Spinal muscular atrophy (SMA) is an autosomal recessive neuromuscular disorder characterized by widespread loss of lower motor neurons from the spinal cord. Lower motor neuron degeneration leads to a progressive decline in motor development, manifesting as muscle atrophy and weakness. It is now well characterised that ubiquitin homeostasis is altered in SMA and that reduction of the ubiquitin-like modifier-activating enzyme 1 (UBA1) is central to this disruption. UBA1 is responsible for activating ubiquitin as the first step in the ubiquitin conjugation process, marking unwanted proteins for degradation by the proteasome. While it is known that therapies targeting UBA1 rescue neuromuscular phenotypes in SMA models, the mechanism by which UBA1 mediates neurodegeneration is unclear. In fact, very little is known about the function of UBA1 beyond its canonical role in the ubiquitin proteasome system. To better understand the role of UBA1 in motor neuron degeneration, a robust set of antibodies for both in vivo and in vitro work to study UBA1 have been identified. This enabled a novel characterisation of UBA1 distribution throughout disease progression in SMA spinal motor neurons to be performed, revealing that UBA1 reduction is an important pre-symptomatic molecular feature of SMA. To identify downstream targets of UBA1 critical for UBA1-mediated degeneration in SMA, label-free proteomics was performed on HEK293 cells after overexpression or knockdown of UBA1. The proteomics data was analysed across multiple platforms, including Biolayout, IPA and DAVID to identify UBA1-dependent pathways and demonstrated that modulation of UBA1 levels lead to disruption of key cellular pathways including translation elongation, nuclear transport, and tRNA synthetases. Validation of target proteins from these UBA1-dependent pathways identified that the tRNA synthetease GARS behaves in a UBA1-dependent manner across a range of model systems in vitro and in vivo. It was then identified that GARS expression is significantly dysregulated across a range of neuronal tissues in a mouse model of SMA. Interestingly, mutations in GARS cause Charcot-Marie-Tooth disease type 2D (CMT2D), an axonal neuropathy, in which a disruption to sensory neuron fate in dorsal root ganglia has recently been identified. In a mouse model of SMA we identified a phenotype consistent with that in the CMT2D mouse model and showed that disruption to sensory neuron fate is reversible and dependent on changes in UBA1 and GARS expression in SMA. In conclusion, modulation of UBA1 levels leads to disruption of key cellular pathways, with dysregulation of tRNA synthetases a prominent feature that is likely to play a role in the pathogenesis of SMA.

La glycyl-ARNt synthétase humaine (GRS) est une enzyme clé dans la traduction des protéines dans le cytosol et la mitochondrie. Chez l’Homme, des mutations de la GRS conduisent à la neuropathie périphérique Charcot-Marie-Tooth (CMT). Bien que l’activité de la GRS soit ubiquitaire, les mutations associées à la CMT n’affectent que les nerfs périphériques, suggérant un rôle supplémentaire de la GRS dans les neurones. Pour comprendre ce rôle, nous avons d’abord élucidé le mécanisme particulièrement complexe qui contrôle l’expression de la GRS mitochondriale et cytosolique à partir du même gène. Nous avons identifié deux ARNm : un codant pour les deux enzymes ; et un autre plus long qui contient une IRES fonctionnelle et un uORF. Cet ARNm complexe, ne génère que la GRS cytosolique et montre que son expression et localisation sont étroitement contrôlées. De plus, nous avons montré une distribution particulière de la GRS dans des neurones, qui est un premier indice sur un rôle non canonique. / Human Glycyl-tRNA synthetase (GRS) is a housekeeping enzyme with a key role in protein synthesis, both in the cytosol and the mitochondria. In human, mutations in GRS cause the Charcot-Marie-Tooth (CMT) peripheral neuropathy. Though GRS activity is required in all cells, the CMT-associated mutations affect only the peripheral nervous system, suggesting an additional non canonical role.To understand how GRS is involved in CMT pathology, we first elucidated the original post-transcriptional regulatory mechanism that controls the expression of both the mitochondrial and the cytosolic GRS from a single gene. We identified two mRNA isoforms: one coding for both enzymes; and a longer one containing a functional IRES and an uORF encoding only the cytosolic GRS, evidence that expression and localization of human GRS are tightly controlled. Furthermore, we found a particular Ca2+ dependant distribution of GRS in neurons, giving us a first clue about a potential non-canonical role in neurons.

