Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2016-04-26T22:28:02Z No. of bitstreams: 1 MarceloJoseSantiagoLisboa_DISSERT.pdf: 3090785 bytes, checksum: 44b33d32778eb8492274ab6f9eccf838 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2016-05-02T22:09:37Z (GMT) No. of bitstreams: 1 MarceloJoseSantiagoLisboa_DISSERT.pdf: 3090785 bytes, checksum: 44b33d32778eb8492274ab6f9eccf838 (MD5) / Made available in DSpace on 2016-05-02T22:09:37Z (GMT). No. of bitstreams: 1 MarceloJoseSantiagoLisboa_DISSERT.pdf: 3090785 bytes, checksum: 44b33d32778eb8492274ab6f9eccf838 (MD5) Previous issue date: 2015-08-26 / A Distrofia Muscular de Duchenne (DMD) ? uma doen?a de car?ter heredit?rio onde ocorre a aus?ncia da prote?na distrofina, levando a quadros de les?o miopatia grave decorrentes do aumento do estresse oxidativo e influxo de Ca+. Les?es na musculatura lisa intestinal podem comprometer a motilidade local devido a altera??es da estrutura da pr?pria t?nica muscular, bem como, a mudan?as morfofuncionais das estruturas do plexo mioent?rico. Este trabalho teve por objetivo avaliar as mudan?as ocorridas na t?nica muscular e nos neur?nios mioent?ricos colin?rgicos do ?leo de camundongos mdx e, os efeitos da suplementa??o com ?cido asc?rbico (AA) nestes dois componentes. Foram utilizados 30 camundongos machos C57BL/10 e 30 C57BL/10Mdx separados em grupos de acordo com a idade e tratamento (n=10): controle com 30 dias de idade (C30); distr?fico com 30 dias de idade (D30); controle com 60 dias de idade (G60); distr?fico com 60 dias de idade (D60); controle com 60 dias de idade suplementados com ?cido asc?rbico (200mg/kg de peso corporal) (CS60) e, distr?fico com 60 dias de idade suplementados com ?cido asc?rbico (200mg/kg de peso corporal) (GDS60). Ap?s o per?odo experimental os animais foram eutanasiados e os ?leos foram coletados e processados seguindo a rotina histol?gica e corados pela t?cnica de Tricr?mico de Masson e, para t?cnica histoqu?mica da Acetilcolinesterase em preparados totais de membrana. Os dados demonstraram que a espessura da t?nica muscular (?m) e a ?rea de m?sculo liso (?m2) do ?leo foi menor nos grupos distr?ficos, especialmente no grupo D30. Nos animais de DS60 a espessura da t?nica muscular foi semelhante aos de C60. Houve redu??o da densidade neuronal colin?rgica do plexo mioent?rico do ?leo foi menor nos animais D30, por?m esta foi semelhante nos animais de 60 dias sem tratamento (C60 e D60) e, maior nos animais DS60. ?rea do perfil do corpo celular (?m2) foi semelhante nos animais de C30-D30 e C60-D60, por?m est? foi maior em DS60. A raz?o ?rea nuclear/?rea citoplasm?tica foi menor em D30 e DS60 e, maior em D60. Desta forma podemos concluir que em camundongos mdx ocorrem altera??es significativas na morfologia da t?nica muscular e, consequentemente, mudan?as morfol?gicas e funcionais dos neur?nios colin?rgicos do plexo mioent?rico do ?leo, bem como, que a suplementa??o com AA teve efeito neuroprotetor nestes animais pois prevenindo a perda neuronal. / The Duchenne muscular dystrophy (DMD) is a hereditary disease where there is a lack of dystrophin protein, leading to severe injury myopathy frames resulting from increased oxidative stress and Ca + influx. Lesions on intestinal smooth muscle can compromise the local motility due to changes in the very muscular layer structure as well as the morphological and functional changes in the myenteric plexus structures. This study aimed to evaluate changes in the muscular wall and myenteric cholinergic neurons of the ileum of mdx mice, and the effects of supplementation with ascorbic acid (AA) in these two components. 