Transactive response DNA-binding protein 43 is enriched at the centrosome in human cells.

The centrosome, as the main microtubule organizing centre, plays key roles in cell polarity, genome stability and ciliogenesis. The recent identification of ribosomes, RNA-binding proteins and transcripts at the centrosome suggests local protein synthesis. In this context, we hypothesized that TDP-43, a highly conserved RNA binding protein involved in the pathophysiology of amyotrophic lateral sclerosis and frontotemporal lobar degeneration, could be enriched at this organelle. Using dedicated high magnification sub-diffraction microscopy on human cells, we discovered a novel localization of TDP-43 at the centrosome during all phases of the cell cycle. These results were confirmed on purified centrosomes by western blot and immunofluorescence microscopy. In addition, the co-localization of TDP-43 and pericentrin suggested a pericentriolar enrichment of the protein, leading us to hypothesize that TDP-43 might interact with local mRNAs and proteins. Supporting this hypothesis, we found four conserved centrosomal mRNAs and 16 centrosomal proteins identified as direct TDP-43 interactors. More strikingly, all the 16 proteins are implicated in the pathophysiology of TDP-43 proteinopathies, suggesting that TDP-43 dysfunction in this organelle contributes to neurodegeneration. This first description of TDP-43 centrosomal enrichment paves the way for a more comprehensive understanding of TDP-43 physiology and pathology.

Clinical and electrophysiological characteristics of women with X-linked Charcot-Marie-Tooth disease.

BACKGROUND: X-Linked Charcot-Marie-Tooth disease type 1 (CMTX1) is characterized by gender differences in clinical severity. Women are usually clinically affected later and less severely than men. However, their clinical presentation appears to be heterogenous. Our aim was to extend the phenotypic description in a large series of women with CMTX1. METHODS: We retrospectively evaluated 263 patients with CMTX1 from 11 French reference centers. Demographic, clinical, and nerve conduction data were collected. The severity was assessed by CMT Examination Score (CMTES) and Overall Neuropathy Limitations Scale (ONLS) scores. We looked for asymmetrical strength, heterogeneous motor nerve conduction velocity (MNCV), and motor conduction blocks (CB). RESULTS: The study included 137 women and 126 men from 151 families. Women had significantly more asymmetric motor deficits and MNCV than men. Women with an age of onset after 19 years were milder. Two groups of women were identified after 48 years of age. The first group represented 55%, with women progressing as severely as men, however, with a later onset age. The second group had mild or no symptoms. Some 39% of women had motor CB. Four women received intravenous immunoglobulin before being diagnosed with CMTX1. CONCLUSIONS: We identified two subgroups of women with CMTX1 who were over 48 years of age. Additionally, we have demonstrated that women with CMTX can exhibit an atypical clinical presentation, which may result in misdiagnosis. Therefore, in women presenting with chronic neuropathy, the presence of clinical asymmetry, heterogeneous MNCV, and/or motor CB should raise suspicion for X-linked CMT, particularly CMTX1, and be included in the differential diagnosis.

Anti-disialosyl-immunoglobulin M chronic autoimmune neuropathies: a nationwide multicenter retrospective study.

BACKGROUND AND PURPOSE: In this retrospective study involving 14 university hospitals from France and Switzerland, the aim was to define the clinicopathological features of chronic neuropathies with anti-disialosyl ganglioside immunoglobulin M (IgM) antibodies (CNDA). RESULTS: Fifty-five patients with a polyneuropathy evolving for more than 2 months and with at least one anti-disialosyl ganglioside IgM antibody, that is, anti-GD1b, -GT1b, -GQ1b, -GT1a, -GD2 and -GD3, were identified. Seventy-eight percent of patients were male, mean age at disease onset was 55 years (30-76) and disease onset was progressive (82%) or acute (18%). Patients presented with limb sensory symptoms (94% of cases), sensory ataxia (85%), oculomotor weakness (36%), limb motor symptoms (31%) and bulbar muscle weakness (18%). Sixty-five percent of patients had a demyelinating polyradiculoneuropathy electrodiagnostic profile and 24% a sensory neuronopathy profile. Anti-GD1b antibodies were found in 78% of cases, whilst other anti-disialosyl antibodies were each observed in less than 51% of patients. Other features included nerve biopsy demyelination (100% of cases), increased cerebrospinal fluid protein content (75%), IgM paraprotein (50%) and malignant hemopathy (8%). Eighty-six percent of CNDA patients were intravenous immunoglobulins-responsive, and rituximab was successfully used as second-line treatment in 50% of cases. Fifteen percent of patients had mild symptoms and were not treated. CNDA course was progressive (55%) or relapsing (45%), and 93% of patients still walked after a mean disease duration of 11 years. CONCLUSION: Chronic neuropathies with anti-disialosyl ganglioside IgM antibodies have a recognizable phenotype, are mostly intravenous immunoglobulins-responsive and present with a good outcome in a majority of cases.

