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1
Huetter, Robert A. "A History of Fort Duchesne, Utah, and the Role of its First Commanding Officer, Frederick W. Benteen." Diss., CLICK HERE for online access, 1990. http://patriot.lib.byu.edu/u?/MTGM,14001.
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Koppaka, Sisir. "Imaging biomarkers for Duchenne muscular dystrophy." Thesis, Massachusetts Institute of Technology, 2015. http://hdl.handle.net/1721.1/106959.
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Thesis: S.M., Massachusetts Institute of Technology, School of Engineering, Center for Computational Engineering, Computation for Design and Optimization Program, 2015.Cataloged from PDF version of thesis.
Includes bibliographical references (pages 75-78).
Duchenne muscular dystrophy (DMD) is the most common muscular dystrophy of childhood and affects 1 in 3600 male births. The disease is caused by mutations in the dystrophin gene leading to progressive muscle weakness which ultimately results in death due to respiratory and cardiac failure. Accurate, practical, and painless tests to diagnose DMD and measure disease progression are needed in order to test the effectiveness of new therapies. Current clinical outcome measures such as the sixminute walk test and North Star Ambulatory Assessment (NSAA) can be subjective and limited by the patient's degree of effort and cannot be accurately performed in the very young or severely affected older patients. We propose the use of image-based biomarkers with suitable machine learning algorithms instead. We find that force-controlled (precise acquisition at a certain force) and force-correlated (acquisition over a force sweep) ultrasound helps to reduce variability in the imaging process. We show that there is a high degree of inter-operator and intra-operator reliability with this integrated hardware-software setup. We also discuss how other imaging biomarkers, segmentation algorithms to target specific subregions, and better machine learning techniques may provide a boost to the performance reported. Optimizing the ultrasound image acquisition process by maximizing the peak discriminatory power of the images vis-à-vis force applied at the contact force is also discussed. The techniques presented here have the potential for providing a reliable and non-invasive method to discriminate, and eventually track the progression of DMD in patients.
by Sisir Koppaka.
S.M.
3
Rabinowitz, Adam Howard. "Antisense therapies for Duchenne muscular dystrophy." Thesis, Imperial College London, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.444590.
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Wakefield, Philip M. "Gene therapy for duchenne muscular dystrophy." Thesis, University of Oxford, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.365743.
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Taktak, Diane M. "A lightweight modular knee-ankle-foot orthosis for Duchenne muscular dystrophy." Thesis, University of Salford, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.261992.
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Al, Majathoub Mohannad. "Development of cryopreservation techniques for strawberry ((Fragaria x ananassa Duchesne)." Thesis, University of Derby, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.427603.
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Dunant, Patrick. "Strategies for Molecular Therapy of Duchenne Muscular Dystrophy." Diss., lmu, 2003. http://nbn-resolving.de/urn:nbn:de:bvb:19-12429.
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Buser, Karen N. Kamiri. "Parental Attitudes Regarding Newborn Screening for Duchenne Muscular Dystrophy." Case Western Reserve University School of Graduate Studies / OhioLINK, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=case1307627473.
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Jara, Peña Enoc Efer. "Evaluación de soluciones hidropónicas para la producción de "fresa" Fragaria x ananassa Duchesne cv. Chandler." Bachelor's thesis, Universidad Nacional Mayor de San Marcos, 1999. https://hdl.handle.net/20.500.12672/6270.
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El documento digital no refiere asesorLa fresa (Fragaria x ananassa Duchesne) es un cultivo de importancia económica que presenta serios problemas de enfermedades con lo que la calidad sanitaria y comercial del fruto baja e implica fuertes pérdidas económicas en su producción. Esta situación puede ser mejorada con plantas que presenten un buen desarrollo, bajo sistemas que mejoren su calidad sanitaria y productividad. Con esta finalidad en el presente trabajo se evaluó a 2 formulaciones de soluciones nutritivas durante la etapa vegetativa y 3 formulaciones durante la etapa fructificación de plantas de la variedad Chandler obtenidas a partir de cultivo in vitro, realizándose el experimento bajo condiciones de "invernadero" usando un sistema hidropónico en grava muy fina en Carabayllo al norte de Lima durante los meses de Diciembre de 1996 a Agosto de 1997. Durante el periodo de cultivo la temperatura fluctuó entre 14.1 °C y 25.6 °C. Se realizaron muestreos destructivos a los 60, 90, 160, 190 y 220 días después de iniciado el tratamiento (ddt) para evaluar crecimiento, desarrollo y análisis químico de los órganos de la planta; adicionalmente en la etapa reproductiva de la planta se evaluaron número de flores, número, peso y contenido de azúcares reductores en los frutos. No se encontró diferencias significativas entre los tratamientos en la etapa vegetativa pero sí en la etapa reproductiva al evaluar la altura de la planta, número de flores, porcentaje de azúcares reductores de los frutos, número y peso de los frutos en la cosecha Tampoco se encontró diferencias significativas en el porcentaje de Nitrógeno, Fósforo, Potasio, Calcio, Magnesio y Hierro. Combinaciones de 200:40:300 ppm de N:P:K en la etapa reproductiva favorecieron el mayor rendimiento en la planta.
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Jara, Peña Enoc Efer. "Evaluación de soluciones hidropónicas para la producción de "fresa" Fragaria x ananassa Duchesne cv. Chandler." Bachelor's thesis, Universidad Nacional Mayor de San Marcos, 1999. http://cybertesis.unmsm.edu.pe/handle/cybertesis/6270.
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La fresa (Fragaria x ananassa Duchesne) es un cultivo de importancia económica que presenta serios problemas de enfermedades con lo que la calidad sanitaria y comercial del fruto baja e implica fuertes pérdidas económicas en su producción. Esta situación puede ser mejorada con plantas que presenten un buen desarrollo, bajo sistemas que mejoren su calidad sanitaria y productividad. Con esta finalidad en el presente trabajo se evaluó a 2 formulaciones de soluciones nutritivas durante la etapa vegetativa y 3 formulaciones durante la etapa fructificación de plantas de la variedad Chandler obtenidas a partir de cultivo in vitro, realizándose el experimento bajo condiciones de "invernadero" usando un sistema hidropónico en grava muy fina en Carabayllo al norte de Lima durante los meses de Diciembre de 1996 a Agosto de 1997. Durante el periodo de cultivo la temperatura fluctuó entre 14.1 °C y 25.6 °C. Se realizaron muestreos destructivos a los 60, 90, 160, 190 y 220 días después de iniciado el tratamiento (ddt) para evaluar crecimiento, desarrollo y análisis químico de los órganos de la planta; adicionalmente en la etapa reproductiva de la planta se evaluaron número de flores, número, peso y contenido de azúcares reductores en los frutos. No se encontró diferencias significativas entre los tratamientos en la etapa vegetativa pero sí en la etapa reproductiva al evaluar la altura de la planta, número de flores, porcentaje de azúcares reductores de los frutos, número y peso de los frutos en la cosecha Tampoco se encontró diferencias significativas en el porcentaje de Nitrógeno, Fósforo, Potasio, Calcio, Magnesio y Hierro. Combinaciones de 200:40:300 ppm de N:P:K en la etapa reproductiva favorecieron el mayor rendimiento en la planta.Tesis
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Thomas, Karen. "The mdx mouse as a model for Duchenne muscular dystrophy." Thesis, University of Cambridge, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.386990.
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Wells, Kim Elizabeth. "Optimisation of constructs for gene therapy of Duchenne muscular dystrophy." Thesis, Imperial College London, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.392669.
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Souza, Mariana Angélica de. "Efeito do uso da ankle-foot orthosis na biomecânica da marcha de pacientes com Distrofia Muscular de Duchenne." Universidade de São Paulo, 2014. http://www.teses.usp.br/teses/disponiveis/17/17152/tde-21012015-092933/.
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O objetivo deste estudo foi avaliar o efeito do uso noturno ou diurno da ankle-foot orthosis (AFO) na biomecânica da marcha de pacientes com DMD. Foram avaliados 20 pacientes deambuladores, do Ambulatório de Miopatias Infantis do CER do HCFMRP-USP, com diagnóstico de distrofia muscular de Duchenne (DMD), com idades entre 4 e 12 anos. Foi realizada a avaliação inicial (Av1) em todos os pacientes e, 7 pacientes foram reavaliados após 6 meses (Av2). Na Av1, os pacientes foram agrupados conforme o uso da órtese: grupo sem órtese (SO; n=7), grupo órtese noturna (ON; n=7), grupo órtese diurna (OD; n=6). Na Av1 e na Av2 foram obtidos dados de massa corporal, altura, composição corporal pela bioimpedância elétrica, escore funcional pela escala medida da função motora, amplitude passiva de movimento articular, força muscular isométrica pelo dinamômetro Handheld e avaliação biomecânica da marcha, na velocidade habitual do paciente. Os pacientes que faziam uso da órtese diurna foram avaliados sem e com órtese, sendo denominados grupos ODs e ODc, respectivamente. Os dados foram analisados de três formas: duas transversais e uma longitudinal. Nas análises transversais, foram realizados dois procedimentos: (i) comparando dados dos grupos SO x ON x ODs; (ii) comparando dados dos grupos SO x ON x ODc. Nestas, foi utilizado o teste ANOVA, considerando um nível de significância de 5%. Na análise longitudinal, foi realizada a análise descritiva comparando os dados obtidos na Av1 e Av2, individualmente para os 7 pacientes reavaliados. Transversalmente, o grupo ODc apresentou maiores picos do ângulo de dorsiflexão e do momento dorsiflexor, menor ângulo de flexão plantar e menor geração de potência de tornozelo (p<0,05) que o grupo SO. Porém, ao caminhar sem a AFO (grupo ODs) estes resultados não foram observados (p>0,05). Em relação ao grupo ON, o grupo ODc obteve menores picos do ângulo de flexão do quadril, de absorção de potência de quadril, do ângulo de flexão plantar e maior pico do momento dorsiflexor (p<0,05), sendo que ao retirar a AFO (ODs) essas diferenças não foram observadas (p>0,05). E ainda, o grupo ON obteve maior pico do ângulo de flexão do joelho e menor momento flexor de quadril (p<0,05) em relação ao grupo ON. Na comparação dos dados entre os grupos SO e ON, o grupo ON obteve maior pico do ângulo de flexão do joelho e maior absorção de potência de quadril (p<0,05). Na análise longitudinal individual foi observado que os 2 pacientes que iniciaram precocemente e mantiveram o uso noturno da AFO apresentaram na Av2 maior velocidade da marcha, maiores momentos extensor de quadril e flexor plantar e maior geração de potência de tornozelo, contrariamente aos paciente que interromperam o uso (noturno ou diurno) da AFO. Conclui-se que o uso diurno da AFO acarretou alterações positivas na biomecânica da marcha, minimizando compensações típicas da DMD na articulação do tornozelo. O uso noturno da AFO, quando iniciado precocemente, também afetou positivamente a marcha dos pacientes. Assim, sugere-se o início precoce e contínuo do uso diurno e noturno da AFO aos pacientes com DMD.The aim of this study was to evaluate the effect of the ankle-foot orthosis (AFO) during nocturnal or daytime usage of the gait biomechanics in patients with Duchenne Muscular Dystrophy (DMD). Twenty ambulant patients from the Myopathies Infant Ambulatory of CER - HCFMRP-USP, were diagnosed with DMD between the ages of 4 and13 years and were evaluated. The initial evaluation (Ev1) was performed in all patients, and 7 patients were reevaluated after 6 months (Ev2). In Av1, patients were grouped according to orthosis use: group without orthosis (NoO, n = 7), group with nocturnal orthosis (NiO, n = 7), group with daytime orthosis (DO, n = 6). In Ev1 and Ev2 data were obtained according to the weight, height, body composition (bioelectrical impedance), functional score (Measure scale of motor function), passive joint range of motion, isometric muscle strength (dynamometer Handheld) and biomechanical gait analyses (usual velocity for the patient). Patients who used the daytime orthosis were evaluated with and without bracing, respectively. The data were analyzed in three ways; the first two were cross-sectional and the other one was longitudinal. In the cross-sectional analyzes, an exploratory analysis of the data from each evaluation was performed, and subsequently, the variables were compared between groups, considering the means and standard deviations. ANOVA test was used, and it was considered a significant level of 5%. In the longitudinal analysis, the description of the data obtained in the evaluation 1 compared to the data obtained in the evaluation 2 was individually performed in the 7 patients who were reevaluated. A cross-sectional analysis compared the data between NoO x NiO x DO groups considering the gait analysis data from the DO group without the orthosis (barefoot), being named DOno. The other cross-sectional analysis compared the data between NoO x NiO x DO groups considering the gait analysis data from the OD group with orthosis, being named DOwith. In individual longitudinal analysis, it was observed that patients who had started early and kept the nocturnal usage of AFO which has been already showed, in six months, an increment of gait velocity, hip extensor and plantar flexor moments and also the increment of ankle power generation, which is the opposite of the patient who has discontinued the AFO usage (daytime or nocturnal). In the cross-sectional analyzes it was observed that, compared to the NoO group, the DOwith group had a higher dorsiflexion angle peak and higher dorsiflexor moment peak (p<0.05). However, when they walked without the device these results were not maintained. There was no difference (p>0.05) between DOno and NoO groups for the kinematic parameters. And, the DOno group had lower plantar flexor moment maximum peak than the SO group (p>0.05). It was concluded that AFO daytime use cause positive changes in gait biomechanics, minimizing typical compensation of DMD in the ankle joint. The night use of AFO, when started early, also positively affected the gait of patients. Thus, it is suggested early prescription of daytime and nocturnal usage of AFO for DMD patients.
