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Thesis (Ph.D.)--Boston University / PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at open-help@bu.edu. Thank you. / The mechanisms by which diets high in saturated fat (HFD) contribute to intestinal adaptation and obesity are unknown. The hypothesis that functional changes in distal portions of small intestine are induced by HFD was tested in C57B1/6 mice. Specifically, it was examined whether the putative fatty acid translocase CD36 was phosphorylated in mouse intestinal epithelial cells and whether dephosphorylation of CD36 increased long chain fatty acid (LCFA) absorption. Co-immunoprecipitation was used to investigate specific intestinal alkaline phosphatases that might interact with CD36. It was also examined whether chronic ingestion of an HFD would lead to upregulation of the CD36 and/or one or more intestinal alkaline phosphatases that may activate CD36. CD36 was found to be phosphorylated on the surface of mouse enterocytes, indicating that there may be a phosphatase-sensitive pool of phospho-CD36 (pCD36) in mouse small intestinal tissue. CD36 was dephosphorylated by alkaline phosphatase and this treatment increased long chain but not short chain fatty acid uptake. Long chain fatty acid uptake was blocked with a specific CD36 inhibitor. CD36 from mouse small intestines physically interacted specifically with global intestinal alkaline phosphatase (gIAP) but not duodenal alkaline phosphatase (dIAP). As expected, HFD increased body weight, adiposity, and plasma triglycerides compared to control mice. CD36 and gIAP but not dIAP protein levels were significantly increased in distal but not proximal regions of intestines of HFD mice. Finally, HFD increased the absorptive capacity of the distal small intestine for LCFA in a CD36-dependent manner. It is concluded that HFD specifically upregulates gIAP protein in epithelial cells of the distal regions of the small intestine of mice, and that one of its substrates is pCD36, which has been implicated in transcellular fat transport. This diet also increases the absorptive capacity of the distal small intestine for LCFAs. Taken together, these results suggest that HFD causes intestinal adaptation that results in an increased capacity to absorb dietary fat. This effect is mediated in part by increasing the expression and activity of the fatty acid transporter CD36 and its regulatory enzyme gIAP. / 2031-01-02

>Magister Scientiae - MSc / The incidence of type 2 diabetes (T2D) is persistently increasing globally. T2D is associated with pancreatic β cell dysfunction and insulin resistance in peripheral tissues such as the liver and skeletal muscle. Skeletal muscle is the major site for insulin stimulated glucose uptake. Maintenance on a gestational high fat diet may programme insulin resistance. Programming is induced by the exposure of organisms to either a stimulus or insult during foetal and/or early neonatal life and alters offspring physiology and metabolism. The aim of the present study was therefore to investigate the effects of maternal diets, varying in fat content, on neonatal hepatic and skeletal muscle gene (mRNA) and protein (immunoreactivity) expression of proximal insulin signalling factors: insulin receptor alpha (IRα), insulin receptor substrate 2 (IRS2) and phosphoinositide 3-kinase-p110 alpha (PI3K-p110α), and to assess the therapeutic potential of Aspalathus linearis extract after high fat programming. Pregnant rats were randomised into groups maintained on diets with varying fat proportions: 10% (control), 20% (20F), 30% (30F) and 40% (40F) fat as energy throughout gestation. Neonatal liver and skeletal muscle were collected to determine the proximal insulin signalling expression profiles of the target factors: IRα, IRS2 and PI3K-p110α. Quantitative polymerase chain reaction (qPCR) was applied to determine mRNA expression of these target insulin signalling factors. Immunostaining of the target proteins in the liver and skeletal muscle was performed followed by relative quantification with image analysis software. Further, Aspalathus linearis (Al) extract was orally administered to mothers during gestation in the 10% (Control-Al) and 40% (HFD-Al) diets at a dose of 150 mg/kg. Body weight, food intake and blood glucose concentrations were monitored throughout gestation in mothers. Maternal diets, varying in the percentage of fat content, showed no significant effect on neonatal hepatic IR and IRS2 mRNA expression. However, hepatic PI3K mRNA expression was elevated in 30F neonates compared to 20F neonates. Skeletal muscle IR and PI3K mRNA expression were reduced in the 30F and 40F neonates compared to 20F neonates. There was reduced hepatic IRα immunoreactivity in 40F neonates compared to control and 20F neonates. Further, skeletal muscle IRα immunoreactivity was significantly reduced in 30F and 40F neonates compared to control neonates. Therefore foetal high fat programming reduced IRα in both the liver and skeletal muscle which may impair proximal insulin signalling in these glucose recipient organs. Aspalathus linearis had no effect on maternal serum insulin and glucagon concentrations. In addition, maternal caloric intake, body weight and organ weights (liver, brain and pancreas) were not altered amongst the groups. Further, HFD-Al neonates were heavier than control neonates. In conclusion, Aspalathus linearis, at a dose of 150 mg/kg, had neither harmful nor ameliorative effects in pregnant mothers fed high fat diet during gestation. In addition, Aspalathus linearis treatment had no ameliorative effects on neonates from mothers fed high fat diet throughout gestation.

