Physical activity can prevent cardiovascular disease, obesity, type II diabetes and some types of cancer. With only 3.5% of adults meeting the recommended physical activity guidelines, research has focused on the regulatory factors that influence physical activity level. Genetic influence accounts for the majority of physical activity regulation. However, there is limited information on the mechanisms that affect physical activity, in part, because of a lack of reliable methods to silence genes in vivo. The purpose of this dissertation was to identify mechanisms in skeletal muscle that influence physical activity. The methods used to accomplish the purpose of this dissertation were the evaluation of Vivo-morpholinos as a gene silencing tool in skeletal muscle and brain, identification of proteins in skeletal muscle associated with increased physical activity level, and the use Vivo-morpholinos to transiently knockdown the identified skeletal muscle proteins as a means to elucidate mechanisms for the peripheral regulation of physical activity. Overall, this study showed that Vivo-morpholinos effectively silenced genes in skeletal muscle yet required the use of a pharmacological aid to achieve gene silencing in the brain. Additionally proteins associated with calcium regulation (Annexin A6 and Calsequestrin 1) and the Kreb’s (TCA) cycle were found to be over expressed in the high active animals. The knockdown of Annexin A6 and Calsequestrin 1 resulted in a significant decrease in physical activity, thus showing that calcium regulation could influence the physical activity response. While these results provide a potential mechanism for the peripheral regulation of physical activity, a side effect observed was that Vivo-morpholinos can hybridize resulting in increased mortality rates of the treatment animals. Therefore, we developed methods to alleviate the toxic effects of Vivo-morpholinos. Thus, this dissertation refined a technique for determining a gene’s effect in an in vivo model and identified two candidate proteins (Annexin A6 and Calsequestrin 1) that play a role in regulating daily physical activity.

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No. of bitstreams: 1 silvajunior_ot_me_bauru.pdf: 2647418 bytes, checksum: 059c52b3ceacccabfdd11cebbc4b6dae (MD5) Previous issue date: 2017-12-21 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Acredita-se que haja um set point que limita o gasto energético utilizando como estratégia o controle dos níveis de atividade física em nosso organismo visando a manutenção da homeostase energética semelhante a outras variáveis reguladas biologicamente. O presente estudo tem como objetivo investigar se o treinamento periodizado e a dieta hiperlipídica influenciam os níveis de atividade física espontânea e voluntária e a alteração na ingestão alimentar de ratos. Para isso, o estudo foi dividido em duas etapas: na primeira o objetivo foi verificar a acurácia e confiabilidade de um sistema de videogrametria na análise do deslocamento de ratos Wistar, e na segunda etapa, verificar se o aumento da atividade física, por meio do treinamento periodizado, é acompanhado por uma redução compensatória na atividade física e alteração da ingestão alimentar. Foram utilizados Ratos (n=32) da linhagem Wistar (Rattus Norvegicus Albinus Wistar) com idade inicial de 120 dias mantidos em gaiolas coletivas durante 8 semanas, após a separação dos grupos: Grupo controle (GC), Grupo controle com dieta hiperlipídica (GDH), Grupo de treinamento periodizado (TP), e Grupo treinamento periodizado dieta hiperlipídica (TPH), os quais realizaram um protocolo de treinamento periodizado de natação, durante 6 semanas, após 2 semanas de adaptação. O limiar anaeróbio pelo teste de lactato mínimo (Lam) foi realizado para a separação dos animais em grupos homogêneos e para verificação dos efeitos do treinamento. Foram verificados o efeito do treinamento físico periodizado sobre o acúmulo de atividade física espontânea (AFE -deslocamento pela filmagem) e voluntária (AFV - roda de atividades), além da verificação da interferência da dieta hiperlipídica na atividade física espontânea e voluntária, no perfil lipídico (triglicerídeos, HDL), glicemia, comportamento alimentar, medidas e gordura corporal. A análise de variância foi utilizada para verificar as diferenças das variáveis entre os grupos, enquanto que a correlação de Pearson foi utilizada para verificar a associação entre a carga de treinamento com a ingestão alimentar, AFE e AFV, considerando alfa de 5%. Na primeira etapa a acurácia e a confiabilidade do sistema de videogrametria para a análise do deslocamento foi confirmada. Na segunda etapa verificou-se que os valores de glicemia, HDL e TG não apresentaram diferença significativa entre os grupos após 8 semanas de experimento. Os grupos treinados apresentaram maiores valores de Lam em relação ao grupo GC e GDH e não apresentaram ganho excessivo de gordura corporal. Em relação ao consumo alimentar, os grupos alimentados com dieta hiperlipídica apresentaram menor consumo calórico em relação aos grupos alimentados com dieta padrão. O acúmulo de AFV não apresentou diferença entre os grupos, porém