Centronuclear myopathy (CNM) is a genetically heterogeneous disorder associated with general skeletal muscle weakness, type I fiber predominance and atrophy, and abnormally centralized nuclei. Autosomal dominant CNM is due to mutations in the large GTPase dynamin 2 (DNM2), a mechanochemical enzyme regulating cytoskeleton and membrane trafficking in cells. To date, 40 families with CNM-related DNM2 mutations have been described, and here we report 60 additional families encompassing a broad genotypic and phenotypic spectrum. In total, 18 different mutations are reported in 100 families and our cohort harbors nine known and four new mutations, including the first splice-site mutation. Genotype-phenotype correlation hypotheses are drawn from the published and new data, and allow an efficient screening strategy for molecular diagnosis. In addition to CNM, dissimilar DNM2 mutations are associated with Charcot-Marie-Tooth (CMT) peripheral neuropathy (CMTD1B and CMT2M), suggesting a tissue-specific impact of the mutations. In this study, we discuss the possible clinical overlap of CNM and CMT, and the biological significance of the respective mutations based on the known functions of dynamin 2 and its protein structure. Defects in membrane trafficking due to DNM2 mutations potentially represent a common pathological mechanism in CNM and CMT.

No description available.

Introdução: A síndrome SPOAN é uma doença neurodegenerativa, de transmissão genética autossômica recessiva, até o momento reconhecida apenas no Brasil, que caracteriza-se por: paraplegia espástica, de início nos primeiros anos de vida e caráter progressivo; atrofia óptica congênita; neuropatia periférica sensitivo-motora axonal, de início a partir da primeira década de vida; sobressaltos à estimulação sonora, disartria, deformidades de coluna e pés e sinais extra piramidais. A sua caracterização foi feita por nosso grupo, que avaliou clinicamente 71 indivíduos, originários do Rio Grande do Norte. Estudo de ligação mapeou o locus responsável pela síndrome SPOAN em uma região de 2 Mb no cromossomo 11q13. O gene responsável pela síndrome SPOAN permanece desconhecido. A síndrome SPOAN é considerada uma forma complicada de paraplegia espástica. A associação entre neuropatia e paraplegia espástica está relacionada à perda progressiva de axônios longos e tem sido relatada em algumas formas complicadas de neuropatias e paraplegias espásticas hereditárias. Casuística e métodos: Foi realizada a avaliação de 27 pacientes, 20 mulheres, com idade variando entre 4 e 58 anos. Todos os indivíduos compartilhavam o mesmo fenótipo (paraplegia espástica, atrofia de nervo óptico e neuropatia periférica) e tinham o mesmo haplótipo 11q13. Pacientes com história de diabetes mellitus ou alcoolismo foram excluídos do estudo. A avaliação neurológica incluiu a pesquisa dos escores modificados de sintomas e comprometimento neuropáticos. A força muscular foi testada e graduada conforme a escala MRC (Medical Research Council). Foi realizada a pesquisa da sensibilidade dolorosa, térmica, tátil, vibratória e artrestésica. O trofismo foi avaliado pela presença de deformidades na coluna e atrofia nos membros inferiores. Os reflexos profundos e o cutâneo plantar também foram analisados. Os estudos da condução nervosa foram realizados em um aparelho portátil Nicolet - Viking Quest, (Viasys, USA). Para os estudos de condução motora foram analisados os nervos axillar, mediano, ulnar, femoral, tibial e fibular direito. A condução sensitiva foi analisada nos nervos mediano, ulnar, radial, sural e fibular direito. O reflexo H e as ondas F foram avaliados com técnicas padrão. Alguns testes não puderam ser realizados devido à intensa atrofia e deformidades esqueléticas. O coeficiente de correlação de Pearson foi calculado entre a idade e os parâmetros, velocidade de condução, latência e amplitude. Valores de P < 0,05 foram considerados estatisticamente significantes. Resultados: Avaliação clínica: Todos os pacientes obtiveram escore de sinais neuropáticos graves e demonstraram déficit de força e atrofia distal. Deformidades dos pés estavam presentes em todos os pacientes e deformidades na coluna, em 58%. Os reflexos profundos dos membros superiores estavam exaltados em 92% dos casos e o reflexo patelar, em 63%. O reflexo Aquileu estava ausente em todos os pacientes. Todas as modalidades de sensibilidade foram afetadas, principalmente nos membros inferiores. Os dados do exame de sensibilidade na paciente de 4 anos foram desconsiderados. Estudo da condução nervosa sensitiva: Os SNAPs dos nervos mediano, sural e fibular estavam ausentes em todos os pacientes. SNAPs do nervo ulnar estavam ausentes em 96% da amostra e do nervo radial, em 80%. Estudo da condução nervosa motora: As latências motoras dos nervos axilar e femoral estavam normais em todos os pacientes. As amplitudes dos CMAPs estavam reduzidas em 15 e 52% da amostra nos nervos mediano e ulnar, respectivamente. Velocidades de condução estavam reduzidas em 50 e 41% desta casuística nos nervos mediano e ulnar, respectivamente. Velocidades de condução estavam acima de 80% do limite inferior da normalidade, em todos os nervos, exceto em 1 paciente que apresentou redução de 27% no nervo ulnar. Entretanto, este mesmo paciente apresentou amplitude menor