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Abmayr, Simone. "Gene therapy for muscular dystrophy using secondary modifiers of the dystrophic phenotype." [S.l.] : [s.n.], 2005. http://deposit.ddb.de/cgi-bin/dokserv?idn=973452595.
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Burt, Matthew. "Resveratrol as a Novel Therapeutic Agent for Treating Duchenne Muscular Dystrophy." Thèse, Université d'Ottawa / University of Ottawa, 2013. http://hdl.handle.net/10393/26273.
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Duchenne Muscular Dystrophy (DMD) is an x-linked neuromuscular disease that is caused by an absence of dystrophin protein, rendering skeletal muscle more susceptible to contraction-induced damage. One therapeutic strategy focuses on increasing the expression of endogenous utrophin A, a dystrophin homologue. Interestingly, slow muscle is more resistant to the dystrophic pathology and has increased utrophin A expression (Webster 1998; Gramolini 2001b). These observations led researchers to explore the therapeutic potential of stimulating the slow, oxidative myogenic program (SOMP) in the mdx context. Beneficial adaptations were seen with pharmacological activation of PPARδ and AMPK. We treated mdx mice with resveratrol (~100mg/kg/day), a putative SIRT1 activator, for 6-7 weeks and evaluated the activity of phenotypic modifiers that are known to influence the SOMP. SIRT1 activity and protein levels increased significantly, as well as downstream PGC-1α activity. There was evidence of a fibre type conversion as the treated mice had a higher proportion of the slow myosin heavy chain isoforms in both the EDL and Soleus skeletal muscles. Utrophin A protein levels showed modest, but consistent increases with resveratrol treatment. Finally, histological analysis revealed improvements in central nucleation and fibre size variability. These findings were promising, but raised the question of whether modifying the treatment regimen may result in greater therapeutic benefits. Surprisingly, we discovered that an elevated dose of 500mg/kg/day was ineffective in its promotion of the SOMP. SIRT1 was not activated and there was no change in utrophin A levels with resveratrol treatment. Taken together, this study demonstrates that resveratrol has the ability to promote the SOMP through SIRT1 and PGC-1α activation. It also highlights the importance of selecting an appropriate dose of resveratrol to maximize its effectiveness.
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Chadwick, Jessica Ann. "Mineralocorticoid Receptors: A Novel Therapeutic Target for Treating Duchenne Muscular Dystrophy." The Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1468946734.
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Sharma, Dishant. "Development of tolerogenic plasmid vectors for gene therapy of Duchenne muscular dystrophy (DMD)." Thesis, University of Portsmouth, 2017. https://researchportal.port.ac.uk/portal/en/theses/development-of-tolerogenic-plasmid-vectors-for-gene-therapy-of-duchenne-muscular-dystrophy-dmd(55b88eaa-5f23-4ae6-83e7-baed45f82d00).html.
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This project focused on the development of an effective gene replacement therapy for the Duchenne muscular dystrophy (DMD) in its mouse models [(X-linked muscular dystrophy, mdx) and mdx-βgeo mice]. Earlier studies on the DMD replacement therapy (usually using mini-dystrophin) were largely not successful because dystrophin is being recognised as an antigen upon re-expression in dystrophic muscles and initiates the specific immune response. This leads to a short-lived or no expression of mini-dystrophin as was found in both human clinical trials and in animal models. It had been shown that the presence of pre-existing Tcells responding to dystrophin was responsible for this effect and immunosuppressive drug treatments in a canine model of DMD (cxmd) resulted in a stable expression of dystrophin for 2 years. This study investigated the potential of using immunomodulatory factors such as IDO (Indoleamine 2,3-dioxygenase) and TDO (tryptophan 2,3-dioxygenase) to prolong expression of newly synthesised antigenic products in dystrophic mice. IDO and TDO are the rate-limiting enzymes of the tryptophan catabolism pathway, which regulate the production of kynurenines. These enzymes are known to increase the survival of grafts in transplantation by targeting dendritic cells, which play an important role in the T-cell activation. The plasmid with the general CMV promoter was used for expression of these enzymes in cell lines (HEK 293 cells and SC5 dystrophic myoblasts) and in skeletal muscles in vivo. To achieve targeting of the immunomodulatory constructs specifically into dendritic cells, the CD11c minimal promoter has been used. The plasmid driven by the CMV promoter was used for expression of the mini-dystrophin (an intracellular, structural protein) or the E.coli b-galactosidase (cytoplasmic but also secreted, strongly antigenic protein) in cells in vitro and muscles in vivo. Another plasmid construct expressing the minidystrophin gene under the muscle- specific creatine kinase promoter and the myosin light chain 1/3 enhancer combination was also used for studies of the effects of muscle-specific expressions of the transgene. The cloning resulted in the generation of plasmids with the mini-dystrophin driven by MCK or CMV promoters and IDO transcripts under the control of CMV or CD11c minimal promoter (Chapter 3). The Western blotting analyses confirmed the ability of plasmids to drive specific transgenes' expression in HEK 293, mouse myoblasts and RAW 264.7 macrophage cell lines in vitro. The RT-PCR analyses confirmed the expression of specific plasmids (Chapter 4). The single plasmid expression experiment using the mini-dystrophin construct targeted into muscles was analysed by Western blotting, RT-PCR and immunohistochemistry. While RT-PCR confirmed its expression, the Western blotting results were ill-reproducible and immunohistochemistry did not confirm transgene expression. Moreover, there was no significant difference in the expression of mini-dystrophin driven by CMV or the muscle-specific MCK promoter/MLC enhancer combination (Chapter 5). The use of Pluronic SP1017-2 did not yield any significant improvement in the expression profile of plasmids as compared with normal saline. Hence, normal saline was used in subsequent experiments as a vehicle of choice. Moreover, to support the hypothesis, there was a requirement to analyse the fold increase of the target plasmid expression in the presence of immunomodulatory factors. This could not be achieved using the minidystrophin plasmids due to low expression and lack of reproducibility. Therefore, the expression profile of b-galactosidase used as a model transgene was analysed instead. This protein is immunogenic due to its E.coli origin and is a 120 KDa protein, which is very close to 125 KDa size of the mini-dystrophin. The timeline of b-galactosidase expression was established based on the presence of this protein at 7 and 14 days and its absence 21 days post-injection, as assessed by Western blotting. The expression profiles of IDO1 driven by CMV or CD11c were analysed and confirmed using RT-PCR; IDO1 did not show detectable expression in Western blotting. (Chapter 5). The effects of co-injection of β-galactosidase with IDO1 driven by CMV or CD11c were analysed by Western blotting, RT-PCR and qPCR. In the control samples, 25% of muscles expressed b-galactosidase two weeks after the injection. This increased to 42% (5 out of 12 muscles) in samples co-injected with CD11c-driven IDO1 and 69% (11 out of 16 muscles) in samples co-injected with IDO1 driven by the CMV promoter. This confirmed the hypothesis that the presence of IDO1 has a potential to sustain the expression of an immunogenic transgene and indicated that the more widespread rather than targeted expression of IDO1 in antigen-presenting cells was more effective in supporting such an expression (Chapter 5). The RT-PCR data showed IDO1 expression in most samples, also some that were not showing β-galactosidase in Western blots, and confirmed the plasmid-driven IDO1 expression. The qPCR data also confirmed significantly increased expression of b-galactosidase and IDO1 in co-injected samples compared to β-galactosidaseonly controls. This further supported the hypothesis that co-expression of immunomodulatory IDO1 increases the transgene expression (Chapter 5). The X-gal staining identified the expression of b-galactosidase in very few myofibres, which correlated with the Western blotting data and confirmed the low efficiency of the "naked" plasmid uptake by skeletal muscles. The presence of infiltrating immune cells surrounding these β-galactosidase positive myofibres was probed by immunohistochemical methods (Chapter 6). The qPCR analyses of a selection of the immune cell markers showed statistically significantly higher expression of CD4, CD8a, FoxP3 and COX2 in co-injected samples while expressions of IL-10 and IL-12 were statistically significantly lower in co-injected muscles. The levels of antibodies against beta-galactosidase were quantified by ELISA in control, b-galactosidase-only injected samples and IDO1 co-injected samples. The anti-β-galactosidase antibody levels were significantly lower in co-injected samples compared to the controls (Chapter 6). These results indicate that co-expression of genes encoding immunomodulatory enzymes of the kynurenine pathway can be a feasible strategy for preventing loss of expression of transgenes targeted into muscles with pre-existing inflammation. Hypothesis: "Co-expression of immunomodulatory factors (IDO/TDO) with minidystrophin or beta-galactosidase transgenes in skeletal muscles of the mdx mouse prolongs the expression of these transgenes." Aims:1-To prolong expression of immunogenic transgenes in dystrophic muscles withpre-existing inflammation.2-To prevent this loss of transgene by exploiting co-expression of genes encoding enzymes controlling kynurenine pathways instead of global immunosuppression. Objectives:1-To modify and validate expression profile of plasmids expressing target genes(mini-dystrophin and beta-galactosidase) and immunomodulatory genes(IDO1/IDO2/TDO/FoxO3) both in vitro and in vivo.2-To check the effects of immunomodulatory genes on the prolongation of target genes expression in vivo.3-To assess the occurrence of the tolerance induction.
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Dias, Florencio Leite Gabriella. "Recombinant Adeno-Associated Viruses : process development and gene transfer application for muscular dystrophy." Thesis, Université Paris-Saclay (ComUE), 2017. http://www.theses.fr/2017SACLV051/document.
