Hyperhomocysteinemia, an elevation of blood homocysteine levels, is a metabolic disorder associated with dysfunction of multiple organs. Previous studies have shown that hyperhomocysteinemia is related to fatty liver. However, the underlying mechanism remains speculative. The objective of the present study is to investigate the regulatory mechanism of hepatic inflammatory response and cholesterol metabolism during metabolic disorders. In the present study, hyperhomocysteinemia was induced in Sprague-Dawley rats by feeding a high-methionine diet. The mRNA and protein expression of cyclooxygenase-2 (COX-2), a pro-inflammatory factor, were significantly elevated in the liver of hyperhomocysteinemic rats. An activation of NF-B and a stimulation of oxidative stress were observed in the same liver tissue in which COX-2 was induced. Inhibition of NF-B or oxidative stress effectively abolished hepatic COX-2 expression, inhibited the formation of inflammatory foci, and improved liver function. Activity of HMG-CoA reductase, the rate-limiting enzyme of cholesterol biosynthesis, was markedly elevated in the liver of hyperhomocysteinemic rats, which may contribute to the hepatic lipid accumulation induced by hyperhomocysteinemia. Administration of Berberine (5mg/ kg body weight/ day for 5 days) inhibited HMG-CoA reductase activity via upregulating AMP-activated protein kinase (AMPK)-mediated phosphorylation of HMG-CoA reductase. Berberine treatment reduced hepatic cholesterol content and ameliorated liver function. In addition, the regulatory mechanism of HMG-CoA reductase activation was investigated in C57BL/6 mice fed a high-fat diet. There was a significant increase in hepatic HMG-CoA reductase mRNA and protein expression as well as enzyme activity. The DNA binding activity of sterol regulatory element binding protein (SREBP)-2 (a transcription factor of HMG-CoA reductase) and Sp1 (a transcription factor of SREBP-2) were both increased in the liver of mice fed a high-fat diet. The in vitro study in palmitic acid-treated HepG2 cells further confirmed that inhibition of Sp1 by siRNA transfection abolished palmitic acid-induced SREBP-2 and HMG-CoA reductase mRNA expression. In conclusion, the present study have demonstrated that (1) Hepatic COX-2 expression is induced via oxidative stress mediated NF-B activation during hyperhomocysteinemia; (2) Dietary berberine reduces cholesterol biosynthesis by elevating AMPK-mediated HMG-CoA reductase phosphorylation; (3) HMG-CoA reductase is upregulated by Sp1-mediated SREBP-2 activation in the liver during high-fat diet feeding.

Resumo: 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 ...(Resumo completo, clicar acesso eletrônico abaixo) / Abstract: 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) / Orientador: Antonio Carlos Cicogna / Coorientador: Patrícia Chakur Brum / Banca: Alessandra Medeiros / Banca: Carlos Eduardo Negrão / Banca: Célia Regina Nogueira / Banca: Maeli Dal Pai Silva / Doutor

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.

The adolescent period has been proposed to be exquisitely sensitive to the impacts of stress and juvenile stressor exposure is associated with anxiety- and depressive- like characteristics in adulthood. Among adult rats, access to a palatable diet has shown to mitigate the effects of stressors, a form of ‘self-medication.’ The present collection of studies sought to further characterize the long-term consequences of stressor exposure early in the juvenile period, as well as the use of palatable food as a coping strategy. The first study (Chapter 2) highlighted the importance of methodological rigor in the design of experiments employing social stressors. The second study (Chapter 3) provided further evidence that exposure to juvenile social defeat can have long-lasting consequences into adulthood, and access to a palatable diet may impart some resilience to initial stressor exposure. The third study (Chapter 4) demonstrated that access to a palatable diet can mitigate the long-term behavioral consequences of a 3-day sub chronic non-social stressor applied during juvenility in pair housed rats. The fourth study (Chapter 5) sought to replicate these findings in individually housed (purportedly more stressed) animals. Interestingly, access to a palatable diet was sufficient to protect against the neuroendocrine consequences of juvenile stress but did not mitigate the behavioral consequences, raising the question of an effectiveness “threshold” of self-medication via a palatable diet. The final study (Chapter 6) provided some preliminary evidence that exposure to juvenile stress amid access to a palatable diet has long-lasting changes on dopamine receptor expression in the nucleus accumbens, although the functional significance needs further characterization. Collectively, all studies provided further evidence that self-medication with a palatable diet comes at the price of poor metabolic outcomes. The results of this body of work provide further evidence that exposure to stress during juvenility can have protracted effects into adulthood, at the cost of poor metabolic outcomes. It also raises the suggestion of an effectiveness threshold of palatable food to cope with stress. Further understanding of the interplay between stress and diet may serve to inform the development of prevention based programs to mitigate the rising tide of concurrent childhood obesity and levels of perceived stress.