que 2mV. Ondas F apresentavam aumento da latência, de acordo com a altura, em 100% dos casos. CMAPs estavam ausentes em 93 e 84% da amostra nos nervos fibular e tibial, respectivamente. Reflexo H estava ausente em 88% dos pacientes. Não houve correlação entre idade e a velocidade de condução, latência e amplitude dos nervos mediano e ulnar. Discussão: O estudo da condução nervosa neste grupo preencheu critérios para uma neuropatia primária axonal. Nenhum paciente apresentou bloqueio de condução ou dispersão temporal. As alterações encontradas na velocidade de condução provavelmente se devem à perda de fibras nervosas de condução rápida. Fenótipos SPOAN-like foram descritos em famílias com mutações nos genes C12orf65, TFG e OPA1. No entanto, não existem detalhes sobre a condução nervosa nestes pacientes. Neuropatia axonal de início tardio foi relacionada à SPG55 e DOA (dominant optic atrophy), enquanto neuropatia axonal e desmielinizante com leve comprometimento sensitivo foi descrita na família com mutação no gene TFG. Conclusão: Os pacientes com a síndrome SPOAN apresentam uma acentuada neuropatia axonal, sensitivo motora. As alterações encontradas na condução nervosa dos pacientes com síndrome SPOAN não são específicas, no entanto, resultados normais excluem esta condição em adultos. A paciente mais jovem desta casuística já apresentava alterações ao exame, o que pode sugerir um início precoce da neuropatia. Entretanto, não temos dados suficientes para afirmar que este seja um achado comum a todos os pacientes SPOAN / Introduction: SPOAN syndrome (Spastic Paraplegia, Optic Atrophy and Neuropathy) is a progressive neurodegenerative disorder of autosomal recessive inheritance described by our group in a large inbred family from Northeastern Brazil. The clinical picture is characterized by non-progressive congenital optic atrophy, progressive spastic paraplegia, axonal neuropathy, auditory startles, dysarthria, spinal and foot deformities and also extrapyramidal signs. Linkage studies mapped the responsible locus for the syndrome to a 2Mb region on chromosome 11q13. The gene responsible for SPOAN syndrome remains elusive. Materials and Methods: This is a cross sectional study which was conducted from 2009 to 2011. We evaluated 27 patients (20 females), with a0ges ranging from 4 to 58 years. All patients shared the same phenotype (spastic paraplegia, optic atrophy and peripheral neuropathy) and had the same 11q13 haplotype in homozygosis. Patients with history of diabetes mellitus or alcoholism were excluded from this study. All patients were evaluated by the same clinical researcher (SA). Neurological evaluation included determination of modified neuropathy symptoms (NSS) and neuropathy disability (NDS) scores. Motor strength was assessed using MRC scale. Sensibility assessment included small-fiber (pain and temperature) and large-fiber modalities (vibration-128Hz diapason, 10g monofilament and joint position sense). Spine deformities and atrophy in the lower limbs were observed. We also evaluated osteotendineous reflexes and cutaneous plantar reflexes. Nerve conduction studies were performed using a portable Nicolet - Viking Quest, (Viasys,USA). Motor conduction studies included axillary, median, ulnar, femoral, tibial and fibular nerves on the right side. Sensory nerve action potentials of median, ulnar, sural and superficial fibular nerves were recorded using a bar electrode of 3 cm and standard fixed distances. Tibial H-reflex was evaluated with standard technique. Minimal F wave latencies were obtained from ulnar and tibial nerves. A few tests could not be done in every patient due to severe deformities. We calculated Pearson\'s correlation coefficients between age and nerve conduction parameters, including velocities, latencies and amplitudes. P values <0.05 were considered statistically significant. Results: Clinical data: Neuropathic symptoms such as pain and paresthesias were rare. All patients had signs of severe neuropathy. All subjects demonstrated weakness and atrophy that were more significant distally than proximally. Foot deformities were present in all patients and spine deformities were seen in 58%. Upper limb deep tendon reflexes were exalted in 92% and patelar reflex in 63%. Ankle reflex was absent in all patients. In one patient, who was 4 years-old, sensory evaluation was inconsistent and the results were not considered. In all the other ones, sensory modalities were affected and occurred predominantly in the lower limbs. Electrodiagnostic data: Sensory nerve conduction: Median nerve SNAP was absent in all 27 patients. Ulnar nerve SNAPs were absent in 96%, whereas radial nerve SNPAs were absent in 80%. Superficial fibular and sural SNAPs were absent in all patients. Motor nerve conduction: The motor latencies of axillary and femoral nerves were normal in all patients. CMAP amplitudes were reduced in 15% of the median nerves and in 52% of the ulnar nerves. Conduction velocities (CV) were reduced in 50% of the median nerves and in 41% of the ulnar nerves. CV was above 80% of the lower normal limit for all nerves, except for one patient who showed a 27% reduction of ulnar CV, but had also a CMAP amplitude of less than 2 mV. F waves were prolonged according to the height in 100%. Only one patient who