30 male C57BL / 10 and 30 C57BL / 10Mdx separated into groups according to age and treatment (n = 10) were control at 30 days old (C30); dystrophic 30 days of age (D30); control with 60 days of age (C60); dystrophic with 60 days of age (D60); control at 60 days of age supplemented with ascorbic acid (200mg / kg body weight) (CS60) and dystrophic 60 days of age supplemented with ascorbic acid (200mg / kg body weight) (DS60). After the trial period the animals were euthanized and ileus have been collected and processed following the histological routine and stained by Masson's technique of Masson and for immunohistochemical technique of acetylcholinesterase in total membrane prepared. The data demonstrated that the thickness of the muscularis (?m) and smooth muscle area (?m 2) of the ileum was lower in dystrophic groups, especially Group D30. In animal DS60 the thickness of the muscular layer was similar to the C60. Decreased cholinergic neuronal density of the myenteric plexus of the ileum was lower in D30 animals, but this was similar in animals 60 days without treatment (C60 and D60) and higher in DS60 animals. Cell body profile area (?m 2) was similar in animals of C30-C60 and D30-D60, but is was higher in DS60. The nuclear area ratio / cytoplasmic area was lower in D30 and DS60 and higher in D60. Thus we conclude that mdx mice occur in significant changes in the morphology of the muscular layer and, consequently, morphological and functional changes of cholinergic neurons of the myenteric plexus of the ileum and that supplementation with AA had neuroprotective effects in these animals as preventing loss Neuronal.

La transplantation de cellules souches de muscle possède un grand potentiel pour la réparation à long terme du muscle dystrophique. Cependant, la croissance ex vivo des cellules souches musculaires réduit de manière significative l'efficacité de leur greffe puisque le potentiel myogénique est considérablement réduit lors de la mise en culture. La voie de signalisation Notch a émergé comme un régulateur majeur des cellules souches musculaires (MuSCs) et il a également été décrit que la sur-activation de Notch est crucial pour le maintien du caractère souche des MuSC. Cette découverte pourrait être traduite comme un bénéfice thérapeutique potentiel. Des MuSCs murines ont été fraîchement isolées et ensemencées sur des boîtes de culture recouverte de Dll1-Fc, le domaine extracellulaire de Delta-like-1 est fusionné au fragment Fc humain, afin d'activer la voie de signalisation Notch et avec un IgG hu-main comme contrôle. Nous avons utilisé le rAAV afin d’exprimer le Dll1 spécifique-ment dans les muscles de souris. Les souris P3 ont été traitées avec de l’AAV pendant 3 semaines et 6 semaines afin d’étudier l'effet de Dll1 au cours du développement postnatal. Afin d’étudier le processus de régénération, l'AAV a également été injecté dans les muscles de souris mdx alors que les souris de type sauvage ont été utilisées comme contrôle. Un potentiel caractère souche supérieur (marquée avec le Pax7) est observé dans les cultures des MuSCs qui sont recouverte de Dll1-Fc par rapport à leurs homologues contrôles, par contre le taux de proliférer est réduit. Au cours du développement postnatal, la sur-activation de la voie de signalisation Notch par Dll1 sur les fibres musculaires a été en mesure d'élargir le pool des cellules Pax7+, cependant elle entraîne une diminution de la masse musculaire avec réduction de la taille des fibres et ceci sans affecter l'accumulation des myonuclei. Dans les MuSCs quiescentes (de type sauvage), la sur-activation de la voie de signalisation Notch ne présente pas de réel effet. La surexpression de Dll1 dans le muscle mdx a diminué la masse musculaire et agrandit le pool de cellules souches musculaires, ce-pendant le taux de régénération n'a pas été affecté. L’augmentation des MuSCs est attribuée à une différenciation entravée des cellules souches musculaires. En étudiant la stimulation de la voie de signalisation Notch dans les MuSCs à la fois in vitro et in vivo, nous démontrons que sur-activation de Notch préserve le caractère souche des cellules via l’inhibition de la prolifération et de la différenciation myogénique des MuSCs. / Muscle stem cell transplantation possesses great potential for long-term repair of dys-trophic muscle. However expansion of muscle stem cells ex vivo significantly reduces their engraftment efficiency since the myogenic potential is dramatically lost in culture. The Notch signaling pathway has emerged as a major regulator of muscle stem cells (MuSCs) and it has recently been discovered that high Notch activity is crucial for maintaining stemness in