INTRODUÇÃO: Atualmente ainda não está claro se a atividade muscular e o treinamento físico afetam a síntese de carnosina muscular (Mcar) em resposta à suplementação de beta-alanina. OBJETIVO: verificar o impacto da atividade e inatividade muscular sobre o conteúdo de carnosina muscular (Mcarn) e sobre o aumento de Mcarn em resposta à suplementação de beta-alanina e, adicionalmente, verificar seu efeito sobra a capacidade de realizar esforços de alta intensidade. METHODS: Dezesseis homens treinados com lesão medular (LM) (escala ASIA: AIS A ou AIS B) foram divididos em 2 grupos: beta-alanina (BA) (N = 11) e placebo (PL) (N = 5). Amostras de biópsias musculares foram obtidas do músculo vasto lateral inativo e do deltoide ativo, antes e após 28 dias de suplementação com beta-alanina (6.4 g.dia-1). Teste t independente foi aplicado para comparar a Mcar basal e a variação absoluta (PÓS - PRÉ) entre os músculos vasto lateral e deltoide. Um teste supramáximo de carga constante até a exaustão e um teste de Wingate de série única foram aplicados para verificar a capacidade de realizar esforços de alta intensidade. Análises por Modelo Misto foram aplicadas para comparar a Mcar, o tempo até a exaustão e os valores de potência pico intra e entre sujeitos. RESULTADOS: (média ± desvio padrão): a concentração basal de Mcar no vasto lateral foi significativamente maior que no deltoide (32,0 ± 12 vs. 20,5 ± 6,1 mmol.kg-1 de músculo seco; p = 0,02). As variações absolutas na Mcar foram significativamente maiores no grupo BA em comparação com o PL, tanto para o vasto lateral (BA: 17,6 ± 10,4 mmol.kg-1 de músculo seco; PL: 2,5 ± 2,3 mmol.kg-1 de músculo seco; p = 0,002) como para o deltoide (BA: 15,7 ± 6,8 mmol.kg-1 de músculo seco; PL: 1,4 ± 2,7 mmol.kg-1 de músculo seco; p <0,001). As variações absolutas verificadas no vasto lateral e deltoide não foram diferentes entre si (vasto lateral: 17,6 ± 10,4; deltóide: 15,7 ± 6,8 mmol.kg-1 do músculo seco; p = 0,6). Não houve efeito da suplementação sobre o tempo até exaustão e potência pico. CONCLUSÃO: a inatividade muscular crônica ocasionada por lesão medular, não afeta a Mcarn e também não interfere sobre o aumento de Mcarn em resposta à suplementação de BA. Estes resultados sugerem que a atividade muscular ou o estado de treinamento não influenciam a capacidade de síntese de Mcarn em resposta à suplementação de beta-alanina / BACKGROUND: it is currently unclear whether muscle activity and exercise training affect the ability of the skeletal muscle to synthesise carnosine in response to beta-alanine supplementation. PURPOSE: to study the impact of the extremes of muscle activity and inactivity on muscle carnosine content (Mcarn) and Mcarn loading in response to beta-alanine supplementation. METHODS: 16 trained male with spinal cord injury (SCI) (ASIA scale: AIS A or AIS B) were divided into 2 groups: beta-alanine (BA) (N = 11) and placebo (PL) (N = 5). Muscle biopsies samples were obtained from active deltoid and paralysed vastus lateralis at baseline and after 28 days of &#946;-alanine supplementation (6.4 g.day-1). Unpaired t-tests were applied to compare Mcarn at baseline and the absolute pre-post change in vastus lateralis and deltoid. Mixed model was used to compare Mcarn values within- and between-subjects. RESULTS: (mean±SD): Baseline Mcarn concentration in vastus lateralis was significantly higher than in deltoid (32.0±12 vs. 20.5±6.1 mmol.kg-1 dry muscle; p=0.02). Absolute changes in Mcarn was significantly higher in the BA group in comparison with PL for both vastus lateralis (BA: 17.6±10.4 mmol.kg-1 dry muscle; PL: 2.5±2.3 mmol.kg-1 dry muscle; p=0.002) and deltoid (BA: 15.7±6.8 mmol?kg-1 dry muscle; PL: 1.4±2.7 mmol?kg-1 dry muscle; p<0.001). Absolut changes in Mcarn following BA supplementation between inactive vastus lateralis and active deltoid was not different (vastus lateralis: 17.6±10.4; deltoid: 15.7±6.8 mmol?kg-1 dry muscle; p=0.6). CONCLUSION: chronic muscle inactivity due to paralysis in SCI does not affect Mcarn at baseline and does not affect Mcarn loading in response to BA supplementation. These results suggest that muscle activity or training status does not influence Mcarn synthesis capacity in response to beta-alanine supplementation

Thesis (Ph. D.)--University of Missouri--Columbia, 2002. / Typescript. Vita. Includes bibliographical references (leaves 141-160). Also issued on the Internet.

