Clinicopathological correlates in the frontotemporal lobar degeneration-motor neuron disease spectrum.

Amyotrophic lateral sclerosis (ALS) is a devastating motor neuron disease (MND) that shares a common clinical, genetic and pathologic spectrum with frontotemporal dementia (FTD). It is highly heterogeneous in its presentation and features. Up to 50% of patients with MND develop cognitive-behavioural symptoms during the course of the disease, meeting criteria for FTD in 10%-15% of cases. In the absence of a precise biomarker, neuropathology is still a valuable tool to understand disease nosology, reach a definite diagnostic confirmation and help define specific subgroups of patients with common phenotypic, genetic and biomarker profiles. However, few neuropathological series have been published, and the frequency of frontotemporal lobar degeneration (FTLD) in MND is difficult to estimate. In this work we describe a large clinicopathological series of MND patients, analysing the frequency of concurrent FTLD changes and trying to define specific subgroups of patients based on their clinical, genetic and pathological characteristics. We performed an observational, retrospective, multicentre case study. We included all cases meeting neuropathological criteria for MND from the Neurological Tissue Bank of the FRCB-IDIBAPS-Hospital Clínic Barcelona Biobank between 1994 and 2022, regardless of their last clinical diagnosis. While brain donation is encouraged in all patients, it is performed in very few, and representativeness of the cohort might not be precise for all patients with MND. We retrospectively reviewed clinical and neuropathological data and describe the main clinical, genetic and pathogenic features, comparing neuropathologic groups between MND with and without FTLD changes and aiming to define specific subgroups. We included brain samples from 124 patients, 44 of whom (35.5%) had FTLD neuropathologic features (i.e. FTLD-MND). Pathologic TDP-43 aggregates were present in 93.6% of the cohort and were more extensive (higher Brettschneider stage) in those with concurrent FTLD (P < 0.001). Motor symptom onset was more frequent in the bulbar region in FTLD-MND cases than in those with isolated MND (P = 0.023), with no differences in survival. We observed a better clinicopathological correlation in the MND group than in the FTLD-MND group (93.8% versus 61.4%; P < 0.001). Pathogenic genetic variants were more common in the FTLD-MND group, especially C9orf72. We describe a frequency of FTLD of 35.5% in our series of neuropathologically confirmed cases of MND. The FTLD-MND spectrum is highly heterogeneous in all aspects, especially in patients with FTLD, in whom it is particularly difficult to define specific subgroups. In the absence of definite biomarkers, neuropathology remains a valuable tool for a definite diagnosis, increasing our knowledge in disease nosology.

TDP-43 regulates LC3ylation in neural tissue through ATG4B cryptic splicing inhibition.

Amyotrophic lateral sclerosis (ALS) is an adult-onset motor neuron disease with a mean survival time of three years. The 97% of the cases have TDP-43 nuclear depletion and cytoplasmic aggregation in motor neurons. TDP-43 prevents non-conserved cryptic exon splicing in certain genes, maintaining transcript stability, including ATG4B, which is crucial for autophagosome maturation and Microtubule-associated proteins 1A/1B light chain 3B (LC3B) homeostasis. In ALS mice (G93A), Atg4b depletion worsens survival rates and autophagy function. For the first time, we observed an elevation of LC3ylation in the CNS of both ALS patients and atg4b-/- mouse spinal cords. Furthermore, LC3ylation modulates the distribution of ATG3 across membrane compartments. Antisense oligonucleotides (ASOs) targeting cryptic exon restore ATG4B mRNA in TARDBP knockdown cells. We further developed multi-target ASOs targeting TDP-43 binding sequences for a broader effect. Importantly, our ASO based in peptide-PMO conjugates show brain distribution post-IV administration, offering a non-invasive ASO-based treatment avenue for neurodegenerative diseases.

Genetic variability in sporadic amyotrophic lateral sclerosis.

