INTRODUÇÃO: A isquemia da medula espinhal continua sendo uma importante complicação nas cirurgias das doenças da aorta descendente torácica e toracoabdominal. OBJETIVOS: Células-tronco são capazes de promover a regeneração do tecido nervoso. Células-tronco derivadas do cordão umbilical humano (CTCUH) são fortes candidatas para uso nas lesões da medula espinhal devido à sua baixa imunogenicidade e pronta disponibilidade. O estudo avaliou os efeitos da administração de CTCUH na lesão isquêmica da medula espinhal em ratos. MÉTODOS: Quarenta ratos Wistar receberam injeção intratecal de 10 uL de solução de HemoHes (6 %) e albumina humana (20 %) contendo 1x104 CTCUH, 30 minutos antes (grupo Tcpré; n=10) e 30 minutos após (grupo Tcpós n=10) oclusão da aorta torácica descendente através de um balão intraluminal por 12 minutos. Os grupos controle receberam apenas a solução de Hemohes (6 %) e albumina humana (20 %) (grupo Cpré; n=10 e grupo Cpós; n=10). O período observacional, para avaliação da função motora dos animais, foi de 28 dias. Cortes de três segmentos tóraco-lombares da medula espinhal foram submetidos à análise histológica e imunohistoquímica para detecção de apoptose (TUNEL) e quantificação de células-tronco humanas hematopoiéticas CD45 +. RESULTADOS: Todos os grupos mostraram incidência semelhante de paraplegia e mortalidade. A média de pontuação da função motora não mostrou diferença durante o período observacional nos grupos, com exceção do grupo Tcpós o qual melhorou de 8,14 ± 8,6 para 14,28 ± 9,8 (p < 0,01). Número de neurônios viáveis foi maior no grupo Tcpós (p=0,14) e a média de apoptose foi mais baixa nesse mesmo grupo (p=0,048), porém sem diferença estatística significativa em relação ao controle. Foi confirmada a presença de células CD45 + quatro semanas após a injeção intratecal em ambos os grupos terapêuticos, principalmente, no grupo Tcpós. CONCLUSÕES: A injeção intratecal de CTCUH é factível e melhora a função motora da medula espinhal em um modelo de oclusão endovascular da aorta torácica descendente / BACKGROUND: Spinal cord ischemia remains a complication after surgical repair of descending and thoracoabdominal aortic diseases. OBJECTIVES: Stem cells have the potential to induce nervous tissue regeneration processes. Human stem cells derived from the umbilical cord are one of strong candidates used in cell therapy for spinal cord injury due to weak immunogenicity and ready availability. We sought to evaluate the use of Human Umbilical Cord Blood Stem Cells (HUCBSC) attenuates the neurologic effects of spinal cord ischemia. METHODS: Fourty Wistar rats received intrathecally injection of 10 uL Hemohes (6 %) and human albumin (20 %) solution contained 1x104HUCBSC, 30 minutes before (Tcpré group; n=10) and 30 minutes after (Tcpós group; n=10) descending thoracic aortic occlusion by intraluminal balloon during 12 minutes. Control groups received only PBS solution (Cpré group; n=10 and Cpós group; n=10). During a 28-day observational period, animals motor function was assessed. Three segments of thoraco-lombar spinal cord specimens were analyzed for histologic and immunohistochemical assessment for detection and quantification of human hematopoietic cells CD45+ and apoptosis (TUNEL). RESULTS: All groups showed similar incidence of paraplegia and mortality. The mean motor function scores showed no difference during time, excepting for Tpos group which improved from 8.14(8.6) to 14.28(9.8)(p < 0,01). Number of viable neurons was higher in Tcpós group (p = 0.14) and apoptosis average was lower in the same animals (p = 0.048), but showed no difference with its respective control. We confirmed the presence of CD45+ cells four weeks after intrathecal injection in both therapeutic groups but mainly in Tpos group. CONCLUSIONS: Intrathecal transplantation of HUCBSC is feasible and improved spinal cord function in a model of endovascular descending aortic occlusion

