Background. The kidney is one of the major organs involved in whole-body homeostasis and it is well understood that chronic renal impairment is further complicated with deranged carbohydrate metabolism, dyslipidemia, altered abdominal fat distribution and the activation of renin-angiotensin system (RAS). Recently, RAS blockades of angiotensinconverting enzyme inhibitor (ACEI) and angiotensin II receptor blocker (ARB) have been noticed for their potential effects on improve glucose and lipid metabolisms and lowering the risk of new-onset diabetes. However, underlying cellular and molecular mechanisms are not fully established. / Conclusions. (1) UNX induces progressive renal impairment and dysregulation of pancreatic and renal RAS in rats. (2) Pancreatic RAS activation leads to intra-islet fibrosis, insulin-secreting beta-cell deficit and insulin secretory deficiency. (3) Renal cortex RAS dysregulation induces ectopic adipocyte differentiation and lipid infiltration, in combination with lipodystrophy and lipid peroxidation, results to insulin resistance. (4) Pancreatic insulin-secretion deficit and insulin resistance contribute to the development of glucose intolerance and hyperglycemia. (5) Kidney impacting on glucose and lipid metabolism by affecting pancreatic islet and adipocyte, suggesting an essential role of the kidney in maintaining the whole-body homeostasis. (6) RAS blockade with ACEI or ARB may prevent the development of chronic renal impairment and glucose and lipid dysmetabolisms in UNX rats. (7) Common pathways modulating blood pressure, glucose and lipid metabolism warrant future studies for the better management of the global epidemic of metabolic syndrome. / Materials and methods. Chronic renal impairment and RAS disturbance were induced by unilateral nephrectomy (UNX) in adult Sprague-Dawley rats undergoing as long as 10 months of observation. Three-month old male rats were randomized into 4 groups: (1) sham operated control rats (n=10), (2) untreated UNX model rats (n=10), (3) ACEI---lisinopril treated UNX rats (n=10), and (4) ARB-olmesartan treated UNX rats (n=10). Blood glucose levels during fasting and oral glucose tolerance test (OGTT) conditions, lipids, insulin and renal function were measured at 3, 6, 8 and 10 months after operation. Histological changes of kidney, pancreas, liver, and adipose tissue were examined at 10 months post-operation. / Objectives. (1) To set up a rat model with persistent chronic renal impairment and RAS activation. (2) To examine changes of fasting blood glucose, glucose tolerance, blood lipids and insulin sensitivity. (3) To examine changes of pancreatic islets and the factors contributing to pancreatic islet damage such as RAS, transforming growth factor (TGF)-beta and alpha-smooth muscle actin (SMA). (4) To examine changes of systemic and renal adipose tissue and the factors contributing to adipopathy such as RAS, peroxisome proliferator-activated receptor-gamma (PPAR-gamma) and hydroxy-3-methylglutary coenzyme A reductase (HMGCR). (5) To investigate preventive effect of RAS blockades by the ACEI-lisinopril (4 mg/kg body weight) and ARB-olmesartan (4 mg/kg body weight) on the rat model of progressive renal deficiency. / Results. (1) UNX rats developed time-dependent progressive renal functional impairment and marked glomerulosclerosis and tubulointerstitial lesions. (2) UNX rats showed fasting hyperglycemia, progressive glucose intolerance, hyperlipidemia and insulin resistance. (3) UNX rats demonstrated insulin secretory deficiency in parallel to pancreatic islet fibrosis, beta-cell deficit, and overexpression of RAS components, TGF-beta, and alpha-SMA. (4) UNX rats displayed adipopathy evidenced by shifts the subcutaneous and visceral fats to the ectopic fat with lipid accumulation, lipofuscin pigmentation and adipocytes transformation. The adipopathy associated with down-regulation of AT1R and over-expression of angiotensin, AT2R, PPAR-gamma and HMGCR in the remnant kidney. (5) Treatment with lisinopril and olmesartan significantly attenuated the development of chronic renal impairment, RAS dysregulation and aberrant proteins expression, islet damage, adipose redistribution, and glucose and lipid dysmetabolism. / Sui, Yi. / Source: Dissertation Abstracts International, Volume: 70-06, Section: B, page: 3422. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2008. / Includes bibliographical references (leaves 195-220). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts in English and Chinese. / School code: 1307.

The gilthead seabream (Sparus aurata) and European seabass (Dicentrarchus labrax) require n-3 long-chain polyunsaturated fatty acids (LC-PUFA), eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), for optimal growth and health. Due to the rapid growth of global aquaculture the quantity of marine oils used in aquafeeds has been limited, yet the overall quantity of oil in an aquafeed has increased by the addition of vegetable oil (VO) to supply dietary energy. For aquaculture to continue to grow more fish must be produced with less marine ingredients, yet EPA and DHA must be maintained at levels above fish requirements. This project set out to re-evaluate the requirement for EPA and DHA in gilthead seabream and European seabass. Two dose-response studies were designed and executed where juvenile seabream and seabass were fed one of six levels of EPA+DHA (0.2 – 3.2 % as fed). Biometric data were collected and analysed to determine new requirement estimates for EPA+DHA for fish of two weight ranges (24 – 80 g and 80 – 200 g). The effects of the dietary LC-PUFA gradient on lipid composition and metabolism were also considered.