With the advent of gene therapies for amyotrophic lateral sclerosis (ALS), there is a surge in gene testing for this disease. Although there is ample experience with gene testing for C9orf72, SOD1, FUS and TARDBP in familial ALS, large studies exploring genetic variation in all ALS-associated genes in sporadic ALS (sALS) are still scarce. Gene testing in a diagnostic setting is challenging, given the complex genetic architecture of sALS, for which there are genetic variants with large and small effect sizes. Guidelines for the interpretation of genetic variants in gene panels and for counselling of patients are lacking. We aimed to provide a thorough characterization of genetic variability in ALS genes by applying the American College of Medical Genetics and Genomics (ACMG) criteria on whole genome sequencing data from a large cohort of 6013 sporadic ALS patients and 2411 matched controls from Project MinE. We studied genetic variation in 90 ALS-associated genes and applied customized ACMG-criteria to identify pathogenic and likely pathogenic variants. Variants of unknown significance were collected as well. In addition, we determined the length of repeat expansions in C9orf72, ATXN1, ATXN2 and NIPA1 using the ExpansionHunter tool. We found C9orf72 repeat expansions in 5.21% of sALS patients. In 50 ALS-associated genes, we did not identify any pathogenic or likely pathogenic variants. In 5.89%, a pathogenic or likely pathogenic variant was found, most commonly in SOD1, TARDBP, FUS, NEK1, OPTN or TBK1. Significantly more cases carried at least one pathogenic or likely pathogenic variant compared to controls (odds ratio 1.75; P-value 1.64 × 10-5). Isolated risk factors in ATXN1, ATXN2, NIPA1 and/or UNC13A were detected in 17.33% of cases. In 71.83%, we did not find any genetic clues. A combination of variants was found in 2.88%. This study provides an inventory of pathogenic and likely pathogenic genetic variation in a large cohort of sALS patients. Overall, we identified pathogenic and likely pathogenic variants in 11.13% of ALS patients in 38 known ALS genes. In line with the oligogenic hypothesis, we found significantly more combinations of variants in cases compared to controls. Many variants of unknown significance may contribute to ALS risk, but diagnostic algorithms to reliably identify and weigh them are lacking. This work can serve as a resource for counselling and for the assembly of gene panels for ALS. Further characterization of the genetic architecture of sALS is necessary given the growing interest in gene testing in ALS.

Neurodegeneration Biomarkers in Adult Spinal Muscular Atrophy (SMA) Patients Treated with Nusinersen.

The objective of this study is to evaluate biomarkers for neurodegenerative disorders in adult SMA patients and their potential for monitoring the response to nusinersen. Biomarkers for neurodegenerative disorders were assessed in plasma and CSF samples obtained from a total of 30 healthy older adult controls and 31 patients with adult SMA type 2 and 3. The samples were collected before and during nusinersen treatment at various time points, approximately at 2, 6, 10, and 22 months. Using ELISA technology, the levels of total tau, pNF-H, NF-L, sAPPß, Aß40, Aß42, and YKL-40 were evaluated in CSF samples. Additionally, plasma samples were used to measure NF-L and total tau levels using SIMOA technology. SMA patients showed improvements in clinical outcomes after nusinersen treatment, which were statistically significant only in walkers, in RULM (p = 0.04) and HFMSE (p = 0.05) at 24 months. A reduction in sAPPß levels was found after nusinersen treatment, but these levels did not correlate with clinical outcomes. Other neurodegeneration biomarkers (NF-L, pNF-H, total tau, YKL-40, Aß40, and Aß42) were not found consistently changed with nusinersen treatment. The slow progression rate and mild treatment response of adult SMA types 2 and 3 may not lead to detectable changes in common markers of axonal degradation, inflammation, or neurodegeneration, since it does not involve large pools of damaged neurons as observed in pediatric forms. However, changes in biomarkers associated with the APP processing pathway might be linked to treatment administration. Further studies are warranted to better understand these findings.

The Effect of SMN Gene Dosage on ALS Risk and Disease Severity.