Antibodies to the Caspr1/contactin-1 complex in chronic inflammatory demyelinating polyradiculoneuropathy.

Previous studies have described the clinical, serological and pathological features of patients with chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) and antibodies directed against the paranodal proteins neurofascin-155, contactin-1 (CNTN1), contactin-associated protein-1 (Caspr1), or nodal forms of neurofascin. Such antibodies are useful for diagnosis and potentially treatment selection. However, antibodies targeting Caspr1 only or the Caspr1/CNTN1 complex have been reported in few patients with CIDP. Moreover, it is unclear if these patients belong to the same pathophysiological subgroup. Using cell-based assays in routine clinical testing, we identified sera from patients with CIDP showing strong membrane reactivity when both CNTN1 and Caspr1 were co-transfected (but not when CNTN1 was transfected alone). Fifteen patients (10 male; aged between 40 and 75) with antibodies targeting Caspr1/CNTN1 co-transfected cells were enrolled for characterization. The prevalence of anti-Caspr1/CNTN1 antibodies was 1.9% (1/52) in the Sant Pau CIDP cohort, and 4.3% (1/23) in a German cohort of acute-onset CIDP. All patients fulfilled European Federation of Neurological Societies/Peripheral Nerve Society (EFNS/PNS) definite diagnostic criteria for CIDP. Seven (47%) were initially diagnosed with Guillain-Barré syndrome due to an acute-subacute onset. Six (40%) patients had cranial nerve involvement, eight (53%) reported neuropathic pain and 12 (80%) ataxia. Axonal involvement and acute denervation were frequent in electrophysiological studies. Complete response to intravenous immunoglobulin was not observed, while most (90%) responded well to rituximab. Enzyme-linked immunosorbent assay (ELISA) and teased nerve fibre immunohistochemistry confirmed reactivity against the paranodal Caspr1/CNTN1 complex. Weaker reactivity against Caspr1 transfected alone was also detected in 10/15 (67%). Sera from 13 of these patients were available for testing by ELISA. All 13 samples reacted against Caspr1 by ELISA and this reactivity was enhanced when CNTN1 was added to the Caspr1 ELISA. IgG subclasses were also investigated by ELISA. IgG4 was the predominant subclass in 10 patients, while IgG3 was predominant in other three patients. In conclusion, patients with antibodies to the Caspr1/CNTN1 complex display similar serological and clinical features and constitute a single subgroup within the CIDP syndrome. These antibodies likely target Caspr1 primarily and are detected with Caspr1-only ELISA, but reactivity is optimal when CNTN1 is added to Caspr1 in cell-based assays and ELISA.

Engineered Microbes for Producing Anticholinergics.

The tropane alkaloids (TAs) hyoscyamine and scopolamine function as acetylcholine receptor antagonists and are used clinically as parasympatholytics to treat neuromuscular disorders in humans. While TAs are synthesized in a small subset of plant families, these specialized metabolites are only accumulated in limited quantities in plant organs. The complex chemical structures of these compounds make their industrial production by chemical synthesis very challenging, Therefore, the supply of these TAs still relies on intensive farming of Duboisia shrubs in tropical countries. Many adverse factors such as climate fluctuations and pandemics can thus influence annual world production. Based on the landmark microbial production of the antimalarial semi-synthetic artemisinin, the Smolke group recently developed a yeast cell factory capable of de novo synthesizing hyoscyamine and scopolamine, thus paving the way for an alternative production of these compounds.