No description available.

Introduction : Chez les nouveau-nés prématurés, l’hyper-alimentation intraveineuse (HAIV) contribue à leur survie, mais elle est aussi une source importante de molécules oxydantes. L’absence d’une protection adéquate contre la lumière ambiante génère in vitro, via la photo-excitation de la riboflavine, du H2O2, des peroxydes organiques et un dérivé peroxydé de la vitamine C, l’ascorbylperoxyde (AscOOH). Plusieurs données du laboratoire associent l’infusion d’HAIV à des désordres lipidiques dans notre modèle animal. L’hypothèse est donc que l’AscOOH a un pouvoir oxydant et est responsable de certains des effets biologiques observés. Mes objectifs sont les suivants : 1) développer une méthode de dosage de l’AscOOH; 2) démontrer, à l’aide du modèle animal bien établi au laboratoire, des relations entre la concentration tissulaire de cette molécule et des paramètres métaboliques et l’état redox au foie et dans la circulation; et 3) confirmer l’effet physiologique de l’AscOOH dans un modèle cellulaire. Méthode : Différents étalons internes potentiels ont été testés pour le dosage de l’AscOOH par spectrométrie de masse après séparation sur HPLC (LC-MS). Les phases mobiles et conditions chromatographiques ont été optimisées. Pour l’objectif 2, des cobayes de 3 jours de vie (n=11) ont reçu par voie intraveineuse une dose d’AscOOH (entre 0 et 3,3mM). Les animaux ont été sacrifiés au 4e jour de traitement pour le prélèvement de tissus. Les concentrations tissulaires d’AscOOH ont été déterminées au LC-MS. La triglycéridémie et la cholestérolémie ont été mesurées à l’aide d’un kit commercial par spectrophotométrie. Le glutathion oxydé et réduit ont été mesurés par électrophorèse capillaire. Les relations linéaires obtenues sont exprimées par le ratio des carrés (r2), et traitées par ANOVA. Résultats : La validation du dosage de l’AscOOH par LC-MS a été réalisée. Chez les animaux, la concentration urinaire d’AscOOH par créatinine corrèle positivement avec la dose reçue, négativement avec la lipidémie, et négativement avec le redox sanguin et érythrocytaire, indiquant un milieu moins oxydé. Conclusion : La concentration urinaire d’AscOOH peut donc être un reflet de l’oxydation de l’HAIV en clinique. Nos données chez l’animal suggèrent une interaction de l’AscOOH avec le métabolisme hépatique produisant une chute de la concentration plasmatique de cholestérol et de triglycérides. Le modèle cellulaire n’a pas permis d’élucider le mécanisme moléculaire de l’action de l’AscOOH sur le métabolisme. / Introduction: Intravenous hyperalimentation (IVHA) often contributes to the survival of preterm newborns, but it is also an important source of oxidizing molecules. The lack of adequate protection from ambient light generates, in vitro, through the photo-excitation of riboflavin, H2O2, organic peroxides and a peroxidated derivative of vitamin C: ascorbylperoxide (AscOOH). Certain data from our laboratory linked the infusion of IVHA to lipid disorders in our animal model. The hypothesis is that AscOOH is an oxidant that is responsible for some of the biological effects observed. My objectives are: 1) to develop a method for quantitation of AscOOH, 2) to demonstrate, using the guinea pig model used by our laboratory, relations between the tissue concentration of this molecule and metabolic and redox parameters in the liver and plasma, and 3) to confirm the physiological effect of AscOOH in a cell culture model. Method: Different promising internal standards were tested for AscOOH quantitation by mass spectrometry after HPLC separation (LC-MS). Mobile phases and chromatography conditions have been optimized. For objective #2, 3 days old guinea pig pups (n = 11) received an intravenous dose of AscOOH (between 0 and 3.3mM). Animals were sacrificed on the 4th day of treatment for tissue gathering. Tissues AscOOH concentrations were determined by LC-MS. The triglyceride and cholesterol levels were measured by spectrophotometry using a commercial kit. The oxidized and reduced glutathione were measured by capillary electrophoresis. The linear relations obtained are expressed by the square of the correlation coefficient (r2), and processed by ANOVA. Results: The validation of the LC-MS method for AscOOH quantification has been achieved. In animals, the concentration of urinary AscOOH by creatinine correlates positively with the dose received, negatively with blood lipids, and negatively with blood and erythrocyte redox, indicating a less oxidized environment. Conclusion: The urinary AscOOH concentration may be a good indicator of the oxidation state of clinical IVHA. Our data in animals suggest an interaction between AscOOH and liver metabolism producing a drop in plasma concentration of cholesterol and triglycerides. The cell model was not able to clarify the molecular mechanism of AscOOH action on metabolism.