INTRODUÇÃO: A lesão medular isquêmica continua sendo uma complicação devastadora das intervenções cirúrgicas na aorta torácica descendente e aorta toracoabdominal. Relatos das diferenças de desfechos clínicos neurológicos entre os gêneros após lesões cerebrais isquêmicas e traumáticas têm levantado o interesse nas influências hormonais, bem como gerado outros estudos buscando a comprovação dos efeitos neuroprotetores do estradiol e da progesterona. Nossa hipótese foi a de que a administração aguda de 17beta-estradiol ou de progesterona seria capaz de prevenir ou atenuar a lesão medular isquêmica causada pela oclusão transitória da aorta torácica descendente proximal. OBJETIVO: Analisar os efeitos na medula espinhal da administração aguda de 17?-estradiol ou de progesterona em modelo experimental de isquemia-reperfusão medular por oclusão transitória da aorta torácica descente proximal de ratos machos. MÉTODOS: Ratos machos, da linhagem wistar, foram divididos aleatoriamente em 3 grupos para a administração de 280ug/Kg de 17beta-estradiol (n=12) ou de 4mg/Kg de progesterona (n=8) ou do veículo de infusão (grupo controle) (n=12), 30 minutos antes da oclusão transitória da aorta torácica descendente por 12 minutos. A confirmação da oclusão efetiva aórtica deu-se por meio da monitorização contínua da pressão arterial média distal com o uso de cateter colocado na artéria caudal dos animais (mantida em 10mmHg). A oclusão da aorta torácica descendente deu-se por meio do posicionamento de um cateter de Fogarty no. 2, passado no sentido caudal, via dissecção da artéria carótida comum esquerda do animal. A função locomotora dos animais foi avaliada no 1o, 3o, 5o, 7o, e 14o dia pós-operatório. No 14o dia pós-operatório, os animais, após anestesia profunda, foram sacrificados e tiveram suas medulas espinhais retiradas para análise histológica e imunohistoquímica. RESULTADOS: Houve comprometimento significativo da função locomotora inicialmente nos 3 grupos de estudo, com recuperação parcial da mesma ao longo do período de observação, não havendo diferença entre os grupos durante o período de observação. A análise histológica da substância cinzenta evidenciou escassos neurônios viáveis e importante vacuolização celular nos 3 grupos de estudo no 14o dia. A análise imunohistoquímica da substância cinzenta medular com anticorpos anti-Bcl2 e anti-anexina V foi similar nos 3 grupos. Houve marcação positiva de necrose celular com o iodeto de propídio, sendo a mesma semelhante nos 3 grupos estudados. CONCLUSÃO: A administração aguda de estradiol ou de progesterona, 30 minutos antes da oclusão transitória da aorta descendente proximal de ratos machos não foi capaz de prevenir ou atenuar a lesão medular isquêmica, até o 14o dia de observação, do ponto vista funcional ou histológico / BACKGROUND: Spinal cord ischemic injury remains a dreadful complication following thoracic and thoracoabdominal aortic interventions. Reports on gender-related neurological outcomes after ischemic and traumatic brain injuries have raised interest in hormonal influences, and have generated studies into neuroprotective effects of estrogen and progesterone. We hypothesized that the acute pre-operative administration of estradiol or of progesterone would prevent or attenuate spinal cord ischemic injury induced by transitory occlusion of the proximal descending thoracic aorta. OBJECTIVE: Evaluate the spinal cord effects of the acute administration of 17beta-estradiol or of progesterone in a spinal cord ischemia-reperfusion model. METHODS: Male rats were divided to receive 280ug/Kg of 17beta-estradiol (n=12) or 4mg/Kg of progesterone (n=8) or vehicle (control group) (n=12) 30 minutes before transitory occlusion of the proximal descending thoracic aorta, mean distal arterial blood pressure was maintained at 10mmHg during 12 minutes. Hind limb motor function was assessed at 1, 3, 5, 7 and 14 days after reperfusion. At the 14th day, a segment of the thoracolumbar spinal cord was harvested and prepared to histological and imunohistochemical analyses. RESULTS: There was an important hind limb motor function impairment initially in the 3 study groups, with partial improvement along time, but no difference was detected between groups during de observational period. Gray matter analysis showed scarce viable neurons and a marked cellular vacuolation in all three groups, but the number of viable neurons per section areas was not different between study groups at day 14th. Immunostaining of the spinal cord gray matter with antibodies anti-Bcl2 and anti-annexin V was similar among the 3 study groups. There was positive staining for the necrotic marker propidium iodide, with all groups presenting a similar staining pattern. CONCLUSION: We found that a single-dose administration of estradiol or of progesterone, 30 minutes before transitory occlusion of the proximal descending thoracic aorta of male rats, was not able to prevent spinal cord ischemic injury through analysis of functional and histological outcomes at 14 days of observation

