Thesis (Ph. D.)--University of Missouri-Columbia, 2007. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Vita. "December 2007" Includes bibliographical references.

Thesis (Ph. D.)--University of Washington, 2006. / Vita. Includes bibliographical references (leaves 121-147).

As doenças neuromusculares são um grupo heterogêneo de doenças genéticas, causadas por mutações nos genes codificando proteínas musculares sarcolemicas, sarcoméricas, e citosolicas. A Distrofia Muscular de Duchenne (DMD) é uma miopatia degenerativa progressiva, caracterizada pela ausência da proteína distrofina na superfície da membrana da célula muscular. O modelo de cão Golden Retriever Muscular Dystrophy (GRMD) apresenta semelhanças clínicas de DMD devido ao seu tamanho maior e significativa fraqueza muscular, é geneticamente homólogo a DMD humana, sendo considerado modelo experimental para estudos de novas propostas terapêuticas e melhor entendimento da fisiopatogenia da doença. Não existe até o momento uma terapia efetiva em bloquear ou reverter o processo da distrofia muscular. Embora a terapia gênica e o transplante de células tronco possam fornecer a cura para a DMD, resultados positivos podem demorar algum tempo até serem clinicamente viáveis. Neste sentido, a busca de informações fisiopatológicas que podem estar correlacionadas com a distrofia, e, com o avanço da pesquisa há possibilidade de melhora na condição vital do paciente, por retardo da progressão dos sinais clínicos ou cura. Existem poucos estudos endócrinos em animais portadores de distrofia como aves, GRMD e o mdx, assim como no homem. Mediante a falta de dados, houve a necessidade de quantificar e comparar hormônios, ainda não analisados, assim como avaliar a morfologia de glândulas endócrinas no modelo experimental GRMD. Para que fosse possível a correlação na interferência hormonal na evolução da Distrofia Muscular do cão, os exames sanguíneos foram comparados com cães normais e com as portadoras, todos da raça Golden Retriever. A mensuração hormonal de triiodotironina total (T3T), tiroxina total (T4T), a tireotropina (TSH) e o cortisol foram processados através de \"kits\" comerciais para radioimunoensaio, e o tiroxina livre (T4L) com \"kit\" comercial por diálise. As análises morfológicas das adrenais e da tireóide foram feitas através da macroscopia, microscopia de luz e microscopia eletrônica de transmissão de materiais de GRMD e de cães sadios. Anatomica e morfológicamente as glândulas não apresentaram alterações. Os níveis de cortisol não variaram significantemente entre os grupos estudados. Os níveis de T3 total foi semelhante para os animais sadios, portadoras e afetados. T4 total apresentou-se em diferentes níveis em alguns grupos. O T4 livre não variou significantemente entre os grupos estudados. Os níveis séricos de TSH da maioria dos Golden Retriever, afetados, portadores e sadios, apresentaram-se abaixo do limite apresentado pelos valores de referência. / Neuromuscular diseases are a heterogeneous group of genetic diseases caused by mutations in genes encoding proteins muscle sarcolemma, sarcomeric, and cytosol. Duchenne Muscular Dystrophy (DMD) is a progressive degenerative myopathy characterized by absence of dystrophin on the surface membrane of muscle cells. The Golden Retriever Muscular Dystrophy (GRMD) shows clinical similarities of DMD due to its size and significant muscle weakness, is genetically homologous to human DMD, and is considered an experimental model for studies of new therapeutic approaches and better understanding of the pathogenesis of the disease.There is not an effective therapy to block or reverse the process of muscular dystrophy yet. Although gene therapy and stem cell transplantation may provide a cure for DMD, positive results may take some time to be clinically viable.In this sense, the search for pathologic and physiologic information can be correlated with muscular dystrophy, and with the advancement of research there is room for improvement in the vital condition of the patient by delaying the progression of clinical signs or cure.There are a few studies in animals with endocrine dystrophy such as birds, and GRMD mdx as well as in man. By lack of data, there was a need to quantify and compare hormones, not yet analyzed, as well as evaluating the morphology of endocrine glands in experimental model GRMD. To make possible the hormonal correlation in the evolution of muscular dystrophy in dogs, blood tests were compared with normal dogs and carriers, all Golden Retrievers. Measurement of total triiodothyronine hormone (T3T), total thyroxine (T4T), the thyrotropin (TSH) and cortisol were processed using commercial kits for radioimmunoassay, and free thyroxine (FT4) with commercial kit by dialysis.The morphological analysis of adrenal and thyroid were made by macroscopic, light microscopy and transmission electron microscopy of materials GRMD and healthy dogs. Anatomical and morphological glands were unaffected. Cortisol levels did not differ significantly between groups. The levels of total T3 was similar to the healthy animals, carriers and affected. The T4 presented at different levels in some groups. The free T4 did not differ significantly between groups. Serum TSH of most Golden Retriever, affected patients and healthy individuals, were below the limit presented by the reference values.