La première partie de mon travail de thèse a été de définir le rôle joué par le récepteur nucléaire Rev-erb alpha dans les mécanismes de synchronisation par la nourriture d’une horloge circadienne putative, non encore localisée, appelée « horloge alimentaire ». La seconde partie de mon travail a consisté à étudier la participation de Rev-erb alpha dans les régulations des métabolismes glucidique et lipidique. L’ensemble de nos données indique que le répresseur transcriptionnel Rev-erb alpha joue un rôle charnière dans les fonctions circadiennes ainsi que dans le métabolisme. En effet, d’un point de vue circadien, l’absence de Rev-erb alpha altère la synchronisation à l’heure des repas – démontré par une réduction des sorties comportementales et physiologiques de l’horloge alimentaire, ainsi que par l’absence d’ajustement du rythme de la protéine d’horloge PER2 dans l’oscillateur cérébelleux. Sur le plan métabolique, la délétion de ce gène modifie notamment le métabolisme des lipides – démontré par une accumulation excessive de tissu adipeux, une utilisation préférentielle des acides gras, ainsi qu’une perte de contrôle de l’expression de la Lipoprotéine lipase. / The work performed during this PhD thesis aimed at investigating the role of the transcriptional silencer Rev-erbα in both the circadian clockwork of the food-entrainable oscillator and metabolic regulations. Firstly, by evaluating food-anticipatory components in animals fed once a day at the same time, we showed that mice lacking Rev-erbα display a reduction in locomotor activity prior to food access compared to littermate controls. Accordingly, the rises in body temperature and corticosterone that anticipate mealtime are also diminished. Interestingly, daily p-ERK expression in hypothalamic regions and daily PER2 expression in the cerebellum of Rev-erbα KO mice are not phase-adjusted to feeding time. These results indicate that Rev-erbα participates in the integration of feeding signals and in food-seeking behaviors. Secondly, by investigating energy balance in fasted, normal chow or high-fat fed animals, we revealed that Rev-erbα KO mice exhibit greater reliance on lipid fuels as energy substrates, contributing to a mild hyperglycemic state. We also found that Lipoprotein lipase (Lpl) expression, is strongly up-regulated in peripheral tissues of Rev-erbα KO mice, predisposing mice to obesity. In this regard, we uncovered a new molecular pathway that ties clock-driven Lpl expression to energy homeostasis. These findings highlight the significance of daily Rev-erbα oscillations to prevent the appearance of the metabolic syndrome.In conclusion, we provide evidence that REV-ERBα may be a part of the food-entrainable oscillator clockwork that triggers food-anticipatory components, and represents a pivotal player to link the core clock machinery to metabolic pathways.

Made available in DSpace on 2016-06-02T19:22:09Z (GMT). No. of bitstreams: 1 5256.pdf: 6965922 bytes, checksum: 5df2e41fa93348b8ef396978016ce615 (MD5) Previous issue date: 2013-06-06 / Universidade Federal de Sao Carlos / Visceral obesity is an important risk factor cardiovascular diseases. After menopause, fat distribution shifts to visceral depots leading to lipid metabolism disorders and chronic lowgrade inflammation. Resistance training (RT) has beneficial effects in postmenopausal women, but its effectiveness in normalizing visceral adipose tissue disorders is not well understood. To address this questions adult female Sprague-Dawley rats were randomly divided into six groups (n=6 per group): sham-sedentary (SHAM), ovariectomized- sedentary (OVX), SHAM-trained (S-T), OVX-T. Sham or ovariectomy procedure was performed with 250 grams. Three weeks after the surgery, trained groups started 10 weeks of climbing in a 1.1-m vertical ladder with progressive load of 65%, 85%, 95% and 100% of rat s previous maximal overload. All animals were killed 92 days after being ovariectomized and trained animals were killed 48 hours after the last training session. Mesenteric, parametrial and subcutaneous adipose tissues were prepared for morphometric analyses in colidina and osmium tetroxide and part of mesenteric adipose tissue was stored for mRNA analyses by quantitative PCR. Resistance training prevented ovariectomy-induced higher body weight, food intake, feeding efficiency and mesenteric and subcutaneous, but not parametrial adipocyte area. In addition, RT avoided OVX-induced upregulation in the lipogenic genes PPAR-γ, SREBP-1c and its downstream targets SCD-1 and ACC gene expression. Further, RT increases the gene expression of mitochondrial β-oxidation enzyme CPT-1 in OVX rats. RT also prevented the chronic low-grade inflammation in OVX group, demonstrated by the presence of macrophage markers (F4/80 and CD11b), increase in