Rationale: The serotonergic system is involved in the modulation of emotion and plays an important role for personality and vulnerability for psychiatric disorders. In the papers included in this thesis, we investigate three biological factors that have been studied in relation to psychiatric symptoms: Platelet monoamine oxidase B (MAO-B) activity, and variations in the MAO-A and the serotonin transporter (5HTT) genes. We also study intensity dependent auditory evoked potentials (IAEP) as an intermediate phenotype for serotonergic capacity. Platelet MAO-B has been shown to be a biological marker for the properties of monoamine systems, with low activity being associated with vulnerability for high scores of sensation seeking, monotony avoidance, and impulsiveness, as well as for susceptibility for alcoholism. Functional polymorphisms in the promoter of the genes encoding MAO-A and the serotonin transporter result in high- or low- activity alleles that have been associated with numerous psychiatric symptoms. One hypothesis for the shaping of personality is that these genotype variants have prenatal effects on the wiring of the brain. Thus, exploring how the development of the brain is affected by different prenatal serotonin levels is relevant in this context. Observations: (i) Platelet MAOB activity was associated with monoamine metabolites in cerebrospinal fluid from cisterna magna in monkeys, as well as with voluntary alcohol intake, alcohol-induced aggression, and alcohol sensitivity. (ii) The long 5HTTLPR allele was associated with increased IAEP. (iii) The functional MAOA and 5HTT polymorphisms were associated with symptoms of ADHD-related traits in a population based sample of Swedish adolescents. Associations of these candidate genes with ADHD scores were strenghtened when the platelet MAOB activity was combined with genotype. (iv) Our pilot data showed that treatment of pregnant mice with 5HTT blocking antidepressives resulted in more serotonergic cellbodies in lateral wings of dorsal raphe in the offspring, when compared to saline treatment. Conclusions: Our studies support the notion that platelet MAOB activity and IAEP are endophenotypes for monoaminergic capacity and related behaviours. The functional candidate polymorphisms in MAOA and 5HTT were linked to behaviour, however, the cause-relationship is unclear and the explanation for the associations need to be further investigated, possibly with focus on prenatal effects of the polymorphisms.

A polymorphism in the promoter region of the human serotonin transporter (5-HTT)-coding SLC6A4 gene (5-HTTLPR) has been implicated in moderating susceptibility to stress-related psychopathology and to possess regulatory functions on human in vivo 5-HTT availability. However, data on a direct relation between 5-HTTLPR and in vivo 5-HTT availability have been inconsistent. Additional factors such as epigenetic modifications of 5-HTTLPR might contribute to this association. This is of particular interest in the context of obesity, as an association with 5-HTTLPR hypermethylation has previously been reported. Here, we tested the hypothesis that methylation rates of 14 cytosine-phosphate-guanine (CpG) 5-HTTLPR loci, in vivo central 5-HTT availability as measured with [11C]DASB positron emission tomography (PET) and body mass index (BMI) are related in a group of 30 obese (age: 36±10 years, BMI>35 kg/m2) and 14 normal-weight controls (age 36±7 years, BMI<25 kg/m2). No significant association between 5-HTTLPR methylation and BMI overall was found. However, site-specific elevations in 5-HTTLPR methylation rates were significantly associated with lower 5-HTT availability in regions of the prefrontal cortex (PFC) specifically within the obese group when analyzed in isolation. This association was independent of functional 5-HTTLPR allelic variation. In addition, negative correlative data showed that CpG10-associated 5-HTT availability determines levels of reward sensitivity in obesity. Together, our findings suggest that epigenetic mechanisms rather than 5-HTTLPR alone influence in vivo 5-HTT availability, predominantly in regions having a critical role in reward processing, and this might have an impact on the progression of the obese phenotype.:Introduction ............................................................................................................. 3 Homeostatic and hedonic control of food intake...................................................... 3 The obesity epidemic .............................................................................................. 4 The role of 5-HT in energy balance......................................................................... 5 The role of 5-HT in the PFC function....................................................................... 6 The role of the PFC in food intake .......................................................................... 