Indiana University-Purdue University Indianapolis (IUPUI) / Chronic consumption of a high-fat diet is a lifestyle factor that increases the risk for cognitive impairment (Granholm et al., 2008; Greenwood & Winocur, 2005; Mattson, 2004; Winocur & Greenwood, 2005). A high-fat diet appears to facilitate cognitive impairment through the promotion of insulin resistance (Greenwood & Winocur, 2005; Stranahan et al., 2008; Winocur & Greenwood, 2005). A gap in the literature is an established timeframe of the progression and underlying mechanism, which study in animals would better afford. Furthermore, A limited number of studies have investigated the relationship between a high-fat diet and behavioral dysregulation such as anxiety and depression. The 1st aim of the study was to determine if consumption of a high-fat diet leads to cognitive impairment and behavioral dysfunction at 3, 8, or 13 weeks of consumption. The 2nd aim was to determine if cholesterol levels and HBP activity are aberrantly increased in specific regions in mice that display feeding induced cognitive/behavioral dysfunction. Consumption of the experimental specialty diets produced a number of significant behavioral effects. These significant effects began to emerge after only 3 weeks of low-and high-fat feeding with increased anxiety-like behavior displayed higher in the high-fat diet group for the Elevated Plus Maze and Open Field Test. There was increased thigmotactic behavior and floating in the low-fat diet group in the Morris Water Maze (MWM) task, therefore making cognitive assessment uninterpretable. This pattern in the behavioral tasks were more robust in the 8 week group and alleviated in the 13 week group. There was only a significant difference in depression-like symptoms in the Forced Swim (FS) Task in the 3 week group. Cholesterol analysis is still under review in Dr. Elmendorf’s lab to correlate cholesterol levels and cognitive/behavioral impairment.

Metabolic flexibility plays a significant role in energy homeostasis by regulating fuel selection in correspondence to energy demand. Obese and type II diabetic populations have displayed a hindered ability to properly transition from fat oxidation while in a fasted state to carbohydrate oxidation once fed, leading to a buildup of mitochondrial metabolites such as acylcarnitines. Carnitine, essential for fatty acyl-CoA transport through the inner and outer mitochondrial membranes, can be an indicator of mitochondrial distress as elevated levels tend to spill over into plasma suggesting a disruption in oxidation. The current study was designed to examine the effect of short term, high fat feeding on plasma acylcarnitine species diversity and levels and if acylcarnitines are associated with metabolic flexibility. 13 healthy, non-obese, sedentary males, aged 18-40 years participated in this study. Following a 12-hour overnight fast a biopsy was taken from the quadricep before and 4 hours after a high fat meal. Blood draws were obtained pre-biopsy while fasted and every hour for 4 hours post high fat meal consumption. Acylcarnitines from plasma were converted to their butyl esters and analyzed by electrospray ionization tandem mass spectrometry (MS/MS). Changes were observed in acetylcarntine (P=0.0125), glucose oxidation (P=0.0295), C16:1/C16:0 desaturation index (P= 0.0397), and C18:1/C18:0 desaturation index (P=0.0012). We did not find that individual changes in flexibility correlated with circulating acylcarnitine measurements in a fasted state / Master of Science