activated Th1 cells (IFN-γ) and the proinflammatory cytokines (TNF-α, IL-1β and IL-6). RT additionally created an antiinflammatory (IL-10) environment. The present results reflect a powerful prophylactic effects of resistance training in preventing pathological effects of estrogen deficiency, both at physiological and molecular levels. Therefore, we suggest that this may be an advantageous low cost-effective treatment at post-menopausal phase, since appropriately prescribed. / A obesidade visceral é um importante fator de risco para doenças cardiovasculares. Após a menopausa, as gorduras passam a ser depositadas preferencialmente em locais viscerais e surgem desordens no metabolismo lipídico e um quadro inflamatório crônico de baixa intensidade. O treinamento resistido (TR) possui efeitos benéficos na pós-menopausa, mas não se sabe se ele é efetivo para normalizar as desordens do tecido adiposo visceral. Para pesquisar estas respostas, ratas Sprague-Dawley adultas foram divididas em 4 grupos (n=6 por grupo): Sham sedentárias (SHAM), ovariectomizadas sedentárias (OVX), sham treinadas (SHAM-TR) e OVX-TR. A cirurgia foi realizada quando as ratas estavam com 250 gramas. Após 3 semanas, os grupos treinados iniciaram um programa de treinamento de 10 semanas (3x por semana) de escalada (4-9 repetições) em uma escada vertical de 1,1 m de altura com sobrecargas progressivas de 65%, 85%, 95% and 100% da carga máxima do rato. As ratas foram sacrificadas 92 dias após a ovariectomia e 48 horas após a última sessão de treino. Os tecidos adiposos mesentérico, parametrial e subcutâneo foram preparados em colidina e tetróxido de ósmio para análises morfométricas e parte do tecido mesentérico foi armazenado para análises de expressão gênica por PCR quantitativa. O treinamento resistido evitou o aumento de massa corporal provocado pela ovariectomia, consumo alimentar, eficiência alimentar e a área dos adipócitos mesentérico e subcutâneo, mas não do parametrial. Além disso, evitou a alta expressão dos genes lipogênicos PPAR-γ, SREBP-1c e de seus genes alvos ACC e SCD-1. Adicionalmente, o TR, aumentou a expressão do gene da β-oxidação mitocondrial CPT-1 nas ratas OVX-TR. O TR também evitou a condição inflamatória de baixa intensidade instaurada nas ratas OVX, demonstrado pelo acúmulo de macrófagos (F4/80 e CD11b), aumento de células Th1 ativadas (IFN-γ) e de mediadores pó-inflamatórios (TNF-α, IL-1β e IL-6), além de criar um ambiente anti-inflamatório (IL-10). Os resultados deste trabalho refletem um significativo efeito profilático do treinamento resistido sobre parâmetros fisiológicos e moleculares na falta dos hormônios ovarianos. Portanto, podemos sugerir que este pode ser um vantajoso tratamento de baixo custo no período pós-menopausal, se adequadamente prescrito.

Introdução: A Organização Mundial da Saúde estimou em 2015 que 23 milhões de pessoas em todo o mundo sofrem de insuficiência cardíaca (IC), com taxas de mortalidade equivalentes às do câncer. Níveis mais elevados de HDL-colesterol têm sido associados com maior sobrevivência na IC. É consensual que as várias funções protetoras da HDL devem ser exploradas além da concentração de HDL-colesterol. Transferência de lípides para HDL, mediada por proteínas de transferência CETP e PLTP, é uma etapa importante no transporte reverso de colesterol e metabolismo de HDL.,Desenvolvemos um ensaio in vitro para avaliar as transferências de lípides para a HDL, mostrando que esse fenômeno é alterado em várias condições, como na doença arterial coronária, no diabetes mellitus e pelo estilo de vida sedentário. Recentemente, tem sido descrito que a HDL transporta pequenos RNAs não codificadores de proteína, os chamados microRNAs (miRNAs). Alguns miRNAs foram descritos como reguladores críticos do metabolismo das lipoproteínas. O objetivo deste estudo foi comparar lípides plasmáticos, transferência lipídica para HDL, perfil inflamatório, miRNAs relacionados ao metabolismo de lipoproteínas obtidos de pacientes com IC e de pacientes sem IC (sem-IC). Métodos: Quarenta e oito pacientes com IC foram avaliados, 25 em classe funcional NYHA I e II (IC-I/II) e 23 em NYHA III e IV (IC-III/IV), bem como 50 pacientes sem-IC pareados por gênero e idade. Todos os pacientes com IC apresentavam uma fração de ejeção <=40%. Foram determinadas as concentrações plasmáticas de CETP, LCAT, LDL oxidada (LDLox) e atividade de paraoxonase 1 (PON-1). Transferências de lípides para a HDL foi avaliada a partir da incubação de uma nanopartícula artificial com plasma total. A expressão de miRNAs circulantes envolvidos no metabolismo das lipoproteínas também foi analisada. Resultados: Os níveis de colesterol total, LDL e HDL e triglicérides não diferiram entre os três grupos. A concentração da apolipoproteína A-I foi menor no grupo IC-I/II em comparação