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L'intérêt de l’utilisation des vecteurs viraux comme le Adeno-Associated Virus recombinant (rAAV) dans la recherche pour le traitement des maladies génétiques a conduit à une évolution rapide des méthodes de production d'AAV au cours des deux dernières décennies (Ayuso et al., 2010). Leur large biodisponibilité in vivo et leur efficacité à long terme dans les tissus postmitotiques en font de bons candidats pour de nombreuses applications de transfert de gènes. En plus, la spécificité du traitement peut être augmentée lorsque le sérotype correct est choisi pour cibler un tissu spécifique. Parmi les méthodes de production actuellement utilisées, la tri-transfection de cellules embryonnaires humaines rénales 293 (HEK293) reste la plus populaire pour l'échelle de recherche; Et la production de rAAV médiée par des baculovirus pour des échelles plus importantes. L'importance croissante des vecteurs viraux dans l'application pratique de la thérapie génique exige l'amélioration des processus de production, en particulier en ce qui concerne les rendements et la pureté du produit final. Mon travail au cours de ces quatre années a été axé sur deux points principaux: (1) améliorer les processus biotechnologiques employés dans la production de rAAV pour la recherche et les échelles d'étude préclinique et (2) tester in vitro et in vivo les applications pour le rAAV dans le l’édition de genome. L'édition de gènes médiée par des nucléases spécialement conçues offre de nouveaux espoirs pour le traitement de plusieurs maladies héréditaires monogéniques. Récemment découvert, le système CRISPR Cas9 (Clustered Regular Interspaced Short Palindromic Repeats) fournit des outils importants nécessaires pour corriger les mutations par homologie. Notre modèle canonique est la souris mdx, un modèle animal naturel de la dystrophie musculaire de Duchenne (DMD). Les mutations DMD, qui conduisent à l'absence de protéine dystrophine, entraînent une myopathie progressive et fatale. Plusieurs stratégies, allant des stratégies pharmacologiques aux stratégies de saut-d’éxon, ont tenté de renverser le phénotype et ralentisser la progression de la maladie, mais les résultats ne sont pas encore satisfaisants. Ce nouvel et puissant outil d'édition de génome peut être vectorisé par rAAV. Les résultats de la première partie ont été publiés en 2015 et 2016 et seront présentés sous la forme d'articles et pour la deuxième partie, je présenterai les résultats préliminaires et les perspectives du travail qui se poursuivra dans le laboratoireThe interest of recombinant Adeno-Associated Virus (rAAV) vectors for research and clinical purposes in the treatment of genetic diseases have led to the rapid evolution of methods for AAV production in the last two decades (Ayuso et al., 2010). Their broad in vivo biodistribution and long-term efficacy in postmitotic tissues make them good candidates for numerous gene transfer applications. In addition, the specificity of the treatment can be increased when the right serotype is chosen to target a specific tissue. Among the production methods currently in use, tri-transfection of human embryonic kidney 293 (HEK293) cells remains the most popular for research scale; and rAAV production mediated by baculoviruses for larger scales. The increasing importance of viral vectors in the practical application of gene therapy demands the improvement of production processes, especially when it concerns the yields and purity of the final product. My work during these four years was focused in two main points: (1) improve biotechnological processes employed in rAAV production for research and pre-clinical study scales and (2) test in vitro and in vivo the applications for rAAV in the field of genome editing. Gene-editing mediated by engineered nucleases offers new hopes for the treatment of several monogenic inherited diseases. Recently discovered, the CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) Cas9 system provides important tools needed to correct by homology-directed repair mutations. Our canonical model is the mdx mouse, a naturally occurring animal model of Duchenne Muscular Dystrophy (DMD). DMD mutations, which lead to the absence of the protein dystrophin, results in a progressive and fatal myopathy. Several strategies, from pharmacological to exon-skipping strategies, have attempt to revert the phenotype and slow down the disease progress, however results are not yet satisfactory. This new and powerful genome editing tool can be vectorized by rAAV. Results for the first part were published in 2015 and 2016 and will be presented in the form of articles and for the second part I will present preliminary results and perspectives for the work that will be continued in the lab
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Heller, Kristin Noreen. "Alternative to Gene Replacement for Duchenne Muscular Dystrophy using Human Alpha7 Integrin (ITGA7)." The Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1388401639.
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Reza, Mojgan. "Engineering and optimisation of mini-dystrophin constructs for Duchenne muscular dystrophy gene therapy." Thesis, University of Newcastle upon Tyne, 2015. http://hdl.handle.net/10443/2827.
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Muscular dystrophies (MDs) are inherited disorders characterised by muscle weakness and atrophy. One of the most severe forms is Duchenne muscular dystrophy (DMD) which together with the milder allelic form Becker muscular dystrophy (BMD) are known as the dystrophinopathies and result from defects in the X-linked gene encoding dystrophin. Dystrophin is a structural protein of the muscle that connects the internal cytoskeleton of muscle fibres to the extracellular matrix. DMD is also amongst the most common forms of muscular dystrophy, affecting ~1 in 4000 live male birth and manifests as rapidly progressive muscle degeneration leading to loss of ambulation and death in the second or third decade from respiratory or cardiac failure. Currently, there is no cure for this devastating disease. Clinical management of symptoms and complications is limited to stabilising the condition, slowing deterioration over time and palliative care. Since discovery of the DMD gene in 1986, researchers have dedicated substantial effort into vector technologies, facilitating the use of gene therapy to reintroduce a functional copy of the dystrophin gene into muscle fibres, a potential approach to treat DMD patients. However, this approach poses additional challenges relative to many gene therapy approaches since the full-length dystrophin cDNA is ~14 kb, exceeding the packaging capacity of most viral vectors. A number of large internal in-frame dystrophin deletions have been identified in patients that produce a relatively mild phenotype with later age of onset and a slower rate of disease progression than classical DMD. This observation has inspired the construction of internally truncated, but largely functional versions of dystrophin suitable for gene transfer using viral vectors. So far the most widely used miniaturised dystrophin transgenes have been tested in AAV-mediated gene delivery which has identified several limitations indicating the use of more favourable transgenes that have smaller deletions, yet carrying more functional parts of dystrophin. In this study human mini-dystrophin constructs of 4.3-7.7 kb in size were designed that retain key functional elements of dystrophin molecule and their relative functionality investigated in mdx mice. The ultimate aim of this study is the characterisation and optimisation of these mini-dystrophin constructs for gene delivery studies via viral vectors as a therapeutic tool for treatment of Duchenne muscular dystrophy.
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Deol, Jatinderpal. "Development of helper-dependent adenovirus for gene expression in muscle." Thesis, McGill University, 2001. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=33745.
Full textAbstract:
Duchenne muscular dystrophy (DMD) is characterized by necrosis and progressive loss of muscle fibers. DMD patients have a mutation in the gene encoding dystrophin, a large membrane-associated cytoskeletal protein on the cytoplasmic side of the sarcolemma. Gene therapy using fully deleted adenoviral vectors shows great potential for the eventual treatment of DMD and other genetic diseases. These vectors are less immunogenic than their predecessors and have the capacity to carry large DNA inserts such as the full-length dystrophin (12 kb). However, the lack of viral genes results in a weakened and subsiding (short) transgene expression in muscle. Findings in the lung and liver have shown the adenoviral E4 region, in particular E4 open reading frame 3 (ORF3) to contribute to the maintenance of transgene expression. We constructed an adenovirus in which E4 ORF3 was reintroduced into a fully-deleted adenovirus along with full-length dystrophin (AdCBDysORF3). Dystrophin levels produced by AdCBDysORF3 were found to be not sustained in mdx mice, dropping significantly by day 90. However, expression levels did increase when AdCBDysORF3 was complemented with other viral proteins such as EIB. Likewise, increasing the expression of the primary adenovirus receptor (CAR) in muscle also resulted in a higher initial dystrophin expression in myofibers.
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Péladeau, Christine. "Utrophin A Upregulation by FDA-Approved Drugs for the Treatment of Duchenne Muscular Dystrophy." Thesis, Université d'Ottawa / University of Ottawa, 2019. http://hdl.handle.net/10393/39298.
Full textAbstract:
Duchenne Muscular Dystrophy (DMD) is a disorder caused by mutations in the dystrophin gene, preventing the production of the functional dystrophin protein which assures maintenance of the myofiber integrity throughout muscle contraction. A lack of dystrophin results in severe muscle degeneration and regeneration accompanied by a loss of muscle function. Many pre-clinical and clinical studies are focused on developing strategies to counteract the detrimental effects of DMD; however, there is no cure. One such approach consists of upregulating the endogenous protein utrophin A in dystrophic muscle, which, once highly expressed at the sarcolemma, could functionally compensate for the lack of dystrophin. Recent evidence demonstrates that utrophin A expression is regulated at its 3’ and 5’UTR through post-transcriptional and translational events. Therefore, in the work presented here, we hypothesized that repurposing FDA-approved drugs that target the signaling pathways involved in post-transcriptional and translational regulation of utrophin A will be an efficient approach in rapidly bringing new therapeutic interventions for DMD.
In this work, we repurposed four promising FDA-approved drugs able to stimulate utrophin A expression levels in dystrophic muscles: the anti-coagulant drug Heparin, the anti-inflammatory drug Celecoxib, the β-adrenergic receptor blocking agent Betaxolol and the cholesterol-lowering drug Pravastatin. These drugs induce significant improvements in the dystrophic phenotype of mdx mice. This includes amelioration of muscle fiber integrity and muscle function as well as promoting morphological and fiber type changes in mdx mice muscles. Collectively, this thesis describes the potential of a repurposing approach to activate key post-transcriptional and translational pathways involved in utrophin A’s regulation in the hopes of developing new therapeutics for the treatment of DMD.
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Thaker, Rajsi Y. "Potential drug treatment for Duchenne muscular dystrophy which could be through upregulation of lipin1." Wright State University / OhioLINK, 2021. http://rave.ohiolink.edu/etdc/view?acc_num=wright1629996330644397.
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Nascimento, Joyce Aline Paganelli. "Adaptações da marcha em pacientes com distrofia muscular de Duchenne pelo uso de AFO (Ankle-Foot Orthosis) diurna: duplo protocolo com uso progressivo e livre." Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/17/17152/tde-19072018-162819/.