Improved detection of mitochondrial DNA instability in mitochondrial genome maintenance disorders.

PURPOSE: Diseases caused by defects in mitochondrial DNA (mtDNA) maintenance machinery, leading to mtDNA deletions, form a specific group of disorders. However, mtDNA deletions also appear during aging, interfering with those resulting from mitochondrial disorders. METHODS: Here, using next-generation sequencing (NGS) data processed by eKLIPse and data mining, we established criteria distinguishing age-related mtDNA rearrangements from those due to mtDNA maintenance defects. MtDNA deletion profiles from muscle and urine patient samples carrying pathogenic variants in nuclear genes involved in mtDNA maintenance (n = 40) were compared with age-matched controls (n = 90). Seventeen additional patient samples were used to validate the data mining model. RESULTS: Overall, deletion number, heteroplasmy level, deletion locations, and the presence of repeats at deletion breakpoints were significantly different between patients and controls, especially in muscle samples. The deletion number was significantly relevant in adults, while breakpoint repeat lengths surrounding deletions were discriminant in young subjects. CONCLUSION: Altogether, eKLIPse analysis is a powerful tool for measuring the accumulation of mtDNA deletions between patients of different ages, as well as in prioritizing novel variants in genes involved in mtDNA stability.

Effect of familial clustering in the genetic screening of 235 French ALS families.

OBJECTIVES: To determine whether the familial clustering of amyotrophic lateral sclerosis (ALS) cases and the phenotype of the disease may help identify the pathogenic genes involved. METHODS: We conducted a targeted next-generation sequencing analysis on 235 French familial ALS (FALS), unrelated probands to identify mutations in 30 genes linked to the disease. The genealogy, that is, number of cases and generations with ALS, gender, age, site of onset and the duration of the disease were analysed. RESULTS: Regarding the number of generations, 49 pedigrees had only one affected generation, 152 had two affected generations and 34 had at least three affected generations. Among the 149 pedigrees (63.4%) for which a deleterious variant was found, an abnormal G4C2 expansion in C9orf72 was found in 98 cases as well as SOD1, TARBP or FUS mutations in 30, 9 and 7 cases, respectively. Considering pedigrees from the number of generations, abnormal G4C2 expansion in C9orf72 was more frequent in pedigrees with pairs of affected ALS cases, which represented 65.2% of our cohort. SOD1 mutation involved all types of pedigrees. No TARDBP nor FUS mutation was present in monogenerational pedigrees. TARDBP mutation predominated in bigenerational pedigrees with at least three cases and FUS mutation in multigenerational pedigrees with more than seven cases, on average, and with an age of onset younger than 45 years. CONCLUSION: Our results suggest that familial clustering, phenotypes and genotypes are interconnected in FALS, and thus it might be possible to target the genetic screening from the familial architecture and the phenotype of ALS cases.

Genotype-phenotype correlation in French patients with myelin protein zero gene-related inherited neuropathy.

BACKGROUND AND PURPOSE: Preparations for clinical trials of unfolded protein response (UPR) inhibitors (such as Sephin1) that target the upregulated UPR in patients with Charcot-Marie-Tooth disease (CMT) carrying MPZ mutations are currently underway. The inclusion criteria for these trials are still being formulated. Our objective was to characterize the relation between genotypes and phenotypes in patients with CMT caused by MPZ mutations, and to refine the inclusion criteria for future trials. METHODS: Clinical and neurophysiological data of CMT patients with MPZ mutations were retrospectively collected at 11 French reference centers. RESULTS: Forty-four mutations in MPZ were identified in 91 patients from 61 families. There was considerable heterogeneity. The same mutation was found to cause either axonal or demyelinating neuropathy. Three groups were identified according to the age at disease onset. CMT Examination Score (CMTES) tended to be higher in the early (≤22 years) and adult (23-47 years) onset groups (mean CMTESv2 = 10.4 and 10.0, respectively) than in the late onset group (>47 years, mean CMTESv2 = 8.6, p = 0.47). There was a significant positive correlation between CMTESv2 and the age of patients in Groups I (p = 0.027) and II (p = 0.023), indicating that clinical severity progressed with age in these patients. CONCLUSIONS: To optimize the selection of CMT patients carrying MPZ mutations for the upcoming trials, inclusion criteria should take into account the pathophysiology of the disease (upregulated UPR). Recruited patients should have a mild to moderate disease severity and a disease onset at between 18 and 50 years, as these patients exhibit significant disease progression over time.