presented significant motor CV reduction of the ulnar nerve. CMAPs were absent in 93% of the fibular nerves and in 84% of the tibial nerves. A single fibular nerve showed more than 20% of CV reduction, but also had severely reduced CMAP amplitude. H reflex was absent in 88% of the patients. There was no correlation between age and neurophysiological parameters, such as median or ulnar CV, latencies or CMAP amplitudes. Discussion: Nerve conduction studies in this group fulfill criteria for primary axonal neuropathy. No patient showed conduction block or temporal dispersion. Abnormalities seen in CV and F waves are probably related to loss of fast conduction fiber nerves. We could not demonstrate correlation between age and nerve conduction parameters, including velocities, latencies and amplitudes. SPOAN-like phenotype has been found in families with mutations in C12orf65, TFG and OPA1 genes, however there is no detailed report on nerve conduction studies in these conditions. Axonal neuropathy is also described in SPG55 and DOA plus, but usually with a later onset than on SPOAN syndrome. Peripheral neuropathy is also described in the family with mutation in TFG gene, but this presents a different pattern characterized as a mixed axonal demyelinating neuropathy with mild sensory involvement. Although the nerve conduction abnormalities seen in SPOAN syndrome are not specific, normal results seem to rule out this condition, at least in adult patients. The younger patient in our series was 4-years-old, and her neurophysiological study was severely abnormal, suggesting an early-onset neuropathy. However, we do not have a comprehensive study of several young patients to support that this feature is

Introdução: A síndrome SPOAN é uma doença neurodegenerativa, de transmissão genética autossômica recessiva, até o momento reconhecida apenas no Brasil, que caracteriza-se por: paraplegia espástica, de início nos primeiros anos de vida e caráter progressivo; atrofia óptica congênita; neuropatia periférica sensitivo-motora axonal, de início a partir da primeira década de vida; sobressaltos à estimulação sonora, disartria, deformidades de coluna e pés e sinais extra piramidais. A sua caracterização foi feita por nosso grupo, que avaliou clinicamente 71 indivíduos, originários do Rio Grande do Norte. Estudo de ligação mapeou o locus responsável pela síndrome SPOAN em uma região de 2 Mb no cromossomo 11q13. O gene responsável pela síndrome SPOAN permanece desconhecido. A síndrome SPOAN é considerada uma forma complicada de paraplegia espástica. A associação entre neuropatia e paraplegia espástica está relacionada à perda progressiva de axônios longos e tem sido relatada em algumas formas complicadas de neuropatias e paraplegias espásticas hereditárias. Casuística e métodos: Foi realizada a avaliação de 27 pacientes, 20 mulheres, com idade variando entre 4 e 58 anos. Todos os indivíduos compartilhavam o mesmo fenótipo (paraplegia espástica, atrofia de nervo óptico e neuropatia periférica) e tinham o mesmo haplótipo 11q13. Pacientes com história de diabetes mellitus ou alcoolismo foram excluídos do estudo. A avaliação neurológica incluiu a pesquisa dos escores modificados de sintomas e comprometimento neuropáticos. A força muscular foi testada e graduada conforme a escala MRC (Medical Research Council). Foi realizada a pesquisa da sensibilidade dolorosa, térmica, tátil, vibratória e artrestésica. O trofismo foi avaliado pela presença de deformidades na coluna e atrofia nos membros inferiores. Os reflexos profundos e o cutâneo plantar também foram analisados. Os estudos da condução nervosa foram realizados em um aparelho portátil Nicolet - Viking Quest, (Viasys, USA). Para os estudos de condução motora foram analisados os nervos axillar, mediano, ulnar, femoral, tibial e fibular direito. A condução sensitiva foi analisada nos nervos mediano, ulnar, radial, sural e fibular direito. O reflexo H e as ondas F foram avaliados com técnicas padrão. Alguns testes não puderam ser realizados devido à intensa atrofia e deformidades esqueléticas. O coeficiente de correlação de Pearson foi calculado entre a idade e os parâmetros, velocidade de condução, latência e amplitude. Valores de P < 0,05 foram considerados estatisticamente significantes. Resultados: Avaliação clínica: Todos os pacientes obtiveram escore de sinais neuropáticos graves e demonstraram déficit de força e atrofia distal. Deformidades dos pés estavam presentes em todos os pacientes e deformidades na coluna, em 58%. Os reflexos profundos dos membros superiores estavam exaltados em 92% dos casos e o reflexo patelar, em 63%. O reflexo Aquileu estava ausente em todos os pacientes. Todas as modalidades de sensibilidade foram afetadas, principalmente nos membros inferiores. Os dados do exame de sensibilidade na paciente de 4 anos foram desconsiderados. Estudo da condução nervosa sensitiva: Os SNAPs dos nervos mediano, sural e fibular estavam ausentes em todos os pacientes. SNAPs do nervo ulnar estavam ausentes em 96% da amostra e do nervo radial, em 80%. Estudo da condução nervosa motora: As latências motoras dos nervos axilar e femoral estavam normais em todos os pacientes. As amplitudes dos CMAPs estavam reduzidas em 15 e 52% da amostra nos nervos mediano e ulnar, respectivamente. Velocidades