Full textAbstract:
Introdução: A distrofia muscular de Duchenne (DMD) é causada pela deleção ou deficiência do gene que codifica a proteína distrofina. Com a evolução da doença, ocorre um grande comprometimento na marcha com consequente perda da capacidade de deambulação, fato que causa grande impacto na qualidade de vida dos pacientes e de seus cuidadores. Recomendações para uso noturno da órtese suropodálica, também chamada AFO (Ankle-Foot Orthosis), já estão bem estabelecidos na literatura científica, porém o uso durante a deambulação ainda é incipiente. Recentemente, o uso da AFO articulada diurna foi avaliado e indicado como importante aliado no tratamento da reabilitação desses pacientes, capaz de minimizar as compensações características da doença e prolongar a marcha. Ainda assim, questões como o tempo recomendado para uso diário e efeitos do uso do dispositivo, a médio e longo prazo, aguardam investigações mais precisas . Objetivo: Identificar as adaptações cinemáticas e cinéticas da marcha de pacientes com DMD que fizeram uso da órtese tipo AFO articulada diurna durante dois períodos de três meses, pelo uso progressivo e livre, respectivamente. Método: A amostra foi composta por 8 pacientes deambuladores diagnosticados com DMD, de 6 a 10 anos de idade. As avaliações foram compostas por testes de força, medida da amplitude de movimento, teste de caminhada dos 10 metros, testes funcionais cronometrados, aplicação da escala de Medida da Função Motora (MFM) e análise cinética e cinemática da marcha, com órtese (CO) e sem órtese (SO). Cada voluntário participou de 4 avaliações ao longo de um período de 6 meses e fez um auto-relato do número de quedas. No período entre 1ª avaliação (AV1) e a 2ª avaliação (AV2) o paciente fez uso diurno da órtese durante 2 horas/dia que foi incrementado para 4 horas/dia (2º mês) e 6 horas/dia (3º mês), momento que foi realizada a 3ª avaliação (AV3). Entre o 3º e 6º mês, o voluntário ficou livre para usar, ou não, a AFO diurna. Ao final desse período, foi realizada a 4ª avaliação (AV4). Para análise dos dados, foi utilizado o teste de regressão linear com efeitos mistos (efeitos aleatórios e fixos) obtidas com o auxílio do Software SAS® 9.3. Os dados obtidos em nosso estudo foram comparados com dados normativos da literatura. Para os dados cinemáticos e cinéticos da marcha foram obtidas as médias, de 3 avaliações, dos picos máximos e mínimos dos parâmetros de cada fase da marcha (apoio e balanço) para cada paciente com e sem órtese. Posteriormente, foram calculadas as médias e os intervalos de confiança de cada grupo, com e sem órtese. Resultados: Os testes cronometrados demonstraram redução do tempo de subida de 4 degraus, sem órtese, quando comparados os tempos de execução na AV1 em relação à AV3 e na AV1 em relação à AV4 (p<0.05). A análise comparativa das médias de força muscular indicou que houve aumento significativo da força de flexores de joelho da AV3 para AV4, dos extensores de joelho da AV1 para AV3 e dos dorsiflexores da AV1 para AV3 e da AV1 para AV4 (p<0,05). A análise dos parâmetros espaço-temporais indicou diminuição da largura da passada (p<0.05) quando comparada a AV1 em relação à AV4 na situação sem órtese (AV1 vs AV4). Quando comparamos dados do grupo CO em relação ao grupo SO, pode ser observado que o grupo CO apresentou maior tempo do ciclo da marcha na AV1 (p<0.01), maior tempo de duplo apoio na AV1 (p<0.01), na AV2 (p<0.01) e na AV3 (p=0.02). Nas avaliações cinética e cinemática, a análise comparativa entre as condições com e sem órtese, na fase de apoio da marcha, indicou redução significativa dos seguintes 11 parâmetros, para condição CO: amplitude de abdução e adução (p=0.0002) e absorção de potência de quadril (p<0.0001), geração de potência de potência de tornozelo (p<0.0001). Outros parâmetros apresentaram aumento significativo na condição CO quando comparado à condição SO, fase de apoio: máximo momento extensor (p<0.0001) e geração de potência (p=0.0035) de quadril, máximo momento flexor (p<0.0001) e amplitud e de geração e absorção de potência (p<0.0001) de joelho, máximo ângulo de dorsiflexão (p<0.0001), máximo momento flexor plantar (p<0.0001) e absorção de potência (p<0.0001) de tornozelo. Na fase de balanço houve redução significativa para máximo momento extensor (p<0.0001) e geração de potência (p<0.0001) de quadril. Nesta mesma fase foi observado aumento significativo para máximo ângulo de flexão (p<0.0001) do joelho, máximo ângulo de dorsiflexão (p<0.0001), máximo momento flexor plantar (p<0.0001) e amplitude de momento dorsiflexor e flexor plantar (p<0.0001) do tornozelo. Foi observado ainda, na fase de balanço da marcha, aumento significativo na geração de potência de tornozelo (p=0.0251) nas AV1, AV3 e AV4, na condição CO quando comparada à condição SO. O efeito de interação das fases de apoio e balanço também indicou que a condição SO apresentou máximo ângulo de inclinação pélvica superior quando comparado à condição CO, nas AV2 (p=0.0011), AV3 (p=0.0024) e AV4 (p=0.0191). Conclusão: O uso diurno e progressivo da órtese AFO articulada, em situação de carga, provoca alterações biomecânicas positivas na marcha de pacientes com DMD que repercutem minimizando o número de quedas e favorecendo a funcionalidade geral das crianças .Introduction: Duchenne Muscular Dystrophy (DMD) is caused by deletion or deficiency in the gene that encodes the protein dystrophin. The clinical evolution of this disease includes significant gait impairment with consequent loss of walking ability, and this fact causes negative impact on the quality of life of the affected ones and their caregivers. It has already been well established that there are beneficial effects of nocturnal use of Ankle Foot Orthosis (AFO), nevertheless, the discussion about the daytime use of articulated AFO is rare. Recently, the daytime use of AFO was evaluated and indicated as an important ally in the treatment of these patients, capable of minimizing the biomechanical compensations and prolonging gait cycle. Even so, some issues such as the recommended time and effects for daily use, in the medium and long term, await more precise investigation. Objective: To identify the effects of daytime use of articulated AFO on spatiotemporal, kinematic and kinetic gait parameters of DMD patients, during two periods of three months, by progressive and free use, respectively. Methods: Eight walking patients diagnosed with DMD between the ages of 6 and 10 years old were evaluated. The data were obtained according to the isometric muscle strength, joint range of motion, timed functional score, the Motor Function Measure (MFM) scale and gait analysis parameters, with (CO) and without (SO) AFO. Four evaluations were carried out over a period of six months and each volunteer self-reported your number of falls. During the first (AV1) and second (AV2) evaluation, patients u sed the daytime orthosis during two hours per day. This time was increased to four hours per day in the second month and six hours per day in the third month, then when the third (AV3) evaluation was conducted. Between third and sixth month, the use of the orthosis was optional. By the end of month six, the fourth (AV4) evaluation was conducted. The data were analyzed using the mixed linear regression model (Random and Fixed Effects) through the Software SAS® 9.3. The results obtained in the present study are compared with literature data. The means of 3 evaluations for the spatiotemporal, kinematic and kinetic gait data were obtained, for the maximum and minimum peaks of the parameters of each phases in a gait cycle (stance and swing) for each patient with and without orthosis. In the end, the means and the confidence interval were calculated for each group, with and without AFO. Results: The timed tests showed a reduction in time for climbing 4 steps without AFO, when compared the AV1 and AV3 runtime in relation to AV1 and AV4 runtime (p<0.05). The comparation of muscle strength showed a significant increase in knee flexor strength from AV3 to AV4, knee extensors from AV1 to AV3 and dorsiflexors from AV1 to AV3 and from AV1 to AV4 (p<0.05). The analysis of s patiotemporal parameters indicates a decrease in the width stride (p<0.05) between AV1 and AV4 without orthosis (AV1 vs AV4). When comparing CO with SO, CO group presented longer gait cycle in AV1 (p<0.01), longer double support phase in AV1 (p<0.01), AV2 (p<0.01) and AV3 (p=0.02). In the kinetic and kinematic evaluations, the comparative analysis between the conditions with and without orthosis in the gait stance phase indicated a significant reduction of the following parameters for the CO condition: adduction-abduction range of motion (p = 0.0002) and hip power absorption (p<0.0001) and ankle power generation (p <0.0001). Other parameters showed a significant increase in the CO condition when compared to the SO in stance phase: peak extensor moment (p<0.0001) and hip power 14 generation (p=0.0035), peak flexor moment (p<0.0001) and power generation and absorption range of motion (p<0.0001), peak dorsiflexion angle (p<0.0001), peak plantar-flexor (p<0.0001) and ankle power absorption (p<0.0001). In the swing phase, a significant reduction in the extension angle (p<0.0001) and hip power generating (p<0.0001). Also in swing phase, a significant increase for peak knee flexion (p<0.0001), peak dorsiflexion range of motion (p<0.0001), peak plantar flexor moment (p<0.0001) and ankle joint dorsiflexor and plantar-flexor range of motion (p<0.0001) were observed. Besides this, in the gait swing phase was observed significant increase in ankle power generation (p=0.0251) in AV1, AV3 and AV4 in the condition CO when compared to the SO condition. The interaction effect of stance and swing phases also indicated that the SO condition presented higher pelvic tilt angle when compared to the CO condition, in AV2 (p=0.0011), AV3 (p=0.0024) and AV4 (p=0.0191). Conclusion: Thus, the progressive use of Articulated Ankle Foot Orthosis (AFO) in loading response phase can change the gait pattern of patients with Duchenne muscular dystrophy. This result turn to positivity when the temporal, kinematic and kinetic gait parameters are evaluated.
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Ricotti, V. "Evolving natural history in Duchenne muscular dystrophy : implications for standard of care and experimental therapies." Thesis, University College London (University of London), 2016. http://discovery.ucl.ac.uk/1474132/.
Full textAbstract:
Duchenne muscular dystrophy (DMD) with an average global incidence of 1:5000 is an X-linked recessive disease, caused by mutations in the DMD gene encoding dystrophin. Lack of dystrophin isoforms results in progressive muscle weakness and cardiomyopathy, leading to loss of ambulation and premature death secondary to cardiac/respiratory complications. At present, there is no curative treatment. However, implementation of standards of care has significantly shifted life expectancy and the natural history of DMD has considerably evolved. Moreover, a number of promising therapeutic approaches are under development, some reaching phase II-III clinical trials. These experimental therapies will further contribute to the transformation of the disease trajectory. The projects of my thesis intended to address specific research questions, which have an impact not only on the clinical care of DMD patients, but also advice on clinical trial design. I studied the effect of steroid therapy on the motor function in DMD boys >7 years, more specifically profiling benefits and side effects of the most commonly used regimens: intermittent and daily prednisolone. I analysed the impact of starting steroids at an earlier age than what is standard of care. I explored the role of different dystrophin gene (DMD) genotypes in the motor progression of the disease, further defining the genotype-phenotype correlations. All results obtained are of particular interest for clinical trials of pharmaco-gene therapies targeting specific DMD mutations. Dystrophin isoforms also play an important role for the CNS and their lack causes morbidity in DMD. My investigations expanded the genotype-phenotype profile specifically in relation to neuropsychiatric co-morbidities in DMD. In conjunction with the CNS profile of DMD, I characterized abnormalities of retinal function and developed electroretinography as a potential and non-invasive CNS endpoint for future clinical trials. Addressing the non-ambulant DMD population, I studied quantitative magnetic resonance imaging and novel functional measures of the upper limb. These results allow for the first time to evaluate disease progression and response to treatment in non-ambulant DMD. All the results obtained in this thesis therefore enlarge our knowledge of the disease evolution under current standard treatment and contribute to trial readiness by developing new endpoints.
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Betts, Corinne A. "Exon skipping peptide-pmos for correction of dystrophin in mouse models of duchenne muscular dystrophy." Thesis, University of Oxford, 2014. http://ora.ox.ac.uk/objects/uuid:545d586a-ad7b-4089-8537-b2677957b874.
Full textAbstract:
Duchenne muscular dystrophy (DMD) is a fatal, muscle-wasting disorder due to mutations/deletions in the dystrophin gene. Whilst improvements in palliative care have increased the life expectancy of patients, cardiomyopathy and respiratory complications are still the leading causes of death. A potential therapy for the treatment of DMD is antisense oligonucleotides (AOs), which modulate dystrophin pre-mRNA splicing to restore the dystrophin reading frame and generate a truncated functional protein. Conjugation of AOs to cell penetrating peptides (CPP), such as Pip5e-, significantly improves delivery to skeletal muscles and to the heart, which is imperative given the impact of cardiomyopathy to mortality. However, it should be noted that the contribution of skeletal muscles, such as the core respiratory muscle, the diaphragm, in dystrophic cardiopulmonary function is poorly understood. The specific aims of the work in this thesis were to (i) understand the effect of the diaphragm on cardiac function using magnetic resonance imaging (MRI), (ii) screen a number of derivatives of Pip5e (Pip6) in an effort to discover further promising peptides and define the properties integral to heart penetrating capacity, and (iii) assess whether Pip6-PMOs restore cardiac function (MRI) following a repeat, low dose regimen. In short, the specific restoration of dystrophin in the diaphragm of the dystrophic mouse model, the mdx mouse, did not improve cardiac function, highlighting the importance of a body-wide therapy. The screening of multiple Pip5e-PMO derivatives revealed 3 promising peptides with improved cardiac splicing capacity; however, serial deletions of amino acids from the central core resulted in the diminution of dystrophin restoration, possibly due to a reduction in hydrophobicity. Finally, the Pip6-PMO treatment regimen substantially restored dystrophin protein (28% in heart) and stabilised cardiac function, even with an increased work load. In conclusion, this study illustrates the importance of a body-wide treatment, such as the CPP strategy (Pip-PMO). These Pip-PMO conjugates demonstrate high dystrophin restoration in a number of muscles, including cardiac muscle, and have a beneficial effect on cardiac function.
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Call, Jarrod Alan. "Low load endurance activity and green tea extract represent potential therapies for Duchenne muscular dystrophy." Thesis, Virginia Tech, 2007. http://hdl.handle.net/10919/34585.