Guillain-BarrÉ syndrome subtype diagnosis: A prospective multicentric European study.

INTRODUCTION: There is uncertainty as to whether the Guillain-Barré syndrome (GBS) subtypes, acute inflammatory demyelinating polyradiculoneuropathy (AIDP) and acute motor axonal neuropathy (AMAN), can be diagnosed electrophysiologically. METHODS: We prospectively included 58 GBS patients. Electrodiagnostic testing (EDX) was performed at means of 5 and 33 days after disease onset. Two traditional and one recent criteria sets were used to classify studies as demyelinating or axonal. Results were correlated with anti-ganglioside antibodies and reversible conduction failure (RCF). RESULTS: No classification shifts were observed, but more patients were classified as axonal with recent criteria. RCF and anti-ganglioside antibodies were present in both subtypes, more frequently in the axonal subtype. DISCUSSION: Serial EDX has no effect on GBS subtype proportions. The absence of exclusive correlation with RCF and anti-ganglioside antibodies may challenge the concept of demyelinating and axonal GBS subtypes based upon electrophysiological criteria. Frequent RCF indicates that nodal/paranodal alterations may represent the main pathophysiology. Muscle Nerve, 2018.

Determinants of health-related quality of life in anti-MAG neuropathy: a cross-sectional multicentre European study.

Our objective was to assess determinants of quality of life (QoL) in anti-myelin associated glycoprotein antibody (MAG) neuropathy. The SF-36 questionnaire was assessed in 55 patients, from Marseille, Angers (France) and Birmingham (UK). Routine clinical evaluations included Medical Research Council (MRC) sum score, inflammatory neuropathy cause and treatment (INCAT) sensory score, inflammatory Rasch-built overall disability score (I-RODS), ataxia score, Jamar grip dynamometry, timed 10-m walk, neuropathic pain symptom inventory (NPSI) score, and fatigue severity score (FSS). Physical component summary (PCS) and mental component summary (MCS) of the SF36 questionnaire were significantly lower than in reported normal subjects of both countries (p < 0.001). All SF-36 domains correlated with I-RODS, except MCS for which significance was, however, approached (p = 0.056). PCS correlated with MRC sum score, ataxia score, timed 10-m walk, tremor, Jamar grip dynamometry, NPSI pain score, FSS and level of social support. MCS correlated exclusively with FSS and level of social support. In multivariate regression, PCS was associated independently with I-RODS (p < 0.001) and NPSI pain score (p = 0.011), whereas MCS was associated independently with FSS (p = 0.022). QoL is accurately predicted in anti-MAG neuropathy by the I-RODS and FSS, lending support to their use in clinical and research settings. Effective measures to improve QoL should include tremor and neuropathic pain treatment, fatigue management, and improved social support.

Therapeutic plasma exchange in chronic dysimmune peripheral neuropathies: A 10-year retrospective study.

