in miRNA production and a reduced response to hypoxia in SALS fibroblasts. Furthermore, metabolic gene changes seen in SALS, many of which were also evident in PLS fibroblasts, resulted in dysfunctional cellular respiration., Conclusions: The data demonstrate that fibroblasts can act as cellular models for ALS and PLS, by establishing the transcriptional changes in known pathogenic pathways that confer subsequent functional effects and potentially highlight targets for therapeutic intervention., (© 2014 The Authors. Neuropathology and Applied Neurobiology published by John Wiley & Sons Ltd. on behalf of British Neuropathological Society.)

MicroRNAs (miRNAs) are small, abundant RNA molecules that constitute part of the cell's non-coding RNA "dark matter." In recent years, the discovery of miRNAs has revolutionised the traditional view of gene expression and our understanding of miRNA biogenesis and function has expanded. Altered expression of miRNAs is increasingly recognized as a feature of many disease states, including neurodegeneration. Here, we review the emerging role for miRNA dysfunction in Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis (ALS) and Huntington's disease pathogenesis. We emphasize the complex nature of gene regulatory networks and the need for systematic studies, with larger sample cohorts than have so far been reported, to reveal the most important miRNA regulators in disease. Finally, miRNA diversity and their potential to target multiple pathways, offers novel clinical applications for miRNAs as biomarkers and therapeutic agents in neurodegenerative diseases.

Aims: Five to 10% of cases of amyotrophic lateral sclerosis are familial, with the most common genetic causes being mutations in the C9ORF72, SOD1, TARDBP and FUS genes. Mutations in the angiogenin gene, ANG, have been identified in both familial and sporadic patients in several populations within Europe and North America. The aim of this study was to establish the incidence of ANG mutations in a large cohort of 517 patients from Northern England and establish the neuropathology associated with these cases., Methods: The single exon ANG gene was amplified, sequenced and analysed for mutations. Pathological examination of brain, spinal cord and skeletal muscle included conventional histology and immunohistochemistry., Results: Mutation screening identified a single sporadic amyotrophic lateral sclerosis case with a p.K54E mutation, which is absent from 278 neurologically normal control samples. The clinical presentation was of limb onset amyotrophic lateral sclerosis, with rapid disease progression and no evidence of cognitive impairment. Neuropathological examination established the presence of characteristic ubiquitinated and TDP-43-positive neuronal and glial inclusions, but no abnormality in the distribution of angiogenin protein., Discussion: There is only one previous report describing the neuropathology in a single case with a p.K17I ANG mutation which highlighted the presence of eosinophilic neuronal intranuclear inclusions in the hippocampus. The absence of this feature in the present case indicates that patients with ANG mutations do not always have pathological changes distinguishable from those of sporadic amyotrophic lateral sclerosis., (© 2012 The Authors. Neuropathology and Applied Neurobiology © 2012 British Neuropathological Society.)

While some cases of familial ALS can be entirely attributed to known inherited variation, the majority (∼ 90%) are sporadic, where the cause(s) are not entirely understood. Both genetic and environmental factors may contribute to susceptibility. Mitochondrial damage, a common feature of neurodegenerative disease, is observed in most patients and inherited polymorphism in the mitochondrial genome has been suggested as a contributing factor. We used an economic and efficient method to test whether such involvement is probable. We genotyped 22 mtDNA coding region SNPs and sequenced the mtDNA hypervariable region 1 to determine the position of each mitochondrial genome within the genealogy of mitochondrial haplotypes in samples of ALS patients (n = 700) and controls (n = 462) from two European populations. We compared haplotype and haplogroup distribution in cases and controls drawn from the same populations. No statistical difference was observed between cases and controls at either the haplogroup or haplotype level (p = ≥  0.2). In conclusion, it is unlikely that common, shared genetic variants in the mitochondrial genome contribute substantially to ALS. Combining the data with other studies will allow meta-analysis to look for variants with modest effect sizes. The sequencing of complete mitochondrial genomes will be required to assess the role of rare mutations.

The finding of TDP-43 as a major component of ubiquitinated protein inclusions in amyotrophic lateral sclerosis (ALS) has led to the identification of 30 mutations in the transactive response-DNA binding protein (TARDBP) gene, encoding TDP-43. All but one are in exon 6, which encodes the glycine-rich domain. The aim of this study was to determine the frequency of TARDBP mutations in a large cohort of motor neurone disease patients from Northern England (42 non-superoxide dismutase 1 (SOD1) familial ALS (FALS), nine ALS-frontotemporal dementia, 474 sporadic ALS (SALS), 45 progressive muscular atrophy cases). We identified four mutations, two of which were novel, in two familial (FALS) and two sporadic (SALS) cases, giving a frequency of TARDBP mutations in non-SOD1 FALS of 5% and SALS of 0.4%. Analysis of clinical data identified that patients had typical ALS, with limb or bulbar onset, and showed considerable variation in age of onset and rapidity of disease course. However, all cases had an absence of clinically overt cognitive dysfunction.