Skeletal muscle is highly involved in macronutrient metabolism. To maintain proper energy metabolism and physiology, skeletal muscle must adapt to nutrient supply. Thus, diet macronutrient composition is an important modulator of skeletal muscle metabolism. Evidence from rodent and human models show high-fat diets contribute to impaired insulin signaling, as well as decreased fatty acid and glucose oxidation. Utilizing proteomic analysis of metabolic proteins in humans may lead to the mechanism behind skeletal muscle adaption to macronutrient composition, potentially providing the groundwork for characterizing the etiology of high-fat feeding induced metabolic disease. The objective of this study was to compare the substrate oxidation patterns and the levels of metabolic proteins in the fasted skeletal muscle of lean, healthy males that either increased fatty acid oxidation in response to the high-fat diet, termed responders, or males that decreased fatty acid oxidation, termed non-responders. We employed a controlled feeding study design, where the participants served as their own controls. Following a 2-week control diet (30% fat, 55% carbohydrate and 15% protein), participants came to the lab fasted overnight and a muscle biopsy was taken from their vastus lateralis muscle. Participants were then placed on a 5-day high-fat diet (50% fat [45% saturated fat], 35% carbohydrate, and 15% protein). Following this diet, participants again came to the lab fasted overnight and another muscle biopsy was taken from their vastus lateralis muscle. Both the control and the high-fat diets were isocaloric to habitual diets. Muscle from the biopsies were utilized for substrate metabolism measures and mass spectrometry. We did not observe any significant differences in glucose oxidation between responders and non-responders, prior to or following the high-fat diet. Our proteomic analysis identified 81 proteins and protein subunits involved in substrate metabolism but only 6 were differentially regulated by the high-fat diet. Independent of the high-fat diet, compared to non-responders, responders contained an overall higher content of protein subunits belonging to Complex I and ATP synthase. The findings from this study suggest that adaption to high-fat feeding is individual specific and proteomic changes alone cannot explain high-fat feeding induced metabolic changes. / Ph. D. / Skeletal muscle is highly involved in macronutrient metabolism, which consist of the breakdown and utilization of glucose and fatty acids, thus making the foods we ingest a major modulator of skeletal muscle metabolism. Over the last few decades, Americans have increased their ingestion of foods high in saturated fats, which has coincided with the increased prevalence of obesity and type 2 diabetes. Further, evidence suggests these metabolic diseases are associated with the skeletal muscle’s inability to switch between the utilization of glucose and fatty acid in response to nutrient supply. Analyzing metabolic protein content in humans may lead to the mechanism behind skeletal muscle adaption to macronutrient composition, potentially leading to the cause behind the development of high-fat feeding induced metabolic disease. The objective of our controlled feeding study was to compare the macronutrient metabolism and the content of metabolic proteins in the fasted skeletal muscle of healthy males that either increased fatty acid utilization in response to a high-fat diet, termed responders, or males that decreased fatty acid utilization, termed non-responders. Following a 2-week control diet (30% fat, 55% carbohydrate and 15% protein), participants came to the lab fasted overnight and a biopsy was taken from their thigh muscle called the vastus lateralis. Participants then began a 5-day high-fat diet (50% fat [45% saturated fat], 35% carbohydrate, and 15% protein). Following this diet, participants came to the lab fasted overnight and another biopsy was taken from their vastus lateralis muscle. Both the control and the high-fat diets were isocaloric to habitual diets. The muscle samples were used to analyze macronutrient metabolism and identify metabolic protein content. We did not observe differences in glucose utilization between responders and non-responders, prior to or following the high-fat diet. We identified 81 metabolic proteins and protein subunits but only 6 were differentially regulated by the high-fat diet. Independent of diet, responders contained higher levels of subunits from 2 proteins involved in cell energy production, Complex I and ATP synthase. Our findings suggest that adaption to high-fat feeding is individual specific and protein content changes alone cannot explain high-fat feeding induced metabolic changes.