a AFE foi maior no grupo controle em relação ao demais. A associação entre a carga de treino total semanal e o consumo alimentar e carga de treino total semanal e AFV foram fracas. Para associação entre a carga de treino total semanal e a AFE, houve forte associação com o grupo TP . O TP e a dieta hiperlipídica levaram a uma redução da AFE, assim como nas semanas onde os treinamentos foram intensos houve um nível de AFE menor. Nosso resultados indicam que existem fatores biológicos responsivos ao treinamento e a dieta hiperlipídica capazes de interferir nos níveis de AFE dos animais. / It is believed that there is a set point that limits the energy expenditure using as a strategy the control of the levels of physical activity in our organism aiming the maintenance of the energy homeostasis similar to other variables regulated biologically. The present study aims to investigate whether the periodized training and the hyperlipidic diet influence the levels of spontaneous and voluntary physical activity and the alteration in dietary intake of rats. For this, the study was divided in two phases: in the first the objective was to verify the accuracy and reliability of a videogrammetry system in the analysis of Wistar rats displacement and in the second stage, to verify if the increase of the physical activity through periodized training is accompanied by a compensatory reduction in physical activity and alteration in food intake. Rats (n = 32) Wistar (Rattus Norvegicus Albinus Wistar) with initial age of 120 days were kept in collective cages for 8 weeks, after separation of the groups: Control group (CG), Control group with hyperlipidic diet (GHD), Periodized training group (PT), and Periodic training group hyperlipidic diet (PTH), who performed a periodized swimming training protocol for 6 weeks after 2 weeks of adaptation. The anaerobic threshold by lactate minimum (Lam) test was performed to separate the animals into homogeneous groups and to verify the effects of the training. The effect of the periodic physical training on the accumulation of spontaneous physical activity (SPA - displacement by filming) and voluntary (VPA - activity wheel) was verified, as well as the verification of the interference of the hyperlipidic diet in spontaneous and voluntary physical activity, in the lipid profile (triglycerides, HDL), glycemia, eating behavior, measures and body fat. The analysis of variance was used to verify the differences of the variables between the groups, while the Pearson correlation was used to verify the association between training load and food intake, SPA and VPA, considering alpha of 5%. In the first phase the accuracy and reliability of the videogrammetry system for displacement analysis was confirmed. In the second phase, it was verified that the values of glycemia, HDL and TG did not present significant difference between the groups after 8 weeks of experiment. The trained groups had higher values of Lam in relation to the CG and GHD groups and did not present an excessive gain of body fat. In relation to the food consumption, the groups fed with a hyperlipid diet presented lower caloric intake than the groups fed a standard diet. The VPA accumulation did not present difference between the groups, but the SPA was higher in the control group in relation to the others. The association between total weekly training load and total weekly training and feeding load and VPA were weak. For association between the total weekly training load and the SPA, there was a strong association with the PT group. The PT and the hyperlipid diet led to the reduction of SPA, just as in the weeks where the training was intense there was a lower level of SPA. Our results indicate that there are training-responsive biological factors and the hyperlipidic diet capable of interfering with the SPA levels of the animals.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal.

Mémoire numérisé par la Division de la gestion de documents et des archives de l'Université de Montréal

Le stress oxydatif joue un rôle prépondérant en tant que messager secondaire dans la régulation de nombreux processus cellulaires tels que l’apoptose, la survie et la prolifération et serait impliqué dans l’ensemble des étapes de la carcinogenèse pulmonaire. L’activité physique et la nutrition sont deux facteurs pouvant moduler le stress oxydatif et les mécanismes associés. La bétaïne et la C-Phycocyanine sont deux micronutriments reconnus pour avoir des effets antioxydants, anti-inflammatoires et antiprolifératifs. Récemment notre équipe a montré in vitro qu’un traitement en bétaïne et/ou la C-phycocyanine diminuait la viabilité des cellules A549 (carcinome pulmonaire). L’objectif général de ce travail de thèse était d’évaluer l’effet de facteurs nutritionnels (bétaïne, C-PC ou activité physique) sur la croissance tumorale de cellules A549 implantées chez le rat Nude et de déterminer les mécanismes sous-jacents. Dans un premier temps, nous avons étudié l’effet d’une supplémentation nutritionnelle (bétaïne ou C-phycocyanine) associée ou non à la pratique d’une activité physique volontaire (roue d’activité) sur l’équilibre redox et l’inflammation, chez