Introdução: A síndrome Spoan é uma forma de paraplegia espástica complicada de herança recessiva recentemente identificada em indivíduos originários do sudoeste do estado do Rio Grande do Norte. O quadro clínico é caracterizado por atrofia óptica congênita, paraplegia crural espástica de caráter progressivo e neuropatia axonal levando a perda da função motora em membros superiores. A caracterização fenotípica dessa doença não está completa, e não foram realizados estudos quantitativos e funcionais, que poderiam mensurar a intensidade e contribuir para a definição de uma estratégia de reabilitação. Objetivos: caracterizar o desempenho motor e as habilidades funcionais de indivíduos acometidos pela Spoan. Casuística e metodologia: participaram do estudo 61 indivíduos com diagnóstico clínico de Spoan com idade entre 5 e 72 anos. Avaliou-se a força de preensão palmar por meio do dinamômetro hidráulico de Jamar e a sensibilidade a pressão profunda e protetora dos pés e mãos por meio dos monofilamentos de náilon de Semmes-Weinstein. Definiu-se o grau de dependência dos indivíduos afetados por meio do Índice de Barthel modificado. Considerou-se para a descrição do desempenho motor: 1. quantificação da espasticidade, por meio da escala modificada de Ashworth; 2. grau de disfunção, de acordo com a escala ponderada de paraplegia espástica descrita por Schule e a escala funcional de paraplegia espástica hereditária descrita por Fink; 3. grau da capacidade de deambulação, por meio do índice de deambulação 4. grau da capacidade de sentar, por meio da escala de avaliação motora. Resultados: constatou-se fraqueza de preensão manual em todos os indivíduos e os valores obtidos indicam correlação inversa moderada entre a idade e a força manual. A sensibilidade mostrava-se anormal em 100% dos indivíduos avaliados em pelo menos seis pontos dos pés e mãos. O grau de dependência foi mínimo em 3,3%, médio em 23,3%, grave em 46,6% e total em 26,6% dos pacientes. Na escala de Schule, 60% dos indivíduos obtiveram entre 40/52 e 52/52 pontos e na escala de Fink detectou-se grau 5 (máximo) de disfunção em 71% dos pacientes. O grau de espasticidade teve uma distribuição bimodal, em média, de 30,5% com grau 1 e 37,7% grau 4. A capacidade de deambulação mostrou-se reduzida, com 83% dos indivíduos restritos a cadeira de rodas e 11% acamados. A habilidade de sentar-se estava preservada em todos os pacientes, sendo que 53% o faziam apenas com apoio. Conclusão: A síndrome Spoan é uma forma grave de paraplegia espástica hereditária, responsável por incapacidade progressiva e duradoura. / INTRODUCTION: Spoan syndrome is a complex form of spastic paraplegia of recessive inheritance recently identified in individuals from Southwest of Rio Grande do Norte state. Clinical features are characterized by congenital optic atrophy, progressive spastic paraplegia, and axonal neuropathy, resulting in severe handicap. Phenotypic description of this disease is nevertheless not complete; functional and quantitative studies, that would help planning a rehabilitation strategy, have not been undertaken. OBJECTIVES: To evaluate the motor performance and functional abilities of individual with Spoan syndrome. CASUISTIC AND METHODS: 61 individuals with confirmed diagnosis of Spoan, with ages ranging from 5 and 72 years were evaluated. Hand grip strength was measured with a Jamar hydraulic dynamometer and the sensitivity to deep pressure and protective hands and feet with Semmes-Weinstein nylon monofilaments. Functional abilities were verified by the Modified Barthel Index. For motor performance, the following procedures were performed: 1. Spasticity quantification, according to modified Ashworth scale; 2. Dysfunction level, according to the spastic paraplegia rating scale described by Schule and functional scale of hereditary spastic paraplegia described by Fink; 3. Gait ability, verified with deambulation index; 4.Sitting ability, using motor assessment scale. RESULTS: grip hand weakness was reduced in all patients, with a moderate inverse correlation between age and hand strength. Sensibility was abnormal in 100% of evaluated individuals in at least six points of hands and feet. Dependency level was minimum in 3.3%, moderate in 23.3%, severe in 46.6%, and total in 26.6% of individuals. According to Schule s scale , 60% of individuals scored between 40/52 and 52/52 points; in Fink s scale,71% achieved level 5 (maximum) of dysfunction. Spasticity level had a bimodal distribution, with 30,5% achieving level 1 and 37,7% level 4. Gait ability was reduced, with 83% of individuals being wheelchair bound and 11% bedridden. Sitting ability was preserved in all patients, but 53% were able to sit only with support. CONCLUSION: Spoan syndrome is a severe form of hereditary spastic paraplegia that is responsible for progressive and long lasting handicap.