MuSCs. This feature might be exploited and developed into a novel therapeutic approach.Murine MuSCs were freshly isolated and seeded on culture vessels coated with Dll1-Fc, which fused Delta-like-1 extracellular domain with human Fc, to activate Notch sig-naling and with human IgG as a control. The rAAV gene delivery system was em-ployed to express Dll1 in murine muscles. P3 mice were treated with AAV for 3 weeks and 6 weeks to investigate the effect of Dll1 during postnatal development. To investi-gate the regeneration process, AAV were injected into mdx muscles whereas wild-type mice were used as control.Higher potential stemness (marked by Pax7 positivity) was observed in MuSCs grow-ing on a Dll1-Fc surface as compared to their counterparts on the control surface, while their proliferation rate was reduced. During postnatal development, overstimulation of Notch signaling by Dll1 on the mus-cle fibers was able to enlarge the Pax7+ cell pool, while also resulting in decreased muscle mass and smaller muscle fibers without affecting the accretion of myonuclei into the fiber. In quiescent (wild-type) MuSCs, overstimulation of Notch signaling did not have any discernible effect. Overexpression of Dll1 in mdx muscle decreased the muscle mass and enlarged the muscle stem cell pool, while muscle regeneration re-mained unaffected. By investigating Notch stimulation in MuSCs both in vitro and in vivo, we demonstrate that high Notch activity preserves stemness via inhibition of MuSCs proliferation and myogenic differentiation. Our findings point out that the Dll1 molecule, as a canonical Notch ligand, might have a therapeutic potential in cell-based therapies against muscu-lar dystrophies.

Résumé de l'étude in vitro. L'objectif de cette étude in vitro était de créer un fantôme avec la même architecture musculaire (fibre, aponévrose ...) et les mêmes propriétés mécaniques que le muscle en condition passive et active. Deux fantômes homogènes ont été fabriqués avec différentes concentrations de plastisol pour simuler les propriétés élastiques du muscle en condition passive (50% plastisol) et active (70% de plastisol). Pour cela, des fils en Téflon (d = 0,9 mm) ont été insérés dans la partie supérieure du fantôme (50%) pour représenter les fibres musculaires. De plus, une feuille en matière plastique (8 x 15 cm) a également été placée au milieu du fantôme pour imiter la structure de l'aponévrose. Ensuite, des tests ERM ont été effectués à 90 Hz avec deux stimulateurs pneumatiques de différentes formes (tube en silicone, membrane circulaire) pour analyser l'effet du type du stimulateur sur la propagation des ondes. La longueur d'onde a été mesurée à partir des images phase et les propriétés élastiques (module de cisaillement) ont été calculées. Les deux fantômes (50% et 70%) ont montré des propriétés élastiques similaires à celles du muscle à l’état passif (2,40 ± 0,18 kPa) et actif (6,24 ± 0,21 kPa). Le stimulateur en forme de tube a donné des valeurs plus élevées (environ 1,2 kPa à 1,53 kPa). L'analyse du comportement des ondes a révélé un glissement le long de la feuille plastique. Ce phénomène a aussi été observé in vivo le long de l’aponévrose. L'onde a également été sensible à la présence des fils en téflon car des coupures, des trous, ont été identifiés au cours de la propagation de l’onde. Une nouvelle méthode de post-traitement a été créée pour mesurer les paramètres G' et G" à partir de tests ERM réalisés à plusieurs fréquences (60, 80, 100 Hz) et en utilisant des modèles rhéologiques. Cette méthode a été testée sur un fantôme (50%) qui n’avait pas d’inclusion. Les résultats des mesures viscoélastiques (G', G") ont été validés avec la technique HFVS (Hyper-Fréquence viscoélastique Spectroscopy). Des valeurs similaires, G' et G’’, ont été obtenues avec les deux techniques. Ce dernier résultat valide la nouvelle méthode de post-traitement pour mesurer les propriétés viscoélastiques. Résumé de l'étude in vivo. L'objectif de cette étude in vivo a été de développer des protocoles ERM pour caractériser les propriétés élastiques (module de cisaillement) des neuf muscles de la cuisse. Ces tests ont