Full textAbstract:
Duchenne muscular dystrophy (DMD) is a progressive muscle wasting disease affecting 1 in every 3500 boys. The disease is characterized by the absence of the dystrophin protein from the sarcolemma of muscle cells. Muscle cells lacking dystrophin go through cycles of degeneration and regeneration and are considered susceptible to contraction-induced injury 144. Eventually, the satellite cell proliferative capacity is exhausted and the muscle fibers are replaced by connective and adipose tissue that yields a progressive loss of force generating capability. DMD patients typically die by their early 20's, primarily due to respiratory or cardiac failure. The precise role of dystrophin is not presently known. However, its absence suggests that it may play a role in both cellular calcium regulation and oxidative stress 152. Recent studies suggest increased reactive oxygen species (ROS) may precede the initial wave of wasting that marks disease onset 49. Therefore, it is possible oxidative stress may contribute as a pathogenic mechanism of DMD. Strategies to reduce the deleterious effects of oxidative stress could be an effective therapeutic approach. Regular exercise is known to increase antioxidant capacity in humans and mice 146. Green tea extract (GTE) is a powerful antioxidant that is easily supplemented in the diet 83.
The purpose of this study was to test the hypotheses that (1) voluntary endurance exercise alone, (2) a diet supplemented with 0.05% (wt/wt) GTE alone, or exercise and GTE combined will blunt the effects of ROS and improve muscle strength and endurance in young mdx mice. Male mdx mice at age 21-days were randomly divided into one of 4 treatment groups: Run Normal diet (RunNorm; n=8), Sedentary Normal diet (SedNorm; n=8), Run GTE diet (RunGTE; n=10), and Sedentary GTE diet (SedGTE; n=8). RunNorm and RunGTE mice were given free access to a running wheel for 3 weeks while SedNorm and SedGTE mice were restricted to normal cage movement. At the end of 3 weeks, mice in each treatment group were sacrificed and assessed for daily and weekly running distances, content of actin and myosin proteins and fiber type distribution (tibialis anterior), contractile/mechanical and fatigue properties (extensor digitorum longus), creatine kinase levels and antioxidant capacity (serum), lipid peroxidation (gastrocnemius), and citrate synthase and beta-hydroxyacyl-CoA dehydrogenase activities (quadriceps and soleus).
The key findings of this study were: In normal diet running mice (RunNorm), average daily distance run was increased 300% (from 0.5 to 2.1 km/d, P<0.05) from week 1 to week 3. In GTE diet (RunGTE) compared to RunNorm mice, total distance over the 3 weeks was markedly improved 128% (61.2 vs. 26.8 km, P<0.0001). Running, independent of diet increased EDL muscle tetanic stress (18%), serum antioxidant capacity (22%), citrate synthase activity (35%), and beta-oxidation (37%; all P<0.05). GTE, independent of running decreased lipid peroxidation (gastrocnemius:-64%; heart: -29%) and serum creatine kinase (-36%), and increased citrate synthase activity (59%; all P<0.05).
These findings in dystrophic mice suggest that voluntary endurance exercise with or without GTE supplementation blunted the deleterious effects of ROS. If similar positive effects are evident in human DMD patients, then these approaches may be beneficial therapies either alone or in combination.
Master of Science
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Gibson, Barbara Ellen. "Ventilator use for patients with Duchenne muscular dystrophy, an ethical analysis of physicians' beliefs and practices." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1998. http://www.collectionscanada.ca/obj/s4/f2/dsk2/tape17/PQDD_0027/MQ33948.pdf.
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Zeiger, Ulrike [Verfasser]. "Physiological, molecular and biochemical characterization of rodent extraocular muscles: Implications for Duchenne Muscular Dystrophy / Ulrike Zeiger." Greifswald : Universitätsbibliothek Greifswald, 2011. http://d-nb.info/1016772033/34.
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Handley, Kirsten Louise. "Investigations into the mechanisms of degeneration in the MDX mouse : a model for Duchenne muscular dystrophy." Thesis, Imperial College London, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.432066.
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Thorogood, Francesca Clare. "Modulation of dystrophin pre-mRNA splicing by antisense oligonucleotides : a potential therapy for Duchenne muscular dystrophy." Thesis, Royal Holloway, University of London, 2007. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.504809.
Full textAbstract:
Duchenne muscular dystrophy (DMD) is an X-linked muscle wasting disorder caused by mutations in the gene for dystrophin, a 427kDa cytoskeletal protein important for maintaining the integrity of muscle fibres. A number of DMD mutations result in an absence of functional protein due to disruption of the translational reading frame. It has been shown previously that antisense oligonucleotide (AON) reagents can modulate dystrophin pre-mRNA splicing to specifically exclude an out-of-frame exon from the mRNA. This restores the open reading frame resulting in production of a semi-functional internally-truncated dystrophin protein, mimicking what occurs in the milder allelic Becker muscular dystrophy (BMD). Previous work in this laboratory demonstrated successful exclusion of nonsense mutation carrying exon 23 in the mdx mouse model of DMD. This thesis is concerned with extension of the investigation by examining the bioactivity of alternative AON backbone chemistries 2' -O-methyl phosphorothioate (20Me), peptide nucleic acid (PNA) and phosphorodiamidate morpholino oligomer (PMO) targeting the 5'splice site of exon 23 in vitro and in vivo. In cultured murine muscle cells both the 20Me and PMO-based AON reagents induced detectable exon 23 exclusion. In the mdx mouse model intramuscular delivery of the 20Me-based AON reagent resulted in de novo dystrophin expression correctly localised at the sarcolemma that persisted for up to 4 weeks after a single dose. The PMO-based reagent resulted in de novo dystrophin expression that persisted for up to 10 weeks after a single intramuscular dose and for up to 8 weeks after a single intravenous dose. To broaden the investigation nine additional murine dystrophin exons were selected and AON regents designed targeting the 5'splice site and putative exonic splicing enhancer (ESE) sequences. Overall the results demonstrate that the AONs employed here induce detectable, reproducible exclusion of exon 23. The PMO-based reagent is currently superior for modulation of dystrophin pre-mRNA splicing.
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Adkin, Carl F. "A model of human muscle regeneration in vivo to test potential therapies for Duchenne Muscular Dystrophy." Thesis, Imperial College London, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.505313.
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Kalra, Spandan Kaur. "Towards the development of human induced pluripotent stem cell models for Duchenne muscular dystrophy-associated cardiomyopathy." Thesis, University of Nottingham, 2015. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.715123.
Full textAbstract:
Duchenne Muscular Dystrophy (DMD) is a fatal X-linked condition that affects 1 in 4710 boys. Causative mutations in the DMD gene result in a loss of functional expression of the dystrophin protein. Patients usually die in their second or third decade due to cardiac or respiratory failure. Incidence of cardiomyopathy increases with age, such that clinical symptoms are observed in 25% of boys below 6 years of age, but by the teenage years prevalence is 100%. The current treatments are palliative and there are no cures. Numerous cell-based and animal models for DMD exist, but each has its limitations. This includes inter-species differences, incomplete phenocopying of DMD pathophysiology and limited access to diseased human cardiac cells. Therefore, to complement these models, cardiomyocytes (CMs) derived from human induced pluripotent stem cells (hiPSCs) from DMD patients were used to investigate therapeutic strategies and to evaluate the phenotype to better understand the disease. Towards these goals, protocols were optimised for differentiation, characterisation and enrichment of CMs from DMD hiPSC lines. These lines had been derived in our laboratory by four factor lentiviral reprogramming (LIN28, SOX2, NANOG and OCT4) of skin biopsies from boys clinically and genetically diagnosed as having DMD (Dick et al., 2011). Specifically, lines DMD19 and DMD16 carried premature stop codon mutations in exons 35 and 70 respectively, while DMD4 harboured frameshift deletion in exons 48-50 and were used in the study. In the first instance, no CM differentiation was observed. However, manipulation of cell seeding density and BMP, TGFP and WNT signalling pathways enabled efficiencies to be improved to up to 100%. In addition, culture of the hiPSC-CMs in low glucose / high lactate medium lead to preferential survival of the CMs to improve purity to >90%. Improved efficiency of hiPSC-CM production and enrichment provided material to evaluate methods of gene therapy. This included delivery of micro-dystrophin and gene correction of the genomic mutation with RNA guided nucleases (RGNs) in both DMD16 and 19 lines. In each case, restoration of dystrophin protein was observed. In parallel to the gene therapy studies, methods to perform phenotypic analysis were developed. Thus, hiPSC-CMs were analysed for: a) sarcolemma permeability by measuring response of stress on LDH release from cells; b) mitochondrial function in response to different pharmacological challenges by a Seahorse XF assay, and c) Ca2+ profiles, which were quantified using a method termed SALVO (synchronization, amplitude, length and variability of oscillations). Even under severe stress, by treatment with doxorubicin or hypo-osmolarity, no difference was observed in sarcolemma permeability between CMs derived from DMD or healthy hiPSCs. The alterations in mitochondrial function were observed in CMs from DMD hiPSCs relative to healthy CMs. Particularly in DMD16 hiPSCCMs, significant reduction in the ATP production, maximum respiration, spare reserve capacity and non-mitochondrial respiration was observed, while the proton leak was increased. While the data on calcium profile was derived, the number of replicates was not enough to overcome variability in recordings and draw any conclusions. Additional experiments are currently ongoing to confirm whether these phenotypic observations can be rescued by the gene therapy approaches employed. In summary, the work in this thesis has developed optimised protocols for CM differentiation from hiPSCs carrying mutation in the DMD gene, which has been used for pilot investigation into suitability as a supplementary model for investigating phenotype and evaluating genetic therapies. Further development of this in vitro model should lead to increased understanding and progress toward the development of therapies for DMD.
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Kawar, Susannah Louise. "Inhibition of myostatin (GDF-8) via myostatin propeptide minigenes : a potential gene therapy for Duchenne muscular dystrophy." Thesis, Royal Holloway, University of London, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.497826.
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The objective of this project was to evaluate the alleviation of muscle wasting phenotypes by hepatic delivery and expression of a recombinant mutated myostatin propeptide mini gene in wild type mice. Initial work focused on using naked plasmid DNA as a vector for delivery of the therapeutic gene. Various β-galactosidase reporter plasmids were tested including pCMVβ, pCAGGSβ, pA2β and a liver specific pSV40 Albuminβ for systemic gene transfer to liver tissue. Following hydrodynamic intravenous administrations, pCAGGSβ proved to be the optimum plasmid and a cDNA encoding a mutated form of the myostatin propeptide (ProFcD/A) was subsequently cloned into a CAGG based plasmid (pAAVCAGGProFcD/A). The expression and bioactivity of the propeptide were assessed using in vitro cell proliferation experiments which showed the ProFcD/A was capable of increasing both proliferation and differentiation in a range of cell types. An initial in vivo study involved intravenous hydrodynamic injections of pAAVCAGGProFcD/A into MF-1 male mice and monitoring growth rates over time. A significant increase was observed in growth rate of the treated mice compared to controls with an accompanying increase in tibialis anterior (TA) muscle mass. Fibre analysis revealed pAAVCAGGProFcD/A treated mice to contain significantly larger fibres than controls owing to muscle hypertrophy. Transgene product levels were detected strongly 7 days post injection in the serum, which declined steadily over the following 21 days. By day 28, levels of transgene product were barely detectable. There was no indication response to either the vector or transgene product. The pAAVCAGGProFcD/A was then used to produce an AAV serotype 8 viral vector containing the ProFcD/A gene (AAV8ProFc) and assessed in initial intravenous tail vein injections of AAV8ProFc into MF-1 male mice. Growth rates were measured over four weeks with a significant increase in the AAV8ProFc treated mice compared to the pCAGG empty plasmid controls. In addition, significant weight increases in both TA and gastrocnemius muscles were seen in AAV8ProFc treated mice over the pCAGG control mice at week 4. Serum showed high levels of transgene expression after 7 days which was sustained up to 28 days post injection with no indication of immune response. Levels of the myogenic protein MyoD also remained higher in AAV8ProFc than pCAGG treated mice. Future studies may involve intravenous injection of both pAAVCAGGProFcD/A and AAV8ProFc into mdx mice, with subsequent monitoring of growth rates, creatine kinase levels, tissue necrosis and muscle strength over time.
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Jonuschies, J. "Characterisation of lentiviral transgene expression in muscle precursor cells : towards a potential therapy for Duchenne Muscular Dystrophy." Thesis, University College London (University of London), 2012. http://discovery.ucl.ac.uk/1362850/.