INTRODUCTION: Therapeutic plasma exchange (TPE) can be proposed in the treatment of chronic dysimmune peripheral neuropathies (CDPN). Actual guidelines are however based on few studies, and indications and protocols still remain to be clarified. We conducted a 10-year retrospective study in order to assess the effectiveness and tolerance of TPE in CDPN. METHODS: All patients treated for CDPN with TPE from October 2006 to March 2016 in the university hospital of Angers were included. Patients were considered responders when they presented a clinical improvement substantial enough to continue the treatment. The Hughes functional grading score was also determined for each patient before and after TPE initiation. RESULTS: Among the 206 patients who received TPE during the study period, 30 (14.6%) met the diagnostic criteria of CDPN. Four of the five paraprotein neuropathies (PPN) patients (80%) and 8 of the 11 chronic inflammatory demyelinating polyneuropathies (CIDP) patients (72.7%) were responders, with a significant improvement of the Hughes score for the latter (P = 0.013). None of the three Lewis-Sumner and the two POEMS patients showed substantial improvement. Six of the nine anti-MAG neuropathy patients (66.7%) responded to treatment, with a trend towards improvement of the Hughes score (P = 0.072). CONCLUSION: TPE appears to be effective in CIDP and PPN, and ineffective in Lewis-Sumner and POEMS syndromes. Interestingly, anti-MAG neuropathy patients showed a good rate of response to TPE. Regarding these preliminary results, a randomized trial would be very worthwhile in this disease for which there is no evidence based treatment to date.

Increased mitochondrial fusion in a autosomal recessive CMT2A family with mitochondrial GTPase mitofusin 2 mutations.

Charcot-Marie-Tooth type 2A disease (CMT2A) is an inherited peripheral neuropathy mainly caused by mutations in the MFN2 gene coding for the mitochondrial fusion protein mitofusin 2. Although the disease is mainly inherited in a dominant fashion, few cases of early-onset autosomal recessive CMT2A (AR-CMT2A) have been reported in recent years. In this study, we characterized the structure of the mitochondrial network in cultured primary fibroblasts obtained from AR-CMT2A family members. The patient-derived cells showed an increase of the mitochondrial fusion with large connected networks and an increase of the mitochondrial volume. Interestingly, fibroblasts derived from the two asymptomatic parents showed similar changes to a lesser extent. These results support the hypothesis that AR-CMT2A-related MFN2 mutations acts through a semi-dominant negative mechanism and suggest that other biological parameters might show mild alterations in asymptomatic heterozygote AR-CMT2A patients. Such alterations could be useful biomarkers helping to distinguish MFN2 mutations from variants, a growing challenge with the advent of next generation sequencing into routine clinical practice.

Posterior Spinal Cord Syndrome after Flexible Ureteroscopy for Kidney Stones.

A 49-year-old woman was treated for right kidney stones using flexible ureteroscopy and laser lithotripsy. She was readmitted 2 weeks after the treatment with complaints of walking difficulties. On neurologic examination, it was found that she had a bilateral proprioceptive dysfunction. Spinal MRI revealed an intramedullary lesion at the T10-T11 level consistent with an intramedullary cavernoma. The final diagnosis was a spinal compression consecutive to the cavernoma bleeding associated with the prolonged lithotomy position. Symptoms have not been reversible and the patient continued to have walking difficulties. We report for the first time a major complication that arises as a result of using flexible ureteroscopy for treating kidney stones.

Improved locus-specific database for OPA1 mutations allows inclusion of advanced clinical data.

Autosomal-dominant optic atrophy (ADOA) is the most common inherited optic neuropathy, due to mutations in the optic atrophy 1 gene (OPA1) in about 60%-80% of cases. At present, the clinical heterogeneity of patients carrying OPA1 variants renders genotype-phenotype correlations difficulty. Since 2005, when we published the first locus-specific database (LSDB) dedicated to OPA1, a large amount of new clinical and genetic knowledge has emerged, prompting us to update this database. We have used the Leiden Open-Source Variation Database to develop a clinico-biological database, aiming to add clinical phenotypes related to OPA1 variants. As a first step, we validated this new database by registering several patients previously reported in the literature, as well as new patients from our own institution. Contributors may now make online submissions of clinical and molecular descriptions of phenotypes due to OPA1 variants, including detailed ophthalmological and neurological data, with due respect to patient anonymity. The updated OPA1 LSDB (http://opa1.mitodyn.org/) should prove useful for molecular diagnoses, large-scale variant statistics, and genotype-phenotype correlations in ADOA studies.