des rats sains. Nous avons montré que la bétaïne et la C-phycocyanine augmentaient les défenses antioxydantes tandis que l’activité physique volontaire n’avait pas d’effet si elle n’était pas couplée à une supplémentation. Nous avons également mis en évidence que C-phycocyanine inhibait l’augmentation de Cox-2 musculaire induite par l’activité physique. Dans un second temps, nous avons étudié l’effet de la bétaïne et/ou la C-Phycocyanine sur la croissance des cellules A549 implantées chez des rats Nude. Nous avons montré que ces deux micronutriments associés ou non ralentissaient la croissance des tumeurs pulmonaires, au travers de mécanismes communs (activation de NF-kappaB, augmentation de la peroxydation lipidique et de l’expression de cytokine pro-inflammatoire (IL-1-beta, Cox-2 et TNF-alpha) au sein de la tumeur) et de mécanismes propres à chaque micronutriment. La C-phycocyanine a induit une diminution du ratio AKT phosphorylé / AKT total et une augmentation du ratio p38 phosphorylé / p38 total, mécanisme en faveur de l’apoptose et de l’autophagie. La bétaïne associée à la C-phycocyanine a augmenté le ratio caspase-3 / pro-caspase-3. Dans un dernier temps, nous avons évalué l’effet de l’activité physique volontaire sur la croissance tumorale des cellules A549 implantées chez des rats Nude. Nous avons mis en évidence que l’activité physique volontaire ralentissait la croissance des tumeurs pulmonaires induites, sans différence significative avec la bétaïne et/ou la C-phycocyanine. Il apparait que l’augmentation de la peroxydation lipidique, l’activation de la MAPK p38 et de NF-kappaB, et l’inhibition d’AKT, favorisant la mort cellulaire soient impliquées dans cette diminution tumorale. Un régime enrichi en bétaïne et/ou C-phycocyanine ralentit la croissance cellulaire d’adénocarcinome pulmonaire implanté chez le rat, suggérant leur intérêt dans l’action anti-carcinogène pulmonaire. L’activité physique semble jouer sur les mêmes mécanismes. Nos résultats méritent d’être confirmés par des protocoles à plus large échelle et suggèrent de possibles applications chez des patients porteurs de tumeurs pulmonaires. / Oxidative stress seems to play a crucial role as a secondary messenger in the regulation of several cellular processes such as apoptosis, survival and proliferation, and could be involved in all steps of the lung carcinogenesis (i.e. initiation, promotion and progression). Physical activity and nutrition are two factors able to modulate oxidative stress and associated mechanisms. Betaine and C-phycocyanin are two known micronutrients having antioxidant, anti-inflammatory and anti-proliferative effects. Previously, our team showed that betaine and/or C-phycocyanin treatment decreased the viability of A549 cells in vitro (pulmonary adenocarcinoma cell line). The main objective of this work was to evaluate the effect of nutritional factors (betaine, C-phycocyanin) or physical activity on growth of implanted A549 cells in Nude rats and to determine underlying mechanisms.Firstly, we studied the effect of nutritional supplementation (betaine or C-phycocyanin) combined or not with voluntary physical activity (wheel running) on redox balance and inflammation in healthy rats. We showed that betaine and C-phycocyanin increased antioxidant defenses, whereas voluntary physical activity did not have an effect when it was not associated with micronutrient supplementation. We also observed that C-phycocyanin inhibited physical activity-induced muscle Cox-2 activity increase.Secondly, we studied the effect of betaine and/or C-phycocyanin on growth of implanted A549 cells in Nude rats. We showed that these two micronutrients, whether associated or supplied separately, slowed down the lung tumour growth through similar mechanisms (NF-kappaB activation and increase of lipid peroxidation and expression of pro-inflammatory cytokines (IL-1beta, Cox-2 et TNF-alpha) in tumour). Also, some mechanisms were specific for each micronutrient or their combination. C-phycocyanin induced a decrease of phosphorylated AKT / total AKT ratio, and an increase of phosphorylated p38 / total p38 ratio, both mechanisms promoting apoptosis and autophagy. On the other hand, betaine associated with C-phycocyanin increased caspase-3 / pro-caspase-3 ratio.Finally, we studied the effect of voluntary physical activity on growth of implanted A549 cells in Nude rats. We showed that voluntary physical activity slowed down the lung tumour growth, without significant difference if animals were supplied with betaine or/and C-phycocyanin. It seems that the increase of lipid peroxidation, NF-kappaB and p38 activation, and AKT inhibition, all having a role in promotion of a cell death, are responsible for the tumour growth slowdown following the physical activity. In conclusion, diet enriched with betaine or/and C-phycocyanin slows down the growth of pulmonary adenocarcinoma cells implanted in rats, suggesting their interest in anti-cancer activity. Physical activity seems to act on similar mechanisms as these micronutrients. Our results have to be confirmed with further studies, but are already suggesting a potential application in lung cancer patients.