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O aumento da adiposidade corpórea pode gerar diversos mediadores inflamatórios com capacidade de influenciar a proliferação e a diferenciação hematopoética e, consequentemente, a complexa regulação da hematopoese. Por isso, propusemo-nos, neste trabalho, avaliar a influência do aumento da adiposidade corpórea sobre a proliferação e a diferenciação de células hematopoéticas, bem como sua capacidade em sintetizar citocinas. Ratos Wistar, machos foram alimentados com uma dieta rica em lipídios durante 14 semanas. Após esse período foram avaliados hemograma, mielograma, perfil lipídico, concentrações séricas de leptina, insulina e adiponectina. Citômetria de fluxo foi utilizada para avaliação da porcentagem de células CD34+/CD133+, bem como o ciclo celular de células medulares. Células medulares foram utilizadas para avaliar a atividade proliferativa in vitro e a capacidade de diferenciação, in vitro, na presença de IL-3, EPO, GM-CSF e G-CSF. Animais, alimentados com dieta hiperlipídica, apresentaram maiores concentrações de leptina circulante, com aumento de gordura corporal, aumento da concetração de proteína C reativa, colesterol total, LDL, VLDL e triacilglicerol. O hemograma apresentou neutrofilia absoluta e a medula óssea apresentou-se hipercelular com aumento do número de granulócitos maduros e da população celular CD133-/CD34+. Os resultados dos testes in vitro demonstraram aumento da capacidade de síntese de IL-3 e aumento de G-CSF, com aumento do potencial proliferativo, também evidenciado pelo maior número de células medulares na fase S/G2/M, bem como o aumento da diferenciação granulocítica. Esses resultados sugerem que a leucocitose e neutrofilia observadas em situações de aumento da adiposidade corpórea são decorrentes de uma complexa modulação do sistema hematopoético. / The body fat increase can generate various inflammatory mediators, that are capable to influence the proliferation and differentiation of hematopoietic cells and consequently modulate the complex regulation of the hematopoiesis. In this study we have proposed to evaluate the effect of increase body fat on the proliferation and differentiation of hematopoietic cells, as well as its ability to synthesize cytokines. Male Wistar rats were subjected to a high fat diet during a period of 14 weeks. After that period were evaluated hemogram, mielogram, lipid profile and the serum concentrations of leptin, insulin and adiponectin. Flow cytometry was used to evaluate the percentage of CD34+/CD133+, as well as the cell cycle of bone marrow cells. Bone marrow cells were used to perform the proliferation and differentiation capacity in vitro in the presence of IL-3, EPO, GM-CSF and G-CSF. Animals fed high-fat diet had higher concentrations of circulating leptin with increase body fat, and increase of C-reactive protein, total cholesterol, LDL, VLDL and triacylglycerol concentrations. The hemogram showed absolute neutrophilia and a hypercellular bone marrow with increase of granulocytic mature population and CD133-/CD34+ cells. The results in vitro, showed an increase of IL-3 and G-CSF production, and higher proliferative potential with an increase in S/G2/M bone marrow cell cycle phases, as well as an increase of the granulocytic differentiation. The results suggest that leukocytosis and neutrophilia observed in this model of body fat increase are in fact a result of a complex modulation of the hematopoietic system.

Made available in DSpace on 2014-06-11T19:31:04Z (GMT). No. of bitstreams: 0 Previous issue date: 2010-02-18Bitstream added on 2014-06-13T19:40:53Z : No. of bitstreams: 1 leopoldo_apl_dr_botfm.pdf: 2253115 bytes, checksum: 28ca3088b9d995d2eab4a88eb473ac7d (MD5) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Atualmente mais de um bilhão de pessoas apresentam sobrepeso, sendo que, mais de 30% desta população é obesa. Diversas alterações estruturais e funcionais do coração em humanos tem sido frequentemente associadas com a obesidade. Modelos experimentais, por dieta hiperlipídica, têm sido utilizados para estudar a relação obesidade e coração. O trânsito de cálcio miocárdico tem sido extensivamente estudado em diversos modelos experimentais e frequentemente relacionado com disfunção cardíaca. Entretanto, a literatura mostra escassez de estudos que avaliaram a relação entre o tempo de exposição à obesidade por dieta hiperlipídica, o RNAm e as proteínas envolvidas na homeostase de Ca+2 miocárdico. Além disso, pesquisas relatam a influência dos hormônios tireoidianos nestas proteínas, podendo acarretar alterações na contração e relaxamento cardíaco.O objetivo principal desse estudo foi testar a hipótese que o aumento no tempo de exposição à obesidade acarreta diminuição na expressão e/ou fosforilação das proteínas e dos respectivos níveis de RNAm relacionados com o trânsito de Ca+2 miocárdico. Este estudo teve como objetivo secundário constatar se a diminuição na expressão gênica foi acompanhada de redução dos níveis hormonais tireoidianos. Os períodos de dieta