Objetivos: Leptina e grelina são hormonios relacionados a regulação da ingestão alimentar e consequentemente o controle de peso. Como tem sido descrito até agora, os valores normais desses hormônios estão sujeitos a grande variabilidade. Devido a falta de estudos que contemplem os valores totais de grelina total, grelina acilada, leptina, insulina e glicose em crianças, esse estudo tem o objetivo de avaliar esses hormônios e comparar com as variáveis antropométricas em crianças eutróficas de 4 meses a 10 anos. Pacientes e métodos: É um estudo transversal que avaliou 111 crianças de 4 meses a 10 anos de idade. Foram consideradas eutróficas crianças de até 5 anos com valores de IMC para idade entre -2,0 e +2,0 escores-z, e crianças maiores de 5 anos com valores entre -2,0 e +1,0 escores-z, de acordo com a OMS 2009. Os participantes foram selecionados de uma população saudável com pequeno procedimento cirúrgico ou revisão de saúde programada. As crianças foram categorizadas em três grupos: V 24 meses (n = 10); 25-60 meses (n = 22); > 60 meses (n = 27). As amostras de sangue foram coletadas com um jejum mínimo de 3 horas e máximo de 14 horas. As concentrações de grelina total, grelina acilada e leptina foram dosadas pelo kit commercial ELISA (Linco Research, St Charles-MI, US). A concentração de Insulina foi testada com o kit ELISA (Diagnostics System Laboratories, INC Webster-TX, USA) e a de Glicose com o kit comercial GlicosePAP liquiform® (Labtest Diagnóstica – Lagoa Santa-MG, Brasil). Variáveis que apresentaram distribuição normal foram comparadas com o teste t de Student e analizadas com ANOVA, e as que apresentaram distribuição assimétrica foram comparadas pelo teste de Kruskal-Wallis e analizadas pelo teste de Mann-Whitney. As correlações foram testadas pelos coeficientes de Pearson ou Spearman. O nível de significância utilizado foi 0,05. Resultados: A média de idade foi 60 meses (mín-máx 35-91 meses). A mediana (percentis 25-75) dos valores de grelina acilada, grelina total, grelina desacilada, leptina e insulina foram 1,374 pg/ml (949-1,875); 288 pg/ml (212-450); 1,086 pg/ml (711-1,393); 1,53 ng/ml (1,11-2,77) e 15,5 ^U/ml (12,1-23,5), respectivamente. A média ± DP da glicose foi 88,6 ± 13,1. A média do escore-z do IMC/I encontrada foi -0,16 ± 0,78 para meninos e -0,04 ± 0,83 para meninas. Foi encontrada uma correlação inversa da grelina total, acilada e desacilada com a idade [rs= -0,455 (P < 0,001); rs= -0,313 (P = 0,001); rs= -0,484 (P < 0,001)], respectivamente. Ocorreu uma relação direta da leptina com IMC/I e com a idade [(rs = 0,190 (P = 0,046); rs = 0,429 (P < 0,001)], respectivamente. Conclusões: Os resultados observados nas dosagens e as correlações encontradas de grelina com faixa etária e leptina com sexo feminino e parâmetros antropométricos podem ser úteis para comparação com crianças que são portadoras de alterações de apetite. / Objective: Leptin and ghrelin are hormones related to regulation of food intake and consequently body weight control. Normal values of these hormones are subject to a great variability as has been described so far. Due to the lack of studies regarding plasma levels of total and acylated ghrelin and serum levels of leptin, insulin and glucose in young children, this study aimed to assess these hormones and compare with body mass index (BMI) in healthy well-nourish children aged 4 months -10 years old. Methods: Cross-sectional study evaluated 118 children aged 4-129 months old with BMI-for-age values were within -2.0 and +2.0 SD z-scores for children under 5 years and within -2.0 and +1.0 SD for children over five years, according to WHO 2009. Subjects were enrolled from a population referred to minimal surgeries and routine medical check up. All subjects were categorized into 3 groups: d 24 months (n = 10); 25-60 months (n = 22); > 60 months (n = 27). Blood samples were collected following a minimum of 3-hour and a maximum of 14 hours fasting period. Total and acylated ghrelin and leptin concentrations were assessed by ELISA commercial kit (Linco Research, St Charles-MI, US). Insulin was assessed by ELISA commercial kit (Diagnostics System Laboratories, INC Webster-TX, USA) and glucose by ELISA commercial kit GlicosePAP