Maternal lifestyle and nutrient intake during pregnancy can have long-lasting effects on the health of offspring as well as the mother. This dissertation focuses on the impact of maternal exercise during pregnancy on offspring insulin sensitivity and glucose uptake and the maternal effects of exercise during pregnancy. The first aim of this dissertation was to investigate if exercise prior to and during pregnancy and nursing would improve glucose uptake and insulin sensitivity in mice and rats. In both mice and rats, it was concluded that maternal exercise could enhance whole-body insulin sensitivity and increase glucose uptake into skeletal muscle and adipose tissue in adult offspring compared with offspring from sedentary dams. Maternal exercise also positively influenced male but not female adult offspring body composition; male offspring from exercised dams had significantly decreased fat mass and increased lean mass compared with offspring from sedentary dams. The second aim of this dissertation was to test whether exercise during pregnancy would improve glucose disposal in mouse dams with diet-induced obesity. Maternal running was effective in reducing fat mass accumulation and glucose intolerance associated with high fat feeding during pregnancy. In high fat diet mice, exercise was also able to improve insulin sensitivity in adipose tissue compared to tissue from sedentary high fat diet mice. The findings in this dissertation provide new insight into the long-term effects exercise during pregnancy can have on offspring health. Women may be encouraged to start an exercise regimen before and during their pregnancy if they are aware of the life-long benefits it can have for their children. The findings from the second aim present new insight into how exercise can affect pregnancies complicated by maternal obesity and glucose intolerance, and the animal model can be used in the future studies to investigate the offspring effects of maternal exercise during a diabetic pregnancy.

Some aspects of anuran metabolism are examined, with special emphasis on possible limitations to aerobic metabolism and the effect of chronic hypoxia acclimation on maximal rates of aerobic metabolism and the metabolic fate of lactic acid accumulated after anaerobic metabolism.

5'-Deoxy-5'-methylthioadenosine (MTA) is derived from s-adenosylmethionine (AdoMet) during the synthesis of the polyamines spermidine and spermine. Methionine can be regenerated from MTA by one of two mechanisms. In mammalian cells and some microorganisms, MTA is degraded to adenine and 5-methylthioribose-1-phosphate (MTR-1-P) via MTA phosphorylase. In certain other microbes, however, MTA is catabolized in two steps; first to adenine and 5-methylthioribose (MTR) via MTA nucleosidase followed by conversion of MTR to MTR-1-P via MTR kinase. This study was to demonstrate the presence of MTA nucleosidase or MTA phosphorylase in both redia containing cercariae and adult Fasciola hepatica Linnaeus, 1758. If MTA nucleosidase was present, it was wanted to determine if MTR kinase was also present. The phosphate-dependent cleaving activity of MTA phosphorylase was demonstrated in the cell-free extracts of adult Fasciola hepatica along with an unidentified MTR metabolizing activity. Redia containing cercariae showed MTA nucleosidase and MTR kinase activity.

A synthesis is described for a number of 16,16-dimethyl analogs of the prostacyclin and related 6-keto prostaglandin types. Included are l6, l6-dimethyl-PGI2 sodium salt (XLIV), 6α-l6, l6-dimethyl-PGI1(XLV), 6β-l6, l6-dimethyl-PGI1 (XLVII), 6-keto-l6, l6-dimethyl-PGF1-α (XLIX), and 6-keto-l6, l6-dimethyl PGE1 (LV). Done, but not included, is the activity for these analogs in the blood platelet aggregation inhibition assay. This activity was consistently less than for the corresponding 16, 16-dihydro compounds.