Thesis (MSc (Interdisciplinary Health Sciences. Speech-Language and Hearing Therapy))--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: Spinal cord injury is a devastating sudden cause of disability which renders a person paralyzed and dependent on care immediately after the incident. A person who has suffered a spinal cord injury requires an intensive rehabilitation program to achieve physical independence as well as reintegration into the community. The aim of this study was to evaluate the rehabilitation program for patients with complete paraplegia at the Netcare Rehabilitation Hospital, a 120 bed private rehabilitation facility in Johannesburg, where rehabilitation for patients with physical disabilities is offered by an interdisciplinary team. The rehabilitation program was evaluated in terms of effectiveness including the degree of physical independence the patients achieved and how well the program prepared patients for successful integration into the community. The Functional Independence Measure (FIM), a standardized outcome measure that measures certain physical and cognitive functions and the Needs Assessment Checklist (NAC), an instrument designed to assess if the rehabilitation program is geared towards the patients’ individual needs were used as outcome measure. FIM scores were determined on admission and discharge and NAC scores were determined at discharge. A convenient, consecutive sample of sixteen patients with complete paraplegia (ASIA A), with a neurological level between T1 and T12, formed the study population. Consistently high FIM and NAC scores in the area of physical functioning suggested that the rehabilitation program at Netcare Rehabilitation Hospital was effective in terms of providing patients with physical independence. However, NAC results showed that the patients were not completely ready to reintegrate back to their communities. Areas that were identified as particularly problematic were knowledge on follow up health care services in the community, readiness for work and accessibility of the home and work environments. Therefore it is recommended that the rehabilitation team re-evaluate the program and incorporate strategies with the aim to improve it’s effectiveness in terms of preparing patients for community reintegration. It is also recommended that further research is conducted to assess the current success rate with regards to community reintegration and determine challenges to re-integration in order to assist with program panning. / AFRIKAANSE OPSOMMING: ‘n Spinaalkoordbesering is ‘n skielike oorsaak van gestremdheid wat die persoon verlam en afhanklik van sorg laat direk na die voorval. ‘n Persoon wat ‘n spinalkoordbesering opdoen benodig ‘n intensiewe rehabilitasie program om weer fisies onafhanklik te wees en ook om hom/haar voor te berei om weer by die gemeenskap in te skakel. Die doel van hierdie studie was om die rehabilitasie program by die Netcare Rehabilitasie Hospitaal, ‘n 120 bed private rehabilitasie fasiliteit in Johannesburg, waar rehabilitasie vir fisies gestremde persone deur ‘n interdissiplinere span aangebied word, te evalueer. Die program is geevalueer in terme van twee uitkomste naamlik, die graad van fisiese onafhanklikheid wat die pasiënte behaal en tot watter mate die program pasiente voorberei vir herintegrasie in die gemeenskap Die “Functional Independence Measure” (FIM), ‘n gestandaardiseerde uitkoms skaal wat sekere fisiese en kognitiewe funksies meet en die “Needs Assessment Checklist” (NAC), `n instrument wat ontwikkel is om te evalueer of rehabilitasie programme pasiente se spesifieke behoeftes aanspreek. Is gebruik om the rehabilitasie program te evalueer. Die FIM is met toelating en ontslag voltooi en die NAC is met ontslag voltooi. ‘n Steekproef van sestien pasiente met volledige spinaalkoordletsels (ASIA A) tussen T1 en T12 het die studie populasie gevorm. Deurgans hoë FIM en NAC tellings vir fisiese funksionering het daarop gedui dat die rehabilitasie program by die Netcare Rehabilitation Hospital effektief is in terme van fisiese onafhanklikheid van pasiente. Aan die ander kant het die NAC tellings daarop gedui dat die program minder suksesvol