7 The association between central 5-HT transporter (5-HTT) availability and obesity ..................................................................................................................................7 Genetics of obesity ................................................................................................. 8 Epigenetics of obesity ............................................................................................. 9 Objectives and hypothesis of the present work...................................................... 11 Manuscript ..... ....................................................................................................... 12 Summary ............................................................................................................... 20 References ............................................................................................................ 22 Appendices ............................................................................................................I Glossary ................................................................................................................ I Publications ........................................................................................................... IV Selbstständigkeitserklärung................................................................................... V Danksagung .......................................................................................................... VI

La physiopathologie de l'hypertension artérielle pulmonaire (HTAP) implique de multiples mécanismes. Elle est caractérisée, entre autres, par la réduction de la production des facteurs vasodilatateurs, l'augmentation exagérée des facteurs vasoconstricteurs et des facteurs de croissance qui conduisent à la vasoconstruction, la prolifération et le remodelage vasculaire pulmonaire. Les cellules endothéliales pulmonaires secrètent des facteurs paracrines qui contribuent à l'hyperplasie des cellules musculaires lisses pendant la progression de l'hypertension artérielle pulmonaire. Le FGF2 produit par les cellules endothéliales et stocké dans la matrice extracellulaire, est l'un de ces facteurs très bien documenté pour entraîner une prolifération des CML. Cette étude montre que l'inhibition de l'expression pulmonaire du bFGF par l'injection répétée du SiRNA chez les rats traités à la MCT, est corrélée à l'amélioration des paramètres hémodynamiques, du remodelage vasculaire et de l'hypertrophie cardiaque droite, en traitement préventif et curatif. Chez les sujets atteints d'hypertension pulmonaire le niveau de la sérotonine circulant est très élevé. La biosynthèse de la sérotonine dépend de la tryptophane hydroxylase. Nous avons étudié l'impact des variations génétiques de la Tph1 et de la Tph2 sur le développement de l'HTAP chez les souris. Cette étude a montré que la déficience en Tph1 (Tph périphérique) protège les souris contre l'hypoxie alors que les souches de souris porteuses du polymorphisme C1473G au niveau du gène Tph2 (Tph centrale) montrent des phénotypes d'HTAP différents pendant l'hypoxie. En plus de l'importance de la TPH, l'expression du 5-HTT est aussi déterminant dans l'HTAP. Les souris SM22 surexprimant le 5-HTT dans les CML à un niveau proche de celui des CML des patients atteints d'hypertension artérielle pulmonaire développent spontanément une HTAP qui s'aggrave avec l'âge. Du fait de la baisse considérable de l'activité de ces deux enzymes, le NO synthase et la prostacycline synthase, au niveau des cellules endothéliales chez les patients HTAP, le taux de l'AMPc et du GMPc baisse. Les PDEs hydrolysent les deux messagers des agents vasodilatateurs prostacycline et oxyde nitrique, et l'utilisation des inhibiteurs de phosphodiestérases augmentent la concentration intracellulaire de GMPc et AMPc et causent la vasodilatation pulmonaire. Le traitements des rats HTAP avec l'inhibiteur du PDE4 (Roflumilast) avec deux doses 1,5 et 0,5mg/Kg/jour, a montré une régression de l'HTAP, baisse de l'hypertrophie cardiaque et du remodelage vasculaire des rats traités à la monocrotaline ou mises en hypoxie aigüe (10% O2), chez qui un traitement préventif et curatif a été effectué. / The physiopatholy of pulmonary arterial hypertension (PAH) implies multiple mechanisms. It is characterized by the reduction of the production of vasodilatators factors, the overproduction of vasoconstrictor factors and growth factors which lead to the pulmonary vasoconstriction, and the pulmonary vascular remodeling. The pulmonary endothelial cells release paracrine factors which contribute to the hyperplasy of the smooth muscule cells (SMC) during the progression of PAH. The FGF2 produced by the endothelial cells and stored in the extracellular matrix is reproted to be involved in the