Orientador: Marciane Milanski Ferreira / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Aplicadas / Made available in DSpace on 2018-08-27T16:06:12Z (GMT). No. of bitstreams: 1 Reginato_Andressa_M.pdf: 3622201 bytes, checksum: 6c4d49212fa7262052afb4aec8d81f48 (MD5) Previous issue date: 2015 / Resumo: O excesso na ingestão calórica e a obesidade têm afetado um número crescente de pessoas em diferentes países, sendo que a obesidade durante a gestação e lactação desempenha impacto negativo no fenótipo prole. Na vida adulta, a obesidade e a sobrecarga de lipídeos constituem fatores que resultam no comprometimento da autofagia, um processo de degradação lisossomal essencial para a manutenção da homeostase celular. A autofagia é responsável pela degradação e reciclagem de componentes citoplasmáticos como organelas senescentes, proteínas agregadas ou mal formadas, microrganismos invasores e macromoléculas. Apesar do conhecimento acerca do prejuízo na atividade autofágica no contexto da obesidade, alterações na homeostase deste processo na prole de mães obesas ainda não foram investigadas. Neste estudo, foi avaliada a hipótese de que a obesidade materna induzida por dieta hiperlipídica seria capaz de modular proteínas da via autofágica no hipotálamo e no fígado da prole de camundongos. Embora sem nenhuma alteração na atividade de autofagia no hipotálamo, a prole de mães obesas ao nascimento (d0) apresentou prejuízo nos marcadores de autofagia no fígado representado por aumento no conteúdo proteico de p62 e diminuição no conteúdo proteico de LC3-II. Ao desmame (d18), a prole de mães obesas teve comprometimento no conteúdo proteico dos marcadores de autofagia em ambos os tecidos (fígado e hipotálamo) quando comparados à prole de mães magras. Após o desmame, a prole de mãe controle e a prole de mãe obesa receberam dieta controle até a vida adulta (d82). Nessa condição não houve modulação dos marcadores de autofagia em nenhum dos tecidos avaliados, sendo que somente a reexposição à dieta hiperlipídica (dos 42 dias até 82 dias) foi responsável por alterar o conteúdo proteico dos marcadores de autofagia quando comparados aos animais com dieta hiperlipídica sem reexposição. Assim, parece que dieta hiperlipídica é essencial para a modulação negativa dos marcadores de autofagia na prole de mães obesas. Em conclusão, a prole de mãe obesa apresentou comprometimento precoce de marcadores de autofagia no fígado e no hipotálamo, o que poderia estar associado ao desenvolvimento de distúrbios metabólicos na prole na idade adulta / Abstract: The nutritional excess and obesity have affected a growing number of people in different countries, being that obesity during pregnancy and lactation has negative impact on offspring phenotype. In adulthood, obesity and lipids overload constitute factors that result in impairment of autophagy, a lysosomal degradation process essential for maintaining cellular homeostasis. Thus, autophagy is responsible for degradation and recycling of cytoplasmic components as senescent organelles, aggregated proteins or proteins poorly formed, microorganisms invaders and macromolecules. It is known that obesity and the use of high fat diet have a negative impact on cellular homeostasis. However, modulation of autophagy in the offspring of obese mothers has yet to be investigated. This study tested the hypothesis that maternal obesity induced by high fat diet would be able to modulate proteins of autophagy in the hypothalamus and liver of mice offspring. At birth (d0), the offspring exhibited prejudice in autophagy markers in liver and after weaning (d18) both tissues (liver and hypothalamus) had compromised autophagy markers. The animals receiving control diet after weaning until adulthood (d82) had no impairment of autophagy proteins in both tissues examined. However, when the animals were re-exposed to high-fat diet they had alteration in protein content of autophagy, when compared to animals with high fat diet without re-exposure. Thus, high fat diet seems to be essential for negative modulation of autophagy markers. In conclusion, the offspring of obese mothers presented early impairment of autophagy proteins in the liver and hypothalamus, which may be associated with the development of metabolic disorders in the offspring in adulthood / Mestrado / Metabolismo e Biologia Molecular / Mestra em Ciências da Nutrição e do Esporte e Metabolismo

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22365号 / 医博第4606号 / 新制||医||1043(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 岩田 想, 教授 川上 浩司, 教授 横出 正之 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