Muscular dystrophy is a genetic disease which affects the morphology, physiology and biochemical nature of the muscle fiber. This study was designed to examine the progressive effects of muscular dystrophy on the differentiation process of skeletal muscle. Chapter 1 examines the neonatal development of muscle spindles and their intrafusal fibers in the soleus and extensor digitorum longus (EDL) of genetically dystrophic mice according to histochemical, quantitative, and ultrastructural parameters. Despite alterations in the surrounding extrafusal fibers, muscle spindles and their intrafusal fibers appeared enzymatically and histologically unaffected in incipient stages of murine dystrophy. In the second chapter the distribution and concentration of parvalbumin (PV), a calcium-binding protein, in 32 and 2-week-old dystrophic mice was mapped by immunohistochemical and biochemical procedures. The number of parvalbumin-immunoreactive fibers was significantly reduced in the adult dystrophic EDL but slightly increased in the adult dystrophic soleus. No differences between strains were observed in the 2-week samples. These findings were supported by routine myosin ATPase histochemistry. Parvalbumin was isolated on SDS-PAGE gels and the concentration of PV was estimated by a RIA. These results confirmed the immunohistochemical data in that PV content was dramatically reduced in the adult dystrophic EDL and significantly increased in the dystrophic soleus. No changes were detected in the samples of the 2-week-old muscles. The similarity in the distribution and content of PV between the fast and slow dystrophic muscles at 32 weeks of age suggests an alteration in the distribution and phenotypic expression of fiber types in muscular dystrophy and supports the hypothesis that dystrophy alters the normal differentiation process of skeletal muscle. / Medicine, Faculty of / Cellular and Physiological Sciences, Department of / Graduate

Introduction/Aims Duchenne and Becker muscular dystrophies (DBMD) are X-linked neuromuscular disorders characterized by progressive muscle weakness, leading to decreased mobility and multisystem complications. We estimate productivity costs attributable to time spent by a parent caring for a male child under the age of 18 y with DBMD, with particular focus on female caregivers of boys with Duchenne muscular dystrophy (DMD) who have already lost ambulation. Methods Primary caregivers of males with DBMD in the Muscular Dystrophy Surveillance and Research Tracking Network (MD STARnet) were surveyed during 2011–2012 on family quality of life measures, including labor market outcomes. Of 211 respondents, 96 female caregivers of boys with DBMD were matched on state, year of survey, respondent's age, child's age, and number of minor children with controls constructed from Current Population Survey extracts. Regression analysis was used to estimate labor market outcomes and productivity costs. Results Caregivers of boys with DBMD worked 296 h less per year on average than caregivers of unaffected children, translating to a $8816 earnings loss in 2020 U.S. dollars. Caregivers of boys with DMD with ≥4 y of ambulation loss had a predicted loss in annualized earnings of $23,995, whereas caregivers of boys with DBMD of the same ages who remained ambulatory had no loss of earnings. Discussion Female caregivers of non-ambulatory boys with DMD face additional household budget constraints through income loss. Failure to include informal care costs in economic studies could understate the societal cost-effectiveness of strategies for managing DMD that might prolong ambulation.

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No description available.