liquiform® (Labtest Diagnóstica – Lagoa Santa-MG, Brazil). Variables with normal distribution were compared by Student´s t test and analysis of variance (ANOVA). Variables with asymmetric distribution were analyzed by Kruskal-Wallis and Mann-Whitney U tests. Correlation between continuous variables was assessed by Pearson or Spearman correlation coefficient test. The level of significance used was 0.05. Results: Total and acyl ghrelin, leptin and glucose were detected in all samples with a median (P 25-75) concentration in the whole group of 1.374 (949-1.875) pg/mL, 288 (212-450) pg/mL, 1.53 (1.11-2.77) ng/mL and 15.5 (12.1-23.5) ^U/mL, respectively. Glucose media ± SD concentration was 88.6 ± 13.1 mg/dL.Conclusions: Ghrelin showed an inverse correlation between and age and leptin a direct correlation between anthropometric parameters and gender in healthy wellnourished children. Insulin and glucose didn’t correlate with any variable.

No description available.

Listeria monocytogenes est une bactérie à Gram+, ubiquiste, pathogène intracellulaire d’origine alimentaire, responsable chez l’homme, de nombreuses infections telles que les infections foeto-maternelles, des méningo-encéphalites et des septicémies. La bactérie utilise préférentiellement le glucose qui est transporté via le système phosphoenolpyruvate:sucre phsosphotransferase (PTS) et des perméases non-PTS. Les deux principaux transporteurs de glucose chez L. monocytogenes seraient des PTS de la classe mannose. Le premier est codé par l’opéron manLMN (man) et le deuxième, par l’opéron mpoABCD (mpo). Nous avons, dans un premier temps, mis en évidence le transport de glucose par ces PTS chez L. monocytogenes et aussi identifier d’autres transporteurs non-PTS de glucose. Des tests de croissance en milieu minimum (MM) additionné de glucose et des tests de consommation de glucose ont permis de montrer que les mutants ΔmanL (manL code pour l’EIIABMan) et ΔmanM (manM code pour l’EIICMan) utilisent moins vite le glucose que la souche sauvage AML73 ou EGDe (3 à 4 fois moins vite). Le mutant ΔmpoA (mpoA code pour l’EIIAMpo) montre un phénotype similaire à la souche sauvage tandis que le mutant ΔmpoB (mpoB code pour l’EIIBMpo) utilise 4 à 5 fois moins vite le glucose que la souche sauvage. Des tests de qRT-PCR ont par ailleurs permis de montrer que la délétion du gène mpoA permet une expression constitutive de l’opéron man tandis que la délétion du gène mpoB entraîne une inhibition de l’expression de cet opéron. Nous avons aussi montré que l’opéron man est induit par le glucose et l’opéron mpo est exprimé constitutivement. Le PTSMan est le principal système de transport de glucose chez L. monocytogenes et le PTSMpo pourrait fonctionner comme un senseur de glucose qui en présence de ce sucre stimule l’expression de l’opéron man en régulant l’activité de ManR. Le mutant ΔptsI (ptsI code pour la protéine générale EI du PTS) utilise 8 à 10 fois moins vite le glucose que la souche sauvage et présente une très faible expression de l’opéron man. L’utilisation du glucose (bien que faible) par le mutant ΔptsI permet d’affirmer qu’il existerait des transporteurs non-PTS qui permettraient à ce mutant d’utiliser le glucose. Des tests de complémentation hétérologue dans la souche E. coli LJ140 (incapable de transporter le glucose) ont permis de montrer que les trois protéines GlcU (GlcU1, GlcU2 et GlcU3, identifiées par homologie de séquences aux GlcU d’autres firmicutes) permettent le transport de glucose chez L. monocytogenes mais avec une très faible affinité. Un rôle potentiel du PTS et des transporteurs non-PTS dans la régulation de PrfA a également été mis en évidence par des tests de dosage β-D-glucuronidase à partir de cultures bactériennes réalisées en milieux liquides ou sur géloses et aussi par des tests de qRT-PCR (pour l’expression des gènes actA et hly). Ces tests ont été réalisés à partir de la souche L. monocytogenes AML73 (portant la fusion Phly-gus) et des mutants ΔmanL, ΔmanM, ΔmpoB, ΔmpoA, ΔptsI et glcU (construits dans cette souche). Les mutations manL, manM, mpoB, ptsI entraînent une augmentation de l’activité de PrfA (de 2 à 14 fois) et une augmentation de l’expression des gènes de virulence PrfA-dépendants (hly et actA) est également observée dans les mutants ΔmanL, ΔmanM et ΔmpoB. Les mutations glcU et mpoA ne montrent aucun effet sur l’activité de PrfA. Les mutants montrant une forte activité de PrfA contiennent peu ou pas de protéine EIIABMan qui est supposée jouer un rôle dans la régulation de l’activité de PrfA par le glucose. L’effet des mutations PTS observé sur l’expression des gènes de virulence dépend de PrfA car cet effet disparaît quand le gène prfA est délété dans les mutants ΔmanL, ΔmanM et ΔmpoB. Les mutations montrant un effet sur l’activité de PrfA ont également été étudiées in vitro par des infections des cellules épithéliales (Caco-2 et Jeg-3) avec les différents mutants et également in vivo dans la souris. La délétion du gène ptsI montre un effet dans l’infection plus particulièrement dans l’entrée des bactéries dans les cellules / L. monocytogenes is a ubiquitous foodborne pathogenic Gram-positive bacterium, which can multiply in host cells and infect humans causing septicemia, spontaneous abortion and méningoencephalitis. This bacterium transports glucose via phosphoenolpyruvate:sugar phosphotransferase systems (PTS) and non-PTS permeases. Two major glucose-transporting PTSs belong to the mannose class. One is encoded by the manLMN (man) operon and the second by the mpoABCD (mpo) operon. One goal was to study the transport of glucose by the proteins encoded by these operons and to identify non-PTS glucose transporters. Growth studies in MM supplemented with glucose and glucose consumption assays with several mutants revealed that deletion of manL (encodes EIIABMan) or manM (encodes EIICMan) significantly slowed glucose utilization (3- to 4-fold) compared to the WT AML73 or EGDe strain. Deletion of mpoA (encodes EIIAMpo) had no significant effect on glucose utilization (same phenotype as the WT) whereas deletion of mpoB (encodes EIIBMpo) significantly slowed glucose utilization (4- to -5 fold). By using qRT-PCR, we show that expression of the man operon is induced by glucose, whereas the mpo operon is expressed constitutively. Nevertheless, deletion of mpoA causes constitutive man operon expression whereas deletion of mpoB inhibits it. The PTSMpo therefore functions as a constantly synthesized glucose sensor regulating man operon expression. Deletion of ptsI (encodes the general PTS component EI) also inhibits man expression and the ΔptsI mutant was most strongly impeded in glucose utilization. The residual glucose uptake probably owes to three GlcU-like non-PTS transporters. The successful heterelogous complementation of the E. coli LJ140 strain, wich is unable to transport glucose, suggests that the L. monocytogenes GlcU proteins, GlcU1, GlcU2 and GlcU3 (identified by sequences homology to GlcU proteins in other firmicutes) are indeed capable of transporting glucose.A potential role of PTS and non-PTS components in PrfA regulation was studied in the L. monocytogenes AML73 strain (contains a Phly-gus fusion) and in the ΔmanL, ΔmanM, ΔmpoB, ΔmpoA, ΔptsI, glcU mutants derived from it. For that purpose, I carried out β-D-glucuronidase activity tests with bacteria grown either in liquid or on solid medium and qRT-PCR experiments (expression of actA and hly genes). Interestingly, deletion of ptsI, manL, manM and mpoB caused elevated PrfA activity (2- to -14 fold) and elevated expression of virulence gene expression (actA and hly) in the ΔmanL, ΔmanM and ΔmpoB mutants was observed. Nevertheless, glcU inactivation and mpoA deletion had no effect on PrfA activity. The elevated PrfA activity disappeared when the prfA gene was also deleted in the ΔmanL, ΔmanM and ΔmpoB mutants, confirming that the stimulatory effect of the various mutations on virulence gene expression is PrfA-dependent. All mutants exhibiting elevated virulence gene expression contain no or only little unphosphorylated EIIABMan, which we therefore suspect to play a major role in glucose-mediated PrfA inhibition. The effect of the PTS mutations was also tested in in vitro host cells infection assays (Caco-2, Jeg-3 cells) and in an in vivo mouse model. Deletion of ptsI led to elevated infection of the host cells, which probably owes to the elevated synthesis of the InlA protein.