de condução estavam reduzidas em 50 e 41% desta casuística nos nervos mediano e ulnar, respectivamente. Velocidades de condução estavam acima de 80% do limite inferior da normalidade, em todos os nervos, exceto em 1 paciente que apresentou redução de 27% no nervo ulnar. Entretanto, este mesmo paciente apresentou amplitude menor que 2mV. Ondas F apresentavam aumento da latência, de acordo com a altura, em 100% dos casos. CMAPs estavam ausentes em 93 e 84% da amostra nos nervos fibular e tibial, respectivamente. Reflexo H estava ausente em 88% dos pacientes. Não houve correlação entre idade e a velocidade de condução, latência e amplitude dos nervos mediano e ulnar. Discussão: O estudo da condução nervosa neste grupo preencheu critérios para uma neuropatia primária axonal. Nenhum paciente apresentou bloqueio de condução ou dispersão temporal. As alterações encontradas na velocidade de condução provavelmente se devem à perda de fibras nervosas de condução rápida. Fenótipos SPOAN-like foram descritos em famílias com mutações nos genes C12orf65, TFG e OPA1. No entanto, não existem detalhes sobre a condução nervosa nestes pacientes. Neuropatia axonal de início tardio foi relacionada à SPG55 e DOA (dominant optic atrophy), enquanto neuropatia axonal e desmielinizante com leve comprometimento sensitivo foi descrita na família com mutação no gene TFG. Conclusão: Os pacientes com a síndrome SPOAN apresentam uma acentuada neuropatia axonal, sensitivo motora. As alterações encontradas na condução nervosa dos pacientes com síndrome SPOAN não são específicas, no entanto, resultados normais excluem esta condição em adultos. A paciente mais jovem desta casuística já apresentava alterações ao exame, o que pode sugerir um início precoce da neuropatia. Entretanto, não temos dados suficientes para afirmar que este seja um achado comum a todos os pacientes SPOAN / Introduction: SPOAN syndrome (Spastic Paraplegia, Optic Atrophy and Neuropathy) is a progressive neurodegenerative disorder of autosomal recessive inheritance described by our group in a large inbred family from Northeastern Brazil. The clinical picture is characterized by non-progressive congenital optic atrophy, progressive spastic paraplegia, axonal neuropathy, auditory startles, dysarthria, spinal and foot deformities and also extrapyramidal signs. Linkage studies mapped the responsible locus for the syndrome to a 2Mb region on chromosome 11q13. The gene responsible for SPOAN syndrome remains elusive. Materials and Methods: This is a cross sectional study which was conducted from 2009 to 2011. We evaluated 27 patients (20 females), with a0ges ranging from 4 to 58 years. All patients shared the same phenotype (spastic paraplegia, optic atrophy and peripheral neuropathy) and had the same 11q13 haplotype in homozygosis. Patients with history of diabetes mellitus or alcoholism were excluded from this study. All patients were evaluated by the same clinical researcher (SA). Neurological evaluation included determination of modified neuropathy symptoms (NSS) and neuropathy disability (NDS) scores. Motor strength was assessed using MRC scale. Sensibility assessment included small-fiber (pain and temperature) and large-fiber modalities (vibration-128Hz diapason, 10g monofilament and joint position sense). Spine deformities and atrophy in the lower limbs were observed. We also evaluated osteotendineous reflexes and cutaneous plantar reflexes. Nerve conduction studies were performed using a portable Nicolet - Viking Quest, (Viasys,USA). Motor conduction studies included axillary, median, ulnar, femoral, tibial and fibular nerves on the right side. Sensory nerve action potentials of median, ulnar, sural and superficial fibular nerves were recorded using a bar electrode of 3 cm and standard fixed distances. Tibial H-reflex was evaluated with standard technique. Minimal F wave latencies were obtained from ulnar and tibial nerves. A few tests could not be done in every patient due to severe deformities. We calculated Pearson\'s correlation coefficients between age and nerve conduction parameters, including velocities, latencies and amplitudes. P values <0.05 were considered statistically significant. Results: Clinical data: Neuropathic symptoms such as pain and paresthesias were rare. All patients had signs of severe neuropathy. All subjects demonstrated weakness and atrophy that were more significant distally than proximally. Foot deformities were present in all patients and spine deformities were seen in 58%. Upper limb deep tendon reflexes were exalted in 92% and patelar reflex in 63%. Ankle reflex was absent in all patients. In one patient, who was 4 years-old, sensory evaluation was inconsistent and the results were not considered. In all the other ones, sensory modalities were affected and occurred predominantly in the lower limbs. Electrodiagnostic data: Sensory nerve conduction: Median nerve SNAP was absent in all 27 patients. Ulnar nerve SNAPs were absent in 96%, whereas radial nerve SNPAs were absent in 80%. Superficial fibular and sural SNAPs were absent in all patients. Motor nerve conduction: The motor latencies of axillary and femoral nerves were normal in all patients. CMAP amplitudes were reduced in 15% of the median nerves and in 52% of the ulnar nerves. Conduction velocities (CV) were reduced in 50% of the median nerves and in 41% of the ulnar nerves. CV was above 80% of the lower normal limit for all nerves, except for one patient who showed a 27% reduction of ulnar CV, but had also a CMAP amplitude of less than 2 mV. F waves were prolonged according to the height in 100%. Only one patient who presented significant motor CV reduction of the ulnar nerve. CMAPs were absent in 93% of the fibular nerves and in 84% of the tibial nerves. A single fibular nerve showed more than 20% of CV reduction, but also had severely reduced CMAP amplitude. H reflex was absent in 88% of the patients. There was no correlation between age and neurophysiological parameters, such as median or ulnar CV, latencies or CMAP amplitudes. Discussion: Nerve conduction studies in this group fulfill criteria for primary axonal neuropathy. No patient showed conduction block or temporal dispersion. Abnormalities seen in CV and F waves are probably related to loss of fast conduction fiber nerves. We could not demonstrate correlation between age and nerve conduction parameters, including velocities, latencies and amplitudes. SPOAN-like phenotype has been found in families with mutations in C12orf65, TFG and OPA1 genes, however there is no detailed report on nerve conduction studies in these conditions. Axonal neuropathy is also described in SPG55 and DOA plus, but usually with a later onset than on SPOAN syndrome. Peripheral neuropathy is also described in the family with mutation in TFG gene, but this presents a different pattern characterized as a mixed axonal demyelinating neuropathy with mild sensory involvement. Although the nerve conduction abnormalities seen in SPOAN syndrome are not specific, normal results seem to rule out this condition, at least in adult patients. The younger patient in our series was 4-years-old, and her neurophysiological study was severely abnormal, suggesting an early-onset neuropathy. However, we do not have a comprehensive study of several young patients to support that this feature is

Les maladies neurodégénératives, telles que la maladie d'Alzheimer (AD) et Charcot Marie Tooth (CMT), sont des maladies complexes. Leurs mécanismes pathologiques ne sont toujours pas bien compris et les progrès dans la recherche et le développement de nouvelles thérapies potentielles modifiant la maladie sont lents. Les données catégorielles, comme les échelles de notation et les données sur les études d'association génomique (GWAS), sont largement utilisées dans les maladies neurodégénératives dans le diagnostic, la prédiction et le suivi de la progression. Il est important de comprendre et d'interpréter ces données correctement si nous voulons améliorer la recherche sur les maladies neurodégénératives. Le but de cette thèse est d'utiliser la théorie psychométrique moderne: théorie de la réponse d’item pour analyser ces données catégoriques afin de mieux comprendre les maladies neurodégénératives et de faciliter la recherche de médicaments correspondante. Tout d'abord, nous avons appliqué l'analyse de Rasch afin d'évaluer la validité du score de neuropathie Charcot-Marie-Tooth (CMTNS), un critère important d'évaluation principal pour les essais cliniques de la maladie de CMT. Nous avons ensuite adapté le modèle Rasch à l'analyse des associations génétiques pour identifier les gènes associés à la maladie d'Alzheimer. Cette méthode résume les génotypes catégoriques de plusieurs marqueurs génétiques tels que les polymorphisme nucléotidique (SNPs) en un seul score génétique. Enfin, nous avons calculé l'information mutuelle basée sur la théorie de réponse d’item pour sélectionner les items sensibles dans ADAS-cog, une mesure de fonctionnement cognitif la plus utilisées dans les études de la maladie d'Alzheimer, afin de mieux évaluer le progrès de la maladie. / Neurodegenerative diseases, such as Alzheimer’s disease (AD) and Charcot Marie Tooth (CMT), are complex diseases. Their pathological mechanisms are still not well understood, and the progress in the research and development of new potential disease-modifying therapies is slow. Categorical data like rating scales and Genome-Wide Association Studies (GWAS) data are widely utilized in the neurodegenerative diseases in the diagnosis, prediction and progression monitor. It is important to understand and interpret these data correctly if we want to improve the disease research. The purpose of this thesis is to use the modern psychometric Item Response Theory to analyze these categorical data for better understanding the neurodegenerative diseases and facilitating the corresponding drug research. First, we applied the Rasch analysis in order to assess the validity of the Charcot-Marie-Tooth Neuropathy Score (CMTNS), a main endpoint for the CMT disease clinical trials. We then adapted the Rasch model to the analysis of genetic associations and used to identify genes associated with Alzheimer’s disease by summarizing the categorical genotypes of several genetic markers such as Single Nucleotide Polymorphisms (SNPs) into one genetic score. Finally, to select sensitive items in the most used psychometrical tests for Alzheimer’s disease, we calculated the mutual information based on the item response model to evaluate the sensitivity of each item on the ADAS-cog scale.