ao grupo sem-IC (125±23 versus 142±19; p < 0,05), enquanto que a concentração da apolipoproteína B foi menor em ICIII/ IV comparado ao sem-IC (81±35 versus 114±40; p < 0,001). A transferência de colesterol esterificado (5,44±1,76 versus 6,26±0,85), fosfolípides (19,05±2,5 versus 20,21±1,45) e de triglicérides (6,29±2,05 versus 7,40±1,47) foi menor no grupo IC-III/IV do que no grupo sem-IC (p < 0,05). No entanto, não houve diferença nas transferências entre IC-I/II e sem-IC. A concentração de LDLox foi menor em ambos os grupos com IC comparados ao sem-IC (p < 0,0001). A massa de CETP foi menor em IC-III/IV do que em IC-I/II (2,77±1,3 versus 3,78±1,3; p=0,021). A concentração de LCAT e a atividade de PON-1 não foram diferentes entre os grupos. A análise da expressão dos miRNAs circulantes miR-33a, miR-144, miR-185, miR-125, miR-758, miR-26a, miR- 106b, miR-122 e miR-30c, mostrou-se significantemente aumentada nos indivíduos com IC em comparação aos indivíduos sem-IC, ao passo que o miR- 10b foi o único encontrado diminuído na IC comparado com indivíduos sem-IC (p=0,007). Conclusão: Em pacientes com IC mais severa e sintomática da IC, o processo de transferência de lípides para a HDL está deficiente, bem como alguns dos mecanismos que o regulam, e possivelmente estas alterações influenciem no transporte reverso do colesterol e nas funções protetoras da HDL desses pacientes / Background: World Health Organization estimated that there were twentythree million subjects worldwide suffering from heart failure (HF) in 2015, with mortality rates equivalent to those of cancer. Higher HDL-cholesterol levels have been associated with longer survival in HF. It is now consensual that the various protective functions of HDL should be explored beyond HDLcholesterol. Transfer of lipids to HDL, mediated by transfer proteins CETP and PLTP, is an important step in reverse cholesterol transport and HDL metabolism. Previously, we developed an in vitro assay to test those lipid transfers and showed that transfer of cholesterol to HDL is altered in several conditions, such as coronary artery disease (CAD), diabetes and sedentary lifestyle. Recently, HDL transports small non-coding RNA molecule, called micro RNAs (miRNAs). Some miRNA are critical regulators of lipoprotein metabolism. The aim of this study was compare plasma lipids, lipid transfers to HDL, inflammatory profile, miRNAs related to plasma lipids from patients with HF with those from patients with without HF (non-HF). Methods: Forty-eight HF patients were studied, 25 with functional class NYHA I and II (HF I/II) and 23 with NYHA III and IV (HF III/IV), as well as 50 non-HF patients matched for gender, age and BMI. All HF had ejection fraction <= 40%. CETP, LCAT, oxidized LDL (oxLDL) and paraoxonase 1 (PON-1) activity were determined. Transfers of lipids from a donor artificial nanoparticle to HDL was determined by an in vitro assay in which the emulsion was incubated with whole plasma. Expression of circulating miRNAs involved in cholesterol metabolism was also analyzed. Results: Total, LDL and HDL cholesterol and triglycerides did not differ among the 3 groups. Apolipoprotein A-I was lower in NYHA I/II group compared to non- HF (125±23 versus 142±19; p < 0.05) and apo B was lower in NYHA III/IV group compared to non-HF (81±35 versus 114±40, p < 0.001). The transfer of esterified cholesterol (5.44±1.76 versus 6.26±0.85), phospholipids (19.05±2.5 versus 20.21±1.45) and of triglycerides (6.29±2.05 versus 7.40±1.47) to HDL was lower in HF-III/IV than in non-HF (p < 0.05), but lipid transfers were not different between HF-I/II and non-HF. oxLDL was lower in both HF groups compared to non-HF (p < 0.0001). CETP mass was lower in HF-III/IV than in HF-I/II (2.77±1.3 versus 3.78±1.3; p=0.021). LCAT and PON-1 activity was not different among the groups. Regarding to miRNA, miR-33a, miR-144, miR-185, miR-125, miR- 758, miR-26a, miR-106b, miR-122 e miR-30c were significantly increased in HF compared to non-HF subjects, whereas miR-10b was the only one found to be decreased in HF compared to non-HF subjects (p=0.007). Conclusion: In patients with the more severe and symptomatic HF, the lipid transfer to HDL is deficient, as well as some mechanisms that regulate it, and possibly these changes influence reverse cholesterol transport and the protective functions of HDL in these patients