Der ubiquitär exprimierte, multifunktionale Glucosetransporter GLUT8 gehört zur Klasse III der Familie der passiven Glucosetransporter, die aus insgesamt 14 Proteinen besteht. Die fünf Mitglieder der Klasse IIII unterscheiden sich strukturell leicht von den Mitgliedern der Klasse I und II (Joost und Thorens, 2001). GLUT8 besitzt ein N-terminales Dileucin-Motiv, das Teil eines [DE]XXXL[LI] Motivs ist, welches für die Sortierung des Transporters in späte Endosomen und Lysosomen verantwortlich ist (Augustin et al., 2005). Da bis heute kein Signal identifiziert wurde, das eine Translokation des Transporters zur Plasmamembran auslöst, wird eine intrazelluläre Funktion von GLUT8 vermutet (Widmer et al., 2005). Im Rahmen der vorliegenden Arbeit wurde die intrazelluläre Funktion des Transporters in der Regulation der Glucosehomöostase des Körpers durch Analyse einer Slc2a8-knockout-Maus untersucht. Die homozygote Deletion des Transporters erbrachte lebensfähige Nachkommen, die sich augenscheinlich nicht von ihren Wildtyp-Geschwistern unterschieden. Allerdings wurde bei Verpaarungen heterozygoter Mäuse eine verminderte Anzahl an Slc2a8-/--Nachkommen beobachtet, die signifikant von der erwarteten Mendel’schen Verteilung abwich. Da Slc2a8 die höchste mRNA-Expression in den Testes aufwies und die Überprüfung der Fertilität mittels verschiedener homozygoter Verpaarungen eine Störung der weiblichen Fortpflanzungsfähigkeit ausschloss, wurden die Spermatozoen der Slc2a8-/--Mäuse eingehender untersucht. Als Ursache für die verringerte Anzahl von Slc2a8-/--Geburten wurde eine verminderte Prozentzahl motiler Slc2a8-/--Spermien ermittelt, die durch eine unzureichende mitochondriale Kondensation in den Spermien bedingt war. Diese Veränderung war mit einem reduzierten mitochondrialen Membranpotential assoziiert, was eine verminderte ATP-Produktion nach sich zog. Somit scheint GLUT8 in den Spermien an einem intrazellulären Transportprozess beteiligt zu sein, der einen Einfluss auf die oxidative Phosphorylierung der Mitochondrien ausübt. Im Gehirn wurde Slc2a8 besonders stark im Hippocampus exprimiert, der in der Regulation von körperlicher Aktivität, Explorationsverhalten, Erinnerungs- und Lernprozessen sowie Angst- und Stressreaktionen eine Rolle spielt. Außerdem wurde GLUT8 im Hypothalamus nachgewiesen, der unter anderem an der Regulation der Nahrungsaufnahme beteiligt ist. Die Slc2a8-/--Mäuse zeigten im Vergleich zu ihren Slc2a8+/+-Geschwistern eine signifikant gesteigerte körperliche Aktivität, die zusammen mit der von Membrez et al. (2006) publizierten erhöhten Zellproliferation im Hippocampus auf eine Nährstoffunterversorgung dieses Areals hindeutet. Die Nahrungsaufnahme war in Abwesenheit von GLUT8 nicht verändert, was zusammen mit dem nur geringfügig niedrigeren Körpergewicht der Slc2a8-/--Mäuse eine Funktion von GLUT8 im Glucose-sensing der Glucose-sensitiven Neurone des Gehirns ausschließt. Das leicht reduzierte Körpergewicht der Slc2a8-/--Mäuse ließ sich keinem bestimmten Organ- oder Gewebetyp zuordnen, sondern schien durch eine marginale Gewichtsreduktion aller untersuchten Gewebe bedingt zu sein. Zusammen mit den erniedrigten Blutglucosespiegeln und der anscheinend gesteigerten Lebenserwartung zeigten die Slc2a8-/--Mäuse Symptome einer leichten Nährstoffunterversorgung. GLUT8 