La formation de la gaine de myéline est un processus complexe et finement régulé. Une altération de l’expression des gènes codant pour les protéines structurales de cette gaine entraine de graves neuropathies démyélinisantes. Notre objectif est d’identifier de nouvelles voies de signalisation capables de moduler l’expression de ces gènes. Les cellules de Schwann et les oligodendrocytes contiennent et synthétisent de grande quantité de dérivés oxydés du cholestérol : les oxystérols. Ces molécules sont connues pour leurs rôles dans le maintien de l’homéostasie du cholestérol et dans la progression des maladies neurodégénératives. Les oxystérols peuvent être classés en deux groupes : ceux dont l’oxydation a lieu sur la chaine carbonée latérale (25OH) et ceux qui portent une oxydation sur l’un des cycles du cholestérol (7KC). Nous nous sommes tout d’abord intéressés à la première catégorie d’oxystérols. Nous avons montré que le 25OH, réprimait l’expression des gènes de la myéline périphérique P0 et PMP22. Cette activité répressive était le fruit d’un mécanisme direct conduisant à une augmentation de la quantité des LXRs liés à leurs éléments de réponse sur les promoteurs des gènes de la myéline, et d’un mécanisme indirect provoquant une diminution de l’activité de la voie Wnt/β-caténine. En revanche, dans le SNC, nos résultats indiquent que le 25OH active l’expression des gènes de la myéline PLP et MBP. Le traitement, par ces oxystérols, de cultures organotypiques de cervelet démyélinisées par la lysolécithine permet une remyélinisation des axones des cellules de Purkinje. Nous nous sommes ensuite penchés sur le rôle du corégulateur transcriptionnel RIP140. Ce dernier peut soit agir comme un corépresseur soit comme un coactivateur. Il peut interagir avec le LXR. L’invalidation de RIP140 dans le poisson zèbre altère les gaines de myéline. Nous avons montré que RIP140 possédait des rôles bivalents dans la régulation de la myélinisation. En effet, il est capable d’activer mais aussi de réprimer l’activité transcriptionnelle de P0 et de PMP22. Enfin, nous nous sommes intéressés à la seconde catégorie d’oxystérols. Le 7KC est l’oxystérol majoritairement présent dans le SNP et la CS. Il est connu pour moduler l’action du récepteur aux dioxines : le AhR. Ce récepteur a été très largement étudié dans un cadre toxicologique. Cependant ses rôles et ses ligands endogènes restent à ce jour encore assez méconnus. Nos résultats indiquent que le AhR est impliqué dans le contrôle de l’expression des gènes de la myéline périphérique. L’invalidation du AhR, chez la souris, provoque des anomalies structurales de la gaine de myéline conduisant à des déficits moteurs. Cette étude a permis de mieux comprendre les dialogues entre les voies de signalisation gouvernant le processus de myélinisation. Ce travail apporte également de nouvelles perspectives thérapeutiques des maladies neurodégénératives comme la CMT1A ou la sclérose en plaques. / The myelination of axons is a complex process performed by Schwann cells (SC) and by oligodendrocytes (OL) respectively in the peripheral nervous system (PNS) and in the central nervous system (CNS). A slight change in expression of myelin structural proteins has a deep impact on the development and preservation of nerve fibers and their myelin sheaths, as observed for example in Charcot-Marie-Tooth disease or in Pelizaeus-Merzbacher disease. Our aim is to identify new signaling pathways able to control the expression of these structural proteins. SC and OL contain and synthesize high amount of reactive molecules generated from the oxidation of cholesterol: the oxysterols. Their implication in cholesterol homeostasis and in the progression of neurodegenerative disorders is well known but few data are available for their functions in myelination of PNS and CNS. Firstly, we demonstrate that oxysterols inhibit peripheral myelin gene expression: MPZ and PMP22. This downregulation is mediated by two mechanisms: by increasing the binding of LXRs to myelin genes promoters and by inhibiting the Wnt/β-catenin pathway leading to a decrease of b-catenin recruitment at the levels of the MPZ and PMP22 promoters. However, in the CNS, our data demonstrate that activation of LXRS by oxysterols stimulate myelin genes expression (PLP and MBP). Interestingly, by using demyelinated organotipc culture of cerebellum, we show that oxysterols enhance OL differentiation and promote remyelination, via LXRs. Then, we studied the role of the transcriptional coregulatory, RIP140, in myelination. RIP140 is able to act as a corepressor or as a coactivator and can interact with LXRs. In Zebrafish, the knocked down of the orthologue of RIP140 led to a decrease of peripheral and central myelin gene expression and to a defect in myelin sheath ultrastructure. Finally, we focused on impact of AhR in myelination process. AhR is a ligand activated transcription factor mostly known to interact with environmental pollutant like dioxins to mediate their toxic and carcinogenic effect. However, its detoxifying activity is posterior to the apparition of the gene and its physiological roles and endogenous ligands remain elusive. We show that the main oxysterol in the nervous system is 7-ketocholesterol which is an endogenous modulator of AhR. We report that the constitutive absence of AhR in mice leads to defects in locomotion behaviors. We studied the impact of this invalidation on the myelin of sciatic nerve. We observed a severe demyelinating phenotype and deregulation of myelin genes expression. Moreover, we demonstrated a cross-talk between AhR and Wnt/β-catenin pathways. Our data reveal a new endogenous role of AhR in myelination process.