Estudos recentes têm demonstrado uma participação importante do receptor toll-like 4 (TLR4) na evolução de doenças envolvendo desordens metabólicas, como a doença do fígado gorduroso não-alcoólico (NAFLD). No entanto, as alterações do metabolismo lipídico que poderiam ser influenciadas pela ativação do TLR4 são desconhecidas. Neste estudo propomos caracterizar o papel do receptor TLR4 no metabolismo de lipídios no fígado de camundongos deficientes para o receptor de LDL, um modelo que desenvolve NAFLD quando submetido a uma dieta rica em gordura saturada e colesterol. Camundongos controle (C57 black6), deficientes para o receptor de LDL (LDLrKO), deficientes para o receptor TLR4 (TLR4KO) ou deficientes para ambos (duplo KO) receberam dieta controle ou hiperlipídica por quatro, oito ou doze semanas. Após o tratamento e sacrifício dos animais, avaliamos o perfil de lipídios plasmáticos, o conteúdo de lipídios do fígado e a expressão gênica de enzimas relacionadas à síntese e degradação de triglicerídeos (TG) e colesterol no fígado. O perfil inflamatório no fígado também foi avaliado. A dieta hiperlipídica induziu uma hipertrigliceridemia e hipercolesterolemia nos animais LDLr KO e duplo KO, sendo que o grupo duplo KO apresentou níveis séricos inferiores de triglicérides (TG) e ácidos graxos livres a partir de oito semanas de tratamento em comparação aos animais LDLrKO. A dieta hiperlipídica também induziu um aumento significativo no conteúdo de TG e de colesterol no fígado de todos os grupos. Na análise da expressão gênica não foram encontradas diferenças na expressão de proteínas relacionadas à síntese de triglicérides e colesterol (ApoB100, MTTP, GPAT1 e GPAT4) entre os grupos. Porém houve aumento significativo na expressão de proteínas relacionadas à oxidação de ácidos graxos (CPT1, MTP, ACOX, PBE, tiolase) e à síntese de ácidos biliares (CYP7a1) no grupo duplo KO em comparação ao grupo LDLr KO. No perfil inflamatório, a expressão de F4/80 demonstrou infiltração de macrófagos significativamente elevada no grupo LDLrKO tratado com a dieta hiperlipídica comparada a todos os outros grupos. No entanto, houve maior expressão de IL-6, IL-1beta e TNF-alfa no grupo duplo KO em comparação ao grupo LDLr KO. Nossos dados sugerem que a ativação do TLR4 no fígado de animais alimentados com uma dieta hiperlipídica pode contribuir para o acúmulo de lipídios e início da esteatose hepática. Estratégias para a inativação hepática do TLR4 podem diminuir a NAFLD não somente devido a diminuição da inflamação, mas por aumentar a oxidação de ácidos graxos no fígado / Recent studies have shown an important role of toll-like receptor 4 (TLR4) in the evolution of diseases involving metabolic disorders, such as non-alcoholic fatty liver disease (NAFLD). However, changes in lipid metabolism regulated by TLR4 activation are still unknown. In this study, we characterized the role of TLR4 receptor in hepatic lipid metabolism of mice deficient for the LDL receptor, a model that develops NAFLD when exposed to a diet rich in saturated fat and cholesterol. We investigated the role of TLR4 activation in the pathogenesis of diet-induced NAFLD by crossing LDLr KO mice with the TLR4 knockout mice (double KO). Animals were fed for 4, 8 or 12 weeks with high-fat diet (HFD) containing 18% saturated fat and 1.25% cholesterol. We evaluated plasma lipid profile, hepatic lipid content and gene expression of enzymes related to the synthesis and degradation of triglycerides and cholesterol in the liver. Liver inflammatory status was also investigated. We observed that HFD induced hypertriglyceri-demia and hypercholesterolemia in LDLr KO and double KO mice, but double KO animals presented lower serum levels of triglycerides and free fatty acids after eight weeks of treatment. HFD also induced a significant increase in liver contents of triglycerides (TG) and of cholesterol in all groups. We did not find differences in the expression of proteins related to triglycerides and cholesterol synthesis (ApoB100, MTTP, GPAT1, GPAT4) between the groups. However, we observed a significant increase in the expression of proteins related to fatty acid oxidation (CPT1, MTP, ACOX, PBE, tiolase ) and bile acid synthesis (CYP7a1) in double KO group in comparison to LDLr KO. Regarding the inflammatory process, F4/80 expression was elevated in LDLr KO mice fed HFD when compared to all groups. On the other hand, IL-6, IL-1beta e TNF-alfa expression was induced by HFD only in double KO mice. Taken together, our results show that TLR4 activation in liver from mice fed on a high-fat diet may contribute to lipid accumulation and steatosis onset. Strategies regarding localized TLR4 inactivation may increase the oxidation of fatty acids and improve NAFLD not only due to decreased inflammation