O tecido muscular tem uma alta capacidade de regeneração após lesão, que está diretamente ligada à presença de células satélites (SCs). Essas células são as principais células-tronco do músculo e também têm um papel fundamental no desenvolvimento muscular na embriogênese. Embora quiescente nos músculos adultos normais, as SCs podem ser ativadas por sinais específicos após lesão muscular. Em doenças caracterizadas por processo de degeneração crônica, como distrofias musculares, as SCs são constantemente ativadas, e esta condição pode levar à depleção do pool de SCs e consequente falha no processo regenerativo. Nós estudamos as SCs musculares nas linhagens distróficas murinas DMDmdx, Largemyd, DMDmdx/Largemyd, em comparação a camundongos normais, com o principal objetivo de avaliar o comportamento das SCs em músculos distróficos com diferentes graus de degeneração histopatológica. A expressão de genes e proteínas de fatores de transcrição relacionados a SCs foram estudadas no músculo, e os resultados foram comparados com as características histopatológicas de regeneração e degeneração e estado de proliferação de células musculares. Nossos resultados mostraram que o músculo distrófico mantém seu pool de células satélites, expressando PAX7, um importante fator muscular para autorrenovação do pool de SCs, em níveis semelhantes em todas as linhagens distróficas e controle normal. As células isoladas de músculo distrófico apresentaram uma maior proporção de células em proliferação, como observado pela análise dos marcadores de ciclo celular no músculo gastrocnêmio dissociado, com maior número de células na fase G2/M. A cascata de genes de regeneração é ativada no músculo distrófico, com altos níveis de expressão de fatores de regeneração muscular, como MYOD e Myogenin. O músculo distrófico mantém a capacidade de formar novas fibras, observada por um número significativo de fibras recém formadas, que expressam dMHC, em todas as linhagens analisadas. No entanto, essas novas fibras mostram características de maturação incompleta, como tamanho pequeno e pouca variação em seu calibre, que pode ser determinante para sua disfunção. A degeneração muscular é intensa apesar da regeneração, com infiltração significativa de tecido conjuntivo em camundongos distróficos. Em conclusão, nossos achados sugerem que os músculos distróficos, independentemente do grau de degeneração, mantêm o pool de células satélites com capacidade proliferativa e estão prontos para responder aos estímulos regenerativos. Por outro lado, a maturação dessas novas fibras é incompleta e não previne a degeneração do músculo / Muscle tissue has a high regeneration capacity after injury, which is directly linked to satellite cells (SCs). These cells are the main stem cells of the muscle and also have a key role in muscle development in embryogenesis. Although quiescent in normal adult muscles, SCs can be activated by specific signals upon muscle injury. In diseases characterized by chronic degeneration process, such as muscular dystrophies, the SCs are constantly activated, leading to depletion of the SC pool and consequent failure of the regenerative process. We studied muscle SCs in the mouse dystrophic strains DMDmdx, Largemyd, DMDmdx/Largemyd, comparing to wild-type mice, with the main objective to evaluate SCs behavior in dystrophic muscles with different degrees of histopathological degeneration. Gene and protein expression of transcription factors related to SCs were studied in the muscle, and the results were compared to regenerating and degenerating histopathologic pattern and proliferative state of muscle cells. Our results showed that the dystrophic muscle retains its satellite cells pool, expressing PAX7, an important muscle factor for self-renewal of the SCs pool, at similar levels in all dystrophic strains and wild-type. Dystrophic muscle single cells presented a higher proportion of proliferating cells, as observed by the analysis of cell cycle markers in dissociated gastrocnemius muscle, with a greater number of cells in the G2/M phase. The cascade of regeneration genes is activated in the dystrophic muscle, with high levels of expression of muscle regenerating factors, such as MYOD and Myogenin. Dystrophic muscle retains the ability to form new fibers, as observed by a significant number of new fibers expressing dMHC in all dystrophic strains. However, these new fibers show incomplete maturation characteristics, such as small size and no variation in fiber caliber, which could be determinant for its dysfunction. Muscle degeneration is intense in spite of regeneration, with significant more connective tissue infiltration in dystrophic mice than wild-typemice. In conclusion, our findings suggest that dystrophic muscles, independently of the degree of degeneration, retain the pool of satellite cells with proliferating capacity and ready to respond to regenerating stimuli. On the other hand, the maturation of these new fibers is incomplete and do not prevent the degeneration of the muscle