scheint daher am Transport von Zuckerderivaten, die während des lysosomalen/endosomalen Abbaus von Glykoproteinen anfallen, beteiligt zu sein. Die so wiederaufbereiteten Zucker dienen dem Körper offenbar als zusätzliche Energiequelle. / The family of facilitative glucose transporters consists of 14 different members in human, which are divided into three classes (Joost and Thorens, 2001). The class III family member GLUT8 contains an amino-terminal dileucine sorting signal, which is part of the highly conserved [DE]XXXL[LI] motif responsible for the localization of GLUT8 in lysosomes and late endosomes (Augustin et al., 2005). To date there is no stimulus known, which translocates the transporter to the plasma membrane, therefore an intracellular function rather than at the cell surface is considered (Widmer et al., 2005). The aim of the present dissertation was to analyze the intracellular role of GLUT8 in the regulation of whole body glucose homeostasis, by the characterization of the corresponding knockout mice (Slc2a8-/-). Slc2a8-/- mice were viable and showed no obvious disparity to their wild-type littermates. However, analysis of the offspring distribution of heterozygous mating provided a reduced number of born Slc2a8-/- offspring which differed significantly from the expected Mendelian distribution. Because Slc2a8 mRNA is expressed at highest levels in the testis and the female Slc2a8-/- mice showed no alterations in fertility, we further investigated the function of Slc2a8-/- spermatozoa. An impaired mitochondrial condensation in the Slc2a8-/- spermatozoa, which was associated with decreased ATP levels resulted in a reduced number of motile Slc2a8-/- sperm, which appeared to be responsible for the reduced number of born Slc2a8-/- offspring. Therefore in sperm cells GLUT8 seems to be important for an intracellular transport process, which exerts an influence on the oxidative phosphorylation in the mitochondria. In the brain Slc2a8 is expressed at highest levels in the hippocampus, which is important for the regulation of physical activity, exploration behaviour, memory and learning as well as anxiety related behaviour. Additionally, GLUT8 was detected in the hypothalamus, which is amongst others involved in the regulation of food intake. The Slc2a8-/- mice showed a significant increase in locomotor activity, which indicates a moderate undersupply of the hippocampus area. According to this finding the group of Membrez et al. (2006) observed a raised cell proliferation in the hippocampus of Slc2a8-/- mice. The fact that no alterations in food intake and only a moderate reduction in body weight was detected in Slc2a8-/- mice, indicates that GLUT8 is not important for the hypothalamic glucose sensing. The marginal decreased body weight of the Slc2a8-/- mice appeared to be associated with a slightly reduced weight of different tissues. Together with the lowered blood glucose concentrations and the apparently enhanced lifespan, the Slc2a8-/- mice showed symptoms of a moderate undersupply compareable to caloric restriction. Thus, we hypothesize that GLUT8 is important for the transport of sugar derivatives which arise during lysosomal/endosomal degradation of glycoproteins. These recycled sugars may serve as an additional energy source in the cell.