été effectués à une seule fréquence (90 Hz). Différents modules de cisaillement ont été trouvés entre les muscles. Le gracilis a révélé des propriétés élastiques plus élevées que les autres muscles. Ces différentes élasticités peuvent être dues à différentes compositions physiologiques et architecturales entre les tissus. Ensuite, les propriétés viscoélastiques des muscles ischio (ST, SM, et la BC) et du muscle Gr ont été déterminées en appliquant la nouvelle méthode de post-traitement des données (précédemment validée sur le fantôme 50%) avec des tests ERM multi fréquences (70, 90 et 120 Hz) et en utilisant des modèles rhéologiques. Les résultats ont montré que deux modèles rhéologiques, Zener et springpot, peuvent être utilisés pour mesurer les propriétés viscoélastiques des muscles à l’état passif. De plus, des résultats similaires ont été trouvés entre G "/ G ', obtenus expérimentalement à 90 Hz, et la valeur α du modèle de springpot. / Summary of the vitro studies. The objective of this in vitro study was to create a phantom witch the same muscle architecture (fiber, aponeurosis …) and mechanical properties of muscle in passive and active states. Two homogeneous phantoms were manufactured with different concentrations of plastisol to simulate the muscle elastic properties in passive (50% of plastisol) and active (70% of plastisol) muscle conditions. Moreover, teflon tubing pipes (D = 0.9 mm) were thread in the upper part of the phantom (50%) to represent the muscle fibers and a plastic sheet (8 x 15 cm) was also included in the middle of the phantom to mimic the aponeurosis structure. Subsequently, MRE tests were performed at 90Hz with two different pneumatic drivers, tube and round shapes, to analyze the effect of the type of driver on the wave propagation. The wavelength was measured from the phase images and the elastic properties (shear modulus) were calculated. Both phantoms revealed elastic properties which were in the same range as in vivo muscle in passive (2.40 ± 0.18 kPa) and active (6.24 ± 0.21 kPa) states. The impact of the type of driver showed higher values with the tube (range: 1.2 kPa to 1.53 kPa). The analysis of the wave behavior revealed a sliding along the plastic sheet as it was observed for in vivo muscle study. The wave was also sensitive to the presence of the fibers where gaps were identified. A new post processing method was established to measure G’ and G” from experimental multi frequencies (60, 80, 100 Hz) MRE (MMRE) tests and rheological models. This method was tested on the phantom (50%) made without fiber. Cross validation of the viscoelastic (G’, G”) results was made with Hyper-Frequency Viscoelastic Spectroscopy (HFVS). Both techniques showed similar range of values for G’ and G” at the same frequencies. This last result validated our new data processing for the viscoelastic measurement. Summary of the in vivo studies. The objective of this in vivo study was to develop MRE protocols to characterize the elastic properties (shear modulus) of the nine thigh muscles. These tests were performed at a single frequency (90Hz). Different shear moduli were found between the muscles. The gracilis revealed the highest elastic properties compared to all the other muscles. These different elasticities may be due to different physiological and architectural compositions between the tissues. Then the viscoelastic properties of the ischio (ST, SM, and BC) and Gr muscles were determined based on our new data-processing method (validated on the phantom 50%) using MMRE tests (70, 90 and 120Hz) and rheological models. The results revealed that two rheological models, zener and springpot, can be used to measure the viscoelastic properties in passive state. A similar trend was found between the experimental ratios G”/G’ obtained at 90 Hz and the α value of the springpot model. The present MRE muscle protocol, and the viscoelastic data base, could be used as non-invasive diagnostic tools to evaluate tissue alterations, the progression of diseases, and the effect of treatments, such as the ongoing therapeutic trials for Duchenne muscular dystrophy.

No description available.