is wat betref die voorbereiding van pasiente vir gemeeskapsintegrasie. Spesifieke probleem areas sluit in kennis van waar om opvolg gesondheidssorg in die gemeenskap te bekom, gereedheid vir werk en toeganklikheid van die huis en werksomgewing. Daar word aanbeveel dat die rehabilitasie span by Netcare Rehabilitasie Hospitaal die program herevalueer om die effektiwiteit van die program in terme van die voorbereiding vir gemeenskaps herintegrasie te verbeter. Dit word ook aanbeveel dat ‘n opvolg studie gedoen word om die huidige sukses ten opsigte van gemeenskaps herintegrasie te evalueer en sruikelblokke te identifiseer ten einde die span the help met program beplanning.

Au cours des dernières années, la génétique a subi une progression phénoménale suite au développement de nouvelles technologies de séquençage. En effet, le séquençage de l’exome entier chez des familles a permis l’identification de nouveaux gènes impliqués pour plusieurs maladies. La neurologie a d’ailleurs bénéficié de ces avancées et plusieurs gènes ont été mis en évidence comme causatifs pour différents désordres neurologiques. Dans ce travail il sera question de deux désordres du mouvement pour lequel nous avons utilisés des technologies de séquençage traditionnelles, en l’occurrence le séquençage par Sanger, ainsi que de nouvelles technologies pour le séquençage de l’exome entier afin d’identifier de nouveaux gènes causatifs. Le premier désordre du mouvement qui sera décrit est l’ataxie, où ne seront abordées que les ataxies de cause génétiques, à transmission récessive. Le premier chapitre relatera les nouvelles mutations qui ont été trouvées chez des canadiens-français souffrant de l’ataxie de Beauce. Il sera aussi question de nouvelles mutations retrouvées dans deux autres populations, confirmant l’implication du gène SYNE1 dans les cas d’ataxie cérébelleuse à travers le monde. Le second chapitre fera la démonstration qu’il est souhaitable d’utiliser le séquençage de l’exome entier dans le but de poser un diagnostic clinique. En effet, il a été possible de trouver la cause génétique d’une famille comportant deux membres atteints d’atrophie congénitale du cervelet, où le symptôme prédominant est l’ataxie. Le séquençage de l’exome a permis la mise en évidence de mutations dans le gène PMM2, déjà connues pour cause le syndrome des glycoprotéines déficientes en hydrates de carbone. Dans un second temps, il sera question d’un autre désordre du mouvement la paraplégie spastique familiale (PSF). Le chapitre 3 relatera les mutations trouvées dans le gène CYP7B1 dans notre cohorte de patients PSF. / Over the past years, genetics has undergone a phenomenal growth due to the development of new sequencing technologies. Indeed, whole exome sequencing in families led to the identification of new genes involved in many diseases. Neurosciences were also able to benefit from these discoveries, where several new genes have been identified in several neurological diseases. This thesis will covered two different movement disorders for which we have used traditional sequencing technology, in this case by Sanger sequencing, combined with whole exome sequencing for new gene discovery. The first movement disorder that is described is ataxia, which will be focused on autosomal recessive mode of inheritance. The first chapter will relate the new mutations found in French-Canadian with ataxia of Beauce. We will also discuss new mutations found in two other populations, confirming the involvement of the gene SYNE1 in worldwide cases of cerebellar ataxia. The second chapter will demonstrate that it is desirable to use whole exome sequencing in order to make a clinical diagnosis. Indeed, it has been possible to find the genetic cause for a family with two members with congenital cerebellar atrophy, which the predominant symptom is ataxia. Exome sequencing allowed the identification of mutations in the PMM2 gene, already known to cause a syndrome leading to glycosylation deficit of glycoprotein. For the second part, we will cover another movement disorder call hereditary spastic paraplegia (HSP). Chapter 3 relates the mutations found in the CYP7B1 gene in our cohort of HSP patients.