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Duchenne Muscular Dystrophy is an X-linked genetic disorder characterised by progressive muscle degeneration due to the absence of functional dystrophin protein. Damaged muscle fibres are initially regenerated by satellite cells, the principal muscle-resident stem cells, which give rise to committed progenitor cells that differentiate and fuse with the damaged muscle fibre to introduce new functional myonuclei. Satellite cells also self-renew to replenish the stem cell pool. Autologous transplantation of genetically-corrected satellite cells presents an attractive strategy to introduce a functional dystrophin copy into dystrophic muscles and to replenish the stem cell pool with corrected satellite cells for a long-term therapeutic benefit. Lentiviral vectors are suitable gene transfer vectors that integrate their genome into the host chromosome and mediate stable transgene expression in dividing and non-dividing cells. Long-term gene expression can be hampered by promoter inactivation and undesirable position effects. Tissue-specific promoters have been shown to reduce the risk of transcriptional silencing and to increase the overall biosafety of transgene expression. In this study, I investigated the potential of integrating lentiviral vectors to efficiently infect quiescent satellite cells in order to confer stable expression of GFP in vitro and in vivo. The effect of viral transduction and GFP overexpression on stem cell properties was assessed. The performance of tissue-specific promoters was directly compared with the strong viral promoter of the spleen focus-forming virus and revealed the desmin promoter as an attractive non-viral alternative to confer stable, high-level, muscle-specific expression in myoblasts and myofibres. An ubiquitously-acting chromatin opening element (UCOE) has been reported to negate position effects in hematopoeitic cells. Here, the UCOE failed to prevent promoter down-regulation and did not significantly increase transgene expression when it was combined with the desmin promoter. In summary, this work provides useful information on suitable promoters to achieve stable transgene expression in the myogenic linage.
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Tabebordbar, Mohammadsharif. "Improving Stem Cell-Based Therapy and Developing a Novel Gene Therapy Approach for Treating Duchenne Muscular Dystrophy (DMD)." Thesis, Harvard University, 2016. http://nrs.harvard.edu/urn-3:HUL.InstRepos:26718751.
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Genetic mutations in muscle structural genes can compromise myofiber integrity, causing repeated muscle damage that ultimately exhausts muscle regenerative capacity and results in devastating degenerative conditions such as Duchenne Muscular Dystrophy (DMD), Congenital Muscular Dystrophy (CMD) and different forms of Limb Girdle Muscular Dystrophy (LGMD). Gene supplementation and autologous stem cell transplant have been put forward as promising, though still unproven, therapeutic avenues for combatting these genetic muscle diseases. Both strategies aim to compensate expression of the missing or mutated protein. For cell therapy, autologous muscle stem cells (satellite cells) from dystrophic muscles undergo in vitro expansion and gene correction and then are transplanted into diseased tissue, where they fuse with resident myofibers to deliver a functional copy of the gene. One of the major obstacles for the autologous adult stem cell transplantation is that adult satellite cells account for a very rare population in muscle and they need to be expanded in culture, while retaining their engraftment potential, to generate sufficient number of cells for gene correction and transplantation. I tackled this problem by developing a culture condition that allows engraftable mouse satellite cells to expand in culture. This study also provides evidence for the feasibility of in vitro expansion, gene correction and transplantation of dystrophic satellite cells to restore DYSTROPHIN expression in dystrophic muscle.
In gene therapy, engineered gene products are delivered directly to muscle fibers as transgenes carried by viral vectors, such as Adeno Associated Viruses (AAVs). Viral- mediated delivery of a normal copy of the mutated genes into dystrophic muscle fibers holds big promise as a therapeutic avenue for Muscular Dystrophies. However, considering the indispensible role of satellite cells in muscle regeneration, an effective and long-term therapy for genetic muscle diseases requires restoration of gene expression in both dystrophic muscle fibers and satellite cells. Conventional gene therapy approaches lack the potential for long-term restoration of the mutated gene expression in satellite cells. In order to address this limitation, this study provides the proof of concept evidence for the use of a novel gene editing approach, which allows irreversible correction of the mutations in both dystrophic skeletal muscle fibers and satellite cells.Biology, Molecular and Cellular
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Sako, Yukiya. "Development of an orally available inhibitor of CLK1 for skipping a mutated dystrophin exon in Duchenne muscular dystrophy." Kyoto University, 2017. http://hdl.handle.net/2433/226771.
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Zarrouki, Faouzi. "Dystrophine et système nerveux central : vers un traitement par saut d’exon des défauts cognitifs de la myopathie de Duchenne Efficacy and Safety Profile of Tricyclo-DNA Antisense Oligonucleotides in Duchenne Muscular Dystrophy Mouse Model Long term efficacy of AAV9-U7snRNA mediated exon 51 skipping in Mdx52 mice Gene therapy for central nervous system dysfunction in Duchenne muscular dystrophy." Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLV102.
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La dystrophie musculaire de Duchenne, causée par des mutations dans le gène de la dystrophine, se traduit principalement par une dégénérescence de l’ensemble de la musculature squelettique et cardiaque mais également par des déficits cognitifs plus ou moins sévères. L’objectif de ce projet doctoral est de comprendre le rôle de la dystrophine dans le système nerveux central (SNC) à des fins fondamentales, mais aussi thérapeutiques, si les déficits observés peuvent être compensés. Notre laboratoire a développé une nouvelle classe d’oligonucléotides antisens : les tricyclo-DNA (tcDNA). Chez la souris modèle mdx, déficiente en dystrophine dans le muscle et le cerveau, nous avons montré que ces tcDNA, déjà très efficaces au niveau des muscles, ont la propriété de franchir la barrière hémato-encéphalique suite à une administration systémique et de restaurer une dystrophine tronquée fonctionnelle au niveau du SNC. Nous avons utilisé ces tcDNA afin d’explorer le comportement et l’évolution des fonctions cognitives des souris traitées par des injections systémiques ou intracérébroventriculaires. Nous avons montré que suite à une injection de tcDNA dans le cerveau, une restauration de la dystrophine cérébrale est possible et s’accompagne d’une correction de certains déficits émotionnels et cognitifs, suggérant la réversibilité d’une partie des atteintes centrales. Chez les souris traitées par voie systémique en revanche, et présentant une restauration plus limitée de dystrophine, seul le déficit émotionnel a pu être corrigé.Une seconde partie du projet était dédié à l’étude du rôle de la dystrophine dans la formation des clusters des récepteurs synaptiques GABAA, car ce mécanisme encore mal compris représente un biomarqueur pertinent d’une restauration fonctionnelle de dystrophine. Nous montrons que l’absence de dystrophine cérébrale perturbe le niveau d’expression et la formation des clusters de plusieurs sous-unités composant les récepteurs GABAA. Ces résultats prometteurs ouvrent des perspectives intéressantes sur des futurs thérapies permettant la correction des déficits émotionnels et cognitifs observés chez de nombreux patients atteint de dystrophie musculaire de DuchenneDuchenne muscular dystrophy mainly results in a degeneration of the entire skeletal and cardiac musculature but is also associated with various degrees of cognitive impairment. The objective of this doctoral project is to understand the role of dystrophin in the central nervous system (CNS) for both fundamental and therapeutic purposes if the defects observed are found to be reversible. Our laboratory is developing a new class of antisense oligonucleotides: tricyclo-DNA (tcDNA). In the mdx mouse model, we have shown that these tcDNAs have the ability to cross the blood-brain barrier and restore a functional truncated dystrophin in the CNS. We used these tcDNAs to explore the behaviour and evolution of cognitive functions of mice treated with systemic or locoregional injections. We have shown that following an injection of tcDNA into the CNS, a restoration of cerebral dystrophin is possible. In addition, this restoration allows the correction of some emotional and cognitive deficits. Interestingly, in mice treated intravenously and therefore expressing lower levels of dystrophin in the CNS, only the emotional deficit could be corrected.A second part of the project was dedicated to a better understanding of the role of dystrophin in the formation of GABAA receptor clusters. We show that the absence of cerebral dystrophin disrupts the level of expression and the formation of clusters of several subunits composing the GABAA receptors. These promising results open up interesting prospects for future therapies to correct the emotional and cognitive deficits observed in many patients with Duchenne muscular dystrophy
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Rangan, Apoorva. "CRISPR-Cas9 Mediated Restoration of Dystrophin Expression and Inhibition of Myostatin: A Novel Gene Therapy for Duchenne Muscular Dystrophy." Scholarship @ Claremont, 2016. http://scholarship.claremont.edu/cmc_theses/1305.
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Duchenne Muscular Dystrophy (DMD) is an X-linked recessive genetic disease, caused by a frame-shift mutation in the dystrophin gene. Current gene therapies for DMD target dystrophin transcripts in existing skeletal and cardiac muscle, rather than adipose and fibrotic tissues. These approaches may be unable to repair muscle functionality in DMD patients who have already undergone extensive muscle damage and wasting. Thus, successful DMD therapies must consider the underlying genetic cause and pathology. Inhibition of the gene myostatin, a negative regulator of muscle growth, has been shown to ameliorate muscle loss. Here, the CRISPR-Cas9 gene-editing platform is proposed to restore dystrophin expression and inhibit myostatin as a novel gene therapy in DMD patient derived induced pluripotent stem cells. Successful CRISPR-Cas9 mediated gene editing would be determined using PCR amplification, western blot analysis, immunofluorescence staining, and off target sequence analysis in differentiated skeletal muscle cells.
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Fonteyne, Lina Marie [Verfasser], and Eckhard [Akademischer Betreuer] Wolf. "Evaluation of emerging diagnostic and therapeutic strategies in a tailored pig model for Duchenne muscular dystrophy / Lina Marie Fonteyne ; Betreuer: Eckhard Wolf." München : Universitätsbibliothek der Ludwig-Maximilians-Universität, 2020. http://d-nb.info/1220631922/34.
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Aupy, Philippine. "Le développement préclinique des tcDNA pour la Dystrophie Musculaire de Duchenne Evaluating the impact of variable phosphorothioate content in tricyclo-DNA antisense oligonucleotides in a Duchenne Muscular Dystrophy mouse model Identifying and avoiding tcDNA-ASO sequence specific toxicity for the development of DMD exon 51 skipping therapy Long term efficacy of AAV9-U7snRNA mediated exon 51 skipping in mdx52 mice The use of tricyclo-DNA for the treatment of Genetic Disorders Exon-skipping advances for Duchenne Muscular Dystrophy." Thesis, Université Paris-Saclay (ComUE), 2019. http://www.theses.fr/2019SACLV083.