Duchenne Muscular Dystrophy (DMD) is a disorder caused by loss-of-function mutations in dystrophin, a critical protein that maintains muscle fiber integrity. Our lab discovered that dystrophin-deficient skeletal muscle stem cells, also known as satellite cells, cannot generate enough myogenic progenitors for proper muscle regeneration. Previously, we demonstrated that WNT7A, a protein expressed during muscle regeneration, stimulates symmetric division of satellite cells, and gives rise to two daughter satellite cells. Conversely, epidermal growth factor (EGF) induces asymmetric division, which generates one daughter satellite cell and one committed precursor cell. We aimed to investigate these satellite cell division mechanisms following WNT7A or EGF treatment in a human model using healthy and DMD-patient derived hiPSCs differentiated into the myogenic lineage. The presence of satellite-like cells was confirmed in both lines by their characteristic expression of PAX7 and other myogenic markers. Intriguingly, DMD-patient hiPSCs precociously differentiated compared to healthy control human induced pluripotent stem cells (hiPSCs). More notably, WNT7A treatment had a potent effect on the DMD differentiated cells. High content analysis revealed an expansion of the satellite-like cell pool as observed by a higher number of PAX7+ cells within the total population and gene expression analysis demonstrated a significant increase in global PAX7 expression. In contrast, EGF treatment reduced the number of PAX7+ cells and increased the proportion of MYOG+ cells within the myogenic population, indicating an increase in myogenic progenitors. Taken together, WNT7A and EGF can alter the myogenic differentiation program of healthy and DMD-patient derived hiPSCs by modulating the satellite-like cell division dynamics.

<p>  </p> <p>Cardiovascular disease has been the leading cause of death in the United States for over 70 years. To evaluate the extent and progression of cardiovascular disease, non-invasive imaging techniques are frequently used clinically and pre-clinically. Current echocardiographic and cine magnetic resonance approaches rely on measurements that are typically obtained from two-dimensional images, which assumes uniformity of the structure being evaluated. To explore methods to potentially address these shortcomings, our group has developed and validated high frequency four-dimensional ultrasound techniques as well as created a software toolbox that allows for measurement of myocardial kinematics. In this thesis, I assisted in the application of these methods to two murine models of disease states: myocardial infarction and aortic aneurysm. Another study I aided in focused on cardiac magnetic resonance imaging data from patients with Duchenne muscular dystrophy. From our software, we are able to obtain various strain and strain rate estimates that reveal significant functional changes in infarction and Duchenne muscular dystrophy earlier than standard measurement techniques. Furthermore, we are able to identify vascular expansion, transmural thickening, and changes in hemodynamics prior to aneurysm development. Earlier detection and localization allows for more targeted surveillance and interventions, which ultimately may result in improved clinical outcomes. Ideally, these findings can be used to expand the capabilities of cardiac research and the development of clinically applicable imaging techniques and treatments to better address underlying cardiovascular pathophysiology. </p>

Duchenne Muscular Dystrophy (DMD) is an X-linked recessive, progressive muscle-wasting disease characterized by mutations in the dystrophin gene. Duchenne muscular dystrophy is the most common and most severe form of inherited muscle diseases, with an incidence of 1 in 3,500 male births1,2. Mutations in the dystrophin gene result in non-functional dystrophin or the complete absence of the protein dystrophin, resulting in necrosis and fibrosis in the muscle, loss of ambulation, cardiomyopathies, inadequate or failure of respiratory function, and decreased lifespan. Although there has been little research for effective nutritional strategies, dietary intervention may be effective as an adjuvant treatment. In this study, wild type (WT) and mdx animals were provided either a control or elevated branched chain amino acid (BCAA) diet nocturnally for 25 weeks to determine if the elevated BCAAs would attenuate muscle torque loss. Twenty-five weeks of chronic, elevated BCAA supplementation had no impact on muscle function measures. Interestingly, mdx and WT animals had the same torque responses in the low stimulation frequencies (1 Hz – 30 Hz) compared to higher stimulation frequencies. Tetanus was reached at a much lower stimulation frequency in mdx animals compared to WT animals (100 Hz vs +150 