OBJECTIVE: The role of the survival of motor neuron (SMN) gene in amyotrophic lateral sclerosis (ALS) is unclear, with several conflicting reports. A decisive result on this topic is needed, given that treatment options are available now for SMN deficiency. METHODS: In this largest multicenter case control study to evaluate the effect of SMN1 and SMN2 copy numbers in ALS, we used whole genome sequencing data from Project MinE data freeze 2. SMN copy numbers of 6,375 patients with ALS and 2,412 controls were called from whole genome sequencing data, and the reliability of the calls was tested with multiplex ligation-dependent probe amplification data. RESULTS: The copy number distribution of SMN1 and SMN2 between cases and controls did not show any statistical differences (binomial multivariate logistic regression SMN1 p = 0.54 and SMN2 p = 0.49). In addition, the copy number of SMN did not associate with patient survival (Royston-Parmar; SMN1 p = 0.78 and SMN2 p = 0.23) or age at onset (Royston-Parmar; SMN1 p = 0.75 and SMN2 p = 0.63). INTERPRETATION: In our well-powered study, there was no association of SMN1 or SMN2 copy numbers with the risk of ALS or ALS disease severity. This suggests that changing SMN protein levels in the physiological range may not modify ALS disease course. This is an important finding in the light of emerging therapies targeted at SMN deficiencies. ANN NEUROL 2021;89:686-697.

Validation of motor and functional scales for the evaluation of adult patients with 5q spinal muscular atrophy.

BACKGROUND AND PURPOSE: Mos scales currently used to evaluate spinal muscular atrophy (SMA) patients have only been validated in children. The aim of this study was to assess the construct validity and responsiveness of several outcome measures in adult SMA patients. METHODS: Patients older than 15 years and followed up in five referral centres for at least 6 months, between October 2015 and August 2020, with a motor function scale score (Hammersmith Functional Motor Scale Expanded [HFMSE], Revised Upper Limb module [RULM]) were included. Bedside functional scales (Egen Klassification [EK2], Revised Amyotrophic Lateral Sclerosis Functional Rating Scale [ALSFRS-R]) were also collected when available. Spearman's rho correlations (rs) and Bangdiwala's concordance test (B) were used to evaluate the scales' construct validity. Monthly slopes of change were used to calculate their responsiveness of the scales. RESULTS: The study included 79 SMA patients, followed up for a mean of 16 months. All scales showed strong correlations with each other (rs > 0.70). A floor effect in motor function scales was found in the weakest patients (HFMSE < 5 and RULM < 10), and a ceiling effect was found in stronger patients (HFMSE > 60 and RULM > 35). The ALSFRS-R (B = 0.72) showed a strong ability to discriminate between walkers, sitters and non-sitters, and the HFMSE (B = 0.86) between walkers and sitters. The responsiveness was low overall, although in treated patients a moderate responsiveness was found for the ALSFRS-R and HFMSE in walkers (0.69 and 0.61, respectively) and for EK2 in sitters (0.65) and non-sitters (0.60). CONCLUSIONS: This study shows the validity and limitations of the scales most frequently used to assess adult SMA patients. Overall, bedside functional scales showed some advantages over motor scales, although all showed limited responsiveness.

Characterizing SOD1 mutations in Spain. The impact of genotype, age, and sex in the natural history of the disease.

INTRODUCTION: The aim of this study is to describe the frequency and distribution of SOD1 mutations in Spain, and to explore those factors contributing to their phenotype and prognosis. METHODS: Seventeen centres shared data on amyotrophic lateral sclerosis (ALS) patients carrying pathogenic or likely pathogenic SOD1 variants. Multivariable models were used to explore prognostic modifiers. RESULTS: In 144 patients (from 88 families), 29 mutations (26 missense, 2 deletion/insertion and 1 frameshift) were found in all 5 exons of SOD1, including 7 novel mutations. 2.6% of ALS patients (including 17.7% familial and 1.3% sporadic) were estimated to carry SOD1 mutations. Its frequency varied considerably between regions, due to founder events. The most frequent mutation was p.Gly38Arg (n = 58), followed by p.Glu22Gly (n = 11), p.Asn140His (n = 10), and the novel p.Leu120Val (n = 10). Most mutations were characterized by a protracted course, and some of them by atypical phenotypes. Older age of onset was independently associated with faster disease progression (exp(Estimate) = 1.03 [0.01, 0.05], p = 0.001) and poorer survival (HR = 1.05 [1.01, 1.08], p = 0.007), regardless of the underlying mutation. Female sex was independently associated to faster disease progression (exp(Estimate) = 2.1 [1.23, 3.65], p = 0.012) in patients carrying the p.Gly38Arg mutation, resulting in shorter survival compared with male carriers (236 vs 301 months). CONCLUSIONS: These data may help to evaluate the efficacy of SOD1 targeted treatments, and to expand the number of patients that might benefit from these treatments.