Introduction: Autophagy is a complex and highly regulated process responsible for the maintenance of cellular homeostasis and the degradation and recycling of key nutrients. Autophagy is generally believed to serve a protective role, responding to periods of starvation and aging, pathogen invasion, development, improper protein folding, and organelle damage. Defective autophagy, on the other hand, is implicated in disease states such as metabolic syndrome, hepatic steatosis, obesity, and atherosclerosis, among others. Interestingly, in various skeletal muscle diseases associated with either atrophy or dystrophy, the accumulation of autophagosomes within myofibers is a common feature. Additionally, modulation of autophagy in skeletal muscle has been reported to influence energy and lipid metabolism, while also affecting these parameters in organs such as the pancreas, liver, and adipose tissue. Ethanol consumption inhibits protein synthesis while simultaneously inducing proteolysis of skeletal muscle, and a decreased skeletal muscle mass correlates to heightened disease progression in patients such as those diagnosed with alcoholic cirrhosis and various cancers. Due to the correlation between skeletal muscle loss and exacerbated disease progression, the role of autophagy in response to ethanol consumption could highlight new therapeutic modalities that may attenuate disease progression and improve final presentation of various disease states. Further, acute ethanol consumption has shown a selectivity of autophagy toward mitochondria and lipid droplets, rather than proteins that are typically degraded during starvation. Due to ethanol’s damaging effects exerted on mitochondria and lipid peroxidation, removal of mitochondria and lipid droplets via autophagy could represent an attempt at host defense. One limitation of various studies analyzing autophagy in skeletal muscle is the duration of ethanol exposure. Therefore, we aimed to characterize skeletal muscle autophagic flux in both the NIAAA, 10-day ethanol model as well as the chronic, six-week ethanol model. Methods: In order to analyze autophagic flux in response to ethanol consumption, C57BL/6 mice were treated with either a control or ethanol diet (5% EtOH/vol) for a period of ten days (NIAAA model) or six weeks (chronic model). Mice were then euthanized and hindlimb skeletal muscles were collected, snap frozen, and analyzed using an immunoblot assay. Results: Markers indicative of autophagy, such as autophagy-related genes (Atg), Beclin-1, and LC3A/B, are expected to exhibit increased expression in response to ethanol consumption in both the NIAAA and chronic ethanol model.

<p><em>Objective: </em>We evaluated the potential effect of oral glucose supplementation on: (1) exercise performance and tolerance, (2) the concentrations of plasma uric acid and ammonia (NH₃) and (3) blood plasma markers of oxidative stress in patients with McArdle’s disease (MCD) after non-ischemic forearm exercise testing (non-ischemic forearm exercise test). <em>Methods:</em> Blood samples and exercise performance measures were performed on from 16 patients with MCD and 17 control subjects (CON) matched for age, sex and physical activity status. Subjects performed 2 exercise bouts 30 minutes apart and received oral glucose or placebo supplementation between tests. Blood samples were analyzed for concentrations of 8-isoprostanes (8-ISO), malondialdehyde (MDA) and total anti-oxidant capacity (TEAC). Exercise performance was assessed using a handgrip dynamometer to measure force of contraction over time. Exercise tolerance was assessed based on subject’s self-reported perception of pain and perception of exertion during exercise. <em>Results:</em> MCD was associated with greater fatigue, perceived pain, perceived exertion, and higher uric acid during non-ischemic forearm exercise test (P < 0.05), and higher concentrations of plasma NH₃post exercise (P < 0.05). Glucose did not influence plasma uric acid or NH₃and had no effect on exercise measures in MCD patients. Baseline plasma markers of oxidative stress were not different between MCD patients and CON; however, MCD patients who ingested glucose between non-ischemic forearm exercise tests had lower plasma 8-ISO concentrations (P < 0.05). CON who ingested glucose between non-ischemic forearm exercise tests had lower plasma 8-ISO concentrations (P < 0.05) at +1 min post exercise compared to initial non-ischemic forearm exercise test. The TEAC of control subjects was lower following non-ischemic forearm exercise test (P < 0.05), with no change in MCD patients. <em>Conclusion: </em>Glucose may have a protective effect on oxidative stress following exercise that may be due to attenuated flux through xanthine oxidase.</p> / Master of Science (MSc)