Skeletal muscle regeneration is a multifaceted process requiring the spatial and temporal coordination of myogenesis as well as angiogenesis. While these processes are often studied independently, recent evidence from our lab has shown that the resident adult stem cell population within skeletal muscle, called satellite cells, begins secreting soluble growth factors likely to contribute to the proangiogenic response. The overall aim of this study is to investigate the role of pro-angiogenic factors secreted by satellite cells during skeletal muscle regeneration. Results from the study indicate that Hepatocyte Growth Factor (HGF) is a critical protein for the proangiogenic effect of satellite cells. It was also shown that in hypoxic environments, such as those seen in an injury state, it appears that satellite cells decrease their proangiogenic effect if oxygen levels fall below a threshold level. This decrease in pro-angiogenic effect in the hypoxic environment appears to be due to the decrease in HGF expression and protein secretion and is not compensated for by the increase in Vascular Endothelial Growth Factor secretion also seen in the hypoxic response. Furthermore, the regulation of HGF in these hypoxic conditions appears to be in part due to increased levels of hypoxia inducible factor, which are acting on the hypoxia response element site found on the HGF promoter. In the last set of experiments, this injury response was further investigated as the effect of satellite cell mediated angiogenesis was examined in the disease state of muscular dystrophy. Here, we also observed a reduction in angiogenesis from media conditioned by satellite cells from dystrophic muscle compared to healthy muscle. Overall, this study further strengthens the case for satellite cells as important mediators of the angiogenic response in regenerating muscle and may serve as a potential site for therapeutic intervention in the future.

Duchenne muscular dystrophy (DMD) is an X-linked recessive disease caused, in most cases, by the complete absence of the 427 kD cytoskeletal protein, dystrophin. Without dystrophin, the dystrophin-associated protein complex (DAPC) does not form and the plasma membrane is destabilised. There is no effective treatment and affected individuals die from respiratory failure and cardiomyopathy by age 30. This thesis describes experiments in which in vivo cardiac function was measured using non-invasive magnetic resonance imaging in a number of mouse models relevant to muscular dystrophy. As syncoilin forms a link from the DAPC to the cytoskeleton, it was postulated in Chapter 3 that the syncoilin knockout mouse would have cardiac defects similar to those caused by the loss of dystrophin. However, the loss of syncoilin did not alter the protein levels of its binding partners, measured by western blotting, and caused no defect in heart function or structure, measured using histological staining. Similarly, in Chapter 4, a mouse with a mutation in the transient receptor potential channel canonical type 3 (TRPC3), a receptor/stretch-activated cation channel thought to be involved in the pathogenesis of DMD, was found to have no functional or morphological cardiac defect. In the mdx mouse, a mouse model of DMD that lacks dystrophin, cardiomyopathy was prevented by either increasing levels of the dystrophin related protein, utrophin, or of dystrophin, in the diaphragm, which thereby restored diaphragm function. In Chapter 5 it was found that in a transgenic mdx mouse in which utrophin was over-expressed in skeletal muscle and diaphragm, but not in the heart, cardiac function was restored to wild-type levels. However, histologically the transgenic heart showed more fibrosis and immune cell infiltration than that of untreated mdx controls. In Chapter 6 it was found that in mdx mice treated with a peptide-conjugated phosphorodiamidate morpholino oligomer (PPMO), that resulted in high dystrophin restoration in skeletal muscle and diaphragm only, cardiac function was also restored to wild-type levels. In Chapter 6 it was also found that in dystrophin/utrophin-deficient double-knockout (dKO) mice, a more severely affected animal model of DMD, treatment with a PPMO again produced high levels of dystrophin only in skeletal muscle and diaphragm, and once more restored cardiac function to wild-type levels. In the dKO mouse, there was no difference in heart function between treatment of the diaphragm plus the heart and treatment of the diaphragm alone. Restoration of diaphragm and other respiratory muscle function, irrespective of the method used, was sufficient to prevent cardiomyopathy in dystrophic mice. The novel mechanism of treating respiratory muscles to prevent cardiomyopathy in dystrophic mice has implications for the study of heart function in the current DMD mouse models and suggests a new approach to treatment.