Nusinersen in adult patients with 5q spinal muscular atrophy: A multicenter observational cohorts' study.

BACKGROUND AND PURPOSE: The aim was to assess the safety and efficacy of nusinersen in adult 5q spinal muscular atrophy (SMA) patients. METHODS: Patients older than 15 years and followed for at least 6 months with one motor scale (Hammersmith Functional Motor Scale Expanded, HFMSE; Revised Upper Limb Module, RULM) in five referral centers were included. The clinical and patients' global impression of change (CGI-C and PGI-C) were recorded in treated patients at the last visit. Functional scales (Egen Klassification, EK2; Revised Amyotrophic Lateral Sclerosis Functional Rating Scale, ALSFRS-R) and the percentage predicted forced vital capacity were collected when available. RESULTS: Seventy-nine SMA patients (39 treated with nusinersen) were included. Compared with untreated patients, treated patients showed a significant improvement of 2 points (±0.46) in RULM (p < 0.001) after 6 months. After a mean follow-up of 16 months, nusinersen treatment was associated with a significant improvement in HFMSE (odds ratio [OR] 1.15, p = 0.006), the 6-min walk test (OR = 1.07, p < 0.001) and the EK2 (OR = 0.81, p = 0.001). Compared with untreated patients, more treated patients experienced clinically meaningful improvements in all scales, but these differences were statistically significant only for RULM (p = 0.033), ALSFRS-R (p = 0.005) and EK2 (p < 0.001). According to the CGI-C and PGI-C, 64.1% and 61.5% of treated patients improved with treatment. Being a non-sitter was associated with less response to treatment, whilst a longer time of treatment was associated with better response. Most treated patients (77%) presented at least one adverse event, mostly mild. CONCLUSIONS: Nusinersen treatment is associated with some improvements in adult SMA patients. Most severely affected patients with complex spines are probably those with the most unfavorable risk-benefit ratio.

Plasma exchange with albumin replacement and disease progression in amyotrophic lateral sclerosis: a pilot study.

BACKGROUND: Plasma exchange (PE) is used to treat a range of neurological disorders. Based on results demonstrated in Alzheimer's disease, we theorized that PE with albumin replacement (PE-A) might alter the metabolic profile of plasma and cerebrospinal fluid in patients with amyotrophic lateral sclerosis (ALS) by removing disease-inducing molecules. The aim of this study was to evaluate the effect of PE-A on disease progression in ALS. METHODS: In this open-label, non-controlled, single-arm, prospective pilot study, 13 adults with ALS had 6 months' treatment with PE-A 5% and 6 months' follow-up. Primary endpoints were changes from baseline in the Amyotrophic Lateral Sclerosis Functional Rating Scale-Revised (ALSFRS-R) score and forced vital capacity (FVC) through 48 weeks. A post hoc analysis compared individual patient data with the expected ALSFRS-R progression slope. RESULTS: The median ALSFRS-R score declined throughout the study, although the rate of decline was slower than expected in seven patients at treatment end and in five patients at study end. Six patients remained in the same baseline slope progression category, and four patients improved their slope category at treatment end. Median FVC decreased significantly during the study. Treatment was well tolerated. Of 330 PE-A procedures, 0.9% were associated with potentially related adverse events. CONCLUSION: Although functional impairment progressed, about two-thirds of patients showed a slower than expected rate of decline at treatment end. Most patients had unaltered (54.5%) or reduced (36.4%) ALSFRS-R slope progression at treatment end. Further evaluation of PE-A in controlled studies involving more patients is warranted. EUDRACT NUMBER: 2013-004842-40. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT02479802.

Detecting Bulbar Involvement in Patients with Amyotrophic Lateral Sclerosis Based on Phonatory and Time-Frequency Features.