Author: Bc. Pavlikhina Ekaterina Title: Influence of the posture's change on respiratory function in spinal cord injury patients. Objectives: The purpose of this thesis is to analyze change in respiratory function on change of posture in patients with spinal cord injury (SCI). Another objective is to compare the respiratory functions in a health subjects and in a SCI patients. Last but not least is an effort to find out whether SCI patients have a pattern of restrictive pulmonary dysfunction. The findings can help understanding development of respiratory complications after spinal cord injury. It may help in choosing optimal position during respiratory examination and respiratory rehabilitation. Methods: In theoretical part, there were explored the available knowledges about respiratory complications and their effects on posture. In practical part, there were 26 subjects - 16 patients with SCI and 10 healthy subjects. Subjects with SCI were divided into two groups - one with paraplegia and other one with quadriplegia. Measurements were performed with spirometer on each subject during supine, sitting and standing postures. There were collected vital capacity (VC), forced vital capacity (FVC) and forced expiratory volume in first second (FEV1) data from all subjects. All data were processed in...

Orientador: Michael John Brennan / Resumo: A independência funcional para indivíduos com deficiência motora é indispensável para a qualidade de vida, principalmente quando se considera tarefas como o autocuidado (incluindo a higiene pessoal), as transferências e locomoção. Essa pesquisa apresenta um triciclo inovador para pessoas com paraplegia ou alguma deficiência motora dos membros inferiores, desenvolvido com duas forma de acionamento: cíclico manual (handbike) e elétrico. A estrutura tubular foi elaborada com liga de alumínio 6063 - T5. A plataforma ANSYS Academic versão 19.2 foi empregada para uma análise estrutural do chassi do equipamento. O chassi foi submetido num ensaio de carregamento estático progressivo de até 1200 N aproximadamente. O deslocamento onde apoia os pés do usuário foi medido por um relógio comparador de precisão de 0,01 mm. O deslocamento da plataforma inferior de apoio dos pés apresentou uma divergência de aproximadamente 4% se comparado com os dados obtidos pela plataforma ANSYS. Assim, validando a análise teórica pela plataforma ANSYS e confirmando que a estrutura fabricada por dobramento e solda TIG suporta com segurança um usuário de até 100 kg. Parâmetros relacionados ao sistema elétrico de acionamento do motor brushless de 1000 W foram obtidos por meio de sensores de corrente e voltagem, assim como parâmetros de desempenho mecânico foram obtidos empregando uma célula de carga de fundo de escala de 2kN para análise de força de tração e um sensor de velocidade. Os resultados indicam uma... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Functional independence for individuals with motor disabilities is essential for quality of life, especially when considering tasks such as self-care (including personal hygiene), transfers and locomotion. This research presents an innovative tricycle for people with motor deficiency of the lower limbs, developed with two forms of drive: manual cyclic (handbike) and electric. The tubular structure was made with 6063 - T5 aluminum alloy. The ANSYS Academic version 19.2 platform was used for a structural analysis of the equipment chassis. A static loading test of up to 1200 N was performed on the chassis. The deformation of support the user's feet was measured by a dial gauge with a 0,01 mm precision. The deformation of the lower footrest platform showed a divergence of approximately 4% when compared to the data obtained by the ANSYS platform. Thus, validating the theoretical analysis by the ANSYS platform and confirming that the structure manufactured by TIG welding safely supports a user of up to 100 kg. Parameters related to the electric drive system of the 1000 W brushless motor were obtained by means of current and voltage sensors, as well as mechanical performance parameters were obtained employing a load cell of 2kN for force analysis and a speed sensor. The results indicate an energy efficiency of the equipment of up to 92%. The maximum strength of the equipment is 500 N, which makes it possible to perform a climb with slope of 20º, approximately considering a user of 8... (Complete abstract click electronic access below) / Doutor

The purpose of the present investigation was to: 1) Determine the relationship between cardiac output (estimated using the acetylene rebreathing methodology) and oxygen consumption in a homogeneous group of men who are paraplegic. 2) Investigate whether lower limb elevation increases stroke volume and decreases heart rate at rest and during submaximal arm exercise. 3) Investigate whether the application of constant circumferential pneumatic pressure applied to dependent lower limbs increases stroke volume and decreases heart rate at rest and during submaximal arm exercise.