Hz). The mdx mouse consistently had more cage activity in the light cycle X- and Y-planes. Interestingly, animals on the BCAA diet increased X-, Y-, and Z-plane activity in the dark cycles at four weeks while animals on the control diet more Z-plane activity at 25 weeks, although not significant. All three BCAAs were elevated in the plasma at 25 weeks, although only Leu was significantly elevated. The BCAAs had no effect on. The diaphragm and skeletal muscle masses were larger in mdx animals, and WT animals had a significantly larger epididymal fat pad. The active state of BCKDC determined by phosphorylation of the E1α enzyme was greater in WT animals in white skeletal muscle, but not red skeletal muscle. Protein synthesis effectors of the mTORC1 signaling pathway and autophagy markers were similar among groups. Wild type animals had increased mTORC1 effectors and animals on the BCAA diet had decreased autophagy markers, although not significant. Although BCAAs did not affect muscle function, fibrosis, or protein synthesis effectors, this study illustrates the functionality of mdx muscles over time. It would be interesting to see how the different muscle fiber types are affected by DMD, noting the differences between the diaphragm, heart, red muscle, and white muscle fibrosis markers. Although there was no increase in mTORC1 effectors with an elevated BCAA diet, it would be interesting to determine muscle protein synthesis, myofibrillar protein synthesis, and total protein turnover in the mdx mouse with an elevated BCAA diet, although the dietary intervention started when mice arrived at 4 weeks of age, earlier intervention may be beneficial early in the disease process. / Doctor of Philosophy / Duchenne Muscular Dystrophy (DMD) is an X-linked recessive, progressive muscle-wasting disease characterized by mutations in the dystrophin gene. Duchenne muscular dystrophy is the most common and most severe form of inherited muscle diseases, with an incidence of 1 in 3,500 male births1,2. Mutations in the dystrophin gene result in non-functional dystrophin or the complete absence of the protein dystrophin, resulting in necrosis and fibrosis in the muscle, loss of movement and walking ability, cardiomyopathies, inadequate or failure of respiratory function, and decreased lifespan. Although there has been little research for effective nutritional strategies, dietary intervention may be effective as an adjuvant treatment and palliative care. The branched chain amino acids (BCAAs) are known to directly stimulate muscle protein synthesis by direct activation of the mechanistic target of rapamycin complex 1 (mTORC1). This study aimed to illustrate the differences between diseased and healthy mice and determine if BCAAs can reduce muscle torque loss. Twenty-five weeks of chronic, elevated BCAA supplementation had no impact on muscle function measures. Interestingly, mdx and WT animals had the same torque responses in the low stimulation frequencies (1 Hz – 30 Hz) compared to higher stimulation frequencies. Tetanus was reached at a much lower stimulation frequency in mdx animals compared to WT animals (100 Hz vs +150 Hz). The mdx mouse consistently had more cage activity in the light cycle X- and Y-planes. Interestingly, animals on the BCAA diet increased X-, Y-, and Z-plane activity in the dark cycles at four weeks while animals on the control diet more Z-plane activity at 25 weeks, although not significant. All three BCAAs were elevated in the plasma at 25 weeks, although only Leu was significantly elevated. The BCAAs had no effect on. The diaphragm and skeletal muscle masses were larger in mdx animals, and WT animals had a significantly larger epididymal fat pad. The active state of BCKDC determined by phosphorylation of the E1α enzyme was greater in WT animals in white skeletal muscle, but not red skeletal muscle. Protein synthesis effectors of the mTORC1 signaling pathway and autophagy markers were similar among groups. Wild type animals had increased mTORC1 effectors and animals on the BCAA diet had decreased autophagy markers, although not significant. Although BCAAs did not affect muscle function, fibrosis, or protein synthesis effectors, this study illustrates the functionality of mdx muscles over time. It would be interesting to see how the different muscle fiber types are affected by DMD, noting the differences between the diaphragm, heart, red muscle, and white muscle fibrosis markers. Although there was no increase in mTORC1 effectors with an elevated BCAA diet, it would be interesting to determine muscle protein synthesis, myofibrillar protein synthesis, and total protein turnover in the mdx mouse with an elevated BCAA diet, although the dietary intervention started when mice arrived at 4 weeks of age, earlier intervention may be beneficial early in the disease process.