Mitochondrial F0F1-ATP synthase governs the induction of mitochondrial fission.

Mitochondrial dynamics is a process that balances fusion and fission events, the latter providing a mechanism for segregating dysfunctional mitochondria. Fission is controlled by the mitochondrial membrane potential (ΔΨm), optic atrophy 1 (OPA1) cleavage, and DRP1 recruitment. It is thought that this process is closely linked to the activity of the mitochondrial respiratory chain (MRC). However, we report here that MRC inhibition does not decrease ΔΨm nor increase fission, as evidenced by hyperconnected mitochondria. Conversely, blocking F0F1-ATP synthase activity induces fragmentation. We show that the F0F1-ATP synthase is sensing the inhibition of MRC activity by immediately promoting its reverse mode of action to hydrolyze matrix ATP and restoring ΔΨm, thus preventing fission. While this reverse mode is expected to be inhibited by the ATPase inhibitor ATPIF1, we show that this sensing is independent of this factor. We have unraveled an unexpected role of F0F1-ATP synthase in controlling the induction of fission by sensing and maintaining ΔΨm.

Design and application of a customizable relational DataBase to assess clinicopathological correlations and concomitant pathology in neurodegenerative diseases.

The diagnosis of neurodegenerative diseases is made complex by the heterogenous phenotype of the patients and the regular occurrence of concomitant pathology. Studying clinicopathological correlations in autopsy series is a central approach to improve pathological prediction in clinical practice. However, such method requires a wealth of information, and the use of standard spreadsheet software is hardly suitable. To overcome this constraint, we designed a customizable and freely available neuropathology form with 456 data entry fields driven by an open-source DataBase Management Systems (DBMS) using Structured Query Language (SQL). This approach allowed us to optimize the compilation of clinical and pathological data from our brain collection (264 autopsied patients, 22,885 data points). Information was then easily retrieved using general and specific queries, facilitating the analysis of demographics, clinicopathological correlations, and incidental and concomitant proteinopathies. Tau, amyloid-ß and α-synuclein incidental pathology was observed in respectively 78.1%, 42.8%, and 10.7% of all the patients. These proportions increased with age, reaching 100% for Tau pathology after 80. Concomitant proteinopathy was observed in 46.4% of the patients diagnosed with neurodegenerative diseases and prion disease. We observed a particularly high rate of co-pathology in patients with Dementia with Lewy bodies (81.3% of associated Tau and amyloid-ß pathology) and Creutzfeldt-Jakob disease (68.4% of associated Tau pathology). Finally, we used specific queries to identify old cases that could meet newly defined neuropathological criteria and revised the diagnosis of a 90-year-old patient to LATE Stage 2. Increasing our understanding of clinicopathological correlations in neurodegenerative diseases is crucial given the implications in clinical diagnosis, biomarker identification and targeted therapies assessment. The precise characterization of clinical and pathological data of autopsy series remains a central approach but the large amount of generated data should encourage a more systematic use of DBMS.

InFUSing antisense oligonucleotides for treating ALS.

The question of a loss or toxic gain of function in FUS-related amyotrophic lateral sclerosis is still debated. Recently, Korobeynikov et al. argued that FUS mutations lead to a gain of function and showed that lowering wild-type and mutant FUS levels could be a promising therapeutic strategy.

Turning up the Green Light.

Fluorescent proteins (FPs) have become indispensable tools in biological research for labeling cells and proteins and sensing their biochemical activity. By introducing 'folding mutations', Campbell et al. engineered a new GFP variant with dramatically enhanced cellular brightness and stability, facilitating advanced cellular bioimaging applications in neuroscience and beyond.

Towards personalized medicine for amyotrophic lateral sclerosis.

Mohassel et al. provide unprecedented dichotomy of consequences on sphingolipid biosynthesis between pathogenic variants in the SPTLC1 gene, responsible for either amyotrophic lateral sclerosis (ALS) or hereditary sensory and autonomic neuropathy type 1 (HSAN1). Normalization of sphingolipid levels by siRNA selectively targeting the ALS mutant allele mRNA sheds light on new therapeutic approaches.