hiperlipídica, utilizados nesse estudo, foram eficientes em promover obesidade, desde que o índice de adiposidade utilizado para caracterizar os animais como obesos foi 79,5%, 82% e 69,5% maior do que os respectivos controles, após 15, 30 e 45 semanas. O tempo de exposição à dieta hiperlipídica não promoveu alterações na gordura corporal total entre os grupos obesos; este resultado indica que, neste trabalho, não houve aumento na intensidade da obesidade ao longo do tempo. Neste estudo foram visualizadas algumas comorbidades frequentemente associadas com a obesidade... / Currently, greater than one billion people are overweight and 30% of the population is obese. Several structural and functional changes of the heart have often been associated with human obesity. Experimental models for high-fat diets have been used to study the relationship between obesity and the heart. Myocardial calcium (Ca2+) handling has been extensively studied in several experimental models and has often been shown to be related to cardiac dysfunction. However, few studies have evaluated the relationship between the duration of exposure to obesity and a high-fat diet, and mRNA and proteins involved in homeostasis of myocardial Ca2+. Some studies have reported the influence of thyroid hormones on these proteins, which may cause changes in cardiac contraction and relaxation. The main objective of the current study was to test the hypothesis that the increased duration of exposure to obesity leads to a reduction in the expression and/or phosphorylation of proteins and mRNA levels related to myocardial Ca2+ handling. This study had, as additional objective, to verify if the decrease in mRNA expression was accompanied by a reduction in thyroid hormone levels. The periods of exposure to a high-fat diet used in this study were effective in promoting obesity since the adiposity index used to characterize animals as obese was 79.5%, 82%, and 69.5% higher than controls after 15, 30, and 45 weeks, respectively. The duration of exposure to a high-fat diet did not change the total body fat between the obese groups. This result indicates that there was not an increase in the intensity of obesity over time. In this study, some co-morbidities often associated with experimental obesity existed, such as glucose intolerance, insulin resistance, hyperinsulinemia, hyperleptinemia, and dyslipidemia; however, the co-morbidities were not associated with changes in systolic blood pressure ...(Complete abstract click electronic access below)

abstract: The prevalence of obesity and obesity-related disorders have increased world-wide. In the last decade, the intestinal microbiome has become a major indicator of metabolic and gastrointestinal health. Previous research has shown that high-fat diet (HFD) consumption can alter the microbial composition of the gut by increasing the abundance of gram-positive bacteria associated with the onset of obesity and type 2 diabetes. Although, the most common form of obesity and metabolic syndrome intervention is exercise and diet, these recommendations may not improve severe cases of obesity. Thus, an important relevance of my project was to investigate whether the intake of an organometallic complex (OMC) would prevent the onset of metabolic and gastrointestinal complications associated with high-fat diet intake. I hypothesized that the consumption of a HFD for 6 weeks would promote the development of metabolic and gastrointestinal disease risk factors. Next, it was hypothesized that OMC treatment would decrease metabolic risk factors by improving insulin sensitivity and decreasing weight gain. Finally, I hypothesized that HFD-intake would increase the abundance of gram-positive bacteria associated with gastrointestinal disease. My preliminary data investigated the effects of a 6-week HFD on the development of hepatic steatosis, intestinal permeability and inflammation in male Sprague Dawley rats. I found that a 6-week HFD increases hepatic triglyceride concentrations, plasma endotoxins and promotes the production of pro-inflammatory cytokines in the cecum wall. I then investigated whether OMC treatment could prevent metabolic risk factors in male Sprague-Dawley rats fed a HFD for 10 weeks and found that OMC can mitigate risk factors such hyperglycemia, liver disease, impaired endothelial function, and inflammation. Lastly, I investigated the effects of a 10-week HFD on the gastrointestinal system and found an increase in liver triglycerides and free glycerol and alterations of the distal gut microbiome. My results support the hypothesis that a HFD can promote metabolic risk factors, alter the gut microbiome and increase systemic inflammation and that OMC treatment may help mitigate some of these effects. Together, these studies are among the first to demonstrate the effects of a soil-derived compound on metabolic complications. Additionally, these conclusions also provide an essential basis for future gastrointestinal and microbiome studies of OMC treatment. / Dissertation/Thesis / Doctoral Dissertation Biology 2019