Titre de l'écran-titre (visionné le 5 juin 2023) / Les maladies neuromusculaires héréditaires n'ont généralement pas de traitement efficace à ce jour. Deux d'entre elles sont étudiées par le laboratoire, soit l'ataxie de Friedreich (FRDA) et la dystrophie musculaire de Duchenne (DMD). Il existe plusieurs modèles de souris pour la FRDA, mais aucun ne reflétait la dégénération progressive de la maladie au niveau locomoteur et cardiaque. Le but de mes travaux était donc de trouver et caractériser un modèle murin qui rendrait possible l'étude des effets d'un traitement de thérapie génique sur ces symptômes. Deux modèles de souris ont été comparés : 1) le YG8sR auquel un petit ARN en épingle à cheveux (shRNA) ciblant l'ARNm de la frataxine est injecté en IV pour tenter de réduire l'expression de cette protéine pour augmenter la sévérité des symptômes de la maladie et 2) le YG8-800 qui est un nouveau modèle prometteur possédant 800 répétitions de GAA. Le YG8-800 présente un phénotype qui modélise mieux l'évolution de la maladie chez l'humain avec une détérioration graduelle de la coordination corrélée à une diminution de la frataxine plus marquée. En effet, les souris YG8-800 font plus de fautes et nécessitent plus de temps pour traverser une poutre et T inversé ou dentée et restent suspendues moins longtemps sur une grille. Elles présentent aussi significativement moins de frataxine dans tous les organes étudiés que les souris YG8sR. Pour la DMD, le traitement étudié est la greffe de myoblastes qui consiste à injecter une suspension de myoblastes sains dans un muscle n'exprimant pas la dystrophine. Cette méthode vise à briser les fibres musculaires existantes pour y faire fusionner les cellules injectées et de ce fait apporter le matériel génétique pour synthétiser la dystrophine dans la fibre musculaire malade. En moyenne, le taux de survie à la procédure des cellules greffées est de 5% dans la littérature. L'expérience vise donc à optimiser les conditions de greffe pour augmenter la survie des myoblastes après la greffe. Le Célastrol est un composé reconnu pour son effet anti-inflammatoire et protecteur, il a donc été testé pour protéger les cellules. La première greffe indiquait une amélioration significative de la survie des cellules, mais la deuxième n'a pas reproduit cette tendance. D'autres expériences seront nécessaires pour déterminer si le Célastrol peut améliorer la survie des cellules greffées. / Hereditary neuromuscular diseases generally have no effective treatment to date. Two of them are being studied by my laboratory: Friedreich's ataxia (FRDA) and Duchenne muscular dystrophy (DMD). There are several mouse models for FRDA, but none reflected the progressive degeneration of the disease at the musculoskeletal and cardiac level. My goal was thus to find and characterize a model that will make it possible to study the effects of gene therapy treatments on these symptoms. I have compared two mouse models: 1) the YG8sR to which a short hairpin RNA (shRNA) targeting frataxin mRNA was i.v. injected to reduce the expression of frataxin to increase the severity of the symptoms and 2) the YG8-800, a promising new model with 800 GAA repeats. The YG8-800 has a phenotype that better reproduces the disease progression in humans with a gradual deterioration in coordination correlated due to a more pronounced frataxin reduction. In fact YG8-800 mice make more foot faults and need more time to cross the inverted T beam or notched beam. They also hang for a shorter time on the grid and contain significantly less frataxin in all studied organs than YG8sR mice. For DMD, the treatment studied was myoblast transplantation, which consisted of injecting a suspension of healthy myoblasts into a muscle that does not express dystrophin, the absent protein in DMD patients. This method aims to damage the existing muscle fibers to permit the fusion of the injected cells and thus provide the gene to express dystrophin in the diseased muscle fibers. On average, the procedure survival rate of transplanted cells is 5% in the literature. My experiment therefore aimed to improve the transplant conditions to increase myoblast survival. I have evaluated the protective effects of Celastrol is a compound known for its anti-inflammatory. The first transplant indicated a significant improvement of cell survival, but a second did not confirm this effect. Further experiments will be needed to determine if Celastrol can improve the survival of transplanted cells.