Duchenne muscular dystrophy is an inherited, X-linked recessive skeletal muscle disorder that is characterized by mutations in the dystrophin gene [1]. Therefore, the disease affects primarily males and women are typically carriers. 1 in 3500 males in the United States are affected [1]. Dystrophin is a critical, large scaffolding protein in the dystrophin-glycoprotein complex found at the sarcolemma of skeletal muscle [1]. The complex helps maintain sarcolemma integrity and stability during muscle contractions by coupling the extracellular matrix proteins to the intracellular cytoskeleton in skeletal muscle [1]. Loss-of-function mutations in the dystrophin protein affect all skeletal muscle found throughout the human body. The 427 kD protein is also present in cardiac muscle, the brain, and peripheral nerves, thus affecting these tissues over time, as well [1]. One theory suggests the weakened stability of the dystrophin-glycoprotein complex when dystrophin is not expressed results in transient membrane tears during contraction, which permit pathological calcium influx [1]. Damaged skeletal muscle results in repair and regeneration of the tissue however, continual damage over time (referred to as muscle wasting) results in extensive fibrosis and loss of muscle fibers. The purpose of this thesis is to provide a comprehensive review on several molecular mechanisms that underlie Duchenne muscular dystrophy and to investigate current treatments and propose potential therapeutic targets for future research.

Lower motor neuron diseases (LMNDs) include a large spectrum of clinically and genetically heterogeneous disorders, characterized by progressive anterior horn cell degeneration. The aims of this thesis were on the one hand to refine the phenotypic description and the clinical classification of hereditary LMNDs, and on the other hand to improve our knowledge of the genetic bases of these disorders. This work was performed in collaboration with the Centre of Human Genetics of the Necker-Enfants-Malades Hospital in Paris. We focused our researches on autosomal recessive variants of LMNDs. First, we selected patients with Spinal Muscular Atrophy with Respiratory Distress (SMARD or d-HMN VI). This severe variant of autosomal recessive LMND is characterized by neurogenic muscular atrophy associated with early life-threatening respiratory failure due to diaphragmatic dysfunction. SMARD type 1 has been ascribed to mutations in the immunoglobulin mu-binding protein 2 (IGHMBP2) gene on chromosome 11q13-q21. We reported the identification of 9 novel IGHMBP2 mutations in five SMARD1 patients, Seven of them occurred at highly conserved residues of the putative DNA helicase domain, suggesting that this particular domain plays a major role in the SMARD1 disease causing mechanism (Hum Mutat. 2004; 23(5):525-6). Then, we collected families and sporadic patients affected by chronic distal spinal muscular atrophy (d-HMN III/IV), an autosomal recessive variant of LMND characterized by a progressive motor weakness and muscular atrophy, predominating in the distal parts of the limbs. A form of chronic dSMA gene had been mapped to a 10.3 cM interval on chromosome 11q13. By linkage analysis in 12 European chronic dSMA families, we reduced the genetic interval to a 2.6cM region on chromosome 11q13.3 and showed partial linkage disequilibrium between 3 rare alleles and the mutant chromosome in European patients, suggesting that most chronic dSMA chromosomes are derived from a single ancestor (Eur J Hum Genet. 2004;12(6):483-8). Additional experiments are now in progress at Necker-Enfants-Malades Hospital, in order to identify the disease-causing gene. At last, we described the clinical features of a novel variant of autosomal recessive LMND, characterized by childhood onset, generalized muscle involvement, and severe outcome. Studying a large inbred African family, we mapped the disease gene to a 3.9-cM interval on chromosome 1p36 (Neurology.2006;67(1):120-4). We identified a homozygous missense mutation of the PLEKHG5 gene and performed in vitro experiments to clarify the pathogenic function of this mutation. In transiently transfected HEK293 and MCF10A cell lines, we found that wild-type PLEKHG5 activated the NFkB signaling pathway and that both the stability and the intracellular location of mutant PLEKHG5 protein were altered, severely impairing the NFkB transduction pathway. Moreover, we observed aggregates in transiently transfected NSC34 murine motor neurons overexpressing the mutant PLEKHG5 protein. In conclusion, we showed that both loss of PLEKHG5 function and aggregate formation might contribute to neurotoxicity in this novel form of LMND (Am J Hum Genet. 