One of the major healthcare issues found almost worldwide, especially in the United States, is the obesity epidemic. Obesity is known to have deleterious effects on many body/organ systems. C1q TNF-related protein-3 (CTRP3) is effective at preventing high-fat diet-induced fatty liver. With these two factors taken into consideration, this study explores the possible effects of a high caloric diet on the muscle wall of the uterus, i.e., the myometrium, and how over-expression of CTRP3 may modify those effects. We hypothesize that consumption of excessive amounts of fat and sugar will have detrimental effects on the dual layers of the mouse myometrium. For this study, 17 mice were divided into 4 treatment groups: wild type/low fat diet, wild type/high fat diet, CTRP3-overexpressing/low fat diet, and CTRP3-overexpressing/high fat diet. The mice were placed on their respective diets at 7 weeks of age with a feeding duration of 12 weeks. At the conclusion of the feeding protocol, the female reproductive tissues were harvested and fixed in 4% paraformaldehyde and subsequently paraffin embedded. The uterine horns of each mouse were painstakingly paraffin embedded in a vertical position so that cross sections of the uterus could be obtained and measured. These 4µ sections were stained using standard H&E staining techniques and visualized under light microscopy. A randomization grid was utilized to determine measurement locations on the tissue. For each animal, 15 measurements were made of the outer longitudinal layer of the uterine horn, as well as 15 measurements of the inner circular layer, and 15 measurements of the thickness of the two layers combined. Two-way ANOVA was used to determine if any changes seen were statistically significant. At the time of the writing of this abstract, no appreciable differences have been found between the treatment groups, although there will be more data and final statistics completed before the presentation of our findings.

C1q TNF-related protein-3 (CTRP3) is effective at preventing high-fat diet-induced fatty liver; Recent studies have shown that overexpression of C1q TNF-related protein-3 (CTRP3) in mice fed a high alcohol diet can protect the animal from developing fatty liver disease, and therefore may be a possible treatment for alcoholic fatty liver disease (ALD). However, the possible effects of overexpression of CTRP3 on other tissue has not been widely investigated. If overexpression of CTRP3 proves to be harmful to other tissues, its use as a treatment for ALD would come into question. This study utilized kidney tissue from mice that were fed a high fat diet for 13-14 weeks. The feeding started when the mice were 7 weeks old and continued for 9 weeks. The mice were divided into 4 categories: wild-type/low fat diet, wild-type/high fat diet, transgenic/low fat diet, and transgenic/high fat diet. The kidneys were harvested and fixed in 4% paraformaldehyde and subsequently paraffin embedded. Sections were cut at 4µ and stained using three different staining techniques: standard H&E, Periodic Acid Schiff (PAS), and Masson’s Trichrome Staining. These three methods were utilized to better visualize possible effects on the tissue, i.e. changes in connective tissue deposition or basement membrane thicknesses, etc. Light microscopic examination of the tissues to date has revealed abnormalities in some of the kidney tubules in the transgenic high fat diet group. These same abnormalities have not been observed in the other treatment groups. This study is still in its early stages and much more in-depth investigation is needed to determine which of the tubules of the nephron are affected, and what this effect is. If this study confirms that overexpression of CTRP3 coupled with a high fat diet is harmful to kidney tissues, the use of CTRP3 in the treatment of ALD would require careful monitoring of the patient’s diet.

京都大学 / 0048 / 新制・課程博士 / 博士(人間健康科学) / 甲第18194号 / 人健博第11号 / 新制||人健||1(附属図書館) / 31052 / 京都大学大学院医学研究科人間健康科学系専攻 / (主査)教授 細田 公則, 教授 高桑 徹也, 教授 任 和子 / 学位規則第4条第1項該当 / Doctor of Human Health Sciences / Kyoto University / DFAM

Exocrine pancreatic insufficiency (EPI) is a major complication of cystic fibrosis. Conventional treatment involves the replacement of pancreatic enzymes and intake of a low fat diet. However, contrary to previous therapeutic strategies, a high fat diet may be beneficial in EPI patients. The present study investigated the effects of dietary supplementation with Creon 10 000 a pancreatic enzyme preparation, in conjunction with a high-fat diet, on growth performance, digestibility and absorption of fat in a pig model of EPI by the surgical ligation of the pancreatic duct in 6 male pigs (Swedish Landrace X Yorkshire X Hampshire). Following surgery, and for the duration of the experimental period, pigs were fed a high fat diet (twice daily). The experimental period lasted for 15 days during which blood, urine and faecal samples were collected. In the last 7 days of the experimental period (days 8-14), Creon 10 000 was included in the high fat meals. Urine and faecal samples were analysed for dry matter, crude protein and fat content. Plasma was used to assess the lipaemic index and the plasma lipid profiles. Treatment with Creon 10 000 significantly increased body mass (P = 0.016) and the digestibility of dry matter, crude protein as well as the co-efficient of fat absorption were also significantly improved following treatment (P<0.05). Creon 10 000 improved the lipaemic index values and significant changes in plasma free fatty acid and triglyceride concentrations were observed but not in cholesterol or high and low density lipoproteins. This study supports previous reports that the administration of pancreatic enzyme preparations together with a high fat meal is a beneficial strategy for the nutritional management of EPI.