The term "bulbar involvement" is employed in ALS to refer to deterioration of motor neurons within the corticobulbar area of the brainstem, which results in speech and swallowing dysfunctions. One of the primary symptoms is a deterioration of the voice. Early detection is crucial for improving the quality of life and lifespan of ALS patients suffering from bulbar involvement. The main objective, and the principal contribution, of this research, was to design a new methodology, based on the phonatory-subsystem and time-frequency characteristics for detecting bulbar involvement automatically. This study focused on providing a set of 50 phonatory-subsystem and time-frequency features to detect this deficiency in males and females through the utterance of the five Spanish vowels. Multivariant Analysis of Variance was then used to select the statistically significant features, and the most common supervised classifications models were analyzed. A set of statistically significant features was obtained for males and females to capture this dysfunction. To date, the accuracy obtained (98.01% for females and 96.10% for males employing a random forest) outperformed the models in the literature. Adding time-frequency features to more classical phonatory-subsystem features increases the prediction capabilities of the machine-learning models for detecting bulbar involvement. Studying men and women separately gives greater success. The proposed method can be deployed in any kind of recording device (i.e., smartphone).

Elevated Cerebrospinal Fluid Proteins and Albumin Determine a Poor Prognosis for Spinal Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a heterogeneous disease, both in its onset phenotype and in its rate of progression. The aim of this study was to establish whether the dysfunction of the blood-brain barrier (BBB) and blood-spinal cord barrier (BSCB) measured through cerebrospinal fluid (CSF) proteins and the albumin-quotient (QAlb) are related to the speed of disease progression. An amount of 246 patients diagnosed with ALS were included. CSF and serum samples were determined biochemically for different parameters. Survival analysis based on phenotype shows higher probability of death for bulbar phenotype compared to spinal phenotype (p-value: 0.0006). For the effect of CSF proteins, data shows an increased risk of death for spinal ALS patients as the value of CSF proteins increases. The same model replicated for CSF albumin yielded similar results. Statistical models determined that the lowest cut-off value for CSF proteins able to differentiate patients with a good prognosis and worse prognosis corresponds to CSF proteins ≥ 0.5 g/L (p-value: 0.0189). For the CSF albumin, the QAlb ≥0.65 is associated with elevated probability of death (p-value: 0.0073). High levels of QAlb are a bad prognostic indicator for the spinal phenotype, in addition to high CSF proteins levels that also act as a marker of poor prognosis.

Deep Molecular Characterization of Milder Spinal Muscular Atrophy Patients Carrying the c.859G>C Variant in SMN2

Spinal muscular atrophy (SMA) is a severe neuromuscular disorder caused by biallelic loss or pathogenic variants in the SMN1 gene. Copy number and modifier intragenic variants in SMN2, an almost identical paralog gene of SMN1, are known to influence the amount of complete SMN proteins. Therefore, SMN2 is considered the main phenotypic modifier of SMA, although genotype−phenotype correlation is not absolute. We present eleven unrelated SMA patients with milder phenotypes carrying the c.859G>C-positive modifier variant in SMN2. All were studied by a specific NGS method to allow a deep characterization of the entire SMN region. Analysis of two homozygous cases for the variant allowed us to identify a specific haplotype, Smn2-859C.1, in association with c.859G>C. Two other cases with the c.859G>C variant in their two SMN2 copies showed a second haplotype, Smn2-859C.2, in cis with Smn2-859C.1, assembling a more complex allele. We also identified a previously unreported variant in intron 2a exclusively linked to the Smn2-859C.1 haplotype (c.154-1141G>A), further suggesting that this region has been ancestrally conserved. The deep molecular characterization of SMN2 in our cohort highlights the importance of testing c.859G>C, as well as accurately assessing the SMN2 region in SMA patients to gain insight into the complex genotype−phenotype correlations and improve prognostic outcomes.

Nuclear lipidome is altered in amyotrophic lateral sclerosis: A pilot study.