<p><strong>Background:</strong> Inadequate nutrition during the postnatal period may be associated with adverse outcomes in later life. Tailoring nutrition to promote optimal growth requires monitoring body composition (BC). This could guide nutritional strategies to promote optimal outcomes, however preterm infant BC data is lacking. <strong></strong></p> <p><strong>Objectives: </strong>To introduce and assess the feasibility of air displacement plethysmography (PEA POD) as a bedside tool in the NICU; Second, to longitudinally measure preterm infant BC in hospital.</p> <p><strong>Methods</strong>: This was a longitudinal, observational study. Inclusion criteria: infants 24-37 weeks gestational age (GA); informed consent. Exclusion criteria: chromosomal/congenital abnormalities; hydrops fetalis. Preterm infant BC assessed by PEA POD; anthropometric measures were collected. Infants assessed from study inclusion to hospital discharge. For reference, 23 term infants were measured.</p> <p><strong>Results</strong>: PEA POD was a feasible bedside tool to measure preterm infant BC that was accepted by parents and free of adverse events. Longitudinal measures demonstrated preterm infants (n=65) gained fat mass (FM) and fat free mass (FFM) at differing rates, leading to an overall %FM increase in hospital. Our data suggests that, at term corrected age, preterm infants may achieve a BC similar to full term infants at birth.</p> <p><strong>Discussion</strong>: This study established PEA POD as a useful bedside clinical tool for preterm infants. Longitudinal BC changes in preterm infants using PEAPOD in hospital is described for the first time. Future application of PEA POD can expand longitudinal measures to create reference preterm infant BC data for evaluating quantity and quality of growth in relation to nutritional management.</p> / Master of Science in Medical Sciences (MSMS)

<p>Obesity is a major global health concern and is associated with the development of numerous non-communicable diseases. A thorough understanding of the onset of obesity is critical to the development of effective therapeutic strategies against this disease state. Recently, obesity has been described as a complex disease characterized by chronic low grade inflammation. Abnormal adipose tissue expansion is accompanied by an increased presence of proinflammatory immune cells, dysregulated adipokine expression, oxidative stress, and is associated with significant changes in the bacterial composition of the gut. While interleukin-15 (IL-15) has been studied extensively for its immunological effects, this cytokine has recently been shown to influence body weight and fat mass. The focus of this thesis was to elucidate the role of and mechanism by which IL-15 modulates adipose tissue. Our first study demonstrated that low levels of IL-15 expression are associated with adiposity and promotes an obese state in IL-15-/- mice and human subjects, while IL-15 overexpression was associated with a lean phenotype in IL-15tg mice when compared to appropriate controls. To uncover the underlining mechanisms by which IL- 15 mediates differences in body weight, we subsequently determined that IL-15 mediated weight loss occurred independently of lymphocytes. In another study, we showed that IL-15tg mice had increased mitochondrial activity and mass specific to adipose tissue compared to IL-15-/- and B6 mice, while acute IL-15 administration induced the expression of FAO markers in adipose tissue. Lastly, IL-15 treatment increased mitochondrial membrane potential and decreased lipid deposition in cultured adipocytes, suggesting that IL-15 may mediate its effects directly on adipose tissue. The experimental results presented in this thesis demonstrate that IL-15 is an important regulator of adipose tissue and body weight. Future studies examining the effects of IL- 15 on adipose tissue will further our knowledge on IL-15 biology, and may contribute to novel therapeutic strategies for the treatment and prevention of obesity.</p> / Doctor of Philosophy (PhD)