2007;81(1):67-76). Further experiments should now be planned, in particular to understand the role of aggregates in neurodegeneration, and to precise the links between the PLEKHG5 gene and the other LMNDs-causing genes. In conclusion, we hope that this work, contributing to a better understanding of the molecular mechanisms involved in motor neuron degeneration, will open the way to new therapeutic strategies. / Les amyotrophies spinales (SMA) se caractérisent par une dégénérescence des motoneurones des cornes antérieures de la moelle épinière ou des noyaux du tronc cérébral. Elles comprennent de nombreuses entités, très hétérogènes tant sur le plan clinique que sur le plan génétique. Cette thèse vise à préciser les caractéristiques phénotypiques et génétiques des différentes formes d’amyotrophie spinale, en particulier celles des variants de transmission autosomique récessive. En premier lieu, nous avons sélectionné une cohorte de patients dont le tableau clinique était compatible avec la variante SMARD d’amyotrophie spinale (pour Spinal Muscular Atrophy with Respiratory Distress). Il s’agit d’une forme très sévère d’amyotrophie spinale autosomique récessive, qui associe une faiblesse musculaire à prédominance distale et une détresse respiratoire précoce secondaire à une paralysie diaphragmatique. Nous avons décrit 9 nouvelles mutations au sein du gène IGHMBP2 (pour immunoglobulin µ-binding protein 2) chez 5 patients atteints et confirmé ainsi le rôle pathogène de ce gène. Sept des 9 mutations décrites concernent des acides aminés conservés dans les espèces et localisés dans le domaine hélicase. Ce domaine pourrait donc jouer un rôle essentiel dans la physiopathologie de la maladie (Hum Mutat. 2004; 23(5):525-6). Deuxièmement, nous avons rassemblé des patients atteints d’amyotrophie spinale chronique distale (d-HMN III/IV), de transmission autosomique récessive. Cette affection se définit par une amyotrophie et une faiblesse musculaire progressive qui prédomine au niveau des pieds et des mains. Grâce à des analyses de liaison réalisées dans 12 familles européennes, nous avons restreint la localisation génétique sur le chromosome 11 (en 11q13.3) à un intervalle de 2.6 cM. Nous avons également mis en évidence un déséquilibre de liaison entre 3 allèles rares et le locus génétique, ce qui suggère un phénomène d’effet fondateur dans la population caucasienne (Eur J Hum Genet. 2004;12(6):483-8). Des études complémentaires sont actuellement en cours à l’Hôpital Necker-Enfants-malades à Paris pour identifier le gène responsable de cette forme chronique d’amyotrophie spinale distale. Troisièment, nous avons décrit un nouveau variant d’amyotrophie spinale de transmission autosomique récessive. L’amyotrophie et la faiblesse musculaire débutent vers l’âge de 3 ans et concernent rapidement l’ensemble de la musculature. Le pronostic est sévère, avec perte de la marche durant l’enfance et altération de l’autonomie respiratoire à l’adolescence. L’étude d’une grande famille Malienne consanguine nous a permis de localiser le gène responsable de la maladie dans un intervalle de 3.9 cM sur le chromosome 1, en 1p36 (Neurology. 2006;67(1):120-4). Nous avons ensuite mis en évidence une mutation faux-sens à l’état homozygote dans le gène PLEKHG5 chez les patients atteints et avons prouvé le caractère pathogène de cette mutation grâce à une série d’études fonctionnelles. Nous avons montré que la protéine PLEKHG5 sauvage a une distribution cytoplasmique homogène dans des cellules rénales (HEK293) et mammaires (MCF10A) humaines transfectées et qu’elle y active la voie de signalisation NF-kappaB. La protéine PLEKHG5 mutée est quant à elle instable, ce qui entraîne une perte de sa fonction activatrice sur NF-kappaB. De plus, grâce à des études de transfection transitoire de motoneurones murins (cellules NSC34), nous avons montré que la protéine PLEKHG5 mutée entraîne la formation d’importants agrégats cytoplasmiques. Dans cette nouvelle forme d’amyotrophie spinale, la perte de la fonction activatrice de la voie de signalisation NF-kappaB et la formation d’agrégats pourraient toutes deux contribuer à la neurotoxicité de la protéine PLEKHG5 mutée et conduire ainsi à la dégénérescence des motoneurones (Am J Hum Genet. 2007;81(1):67-76). En conclusion, nous espérons que ces résultats, qui contribuent à améliorer la connaissance des mécanismes physiopathologiques responsables de la dégénérescence des motoneurones, ouvriront à l’avenir la voie vers de nouvelles perspectives thérapeutiques.

The importance of touch-related sensations as a kinesthetic perceptual system through the observation of the subject/object phenomenon is explored through defining aspects of movement learning experiences associated with dance training.

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