Fundação Carlos Chagas Filho de Amparo a Pesquisa do Estado do Rio de Janeiro / Este trabalho teve como objetivo investigar os efeitos da telmisartana (agonista PPAR-gama parcial), losartana (puro bloqueador do receptor AT1 da angiotensina II) e rosiglitazona (agonista PPAR-gama) em modelo experimental de síndrome metabólica. Os alvos do estudo foram a pressão arterial, metabolismo de carboidratos, resistência insulínica, inflamação, tecido adiposo e fígado. Camundongos C57BL/6 (a partir de 3 meses de idade) foram alimentados com dieta padrão (SC, n = 10) ou dieta hiperlipídica rica em sal (HFHS, n = 40) por 12 semanas. Após esse tempo, os animais do grupo HFHS foram subdivididos em 4 grupos (n = 10): HFHS (sem tratamento), ROSI (HFHS tratado com rosiglitazona), TELM (HFHS tratado com telmisartana) e LOS (HFHS tratado com losartana) por 5 semanas. O grupo HFHS apresentou um significante ganho de peso e aumento da pressão arterial sistólica, hiperinsulinemia com resistência insulínica, hiperleptinemia, hipertrofia de adipócitos bem como um quadro de esteatose hepática e níveis aumentados da citocina inflamatória interleucina-6 (IL-6). Os animais tratados com telmisartana chegou ao final do experimento com massa corporal similar ao grupo SC, com reversão do quadro de resistência insulínica, com pressão arterial normal, adipócitos de tamanho normal e sem apresentar esteatose hepática. Além disso, o tratamento com telmisartana aumentou a expressão de PPARγ e adiponectina no tecido adiposo epididimal. A expressão da proteína desacopladora-1 (UCP-1) no tecido adiposo branco (TAB) também foi aumentada. O tratamento com losartana diminuiu a pressão arterial para valores normais, porém com menores efeitos nos parâmetros metabólicos dos animais. O presente modelo experimental de ganho de peso e hipertensão induzidos por dieta mimetiza a síndrome metabólica humana. Neste modelo, a telmisartana aumentou a expressão de UCP-1 no TAB, preveniu o ganho de peso e melhorou a sensibilidade à insulina e a esteatose hepática dos camundongos C57BL/6, provavelmente devido à ativação PPAR-gama. / The study aimed to investigate the effects of telmisartan (a partial PPAR gamma agonist), losartan (a pure angiotensin II receptor blocker) and rosiglitazone (PPAR gamma agonist) in a mice model of metabolic syndrome (MetS). The targets of this study were blood pressure (BP), carbohydrate metabolism, insulin resistance, inflammation, white adipose tissue (WAT) and liver. Male C57BL/6 mice were studied over 17 weeks after being separated into two major groups according to diet: standard chow (SC, 10% fat, n = 10) or high-fat high-salt chow (HFHS, 60% fat, 7% salt, n = 40). In the last 5 weeks of the experiment, the HFHS group was divided into four groups (n = 10): untreated HFHS, ROSI (HFHS plus rosiglitazone), TELM (HFHS plus telmisartan), and LOS (HFHS plus losartan). The HFHS group had significantly greater body mass and BP, in addition to hyperinsulinemia with insulin resistance, hyperleptinemia, adipocyte hypertrophy and hepatic steatosis as well as increased inflammatory cytokine levels. Animals treated with telmisartan had body weights similar to the SC group, in addition to reversed insulin resistance, reduced hypertension, reduced adipocyte hypertrophy, ameliorates hepatic steatosis and decreased IL-6. Telmisartan increased PPARγ and adiponectin expression in white adipose tissue. Interestingly, the expression of UCP-1 in white adipose tissue was also increased by treatment with telmisartan. Losartan decreased BP but had smaller effects on metabolic parameters. The present model of diet-induced weight gain and hypertension in mice mimics human features of MetS. In this model, telmisartan enhances UCP-1 expression in WAT, prevented weight gain and ameliorates insulin sensitivity and hepatic steatosis in C57Bl/6 mice, probably due to PPAR gamma activation.

No description available.

Pubertal timing displays wide, normally distributed variation in a healthy population of sexually maturing adolescents. However, like many complex traits, factors contributing to the variation are not well understood. Epigenetic regulation may contribute to some of the population variation. The role that epigenetics, specifically DNA methylation and histone acetylation, may play in regulating pubertal timing was investigated in C57BL/6 female mice: investigating whether population variation in pubertal timing among inbred mice could be explained by environmental factors; whether perturbing the epigenome using a histone deacetylase inhibitor or methyl-donor would alter pubertal timing; and examining genome-wide methylation patterns in hypothalami of early versus late maturing mice. Results demonstrate that measurable micro-environmental factors have only negligible effects on pubertal timing; pubertal timing was significantly altered by administration of epigenetic modifying agents; differences in methylation patterns are subtle. This initial evidence supports the involvement of epigenetic mechanisms in regulating pubertal timing.

Pubertal timing displays wide, normally distributed variation in a healthy population of sexually maturing adolescents. However, like many complex traits, factors contributing to the variation are not well understood. Epigenetic regulation may contribute to some of the population variation. The role that epigenetics, specifically DNA methylation and histone acetylation, may play in regulating pubertal timing was investigated in C57BL/6 female mice: investigating whether population variation in pubertal timing among inbred mice could be explained by environmental factors; whether perturbing the epigenome using a histone deacetylase inhibitor or methyl-donor would alter pubertal timing; and examining genome-wide methylation patterns in hypothalami of early versus late maturing mice. Results demonstrate that measurable micro-environmental factors have only negligible effects on pubertal timing; pubertal timing was significantly altered by administration of epigenetic modifying agents; differences in methylation patterns are subtle. This initial evidence supports the involvement of epigenetic mechanisms in regulating pubertal timing.