Nucleocytosolic transport, a membrane process, is impaired in motor neurons in amyotrophic lateral sclerosis (ALS). This study analyzes the nuclear lipidome in motor neurons in ALS and examines molecular pathways linked to the major lipid alterations. Nuclei were obtained from the frozen anterior horn of the lumbar spinal cord of ALS patients and age-matched controls. Lipidomic profiles of this subcellular fraction were obtained using liquid chromatography and mass spectrometry. We validated the mechanisms behind presumable lipidomic changes by exploring ALS surrogate models including human motor neurons (derived from ALS lines and controls) subjected to oxidative stress, the hSOD-G93A transgenic mice, and samples from an independent cohort of ALS patients. Among the differential lipid species, we noted 41 potential identities, mostly belonging to phospholipids (particularly ether phospholipids, as plasmalogens), as well as diacylglycerols and triacylglycerides. Decreased expression of alkyldihydroxyacetonephosphate synthase (AGPS)-a critical peroxisomal enzyme in plasmalogen synthesis-is found in motor neuron disease models; this occurs in parallel with an increase in the expression of sterol carrier protein 2 (SCP2) mRNA in ALS and Scp2 levels in G93A transgenic mice. Further, we identified diminished expression of diacylglycerol-related enzymes, such as phospholipase C ßI (PLCßI) and protein kinase CßII (PKCßII), linked to diacylglycerol metabolism. Finally, lipid droplets were recognized in the nuclei, supporting the identification of triacylglycerides as differential lipids. Our results point to the potentially pathogenic role of altered composition of nuclear membrane lipids and lipids in the nucleoplasm in the anterior horn of the spinal cord in ALS. Overall, these data support the usefulness of subcellular lipidomics applied to neurodegenerative diseases.

Lipid alterations in human frontal cortex in ALS-FTLD-TDP43 proteinopathy spectrum are partly related to peroxisome impairment.

AIM: Peroxisomes play a key role in lipid metabolism, and peroxisome defects have been associated with neurodegenerative diseases such as X-adrenoleukodystrophy and Alzheimer's disease. This study aims to elucidate the contribution of peroxisomes in lipid alterations of area 8 of the frontal cortex in the spectrum of TDP43-proteinopathies. Cases of frontotemporal lobar degeneration-TDP43 (FTLD-TDP), manifested as sporadic (sFTLD-TDP) or linked to mutations in various genes including expansions of the non-coding region of C9ORF72 (c9FTLD), and of sporadic amyotrophic lateral sclerosis (sALS) as the most common TDP43 proteinopathies, were analysed. METHODS: We used transcriptomics and lipidomics methods to define the steady-state levels of gene expression and lipid profiles. RESULTS: Our results show alterations in gene expression of some components of peroxisomes and related lipid pathways in frontal cortex area 8 in sALS, sFTLD-TDP and c9FTLD. Additionally, we identify a lipidomic pattern associated with the ALS-FTLD-TDP43 proteinopathy spectrum, notably characterised by down-regulation of ether lipids and acylcarnitine among other lipid species, as well as alterations in the lipidome of each phenotype of TDP43 proteinopathy, which reveals commonalities and disease-dependent differences in lipid composition. CONCLUSION: Globally, lipid alterations in the human frontal cortex of the ALS-FTLD-TDP43 proteinopathy spectrum, which involve cell membrane composition and signalling, vulnerability against cellular stress and possible glucose metabolism, are partly related to peroxisome impairment.

Evaluation of Dysphagia in Motor Neuron Disease. Review of Available Diagnostic Tools and New Perspectives.

Oropharyngeal dysphagia (OD) is highly prevalent (up to 80%) in patients with motor neuron disease (MND), influencing the prognosis of the disease. The clinical assessment of dysphagia is complex. There are assessment scales and screening questionnaires, but they have not been tested in patients with MND. In a sample of 46 patients with MND, the sensitivity and specificity of the EAT-10 and SwalQoL questionnaires, as well as the ALS-SS and FOIS scales, were tested and compared to the gold standard technique (videofluoroscopy, VFS). The patients were stratified using the DOSSc variable according to the video fluoroscopic examination with (n = 37) or without (n = 8) signs of dysphagia, and the results were compared with the scores obtained in the dysphagia questionnaires. None of the studied questionnaires was more sensitive than the others, but one stood out for its high specificity (= 1): the SwalQoL revised FS. The symptom frequency section of the SwalQoL questionnaire with some modifications, (SwalQoL revised FS) may be a useful tool in the clinical assessment of dysphagia because it's capable to detect the patients that really don't have dysphagia. The ALS-SS showed the greatest validity as a severity scale of dysphagia among the sample studied. A specific questionnaire for screening for dysphagia in MND needs to be developed. Until that time, the proposal is to use a combination of the existing questionnaires for other pathologies (EAT-10 and SwalQoL) and the specific scale for MND, the ALS-SS, to make an accurately clinical assessment of OD in MND patients before to perform a videofluoroscopy.