Introdução: A Distrofia Muscular de Duchenne (DMD) é uma doença genética que causa limitações físicas e motoras progressivas, além de alterações da função pulmonar em fase mais tardia. A escala da medida da função motora (MFM) e do teste de caminhada de 6 minutos (TC6) são ferramentas confiáveis e precisas para avaliar pacientes com DMD. A capacidade de realização desses testes pode ser influenciada pela condição cardiorrespiratória, resistência e força muscular, mas ainda não está bem definido qual, ou quais, destas variáveis podem interferir de forma mais significativa na função motora e na capacidade funcional de exercício. Objetivo: Avaliar a função motora por meio da MFM e correlacionar com a função pulmonar e capacidade funcional de exercício em pacientes com DMD. Métodos: Trata-se de um estudo transversal que envolveu 61 voluntários com DMD submetidos a um protocolo de avaliação composto por informações pessoais, antropometria, escala MFM, espirometria (CVF,VEF1,VEF1/CVF, FEF25-75, PFE, CVL, VVM), ventilometria (VC, VM e CVL), pico de fluxo expiratório (PFE) e pico de fluxo de tosse (PFT) no medidor portátil, pressão inspiratória máxima (PImax) e pressão expiratória máxima (PEmax), pressão inspiratória nasal (SNIP), oscilometria de impulso (R5, R20, R5-R20, X5) e TC6. Foi realizada a análise de correlação entre as variáveis e posteriormente a comparação entre os grupos de voluntários deambuladores e não deambuladores. Resultados: A média de idade dos voluntários foi de 13,70±3,93. Na espirometria, houve correlação positiva do escore total, domínios 1(D1) e 2 (D2) da MFM com a porcentagem do previsto da CVF% (r=0,58, r=0,51, r=0,57, respectivamente), correlação negativa do escore total, domínios 1, 2 e 3 com VEF1/CVF (r=-0,61, r=-0,57, r=-0,49 e r=-0,49, respectivamente), correlação negativa do escore total, domínio 1 e domínio 3 com o FEF25-75% (r=-0,48, -0,47, -0,42, respectivamente). Na manobra de PFE realizada no medidor portátil, houve correlação do PFE% com o escore total e todos os domínios da MFM. O escore total, domínio 1, 2 e 3 se correlacionaram positivamente com os valores percentuais da PEmax (r=0,67, r=0,60, r=0,63 e 0,42, respectivamente) e SNIP (r=0,56, r=0,42, r=0,64 e r=0,45, respectivamente). Na oscilometria de impulso (IOS), houve correlação da resistência total (R5) com o escore total, domínio 1 e 2 (r=0,55; r=0,52 e r=0,50, respectivamente) e resistência central (R20) com o escore total, domínios 1 e 2 (r=0,52; r=0,51 e r=0,45, respectivamente); correlação da resistência periférica (R5-R20) com o escore total (r = 0,46) e domínio 2 (r=0,49), e da reatância (X5) com o escore total, D1 e D2(r=-0,43; r=- 0,40 e r=-0,36, respectivamente). No TC6, a distância em metros foi de 294,75±96,97 e se correlacionou fortemente com a MFM, e a percepção da dispneia relatada no TC6 se correlacionou negativamente com o domínio 3 daMFM (r=-0,75). Na análise comparativa entre os grupos de deambuladores (D) e não deambuladores (ND), o IOS demonstrou valores obtidos significativamente menores no grupo ND de R5, R20, R5-R20, X5 e da porcentagem do previsto de R5-20. No mesmo grupo (ND), a espirometria demonstrou valores obtidos significativamente maiores de VVM, da VEF1/CVF, FEF25-75 e porcentagem do previsto de VEF1/CVF, além da diminuição da porcentagem do previsto da CVF, PFE no medidor portátil, PEmax, SNIP e MFM (escore total e domínios 1,2 e 3). Conclusão: A diminuição da MFM se correlacionou com a diminuição da resistência e reatância das vias aéreas, com a diminuição da PEmax, SNIP e da CVF. Além desses parâmetros, houve correlação da diminuição da MFM com o aumento dos valores de PFE, FEF25-75 e VEF1/CVF. Na comparação entre os grupos, os resultados confirmam o que foi encontrado nas correlações no grupo de pacientes que não deambulavam, com diminuição das resistências e reatância das vias aéreas, da força muscular respiratória e valores maiores de volume (VEF1/CVF) e fluxo pulmonar (FEF25-75). Apesar da forte correlação entre a distância percorrida no teste de caminhada com a função motora, não houve alteração dos dados vitais, o que pode indicar que os pacientes não atingiram o esforço submáximo estimado / Introduction: Duchenne Muscular Dystrophy (DMD) is a genetic disease which causes progressive physical and motor limitations, including alterations to the pulmonary function in later