Background: Amyotrophic lateral sclerosis (ALS), a common late-onset neurodegenerative disease, is associated with fronto-temporal dementia (FTD) in 3-10% of patients. A mutation in CHMP2B was recently identified in a Danish pedigree with autosomal dominant FTD. Subsequently, two unrelated patients with familial ALS, one of whom also showed features of FTD, were shown to carry missense mutations in CHMP2B. The initial aim of this study was to determine whether mutations in CHMP2B contribute more broadly to ALS pathogenesis., Methodology/principal Findings: Sequencing of CHMP2B in 433 ALS cases from the North of England identified 4 cases carrying 3 missense mutations, including one novel mutation, p.Thr104Asn, none of which were present in 500 neurologically normal controls. Analysis of clinical and neuropathological data of these 4 cases showed a phenotype consistent with the lower motor neuron predominant (progressive muscular atrophy (PMA)) variant of ALS. Only one had a recognised family history of ALS and none had clinically apparent dementia. Microarray analysis of motor neurons from CHMP2B cases, compared to controls, showed a distinct gene expression signature with significant differential expression predicting disassembly of cell structure; increased calcium concentration in the ER lumen; decrease in the availability of ATP; down-regulation of the classical and p38 MAPK signalling pathways, reduction in autophagy initiation and a global repression of translation. Transfection of mutant CHMP2B into HEK-293 and COS-7 cells resulted in the formation of large cytoplasmic vacuoles, aberrant lysosomal localisation demonstrated by CD63 staining and impairment of autophagy indicated by increased levels of LC3-II protein. These changes were absent in control cells transfected with wild-type CHMP2B., Conclusions/significance: We conclude that in a population drawn from North of England pathogenic CHMP2B mutations are found in approximately 1% of cases of ALS and 10% of those with lower motor neuron predominant ALS. We provide a body of evidence indicating the likely pathogenicity of the reported gene alterations. However, absolute confirmation of pathogenicity requires further evidence, including documentation of familial transmission in ALS pedigrees which might be most fruitfully explored in cases with a LMN predominant phenotype.

Iron misregulation promotes oxidative stress, a proposed pathological mechanism in neurodegenerative disease. The aim of this study was to evaluate serum iron metabolism indicators in 60 amyotrophic lateral sclerosis (ALS) patients and 44 age matched controls. Serum ferritin levels were significantly increased in ALS patients compared to controls (p < 0.001), while no differences in the levels of serum iron, transferrin, iron saturation or total iron binding capacity were found. Likewise no differences in C reactive protein (CRP) or caeruloplasmin were detected, suggesting that the elevated ferritin levels in ALS did not merely indicate an acute phase response. The increased ferritin level may reflect a general increase in stored iron or be a consequence of ongoing muscle degeneration.

There is increasing evidence of a clinical, neuropathological and genetic overlap between frontotemporal dementia (FTD) and amyotrophic lateral sclerosis (ALS). We conducted a case-control study using a UK dataset to test the hypothesis that polymorphisms in two FTD-related genes (GRN and FT74) are associated with increased susceptibility to ALS. We evaluated the majority of known genetic variability in IFT74 and GRN. The results revealed that the common variations in IFT74 and GRN neither constitute strong ALS risk factors nor modify the age-at-onset. However, the possibility of a modest risk effect remains to be assessed in large datasets.

Mutations in the progranulin gene (GRN) are a major cause of frontotemporal lobar degeneration with ubiquitin-positive, tau-negative inclusions (FTLD-U) but the distinguishing clinical and anatomical features of this subgroup remain unclear. In a large UK cohort we found five different frameshift and premature termination mutations likely to be causative of FTLD in 25 affected family members. A previously described 4-bp insertion mutation in GRN exon 2 comprised the majority of cases in our cohort (20/25), with four novel mutations being identified in the other five affected members. Additional novel missense changes were discovered, of uncertain pathogenicity, but deletion of the entire gene was not detected. The patient collection was investigated by a single tertiary referral centre and is enriched for familial early onset FTLD with a high proportion of patients undergoing neuropsychological testing, MRI and eventual neuropathological diagnosis. Age at onset was variable, but four mutation carriers presented in their 40s and when analysed as a group, the mean age at onset of disease in GRN mutation carriers was later than tau gene (MAPT) mutation carriers and duration of disease was shorter when compared with both MAPT and FTLD-U without mutation. The most common clinical presentation seen in GRN mutation carriers was behavioural variant FTLD with apathy as the dominant feature. However, many patients had language output impairment that was either a progressive non-fluent aphasia or decreased speech output consistent with a dynamic aphasia. Neurological and neuropsychological examination also suggests that parietal lobe dysfunction is a characteristic feature of GRN mutation and differentiates this group from other patients with FTLD. MR imaging showed evidence of strikingly asymmetrical atrophy with the frontal, temporal and parietal lobes all affected. Both right- and left-sided predominant atrophy was seen even within the same family. As a group, the GRN carriers showed more asymmetry than in other FTLD groups. All pathologically investigated cases showed extensive type 3 TDP-43-positive pathology, including frequent neuronal cytoplasmic inclusions, dystrophic neurites in both grey and white matter and also neuronal intranuclear inclusions. Finally, we confirmed a modifying effect of APOE-E4 genotype on clinical phenotype with a later onset in the GRN carriers suggesting that this gene has distinct phenotypic effects in different neurodegenerative diseases.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterised by loss of motor neurons. The cause of disease is unknown other than in the rare cases of familial disease arising from mutations in the superoxide dismutase 1 gene. Many theories for pathogenesis have been proposed - including oxidative stress, excitotoxicity, mitochondrial dysfunction and abnormal protein aggregation - based on studies of human post mortem tissue, research on animal models, and in vitro work. Here we review the evidence for the main pathogenic mechanisms and outline how they might interact to cause motor neuron death. Clinical trials have as yet failed to identify any truly effective therapies in ALS, with only riluzole providing a modest improvement in survival. Ongoing trials are exploring the value of antiglutamatergic agents, including the cephalosporin antibiotic ceftriaxone, as well as antioxidants, mitochondrial enhancers and anti-apoptotic drugs. It is likely that effective therapy will involve combinations of agents acting on different mechanisms. Gene therapy with neurotrophic factors will soon be in clinical trials, while work on stem cell therapy remains preclinical. In addition to finding effective therapies, research also needs to identify early disease markers because therapy is likely to be of most benefit when given early in the course of disease.