Abstract Uterine leiomyomas are the most common benign tumours in females. They are myometrial neoplasms, may present single or multiple, and may be located in various sites of the uterus. Leiomyomas distort the uterine cavity and the uterus itself, causing abnormal vaginal bleeding, reduced fertility and also pelvic pressure and pain symptoms. The aim of this study was to elaborate current knowledge on familial uterine leiomyomas and to explore the possible association between uterine leiomyoma and cardiovascular disease risk factors, and also the association between leiomyomas and endometriosis. The natural history of familial uterine leiomyoma study showed significant differences between familial and non-familial leiomyoma cases, familial cases having more severe clinical characteristics. They presented with multiple uterine leiomyomas and were more often symptomatic. They were also diagnosed at a younger age. The prevalence study on uterine leiomyomas and endometriosis offered confirmation of an association between the diseases. Uterine leiomyomas and endometriosis seem to decrease female fertility independently of each other. Uterine leiomyomas related to the hereditary leiomyomatosis and renal cell cancer (HLRCC) tumour syndrome were studied in regard to their clinical characteristics and immunophenotype. The study provided evidence that women with HLRCC may be identified through distinct leiomyoma clinical characteristics, and routine-use IHC of CD34 and Bcl-2. Distinguishing these leiomyoma cases from sporadic ones may identify families affected by fumarate hydratase (fumarase, FH) mutation. Uterine leiomyoma and cardiovascular disease risk factors were studied in The Northern Finland Birth Cohort 1966 (NFBC1966). The study showed an association between leiomyomas and raised cardiovascular disease risk factors, serum lipids and metabolic syndrome in particular. These findings may suggest that there are shared predisposing factors underlying both uterine leiomyomas and adverse metabolic and cardiac disease risks, or that metabolic factors have a role in biological mechanisms underlying leiomyoma development. This study provides novel information on clinical characteristics of familial uterine leiomyomas and on the immunophenotype of HLRCC-related leiomyomas. The study also offers significant confirmation of associations between uterine leiomyomas and both endometriosis and several CVD risk factors. / Tiivistelmä Kohdun leiomyoomat ovat naisten yleisin hyvänlaatuinen kasvain. Ne ovat myometriumin neoplastisia muutoksia ja ne ilmenevät joko yksittäisinä tai monilukuisina, ja ne voivat sijaita missä tahansa kohdun myometriumia. Leiomyoomat muuttavat kohdun ja kohtuontelon säännöllistä muotoa. Lisäksi ne aiheuttavat vuotohäiriöitä, alentunutta hedelmällisyyttä, ja lantion alueen painetta ja kipua. Tämän tutkimuksen tavoitteena oli laajentaa nykyistä tietämystä suvuittain esiintyvistä kohdun leiomyoomista ja selvittää mahdollista leiomyoomien ja kardiovaskulaaritautiriskin assosiaatiota, ja lisäksi selvittää leiomyoomien ja endometrioosin assosiaatiota. Suvuittain esiintyvien kohdun leiomyoomien taudinkulkua selvittävässä tutkimuksessa osoitettiin merkittäviä eroja suvuittain ja ei-suvuittain esiintyvien leiomyoomien välillä. Suvuittain esiintyvien leiomyoomien kliininen taudinkuva oli vaikeampi, leiomyoomia oli kohdussa useampia ja ne aiheuttivat useammin oireita ja lisäksi ne diagnosoitiin nuoremmalla iällä. Kohdun leiomyoomien ja endometrioosin yleisyyttä selvittävä tutkimus antoi lisävahvistusta sille havainnolle, että nämä taudit assosioivat keskenään. Tutkimustuloksen mukaan leiomyoomat ja endometrioosi vähentävät naisen hedelmällisyyttä toisistaan riippumatta. Perinnöllinen kohdun leiomyomatoosi ja munuaissyöpä (hereditary leiomyomatosis and renal cell cancer, HLRCC) -kasvainoireyhtymään liittyvän kohdun leiomyoomia selvittävän tutkimuksen tuloksien mukaan HLRCC-naisten kohdun leiomyoomien kliiniset ominaisuudet poikkeavat satunnaisesti esiintyvien leiomyoomien ominaisuuksista. Naisella HLRCC voitaisiinkin tunnistaa näiden poikkeavien ominaisuuksien perusteella, sekä immunohistokemiallisilla värjäyksillä CD34 ja Bcl-2. Fumaraattihydrataasi (fumaraasi, FH) -geenin mutaatiota kantava suku voitaisiin siten tunnistaa yksittäisen HLRCC leiomyoomatapauksen avulla. Pohjois-Suomen syntymäkohortti 1966 (Northern Finland Birth Cohort 1966, NFBC1966) tutkittiin kohdun leiomyoomia ja kardiovaskulaarisairauden riskitekijöitä. Tutkimustuloksien perusteella kohdun leiomyoomat assosioivat koholla olevien kardiovaskulaarisairauden riskien kanssa, erityisesti seerumin lipidien ja metabolisen syndrooman suhteen. Näiden tutkimustulosten perusteella voidaan esittää, että leiomyoomien ja terveydelle epäedullisen metabolian ja kardiovaskulaaritaudin riskien taustalla on mahdollisesti joitain yhteisiä altistavia tekijöitä, tai että metabolisilla tekijöillä on rooli kohdun leiomyoomien tautimekanismissa. Tämä tutkimus on tuottanut uutta tietoa suvuittain esiintyvien kohdun leiomyoomien kliinisestä taudinkuvasta ja HLRCC:n liittyvien leiomyoomien immunofenotyypistä. Lisäksi tämä tutkimus esittää lisävahvistusta kohdun leiomyoomien ja endometrioosin assosiaatiolle sekä useille kardiovaskulaaririskitekijöille.