O nosso objetivo foi de avaliar evolutivamente a perda da força muscular e das habilidades motoras, bem como a progressão da distribuição da fraqueza muscular nos diferentes segmentos dos membros superiores em pacientes com distrofia muscular de Duchenne em corticoterapia. Selecionamos seguintes testes de fácil aplicação: Teste de força Manual Muscular Escala Medical Research Council; Teste ABC provas de coordenação visual-motora e fatigabilidade provas n° 1, 3, 7 e 8; Grau funcional de Brooke; Índice de Barthel. Os testes foram aplicados em 40 pacientes com idades entre 5 e 15 anos, deambulantes e não deambulantes, os quais foram avaliados três vezes, com intervalos de seis meses entre cada avaliação. Os resultados mostraram que a progressão da distribuição da força muscular nos membros superiores ocorreu dos segmentos proximais para os distais em todos os pacientes e foi maior nos pacientes não deambulantes e com maior idade. O ato de escrever (pegada no lápis e/ou caneta) não foi influenciado pela progressão; no entanto, o aumento da fatigabilidade foi um fator limitante contribuindo para a redução do ritmo e da qualidade da escrita. O grau funcional de Brooke confirmou a variação das medidas de força muscular e nas atividades de vida diária o nível de dependência foi maior nos pacientes com maior idade, acontecendo nestes compensações funcionais importantes que lhes permitiram realizar a atividade de forma adaptada. / Our aim was to analyze the progression of the involvement of muscle strength and functional motor hability as well as the progression of the weakness pattern in the upper limbs of children with Duchenne muscular dystrophy who were receiving steroid therapy. We evaluated 40 patients with DMD, aged 5-15 years, by using the following simple tests: MRC score based on the upper limbs muscles; ABC tests number 1, 3, 7 and 8 for assessing visual motor coordination and fatigability; Brooke functional ability scale and modified Barthel index. All boys were evaluated every 6 months along a period of 18 months. The loss of muscular strength showed a proximal to distal progression and was greater in non- ambulant and in older patients. The act of writing (to hold pencil or pen) was not influenced by the progression; however, the fatigability increased and was a limiting factor for the speed and the quality of the hand writing. The Brooke functional ability scale confirmed the changes in the muscular strength. The performance in daily activities showed a greater dependence in the older patients who adopted functional compensations for performing the activities in an adapted way.

O nosso objetivo foi de avaliar evolutivamente a perda da força muscular e das habilidades motoras, bem como a progressão da distribuição da fraqueza muscular nos diferentes segmentos dos membros superiores em pacientes com distrofia muscular de Duchenne em corticoterapia. Selecionamos seguintes testes de fácil aplicação: Teste de força Manual Muscular Escala Medical Research Council; Teste ABC provas de coordenação visual-motora e fatigabilidade provas n° 1, 3, 7 e 8; Grau funcional de Brooke; Índice de Barthel. Os testes foram aplicados em 40 pacientes com idades entre 5 e 15 anos, deambulantes e não deambulantes, os quais foram avaliados três vezes, com intervalos de seis meses entre cada avaliação. Os resultados mostraram que a progressão da distribuição da força muscular nos membros superiores ocorreu dos segmentos proximais para os distais em todos os pacientes e foi maior nos pacientes não deambulantes e com maior idade. O ato de escrever (pegada no lápis e/ou caneta) não foi influenciado pela progressão; no entanto, o aumento da fatigabilidade foi um fator limitante contribuindo para a redução do ritmo e da qualidade da escrita. O grau funcional de Brooke confirmou a variação das medidas de força muscular e nas atividades de vida diária o nível de dependência foi maior nos pacientes com maior idade, acontecendo nestes compensações funcionais importantes que lhes permitiram realizar a atividade de forma adaptada. / Our aim was to analyze the progression of the involvement of muscle strength and functional motor hability as well as the progression of the weakness pattern in the upper limbs of children with Duchenne muscular dystrophy who were receiving steroid therapy. We evaluated 40 patients with DMD, aged 5-15 years, by using the following simple tests: MRC score based on the upper limbs muscles; ABC tests number 1, 3, 7 and 8 for assessing visual motor coordination and fatigability; Brooke functional ability scale and modified Barthel index. All boys were evaluated every 6 months along a period of 18 months. The loss of muscular strength showed a proximal to distal progression and was greater in non- ambulant and in older patients. The act of writing (to hold pencil or pen) was not influenced by the progression; however, the fatigability increased and was a limiting factor for the speed and the quality of the hand writing. The Brooke functional ability scale confirmed the changes in the muscular strength. The performance in daily activities showed a greater dependence in the older patients who adopted functional compensations for performing the activities in an adapted way.