PhD (Microbiology) / Department of Microbiology / Diarrhoea is still a major problem worldwide. Enteric pathogens such as Enteroaggregative E. coli (EAEC), Enteropathogenic E. coli (EPEC) and Enterotoxigenic E. coli (ETEC) have been reported to cause diarrhoea in children under the age of 5 years. The incidences of these pathogens are due to factors such as poor water quality, sanitation and hygiene practices. Infections with these pathogens result in diarrhoea and have been reported to result in severe disease outcomes more especially in children under 2 years of age. EPEC infections have been well studied using in vitro analyses, with studies highlighting the adherence traits, proteins and virulence genes involved in pathogenesis and inflammatory responses. EPEC is characterized by localized adherence with microcolony formation at the site of infection. In vivo studies have reported on human EPEC infection. However, the current animal models have not been able to replicate clinical outcomes (such as diarrhoea and weigh loss) of EPEC infection similar to humans. Therefore, there is still a need for a suitable small animal model that mimic clinical outcomes of human EPEC infections in vivo. Children living in poor environmental conditions are more susceptible to diarrhoeal pathogens. Furthermore, the incidences of children being exposed to co-infections (more than one pathogen at the same time) is relatively high. The EAEC/EPEC (A/P) and EPEC/ETEC (P/T) co-infections have been increasingly detected in children with and without diarrhoea. It has been suggested that patients infected with these co-infections might result in severe disease outcome than those infected with single pathogens. Pathogens are constantly evolving and the microbe-microbe interaction in the host can result in these pathogens competing for the same niche and thus result in increased virulence. Interaction of co-infections can lead to increased inflammatory responses thus affecting the infected host. The first objective of this study was to develop an EPEC murine model using weaned C57BL/6 mice that have been pretreated with antibiotic cocktail. Mice were orally infected with wild-type (WT) typical EPEC, bfp- and escN mutant strains. The WT had transient weight loss and wet stools with mucous; and the bfp- infected mice also had transient weight loss and bloody stool appearance. Increase in inflammatory biomarkers MPO, LCN-2, CRP, IL-6 and SAA were observed in the WT and bfp- infected mice. The mice infected with escN mutant did not exhibit any weight changes and the stools were similar to the uninfected mice. Furthermore, no inflammatory biomarkers were observed in mice infected with the escN mutant. Metabolic perturbations were observed in WT EPEC infected mice at day 3 post infection with the TCA cycle metabolites (reduced succinate, citrate, fumarate, cis-aconitate) being excreted at lower quantities indicating that the energy production in the infected mice was greatly affected. The second objective of this study was to determine the interaction between the P/T coinfections using in vitro and in vivo analyses. In vitro, human colorectal tumour 8 (HCT-8) cells were infected with single strains of ETEC, EPEC and both the pathogens and incubated for 3 hours. After infection the cells were analysed for bacterial adherence using real-time PCR. The single strains adhered at the same rate similar to the P/T coinfected cells. IL-8, as a marker of inflammatory response, was measured using ELISA. The results indicated that the P/T co-infected cells had a significant increase in IL-8 response higher than the single infections. The P/T co-infections were further analysed in vivo using the EPEC murine model developed in this study. Interestingly, mice infected with P/T co-infections developed severe diarrhoea accompanied with significant increased weight loss and some mice died during the 3-day infection period. The inflammatory responses MPO, LCN-2 and SAA were higher in the co-infected mice indicating a synergistic effect. The bfp and eltA virulence genes were significantly increased in the P/T co-infections. The third objective of this study was to determine the interaction between A/P coinfections using in vitro and in vivo analyses. HeLa cells and HCT-8 cells were infected with EAEC, EPEC and both the pathogens at the same time in order to determine adherence and inflammatory responses. EAEC adherence was higher than EPEC and A/P co-infections adherence. A/P co-infections did not have increased IL-8 response in HCT-8 cells when compared to EAEC alone. The virulence genes involved in EPEC adherence and Type 3 Secretion System (bfp, eae, tir, ler, per, espB and espA) were significantly reduced in A/P co-infected cells. An interesting adherence trait was observed between the A/P co-infections in HeLA cells, EAEC was found to adhere around EPEC altering the localized adherence pattern. The A/P co-infections were further analysed using the EPEC murine model developed in this study. The A/P infected mice had diminished weight changes and EAEC shedding was enhanced when EPEC was present. Faecal inflammatory biomarkers MPO and LCN-2 in A/P infected mice did not have any additive effect. The findings of this study contributed significantly to the knowledge of human EPEC infection in weaned C57BL/6 mice, highlighting clinical outcomes, inflammatory responses and metabolic perturbations. Furthermore, this study also highlighted the interaction of P/T and A/P co-infections using in vitro and in vivo analyses in order to determine the disease severity and outcomes. It was observed in this study that coinfections can result in either synergistic or antagonistic effects. Further studies are therefore, required in order to understand the underlying mechanisms that are involved during co-infections and this can further assist in the development of therapeutic interventions. / NRF