stages. The scale of the motor function measure (MFM) and of the six minute walking test (TC6) are reliable and precise tools to assess DMD patients. The possibility of performing these tests may be influenced both by the patient\'s cardiopulmonary condition, resistance and muscular strength. However, it has not been yet demonstrated which of these variables can have the most significant impact on the patient\'s motor function and functional capacity of exercise. Objective: To assess the motor function though the application of the MFM scale and correlate it with the pulmonary function and functional capacity of exercise on DMD patients. Methodology: The approach adopted in this study consisted of a transversal analysis involving sixty one DMD patients who have volunteered to submit themselves to an evaluation protocol which includes the following parameters: personal information, anthropometry, MFM scale, spirometry (CVF,VEF1,VEF1/CVF, FEF25-75, PFE, CVL, VVM), ventilometry (VC, VM e CVL), peak expiratory flow (PFE) and peak cough flow (PFT) according to the portable meter measurements, maximum inspiratory pressure (PImax), maximum expiratory pressure (PEmax), nasal inspiratory pressure (SNIP), impulse oscillometry (R5, R20, R5-R20, X5) and TC6. The study focused on the analysis of the correlation between variables, followed by a comparison between two groups of patients: wanderers and non-wanderers. Results: The average age of patients was 13,70±3,93. On spirometry, there was a positive correlation between the total score, domains 1(D1) and 2 (D2) of the MFM and the expected CVF percentage (r=0,58, r=0,51, r=0,57, respectively); there was a negative correlation between the total score, domains 1, 2 and 3 and VEF1/CVF (r=-0,61, r=-0,57, r=-0,49 and r=-0,49, respectively), as well as a negative correlation between domain 1, domain 3 and FEF25-75% (r=-0,48, -0,47, -0,42, respectively). On the PFE maneuver performed using the portable meter, there was a correlation between the expected PFE percentage, the total score and all MFM domains. The total score and domains 1, 2 and 3 had a positive correlation with the percentage values of PEmax (r=0,67, r=0,60, r=0,63 and 0,42, respectively) and SNIP (r=0,56, r=0,42, r=0,64 and r=0,45, respectively). Regarding the impulse oscillometry system (IOS), there was a correlation between the total resistance (R5), the total score and domains 1 and 2 (r=0,55; r=0,52 and r=0,50, respectively); and between the central resistance (R20) and total score, domains 1 and 2 (r=0,52; r=0,51 and r=0,45, respectively); in addition, there were correlations between the peripheral resistance (R5-R20), the total score (r = 0,46) and domain 2 (r=0,49), as well as between the reactance (X5) and total score, D1 and D2(r=-0,43; r=-0,40 and r=-0,36, respectively). Thedistance in meters reached in the TC6 was of 294,75±96,97 and had a strong correlation with the MFM. The perception of dyspnoea reported through the TC6 had a negative correlation with domain 3 of the MFM (r=-0,75). On the comparative analysis between groups of wanderers (D) and non-wanderers (ND), the IOS has shown significantly lower values in the group of ND (R5, R20, R5- R20, X5), as well as on the expected percentage of R5-20. In the same group (ND), the spirometry has shown significantly higher values of VVM, VEF1/CVF, FEF25-75 and the expected percentage of VEF1/CVF; in addition to a reduction of the expected percentage of CVF and PFE on the portable meter, PEmax, SNIP e MFM (total score and domains 1, 2 and 3). Conclusion: The reduction of the MFM correlates with the reduction of airways resistance and reactance, and with the reduction of PEmax, SNIP and CVF. In addition to these parameters, there was a correlation between the MFM reduction and the increase of the PFE, FEF25- 75 e VEF1/CVF values. In the comparison between groups, the results confirm the findings in the correlations of the non-wanderers group, with reduction of the airways resistance and reactance, of the respiratory muscular strength and higher volume values (VEF1/CVF) and pulmonary flow (FEF25-75). Despite the strong correlation between the distance covered during the walking test and the motor function, there was no alteration on vital indicators, which may point out to the fact that the patients did not achieve the expected sub-maximum effort level