Cell Stress Induces Mislocalization of Transcription Factors with Mitochondrial Enrichment.

Previous evidence links the formation of extranuclear inclusions of transcription factors, such as ERK, Jun, TDP-43, and REST, with oxidative, endoplasmic-reticulum, proteasomal, and osmotic stress. To further characterize its extranuclear location, we performed a high-content screening based on confocal microscopy and automatized image analyses of an epithelial cell culture treated with hydrogen peroxide, thapsigargin, epoxomicin, or sorbitol at different concentrations and times to recreate the stresses mentioned above. We also performed a subcellular fractionation of the brain from transgenic mice overexpressing the Q331K-mutated TARDBP, and we analyzed the REST-regulated mRNAs. The results show that these nuclear proteins exhibit a mitochondrial location, together with significant nuclear/extranuclear ratio changes, in a protein and stress-specific manner. The presence of these proteins in enriched mitochondrial fractions in vivo confirmed the results of the image analyses. TDP-43 aggregation was associated with alterations in the mRNA levels of the REST target genes involved in calcium homeostasis, apoptosis, and metabolism. In conclusion, cell stress increased the mitochondrial translocation of nuclear proteins, increasing the chance of proteostasis alterations. Furthermore, TDP-43 aggregation impacts REST target genes, disclosing an exciting interaction between these two transcription factors in neurodegenerative processes.

[Amyotrophic lateral sclerosis (ALS) from the perspective of Primary Care. Epidemiology and clinical-care characteristics]. / La esclerosis lateral amiotrófica (ELA) desde la Atención Primaria. Epidemiología y características clínico-asistenciales.

Amyotrophic Lateral Sclerosis (ALS) is a rare disease in primary care (PC), it represents a challenge for the family doctor, especially in home care. OBJECTIVE: To know the incidence and prevalence of ALS in an area of ??PA management, the clinical characteristics and use of health resources. DESIGN: Observational study. LOCATION: PC-Direction Costa de Ponent, South Metropolitan Health Region, Barcelona, Catalonia, Spain. PARTICIPANTS: Patients with ALS ≥18 years diagnosed until 03/01/2017. Main measurements Age, sex, characteristics: form of appearance (spinal, bulbar, others), interval between onset of symptoms and diagnosis, percutaneous gastrostomy carriers, ventilation non-invasive or invasive. Identification in PC as a Complex Chronic Patient or with palliative needs (CCP). Inclusion in home care programs (PAD). Model of attention hospitable. RESULTS: 81 patients, mean age 65.6 years (± 11.7), men 49.4%. Shape of onset: spinal 69%, bulbar 21%, another 4%. Interval between the onset of symptoms and diagnosis 12 months. Identified as a CCP 13.6%, 29 patients (35.8%) included in PAD. Attended in comprehensive hospital model 79 patients (97.5%). Prevalence 6.1/100,000 inhabitants in 2017. Annual incidence between 1.2 cases/100,000 inhabitants/year in 2012 and 3.5 cases/100,000 inhabitants/year in 2016. CONCLUSIONS: The use of percutaneous gastrostomy in ALS favors the identification as CCP or with palliative needs and inclusion in PAD. The use of non-invasive ventilation favors inclusion in PAD. The incidence and prevalence data for ALS are higher than those described above in the same area. Early identification is necessary of these patients in the chronic care models in PC teams.

Increased C-X-C Motif Chemokine Ligand 12 Levels in Cerebrospinal Fluid as a Candidate Biomarker in Sporadic Amyotrophic Lateral Sclerosis.