<p>The study introduces and puts forward Sector Policing as a model to expand community Policing and to broaden the scope of crime prevention. It also demonstrates how Sector Policing can be utilised to decentralise policing and deepen community participation.</p>

Chlamydiae are important human bacterial pathogens with an intracellular life cycle that consists of two distinct bacterial forms, an infectious form (EB) that infects the eukaryotic host cell, and a non-infectious form (RB) that allows intracellular proliferation. To be successful, chlamydiae need to alternate between EB and RB to generate infectious EB’s which are competent to infect new host cells. Chemical genetics is an attractive approach to study bacterial pathogenesis; in principal this approach relies on an inhibitory compound that specifically inhibits a protein of interest. An obstacle in using this approach is target identification, however whole genome sequencing (WGS) of spontaneous mutants resistant to novel inhibitory compounds has significantly extended the utility of chemical genetic approaches by allowing the identification of their target proteins and/or biological pathways. In this thesis, a chemical genetics approach is used, I have found that heme and glucose metabolism of C. trachomatis is specifically important for the transition from the RB form to the infectious EB form. Heme and glucose metabolism are both coupled to energy metabolism, which suggests a common link between the RB-to-EB transitions. In connection with the above findings I have developed strategies that enable the isolation of isogenic C. trachomatis mutant strains. These strategies are based on WGS of spontaneous mutant populations and subsequent genotyping of clonal strains isolated from these mutant populations. Experiments with the mutant strains suggest that the uptake of glucose-6-phosphate (G-6-P) regulates the RB-to-EB transition, representing one of the first examples where genetics has been used to study C. trachomatis pathogenesis. Additional experiments with the mutant strains indicate that G-6-P promotes bacterial growth during metabolic stress. In concert with other findings presented in this thesis, I have fine-tuned methods that could be employed to reveal how novel inhibitory chemical compounds affect chlamydiae. In a broader context, I suggest that C. trachomatis could be used as a model organism to understand how new inhibitory drugs affect other bacterial pathogens. In addition, I observed that C. pneumoniae infections resulted in generalized bone loss in mice and that these mice display a cytokine profile similar to infected bone cells in vitro. Thus, this study indicates that C. pneumoniae potentially can infect bone cells in vivo, resulting in bone loss, alternatively, the inflammatory responses seen in vivo could be the causative factor of the bone loss observed.

Background Surgery induces insulin resistance that might be alleviated by a nutritional drink given preoperatively. The authors hypothesized that some of the beneficial effects of the drink could be attributed to the volume component (approximately 1 L) rather than to the nutrients. Methods Sixty patients scheduled for elective total hip replacement under spinal anesthesia were recruited to a clinical trial, and randomly allocated to preoperative fasting, to oral ingestion of tap water, or to oral ingestion of a carbohydrate drink. An intravenous glucose tolerance test calculated glucose clearance and insulin sensitivity on the day before surgery, in the postoperative ward, and on the day after surgery. Other parameters were stress (cortisol in plasma and urine), muscle catabolism (urinary 3-methylhistidine), and wellbeing. Results Fifty-seven patients completed the study. In the postoperative ward, the glucose clearance and the insulin response had decreased from the previous day by 23% and 36%, respectively. Insulin sensitivity did not decrease until the next morning (−48%) and was due to an increased insulin response (+51%). Cortisol excretion was highest on the day of surgery, while 3-methylhistidine increased 1 day later. Follow-up on the third postoperative day showed an average of 1.5 complications per patient. Wellbeing was better 2 weeks after than before the surgery. None of the measured parameters differed significantly between the study groups. Conclusions Preoperative ingestion of tap water or a nutritional drink had no statistically significant effect on glucose clearance, insulin sensitivity, postoperative complications, or wellbeing in patients undergoing elective hip surgery. / <p>Funding Agencies|Olle Engkvist Byggmastare Foundation||Stockholm County Council|2009-0433|</p>

SiOx nanowires grown through the VLS mechanism were electrophoretically deposited on top of Au electrodes. GOx was immobilized using APTES and the EDC-NHS chemistry. Cyclic Voltammetry was used as the method to characterize the electrodes through their processing steps, and CV was also used to detect glucose in a PBS based solution. Ferro-Ferri Cyanide couple was used as the mediator.

<p>The study introduces and puts forward Sector Policing as a model to expand community Policing and to broaden the scope of crime prevention. It also demonstrates how Sector Policing can be utilised to decentralise policing and deepen community participation.</p>

We report controllable synthesis of Pd aerogels with high surface area and porosity by destabilizing colloidal solutions of Pd nanoparticles with variable concentrations of calcium ions. Enzyme electrodes based on Pd aerogels co-immobilized with glucose oxidase show high activity toward glucose oxidation and are promising materials for applications in bioelectronics.