<p>The endoplasmic reticulum (ER) is a specialized organelle that facilitates correct protein folding and maturation. Disruptions in ER homeostasis lead to ER stress and activation of a series of signal transduction cascades known as the unfolded protein response (UPR), which acts to restore ER homeostasis. In recent years, ER stress and UPR dysfunction have been linked to obesity, fatty liver and insulin resistance. Lipid-laden adipocytes, the main cellular component of white adipose tissue (WAT), play a critical role in whole body energy homeostasis as well as lipid and carbohydrate metabolism. Mature adipocytes, which are metabolically active endocrine cells, differentiate from precursor fibroblast-like preadipocytes, through a process called adipogenesis, leading to formation of cells capable of secreting numerous proteins, cytokines and hormones. ER homeostasis and UPR activation are essential to the function/differentiation of highly secretory cells, however, the role of ER stress/UPR activation in adipogenesis had previously not been examined. We hypothesized that<em> adipogenesis may rely on physiological UPR activation to accommodate the demand on the ER for increased folding and secretion of proteins.</em></p> <p>Initial experiments examining UPR activation during 3T3-L1 adipogenesis identified that expression of ER stress/UPR markers was modulated during adipocyte differentiation. Furthermore, inhibition of ER stress/UPR activation by the chemical chaperone, 4-phenyl butyric acid (4-PBA), inhibited adipogenesis and blunted high fat-diet induced weight gain in 4-PBA supplemented mice. These findings suggested that UPR activation modulates adipogenesis and adipose tissue metabolism.</p> <p>Subsequently, we sought to identify novel candidate ER stress/UPR responsive genes that may be involved in adipogenesis and WAT metabolism. The expression of a recently recognized ER stress-responsive gene, T-cell death associated gene 51 (TDAG51) was identified to be differentially regulated during adipogenesis. However, the function of TDAG51 in adipogenesis or energy regulation was not known. Studies from this thesis showed that TDAG51 protein expression is attenuated by ER stress/UPR activation in preadipocytes and declines during adipogenesis. Based on these results, and given the importance of adipogenesis in WAT function and whole body energy metabolism, it was<em> </em>hypothesized that<em> TDAG51 may be a novel regulator of adipogenesis and energy homeostasis.</em> Indeed, as reported here, knock-down or absence of TDAG51 (<em>TDAG51^-/-</em>) in pre-adipocytes increased lipogenesis and lead to earlier and more potent expression of adipogenic markers.</p> <p>Finally, we investigated whether absence of TDAG51 in mice affected adiposity and metabolic outcomes. Consistent with the <em>in vitro </em>results, we found that <em>TDAG51^-/-</em>mice fed a standard chow diet, exhibited an age-associated increase in WAT, developed fatty liver, and exhibited insulin resistance as compared to wild-type mice.</p> <p>Taken together, the findings in this thesis indicate that physiological UPR activation and the UPR-responsive gene TDAG51 play important roles in regulating adipogenesis, lipogenesis and whole-body energy metabolism. Thus, therapeutic approaches aimed at modulating ER folding capacity, UPR activation and/or TDAG51 expression may have great potential in the treatment of obesity and its co-morbidities.</p> / Doctor of Philosophy (PhD)

Os objetivos da tese são, investigar (i) as estruturas terciária e quaternária de diversas proteínas em solução e (ii) as mudanças conformacionais de proteínas induzidas por ligantes e/ou temperatura, variando-se a concentração dos mesmos, etc. A principal técnica utilizada foi a de espalhamento de raios X a baixos ângulos (no inglês SAXS de mall-angle X-ray scattering), associada a outras metodologias como cristalografia e modelagem de proteínas. As contribuições dos estudos realizados de várias proteínas de interesse científico foram as seguintes: (i) propôs-se pela primeira vez um modelo de um receptor nuclear (RXR) composto por dois domínios, um de ligação ao ligante e outro ao DNA; (ii) elucidou-se o estado configuracional da fosfo-enol-piruvato carboxoquinase em solução; (iii) foi determinado o efeito de agregação e mudança conformacional induzido por ligantesna isoenzima fosfofrutoquinase e (iv) determinou-se a estrutura em solução da interleucina humana 22 e propôs-se modelos de interação com seus receptores. Além disso, (i) foi desenvolvido um método para determinar o estado oligomérico de uma proteína em solução, em quaisquer condições de tampão e concentração, utilizando medidas de SAXS em escala relativa, e (ii) mostrou-se que a densidade das proteínas é uma função do seu peso molecular, contrariando conceitos clássicos, tendo as proteínas pequenas (< 20kDa) uma densidade mais elevada que as maiores. / The purpose of this thesis is to investigate (I) the tertiary and quaternary structure of several proteins in solution and (II) conformational changed that such proteins undergo when they bind to ligands or temperature, pH, etc. conditions are varied. The main experimental technique utilized in this study was Small Angle X-Ray Scattering (SAXS) associated to other techniques and approaches like protein crystallography and homology modeling. The main contributions related to this work are: (i) for the first time a model for a nuclear receptor (RXR) containing two domains was proposed, one DNA binding domain and one ligand binding domain; (ii) the oligomeric state of the phosphor-enol-pyruvate caboxykynase in solution was detennined; (iii) the conformation changed induced by ligants to the isoenzyme phosphofructokinase was determined and (iv) the solution structure of human interleukin revealed and a model with its receptor proposed. Additionally, (i) a method for determining the oligomeric state of a protein in solution under any buffer conditions and concentration using SAXS in relative scale is proposed and (ii) it was also shown that proteins density is a molecular weight dependent function, where small proteins with less than 20kDa have in average a larger density than bigger ones.