Sporadic amyotrophic lateral sclerosis (sALS) is a fatal progressive neurodegenerative disease affecting upper and lower motor neurons. Biomarkers are useful to facilitate the diagnosis and/or prognosis of patients and to reveal possible mechanistic clues about the disease. This study aimed to identify and validate selected putative biomarkers in the cerebrospinal fluid (CSF) of sALS patients at early disease stages compared with age-matched controls and with other neurodegenerative diseases including Alzheimer disease (AD), spinal muscular atrophy type III (SMA), frontotemporal dementia behavioral variant (FTD), and multiple sclerosis (MS). SWATH acquisition on liquid chromatography-tandem mass spectrometry (LC-MS/MS) for protein quantitation, and ELISA for validation, were used in CSF samples of sALS cases at early stages of the disease. Analysis of mRNA and protein expression was carried out in the anterior horn of the lumbar spinal cord in post-mortem tissue of sALS cases (terminal stage) and controls using RTq-PCR, and Western blotting, and immunohistochemistry, respectively. SWATH acquisition on liquid chromatography-tandem mass spectrometry (LC-MS/MS) revealed 51 differentially expressed proteins in the CSF in sALS. Receiver operating characteristic (ROC) curves showed CXCL12 to be the most valuable candidate biomarker. We validated the values of CXCL12 in CSF with ELISA in two different cohorts. Besides sALS, increased CXCL12 levels were found in MS but were not altered in AD, SMA, and FTD. Therefore, increased CXCL12 levels in the CSF can be useful in the diagnoses of MS and sALS in the context of the clinical settings. CXCL12 immunoreactivity was localized in motor neurons in control and sALS, and in a few glial cells in sALS at the terminal stage; CXCR4 was in a subset of oligodendroglial-like cells and axonal ballooning of motor neurons in sALS; and CXCR7 in motor neurons in control and sALS, and reactive astrocytes in the pyramidal tracts in terminal sALS. CXCL12/CXCR4/CXCR7 axis in the spinal cord probably plays a complex role in inflammation, oligodendroglial and astrocyte signaling, and neuronal and axonal preservation in sALS.

Spatial Assessment of the Association between Long-Term Exposure to Environmental Factors and the Occurrence of Amyotrophic Lateral Sclerosis in Catalonia, Spain: A Population-Based Nested Case-Control Study.

BACKGROUND: It is believed that an interaction between genetic and non-genetic factors may be involved in the development of amyotrophic lateral sclerosis (ALS). With the exception of exposure to agricultural chemicals like pesticides, evidence of an association between environmental risk factors and ALS is inconsistent. Our objective here was to investigate the association between long-term exposure to environmental factors and the occurrence of ALS in Catalonia, Spain, and to provide evidence that spatial clusters of ALS related to these environmental factors exist. METHODS: We carried out a nested case-control study constructed from a retrospective population-based cohort, covering the entire region. Environmental variables were the explanatory variables of interest. We controlled for both observed and unobserved confounders. RESULTS: We have found some spatial clusters of ALS. The results from the multivariate model suggest that these clusters could be related to some of the environmental variables, in particular agricultural chemicals. In addition, in high-risk clusters, besides corresponding to agricultural areas, key road infrastructures with a high density of traffic are also located. CONCLUSION: Our results indicate that some environmental factors, in particular those associated with exposure to pesticides and air pollutants as a result of urban traffic, could be associated with the occurrence of ALS.

The increasing importance of environmental conditions in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is the most common neurodegenerative disease affecting motor neurons (MNs). Although a small percentage of ALS has a familial origin, the vast majority of cases are sporadic in which genetic factors and environment interact with each other leading to disease onset in genetically predisposed individuals. In the current model of the disease, each individual has a determined genetic load, some degree of cell degeneration related to age and several risky environmental exposures. In this scenario, MN degeneration would occur when the sum of these factors reach a certain threshold. To date, an extensive list of environmental factors has been associated to ALS, including different categories, such as exposure to heavy metals and other toxicants, cyanotoxins or infectious agents. In addition, in recent years, lifestyle and other demographic parameters are gaining relevance in the genesis of the disease. Among them, physical activity, nutrition, body mass index, cardiovascular risk factors, autoimmune diseases and cancer are some of the conditions which have been related to the disease. In this review, we will discuss the potential mechanisms of environmental conditions in motor neuron degeneration. Understanding the role of each one of these factors as well as their interactions appears as a crucial step in order to develop new preventive, diagnostic and therapeutic approaches for ALS patients.