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La Dystrophie Musculaire de Duchenne est une maladie génétique mortelle qui touche un garçon sur 3500. Elle se manifeste par une faiblesse musculaire progressive conduisant à une perte de la marche autour de l’âge de 10 ans, puis des problèmes respiratoires et cardiaques. Elle est due à des mutations dans le gène DMD conduisant à une absence de la protéine dystrophine. Il n’existe à l’heure actuelle aucun traitement satisfaisant. L’une des stratégies thérapeutiques les plus prometteuses pour cette maladie consiste à moduler l’épissage de l’ARN pré-messager. Cette stratégie appelée aussi « saut d’exon » utilise principalement des oligonucléotides antisens qui vont permettre de restaurer le cadre de lecture et ainsi entrainer la production de protéine.Le laboratoire Biothérapie des Maladies du Système Neuromusculaire a développé une nouvelle chimie d’oligonucléotide antisens, les tricyclo-DNA (tcDNA), ayant fait leur preuve pour effectuer un saut de l’exon 23 efficace dans des modèles murins de la DMD. En effet, les chercheurs de l’équipe ont pu démontrer la présence de saut d’exon et de restauration de dystrophine dans l’ensemble de la musculature et dans le système nerveux central, permettant d’obtenir une amélioration fonctionnelle. Lors de ma thèse, je me suis intéressée au développement pré-clinique d’un tcDNA ciblant l’exon 51 humain, puisqu’il s’agit de l’exon permettant de traiter la plus grande proportion de patients (13%).La première partie de mon projet a été consacrée à l’amélioration de la tolérabilité des tcDNAs à travers deux approches : la modification de la séquence et la modification du design de la molécule. En effet, la cause principale de la toxicité des tcDNAs est la formation de structures homodimériques associée à la présence de liens phosphorothioates (PS). Cette première étude a permis, d’une part, de démontrer qu’une modification de la séquence entraine une élimination des structures homodimériques et permet ainsi d’obtenir une meilleure tolérabilité de la molécule. D’autre part nous avons pu mettre en évidence qu’une diminution du contenu en liens PS permet de limiter l’apparition d’une toxicité à long terme sans impacter significativement l’efficacité.La deuxième partie de mon projet de thèse a été consacrée à l’optimisation de l’efficacité des tcDNAs. Pour cela deux approches ont été investiguées : d’une part l’amélioration de la biodisponibilité de la molécule et d’autre part l’optimisation de la séquence cible. Nous avons ainsi pu démontrer que la conjugaison d’un acide gras à un tcDNA entraine une amélioriation significative de sa biodistribution et de l’efficacité du tcDNA. En parallèle, un criblage de nombreuses séquences ciblant différentes régions de l’exon 51 a permis de sélectionner une séquence candidate présentant une efficacité nettement supérieure à celle de la séquence initiale. Cette séquence, conjuguée à un acide palmitique, a démontrée des résultats extremement encourageants pour les futurs essais cliniques et est actuellement en phase finale de développement précliniqueDuchenne Muscular Dystrophy is a fatal genetic disorder affecting 1/3500 newborn males. It is characterized by progressive muscle weakness causing loss of ambulatory functions and respiratory and cardiac failures. This disease is due to mutations in the DMD gene leading to complete loss of protein expression. There is currently no satisfactory treatment but one of the most promising therapeutic strategy is splicing modulation. This strategy also called “exon skipping” is achieved through the use of antisense oligonucleotides allowing a restoration of the reading frame, and thus leading to protein rescue.The laboratory Biothérapie des Maladies du Système Neuromusculaire has developped a new chemistry of antisense oligonucleotide, tricyclo-DNA (tcDNA). They have demonstrated the therapeutic potential of tcDNA in different mouse models of DMD. Indeed, after systemic treatment significant exon 23 skipping and dystrophin restoration were found in all tested muscles as well as in the central nervous system, leading to functional improvement. During my phD project, I worked on the pre-clinical development of a tcDNA targeting human exon 51, which could be applicable to a large proportion of DMD patients (13%).The first part of my project was dedicated to the improvement of tcDNA tolerability through the modification of the sequence itself and the modification of the chemical design. Indeed, the major cause of tcDNA toxicity is the formation of homodimeric structure associated with the presence of phosphorothioates linkages (PS). In this study, we were able to demonstrate that modification of the toxic sequence impairs homodimerization, thus suppressing toxicity. Moreover, we have demonstrated that a decrease in the PS content prevent the apparition of long term toxicity without impairing significatively exon skipping efficacy but.The second part of my project focused on the optimisation of tcDNA efficacy through improvement of their biodisponibility and optimisation of the targeted sequence. We first demonstrated that fatty acid conjugation to tcDNA significantly improves biodistribution and efficacy. In parallel, we screened numerous sequences targeting different regions of the exon 51 and selected a novel sequence with a significantly higher efficacy than the initial sequence. This novel tcDNA sequence, once conjugated with palmitic acid demonstrated extremely encouraging results for the treatment of DMD and we are currently finalizing its development for future clinical trials
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Condin, Christopher J. "Families’ experiences with medical research for pediatric rare diseases : a qualitative ethnographic study of parents and children participating in clinical trials for Duchenne muscular dystrophy (DMD)." Thesis, University of British Columbia, 2014. http://hdl.handle.net/2429/50780.
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The biopharmaceutical industry has recently expanded its focus on developing new cures for rare diseases. As a growing number of personalised genomic treatments are tested in clinical trials, there is uncertainty about how to account for patient perspectives, and how to measure functional changes reported by patients and caregivers. The illness experiences of patients and families are also being reshaped as they adopt roles as collaborative stakeholders and participants in clinical studies.
This dissertation examines these changes using data from qualitative ethnographic research conducted with families of children with Duchenne muscular dystrophy, a progressive and fatal genetic disease diagnosed in boys. Canadian and American families were followed using semi-structured interviews and observational methods as they participated in clinical trials testing a genomic treatment for DMD, called ataluren (formerly known as PTC124). Ethnographic work was also carried out with physicians, patient-advocates, and other professionals engaged in clinical neuromuscular research.
The dissertation contributes to scholarly understanding of families’ everyday experiences in the clinical trial, the significance and meaning of investigational treatments from the patient perspective, and the social context in which pharmaceutical development for rare diseases occurs. I show how genetic research is reconfiguring patient communities and altering moral sensibilities about treatment and care, by revealing “lucky mutations” and new axes of biosocial commonality and difference. I explore the paths families take to the clinical trial, and the “stories of waiting” they tell about their experience in it. Finally, I examine how families navigate the uncertainty and liminality of their experience as trial subjects. I discuss how the trial unsettles taken-for-granted social roles, constraining clinical relationships and leaving parents to construct the significance of an experimental treatment in the context of limited information. In so doing, parents assemble and tell “narratives of efficacy” while administering study-drug to their children, drawing on their observations and those made by others. Though parents’ narratives are often dismissed as mere anecdote, I suggest they also offer insight for developing more personalised approaches to clinical research and outcome measurement for rare diseases, by restoring focus on the nuance, idiosyncrasy, and context of families’ experiences with investigational treatments.Arts, Faculty of
Anthropology, Department of
Graduate
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Abarca, Barriga Hugo Hernán. "Análisis retrospectivo de las características clínicas y moleculares de 40 pacientes con distrofia muscular de Duchenne y de Becker en el Hospital Nacional Guillermo Almenara Irigoyen, 1997 - 2007." Master's thesis, Universidad Nacional Mayor de San Marcos, 2016. https://hdl.handle.net/20.500.12672/5789.
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Publicación a texto completo no autorizada por el autorEl documento digital no refiere asesor
Determina las características clínicas y moleculares de pacientes con sospecha clínica de la distrofia muscular de Duchenne y de Becker en el Hospital Nacional Guillermo Almenara Irigoyen entre los años 1997 y 2007. Se recolecta la información clínica, análisis de laboratorio y estudios moleculares de 93 pacientes atendidos en dicho hospital.
Tesis
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Castillo, Andrea. ""Immortalizing a transient splendor" and the "beautiful totality" : Goethe's occasional works for Duchess Louise from 1777 to 1784 /." May be available electronically:, 2009. http://proquest.umi.com/login?COPT=REJTPTU1MTUmSU5UPTAmVkVSPTI=&clientId=12498.
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Lindvall, Mattias. "Studies towards a general method for attachment of a nuclear import signal. Stabilization of the m3G-Cap." Thesis, Mälardalen University, School of Sustainable Development of Society and Technology, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:mdh:diva-9728.
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A synthetic pathway towards the cap-structure of 2,2,7-trimethylguanosine containing a methylene modified triphosphate bridge have been investigated. The modification to the triphosphate bridge is hoped to slow down cap degradation and give the connected oligunucleotide an increased lifetime. This could result in an better understanding of nuclear transport of oligonucleotides and could thereby helping to develop new treatments for different diseases. The synthesis relies on a coupling reaction between the 2,2,7-trimethylguanosine 5’phosphate and 2’-O-methyladenosine with a 5’-pyrophosphate where the central oxygen has been replaced by a methylene group. The reaction pathway consists of 9 steps of which 8 steps have been successfully performed. The last step, which includes a coupling reaction, was attempted but without successful identification and isolation of the cap-structure, and will need further attention. The reaction has been performed in a milligram scale with various yields.
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Zalcman, Amy. "The Use of Magnetic Resonance Imaging and Proton Spectroscopy to Identify Critical Tissues in Dogs with Duchenne Muscular Dystrophy for Future Assessment of Therapeutic Intervention| A Pilot Study." Thesis, University of Missouri - Columbia, 2019. http://pqdtopen.proquest.com/#viewpdf?dispub=13850759.
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Duchenne’s Muscular Dystrophy is a debilitating disease that affects skeletal and cardiac muscle of 1 in 5000 male births. In the last thirty years, the gene responsible for the encoding of Dystrophin has been identified, sequenced and the variations of mutations described. There remains a void in the successful treatment of the disease although corticosteroid use has proven useful in delaying progression. Novel therapies are produced in the categories of virus-mediated gene delivery and stem cells, but evaluating their efficacy is hindered by an inability to contemporaneously assess the changes in muscle. The purpose of this pilot study was to characterize the changes in skeletal and cardiac muscle in a clinically advanced population of dogs affected with Duchenne Muscular Dystrophy. Using traditional sequences, delayed gadolinium enhancement, novel sequences and spectroscopy, changes in the investigated muscle were characterized. By establishing the differences between affected and unaffected dogs, the long-term goal of this body of work is to characterize these changes longitudinally and design a non-invasive method for tissue assessment as novel treatments are trialed.
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Lindvall, Mattias. "Studies towards a general method for attachment of a nuclear import signal. Stabilization of the m3G-Cap." Thesis, Mälardalens högskola, Akademin för hållbar samhälls- och teknikutveckling, 2010. http://urn.kb.se/resolve?urn=urn:nbn:se:mdh:diva-9728.
Full textAbstract:
A synthetic pathway towards the cap-structure of 2,2,7-trimethylguanosine containing a methylene modified triphosphate bridge have been investigated. The modification to the triphosphate bridge is hoped to slow down cap degradation and give the connected oligunucleotide an increased lifetime. This could result in an better understanding of nuclear transport of oligonucleotides and could thereby helping to develop new treatments for different diseases. The synthesis relies on a coupling reaction between the 2,2,7-trimethylguanosine 5’phosphate and 2’-O-methyladenosine with a 5’-pyrophosphate where the central oxygen has been replaced by a methylene group. The reaction pathway consists of 9 steps of which 8 steps have been successfully performed. The last step, which includes a coupling reaction, was attempted but without successful identification and isolation of the cap-structure, and will need further attention. The reaction has been performed in a milligram scale with various yields.Presentation utförd
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Simon, Valdecir Antonio. "Qualidade de vida em crianças e adolescentes com doenças neuromusculares e validação de dois questionários para o português: Life Satisfaction Index for Adolescents - LSI-A e Pediatric Quality of Life Inventory Duchenne Muscular Dystrophy Module." Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/5/5138/tde-06092016-160616/.