Local tissue factors, ischemia and infection (which are often the cause of re-ulceration) are the main pathogenetic factors for diabetic foot disease (DFD). Neuropathic bone metabolism disorder leads to Charcot osteoarthropathy (CHOA). The aim of this dissertation was to assess experimentally the effectiveness of new skin substitutes, evaluate local vasculogenesis in different types of cell therapy of DFD, the role of infection in recurrence of DFD and scintigraphic parameters of activity of CHOA. Our studies concerning local pathological processes in DFD experimentally proved that gelatine nanofibers accelerate wound healing and can be suitable scaffolds for cell transfer and skin regeneration and also that acellular porcine dermis is more effective in healing of chronic wounds then xenotransplants. Our studies concerning therapeutic vasculogenesis confirmed that efficacy of stem cells (SC) harvested from bone marrow is similar in efficacy to SC separated from peripheral blood after stimulation. We found no evidence for systemic vasculogenesis by means of a significant increase of pro-angiogenic cytokines, which confirms the paracrine effect of injected SC. We proved a significant correlation between angiogeneisis inhibitor (endostatin) and the number of injected SC, which could be an indicator of...

Neurodegenerative diseases are becoming increasingly prevalent due to the ageing population, and are among the major contributors to disability and disease worldwide. The identification of the gene defects responsible for many of these conditions has played a major role in our understanding of the pathogenic processes involved, and provided opportunity to develop targeted treatment strategies. Cholinergic neurotransmission supports a wide range of physiological and behavioural processes and its dysfunction of cholinergic signalling has been associated with a number of disorders, including myasthenias, cardiovascular disease(1), attention-deficit hyperactivity disorder (ADHD) (2), Alzheimer’s disease (ADi), schizophrenia, addiction(3), and depression(4). SLC5A7 encodes the Na+/Cl- dependent, high-affinity choline transporter (CHT) which represents the rate limiting step in acetylcholine (Ach) synthesis and is critical for normal cholinergic signalling. The work in this thesis details two new inherited disorders, caused by distinct pathogenic disease mechanisms, associated with novel SLC5A7 mutations. Chapter three documents the discovery of two autosomal-dominantly acting SLC5A7/CHT mutations associated with adult onset motor neurone disorders. Initially we identified a frameshift mutation that results in premature truncation of the transporter protein in a large Welsh kindred affected with distal hereditary motor neuropathy type VII (dHMN-VII), in which neurodegeneration and muscle paresis is largely restricted to the distal limb muscles and vocal cords. The mutation responsible results in the dominant-negative interference of the mutant molecule with function of the wild type choline transporter, resulting in significantly reduced (although not completely abolished) transporter activity. This finding is further evidenced by the discovery of a second dHMN family associated with a distinct frameshift SLC5A7 mutation indicative of a similar dominant-negative disease mechanism. Together these findings corroborate a dominant-negative disease mechanism arising from C-terminal truncating SLC5A7 mutations associated with dHMN, and provide further insight into the role of aberrant choline transporter function in neurological disease. Chapter four describes N-terminal missense mutations located in the transmembrane spanning region of SLC5A7/CHT, associated with a severe infantile neuromuscular disorder characterised by predominantly central hypotonia and developmental delay. The phenotypic effects of these mutations are likely to result from the near abolition of CHT-mediated choline transport in homozygous individuals, and are in keeping with those observed in CHT knock-out mouse models(5). The development of a mouse model of the human motor neurone disease arising from SLC5A7 frameshift mutations should allow for further investigation of the mechanism by which truncated CHT leads to the dHMN phenotype. Chapter 5 details treatment hypotheses for dHMN, as well as the generation of a patient-specific knock-in mouse model carrying an Slc5a7 mutation orthologous to that identified in dHMN-VII families in chapter 3, and results from preliminary neurological phenotyping of the mouse model. This model will be crucially important for the exploration of treatment options in dHMN-VII motor neurone disease as a prelude to clinical trials in humans.