Energy homeostasis is an important physiological mechanism essential for balancingenergy flow through the living systems by managing overall metabolism in the body. Thus, energy homeostasis is under a tight control by means of extremely well-regulated energy metabolism. One of the most common metabolic disorders that occur following disruption in energy homeostasis mechanisms is obesity. Obese individuals develop insulin resistance in the peripheral tissues (fat and muscle) and may also include non-alcoholic fatty liver disease. Insulin resistance is the primary factor responsible for the development of type 2 diabetes mellitus (T2D). Towards control and management of T2D condition, insulin, drugs that regulate the insulin sensitivity and drugs that regulate glucose metabolism are widely used. Repeated insulin administration is painful, expensive and requires constant glucose monitoring while other drugs have various limitations and side effects. Therefore, there is wide scope development of new anti-diabetic molecules for effective management of T2D. Studies related to energy metabolism are necessary to understand the cause of such disorders and improve existing methods to manage metabolic abnormalities. Animal models to understand such metabolic disorders have been developed by chemical treatments and genetic modifications, but diet-induced obese (DIO) animal models appear to be the better among all the models reported. DIO animal models are known to most closely mimic the physiological situation. Apart from the experimental model system studies have been conducted under physiological conditions to gain knowledge on possible mechanisms behind energy balance maintained and established during extreme situations such as pregnancy and lactation. To support fetal growth and milk synthesis several metabolic adjustments occur during pregnancy and lactation without the major disruption in the maternal energy homeostasis. In the present study, to gain knowledge on the mother’s body glucose, lipid management and insulin responses throughout the gestation and lactation periods analyses were carried out during at different stages of pregnancy and lactation in rats. It was observed that during pregnancy, the dam developed insulin resistance in peripheral tissues with decreased activation of insulin pathway and reduced glucose utilization while the liver remained unaffected. Although, as soon as the lactation began, peripheral tissue such as muscle developed increased insulin sensitivity associated with increased expression of glucose transporter gene and higher glucose metabolism. The reversal of insulin response in the muscle tissue observed during lactation appears to be a suitable model system for understanding the process by which the body undergoes a transition from insulin resistant state to sensitive state under a physiological condition. Interestingly, early lactation period is known to have much lower levels of insulin available to act upon peripheral tissues. Factors involved in this transition could be potential therapeutic agents for control of T2D, since during early stages of T2D muscle appears to be the first metabolic organ to exhibit resistance to insulin. The undercarboxylated osteocalcin (UNOC) has been reported to function as anti-diabetic molecule. UNOC is released from skeletal system during bone turnover, especially due to resorption process. Experiments were carried out to examine the role of UNOC during the transition from insulin resistant state of pregnancy to sensitive state of lactation period. It was observed that UNOC levels were lower during pregnancy, but increased during early lactation (day 3 to 6 of lactation). The increased UNOC levels seen during early lactation was higher than the levels observed in non-pregnant, non-lactating (NPNL) rats and the UNOC levels decreased following removal of pups immediately after parturition. It was noted that altering UNOC levels during early lactation altered the insulin response of the whole body and muscle transporter-4 expression (glut4) of lactating rats. A significant increase in bone turnover was also observed during lactation compared to NPNL and pregnant rats. The data suggest that increased bone turnover leads to increased UNOC levels in blood during lactation. Estrogen is known as bone protector molecule which acts via its receptors, estrogen receptor α and β (ERα and β). It was reported that ERβ is a dominant regulator of estrogen signaling when both the receptors of estrogen i.e. ERα and ERβ coexist in the target tissue and estrogen levels are relatively higher. Compared to NPNL rats estrogen levels have shown to be higher during late pregnancy and lower during early lactation. It was observed that liver and adipose tissues largely express ERα, but the muscle showed expression of both the receptors in NPNL rats indicating that muscle is the metabolic tissue that may be modulated by both the receptors. It has been reported that ERβ suppresses ERα action on glut4 transcription in the myocytes. It is possible that the altered ERs ratio modulates glut4 expression during late pregnancy and early lactation. The receptor expression ratio data indicated that muscle is an ERβ dominant during late pregnancy, while it is ERα dominant during early lactation. Further, alteration in UNOC levels during early lactation changed ERs ratio but not sufficient enough to alter the ER dominance, indicating lack of effect of UNOC on ER dominance during early lactation. Experiments were conducted to alter insulin sensitivity during early lactation to extrapolate physiological findings to a pathological condition of the DIO model by feeding rats with high-fat diet (HFD). During early lactation, HFD dams had lower insulin response, lower circulatory UNOC level and lower UNOC receptor (GPRC6A) expression in the muscle. Gene expression of muscle glut4 was lower in HFD rats and the tissue remained ERα dominant indicating no role of HFD on ERs ratio in muscle during early lactation. UNOC has been found to have negative effect on lipid accumulation. During pregnancy, lipid accumulation is one of the first events essential for proper fetal development. Since UNOC levels were suppressed during pregnancy, experiments were carried out to examine relevance of UNOC suppression on lipid accumulation during early pregnancy. For this purpose, pharmacological approaches were utilized to alter UNOC levels during early pregnancy. It was observed that the transient elevation of UNOC levels caused decrease in maternal fat depots without changing circulatory triacylglyceride (TAG) levels. In experiments that decreased UNOC levels in NPNL state to mimic lower levels of UNOC present during early pregnancy, it was found fat storage was higher and TG was found to be lowered in the circulation. These results indicate that UNOC can cause a reduction in fat accumulation and TG levels but UNOC effects on TG levels, was not observed during pregnancy. The data taken together suggest that suppression of UNOC is required for better fat deposition in the mother’s body. Although, some studies have indicated an insulin response transition occurring during pregnancy to lactation, but the factors involved in this transition have not been reported. This report discusses about the factors such as UNOC and ERs and their involvement in the transition process. UNOC role has been studied in genetically modified models and in metabolic disorders such as obesity model system and evidence for physiological role of UNOC would further support its candidature as anti-diabetic molecule. The present research work is the first report to detail relevance of UNOC in physiological conditions such as pregnancy and lactation for glucose and lipid management.