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INTRODUÇÃO: As distrofias musculares progressivas e a amiotrofia espinhal progressiva (AEP) são doenças neuromusculares (DNM) caracterizadas pela degeneração irreversível das fibras musculares, a qual leva à fraqueza muscular e à incapacidade motora. Qualidade de Vida Relacionada à Saúde (QVRS) inclui subjetividade, multidimensionalidade, aspectos negativos e positivos diante da percepção e da expectativa individual de vida; sofre influência cultural. JUSTIFICATIVA: A avaliação da QVRS é essencial para definir a resposta ao tratamento multidisciplinar ou efetivo do paciente com DNM e para sinalizar medidas destinadas a incrementar o sucesso terapêutico. OBJETIVOS: Validar os questionários Life Satisfaction Índex for Adolescents (LSI-A) versão pais e versão paciente e Pediatric Quality of Life Inventory Duchenne (PedsQL DMD) versão pais e versão paciente para o português; avaliar a QVRS dos pacientes com distrofia muscular de Duchenne (DMD), amiotrofia espinhal progressiva (AEP) ou distrofia muscular de cinturas (DMC); avaliar a QV familiar e da mãe/cuidadora. METODOLOGIA: Os questionários LSI-A e PedsQL DMD foram validados obedecendo às etapas de adaptação cultural e validação. Após validação, o questionário LSI-A foi aplicado a pacientes com DMD, AEP ou DMC; o PedsQL Duchenne foi aplicado aos pacientes com DMD e o PedsQL NM a pacientes com AEP ou DMC. Os pais dos pacientes responderam ao FQoL e as mães/cuidadoras ao WHOQOL-Bref. Para cálculo estatístico utilizaram-se: testes alfa de Cronbach, CIC, Pearson, Curva ROC para a validação, e Mann Whitney, Friedman e Dunn para a aplicação. RESULTADOS: Quanto à validação: Probe final do LSI-A versão pais, 97% e versão paciente, 95%; PesdQL DMD versão pais, 99% e versão paciente, 97%, sinalizando compreensão excelente; o teste ? de Cronbach no LSI-A versão pais e paciente, respectivamente, obteve escore geral 0.87 e 0.89; no PesdsQL versão pais e versão paciente, respectivamente, escore geral 0.87 e 0.84. Em ambos foi evidenciado alta confiabilidade dos itens. Quanto à aplicação do LSI-A versão pais e pacientes para avaliação da QVRS, quando comparada aos controles, houve maior número de domínios significantes em pacientes com DMD, AEP ou DMC, nesta ordem. A QVRS mediante aplicação do questionário PedsQL módulo DMD e NM obedeceu a esta mesma sequência. CONCLUSÕES: Conforme os dados psicométricos, os questionários são válidos para serem aplicados a pacientes com DNM e respectivos pais, como segue: LSI-A versão pais e versão paciente a pacientes com DMD, AEP e DMC e respectivos pais, e PedsQL 3.0 Duchenne versão pais e paciente a pacientes com DMD e respectivos pais. A QVRS apresentou-se mais satisfatória nos pacientes com DMC, seguidos pelos pacientes com AEP tipo II e III e, por último, pelos pacientes com DMD. A QV da família apresentou-se reduzida quanto aos aspectos relativos ao bem estar material, particularmente no caso das famílias dos pacientes com DMD. A QV das mães/cuidadoras, decresceu conforme o aumento da idade dos pacientes, quanto aos aspectos psicológicos, sociais e ambientais, em especial a das mães/cuidadoras dos pacientes com AEPINTRODUCTION: Progressive muscular dystrophies and spinal muscular atrophy (SMA) are neuromuscular diseases (NMD) characterized by irreversible degeneration of muscle fibers which leads to muscle weakness and motor disability. Health-related quality of life (HRQoL) includes subjectivity, multidimensionality, negative and positive aspects on the perception and individual life expectancy; in addition, it suffers cultural influences. BACKGROUND: The assessment of HRQoL is essential to define the response to the multidisciplinary or effective treatment of patients with NMD and to indicate measures to increase the therapeutic success. OBJECTIVES: to validate to the Portuguese the following HRQoL instruments for patients with NMD: Life Satisfaction Index for Adolescents (LSI-A) and Pediatric Quality of Life Inventory Duchenne (PedsQL Duchenne); to evaluate the HRQoL of patients with Duchenne muscular dystrophy (DMD), spinal muscular atrophy (SMA) or limb girdle muscular dystrophy (LGMD), and to assess the family and caregiver QoL. METHODOLOGY: The LSI-A and PedsQL Duchenne questionnaires were validated obeying the stages of cultural adaptation and validation. After validation, the LSI-A questionnaire was administered to patients with DMD, SMA or LGMD, the PedsQL Duchenne to patients with DMD, and the PedsQL NM to patients with SMA or LGMD. Parents of patients responded to FQoL and mothers/caregiver to WHOQOL-Bref. For statistical calculations were used: ? test Cronbach, CIC, Pearson, ROC curve for validation, and Mann Whitney, Friedman and Dunn for the application. RESULTS: Validation: the final \"Probe\" of the LSI-A parents version was 97% and patient version, 95%; PesdQL DMD parents version, 99% and patient version, 97%, indicating excellent comprehension; Cronbach\'s alfa test at LSI-A parents and patients version, respectively, achieved overall score 0.87 and 0.89; at PesdsQL parents and patient version, respectively, were obtained overall score 0.87 and 0.84. At both it was demonstrated high reliability of the items. At the application of LSI-A parents and patients version to measure HRQoL compared to controls, there was a greater number of significant dominions in DMD, SMA and LGMD, in that order. The PedsQL DMD module and NM followed the same sequence. CONCLUSIONS: According with the psychometric data, questionnaires are valid to be applied to parents and patients with NMD, as follows: LSI-A to parents and patients with DMD, SMA and LGMD, and PedsQL 3.0 Duchenne to patients with DMD and parents. The HRQL was more satisfactory in patients with LGMD, followed by patients with SMA and, finally, by DMD patients. The family QOL presented reduction of the aspects concerning material well-being, particularly for families of patients with DMD. The QoL of mothers decreased with the increase of the patients\' age, concerning the psychological, social and environmental aspects, in particular for the mothers of patients with SMA
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Douglas, Andrew Graham Lim. "Oligonucleotide-based therapies for neuromuscular disease." Thesis, University of Oxford, 2015. http://ora.ox.ac.uk/objects/uuid:d14706a3-c436-46ff-87c4-40bbbad6dc01.
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Vianello, Sara. "N-Butyryl arginine and 3-Hydroxybutyrate arginine, for the treatment of DMD through oral administration." Thesis, Paris 11, 2013. http://www.theses.fr/2013PA11T046/document.
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La dystrophie musculaire de Duchenne est une maladie neuromusculaire qui touche 1 enfant sur 3500, liée au chromosome X, caractérisée par l’absence de dystrophine, protéine située sous le sarcolemme qui confère stabilité à la membrane cellulaire en connectant l’actine du cytosquelette avec la matrice extracellulaire. Elle fait partie d’un complexe multi protéique, nommé « dystrophin associated protein complex (DAPC)», qui contient, entre autre, le -dystroglycane et l’oxyde nitrique synthase (NOS). Son absence cause la dérégulation de l’homéostasie calcique, la nécrose tissulaire, l’accumulation de tissu graisseux et fibreux, l’incapacité de mouvement et des déficits cardiaques et respiratoires qui aboutissent au décès des patients. Mon travail avait comme objectif l’amélioration de différents aspects du phénotype dystrophique. J’ai utilisé des molécules capables d’activer deux voies de signalisations (la voie du NO et l’inhibition des histones deacetylase (HDAC)), connues pour induire l’amélioration du phénotype dystrophique chez la souris mdx, modèle de la maladie. Plus particulièrement, j’ai testé chez la souris, deux mode d’administration du butyrate d’arginine (AB), la drogue de référence car déjà utilisée en clinique sur des jeunes patients pour une autre indication, par gavage et par injection intrapéritonéale. J’ai étudié aussi deux nouvelles molécules dérivées du AB, qui pourraient être administrées par voie orale et être efficace à faible dose : le 3-Hydroxybutyrate arginate (ABE) et le N-butyril arginine (ABA). AB, ABE et ABA ont été testés in vitro sur les myotubes de patients dystrophiques et in vivo sur des souris mdx. L’administration orale du AB a les mêmes effets positifs que l’injection intrapéritonéale chez les souris mdx. Ces résultats démontrent que l’administration par voie orale doit être prise en considération lors des futurs essais cliniques. Dans un deuxième temps, je me suis focalisée sur les défauts cardiaques. Un suivi par échocardiographie mensuelle a été réalisé sur des souris de 8 mois traitées avec du AB. En parallèle nous avons analysé les effets de l’administration par voie orale du AB sur les déformations de la colonne vertébrale. Enfin, les altérations des signaux de l’électromyogramme (réalisé avec une méthode non invasive développée en clinique et appliquée pour les animaux) ont été également analysées. L’ensemble des résultats obtenus montre que le AB est capable de préserver l’activité cardiaque, d’empêcher la déformation de la colonne vertébrale et de rétablir les paramètres d’excitabilité axonale mesurés chez les souris traitées.Différentes concentrations des ABE et ABA ont été testé in vivo et observé à des faibles doses les mêmes résultats bénéfiques sur de nombreux paramètres structuraux et fonctionnels, que ceux obtenu avec une dose importante de AB (800mg/kg/j). Les deux nouvelles drogues peuvent être administrées à une dose 10 fois inferieur que la dose de AB pour obtenir les mêmes effets. J’ai testé aussi in vitro, sur des cellules musculaires humaines, la capacité des deux produits à induire une augmentation des niveaux intracellulaires d’utrophine et des protéines associées (β-dystroglycan et la myosine embryonnaire). J’ai aussi démontré qu’une augmentation de l’expression de l’utrophine et des protéines associées pouvait être induite par les inhibiteurs d’HDAC (le butyrate, la trichostatine A, l’acide valproique et l’isobutyramide). Enfin, une étude portant sur l’homéostasie calcique a été réalisé car des altérations de cet équilibre sont en partie responsables de la nécrose/dégénérescence du tissue musculaire. En particulier, l’activité spontanée du Ca2+, enregistrée sur le myotubes humaine, été fortement réduite après un traitement agissant sur la voie d’activation du NO et/ou par des inhibiteurs des HDAC. L’ensemble des résultats obtenus apportent la preuve des effets bénéfiques du AB et de ses dérivés sur la DMD, a travers la voie du NO et en inhibant les HDACDuchenne muscular dystrophy is a X-linked progressive neuromuscular disease affecting 1:3500 boys at birth. It is caused by the absence of dystrophin, a subsarcolemmal protein that confers membrane stability linking cytoskeletal actin to the extracellular matrix. It is part of a multi-protein complex called dystrophin associated protein complex (DAPC), which contains, among the other components, -dystroglycan and nitric oxide synthase (NOS).The consequences of the absence of dystrophin are: deregulation of calcium homeostasis, tissues necrosis, progressive accumulation of fat and fibrosis, inability of the movements and cardiac and respiratory failures that lead to patient’s death, around the age of 20-30 years.The objective of my PhD work is to ameliorate different aspects of dystrophic phenotype. In particular I have tested two different ways of administration of arginine butyrate (AB), the reference drug, through feeding-force and intraperitoneal injection. Meanwhile I have studied two new pharmacological molecules, AB derived, which could be administered orally to DMD patients. These compounds are: 3-Hydroxybutyrate arginate (refer as ABE) and N-butyryl arginine (refer as ABA). All of these molecules partially restore dystrophic phenotype activating two independent pathways (both the nitric oxide pathway and the inhibition of the histone deacetilase), which are known to be beneficent for mdx mice.AB, ABE and ABA have been tested in vitro on human DMD myotubes and in vivo on the mdx mice. The first goal of my project is the observation that the positive effects obtained after intraperitoneal injections of AB can be detected also after oral protocol, promoting the idea that the oral way has to be developed for future clinical trials. I have focused my attention on heart defaults; in particular, starting from the 8th month, a monthly study on heart activity based on echocardiography has been performed on mdx mice treated with AB. We addressed the potential profits of the oral administration of arginine butyrate on vertebral column deformation and electromyogram defaults, with a non-invasive automatized method developed in clinic and then applied to animals. The results collected from these experiments show that AB preserve heart activity, reverse vertebral column deformity and all the axonal excitability parameters that were modified in saline-treated mdx mice.In complement, I have tested different concentrations of ABE and ABA in vivo. The positive effects on many structural and functional dystrophic parameters, previously obtained with high dose of AB administered per os (800 mg/kg/d), has been observed with doses 10 times lower with both new compounds.In parallel, both products were tested in vitro on human muscular cells cultures to investigate their capacity to increase utrophin level. Moreover, the potential ability of histone deacetylase inhibitors (byturate, valproic acid, trichostatin A and isobutyramide) to increase the expression of utrophin and related proteins (-dystroglycan and embryonic myosin) has been studied. Finally, the alteration of calcium homeostasis, largely implicated in the cascades resulting in muscle necrosis/degeneration, was investigated. The spontaneous Ca2+ activity recorded in patient myotubes, i.e. without sarcolemmal integrity was strongly reduced after treatment acting on the NO-pathway activation and/or with HDAC inhibitors. All together, these data constitute a proof of principle of the beneficial effects of arginine butyrate and its derivates on muscular dystrophy, by enhancing NO pathway and inhibiting HDAC