How organelles acquire their unique shapes is a fundamental question of cell biology. The peripheral endoplasmic reticulum (ER) consists of a vast network of membrane sheets and tubules, the formation of which requires homotypic membrane fusion. Previous studies suggest that the dynamin-like GTPase, atlastin (ATL), mediates ER fusion, but the mechanism by which this occurs is unclear. In this study, I investigate 1) the role of dimerization and conformational changes in the N-terminal domain of ATL, 2) how the C-terminal amphipathic helix and the transmembrane domain of ATL cooperate with the N-terminal domain, and 3) the formation of cis and trans ATL dimers in the fusion mechanism. ATL has a cytosolic N-terminal domain, consisting of a GTPase domain and three-helix bundle (3HB), followed by two transmembrane segments (TMs) and a cytosolic C-terminal tail (CT). Crystal structures of ATL and biochemical experiments suggest that nucleotide-dependent dimerization between ATL molecules sitting in different membranes can tether the membranes together. A subsequent conformational change triggered by GTP hydrolysis could pull the membranes toward one another for fusion. This mechanism is supported by in vitro membrane tethering and fusion assays using vesicles containing full-length Drosophila ATL. The CT and TMs of ATL are also required for efficient membrane fusion. A synthetic peptide corresponding to a conserved amphipathic helix in the CT can act in trans to restore the fusion activity of a tailless ATL mutant. We characterize CT mutants to show that the C-terminal helix promotes fusion by perturbing the lipid bilayer. The TMs of ATL also mediate nucleotide-independent oligomerization, which may allow ATL molecules in the same membrane to synchronously undergo the conformational change leading to fusion. Lastly, we show that continuous GTP hydrolysis is required for membrane tethering, occasionally resulting in fusion. The N-terminal cytosolic domain mediates trans dimer formation between ATL molecules on different membranes. GTP binding induces dimerization through the GTPase domains and 3HBs. We propose that GTP hydrolysis and phosphate release are required not just to drive fusion, but also to dissociate cis dimers that form on the same membrane, thus allowing ATL molecules to form trans dimers.

The purpose of this thesis was to examine coronary heart disease (CHD) risk factors and secondary complications in individuals with spinal cord injury (SCI). In particular, this thesis was organized around the central theme of adiposity, which is a prevalent complication following SCI. Study 1 focused on understanding the relationships between activities of daily living (ADL) and CHD risk factors including central adiposity, lipoproteins, and triglycerides. Using generalized linear models, while controlling for pertinent covariates such as sex, age, and leisure time physical activity (LTPA), it was found that Mobility ADL (wheeling and transferring) were negatively associated with total and LDL-cholesterol. Study 2 examined whether individuals who considered themselves to be overweight subsequently had less favourable subjective well-being, and were more likely to report specific secondary complications than individuals who did not consider themselves to be overweight. Logistic regression analysis and partial correlations controlling for pertinent covariates such as sex, age, and injury severity, revealed that individuals who considered themselves to be overweight reported greater pain, depression, overuse injuries, and fatigue, and less satisfaction with life than individuals who did not consider themselves to be overweight. In summary, the findings suggest that a) participation in specific types of ADL (i.e. Mobility ADL) are associated with a lower CHD risk and should be further explored and that b) elevated perceived adiposity is associated with specific secondary complications and lower subjective well-being. Overall thesis findings support the overwhelming evidence of the benefits of daily physical activity and maintaining a healthy bodyweight in the SCI population. / Thesis (Master, Kinesiology & Health Studies) -- Queen's University, 2009-05-28 11:40:32.574