SMC proliferation. This study shows that inhibition of the pulmonary expression of FGF2 by the repeated injection of SiRNA in the rats treated with monocrotaline (MCT), is correlated with the improvment of the hemodynamic parameters, vascular remodeling and right ventricular hypertrophy, in both, a preventive and a curative treatment. In pateints with PAH, it has been shown that the circulating level of serotonin is very high as compared to control patients. The biosynthesis of serotonin depends on tryptophan hydroxylase. Therefore, we studied the impact of the genetic variations of Tph1 and Tph2 on the development of PAH in mice. This study showed that deficiency in Tph1 (peripheral Tph) protects mice against the hypoxia-induced PAH severity during hypoxia. In addition to the importance of the Tph, the serotonin transporter (5-HTT) expression is also a determining factor in the development of PAH. Mice overexpressing 5-HTT (SM22-5-HTT+ mice) specifically in SMC develop spontaneously PAH in normoxia, which worsens with age. We observed a significant decrease int he rate of cAMP and cGMP produced in the endothelial cells from patients with PAH, which has been reported to be due to low activities of NO synthase and prostacyclin synthase. Phosphodiesterases (PDE)hydrolize the two messengers (cAMP and cGMP), and the use of specific inhibitors of PDE increases the intracellular concentration of cGMP and cAMP and causes pulmonary vasodilatation. A daily oral administration of Roflumilast (1,5 and 0,5 mg/kg/day) a specific inhibitor of PDE4, was effective in preventing and in reversin the PAH in both experimental model of PAH, the MCT and chronic hypoxia (10%O2,2 weeks)

Evidence suggests that certain genotypic variants associated with novelty-seeking and aggressiveness, such as the 7-repeat dopamine D4 receptor variant (DRD4-7R), short (s) allele of the serotonin transporter (5-HTT), and the low-activity variant of the MAOa promoter (MAOa-L), are more prevalent in human groups that radiated out of Africa than human groups that remained in Africa. Rhesus macaques (Macaca mulatta), like humans, are a widespread species of primates that needed to adapt to different regional environments with one group, Indian-derived rhesus macaques, largely occupying predictable and resource-rich environments, while the other group, the Chinese-derived rhesus macaques, has come to occupy less predictable and resource-abundant environments. Rhesus macaques possess orthologues of these trait-related genes, making it possible to compare the frequency of genotypes associated with these traits between members of two strains. DNA was obtained from N=212 rhesus macaques (n=54 Chinese-derived, n=158 Indian-derived) and genotyped for DRD4 (n=98), 5-HTT (n=190), and MAOA (n=97). Analyses showed that Chinese-derived subjects exhibited higher frequencies of the DRD4-7R and 5-HTT-s-allele when compared to Indian-derived subjects. There were no strain differences in MAOA-L genotype groupings, but the Chinese-derived subjects exhibited a more frequent high-activity (MAOA-H-6R) allele when compared to the Indian-derived subjects. The results suggest that the Chinese-derived rhesus macaques possess a higher frequency of alleles associated with novelty-seeking, impulsivity, and aggressiveness compared to their Indian-derived peers and that those genotypically-mediated traits may have beneficial to both humans and rhesus macaques as they spread into novel and unfamiliar environments.

Evidence suggests that certain genotypic variants associated with novelty-seeking and aggressiveness, such as the 7-repeat dopamine D4 receptor variant (DRD4-7R), short (s) allele of the serotonin transporter (5-HTT), and the low-activity variant of the MAOa promoter (MAOa-L), are more prevalent in human groups that radiated out of Africa than human groups that remained in Africa. Rhesus macaques (Macaca mulatta), like humans, are a widespread species of primates that needed to adapt to different regional environments with one group, Indian-derived rhesus macaques, largely occupying predictable and resource-rich environments, while the other group, the Chinese-derived rhesus macaques, has come to occupy less predictable and resource-abundant environments. Rhesus macaques possess orthologues of these trait-related genes, making it possible to compare the frequency of genotypes associated with these traits between members of two strains. DNA was obtained from N=212 rhesus macaques (n=54 Chinese-derived, n=158 Indian-derived) and genotyped for DRD4 (n=98), 5-HTT (n=190), and MAOA (n=97). Analyses showed that Chinese-derived subjects exhibited higher frequencies of the DRD4-7R and 5-HTT-s-allele when compared to Indian-derived subjects. There were no strain differences in MAOA-L genotype groupings, but the Chinese-derived subjects exhibited a more frequent high-activity (MAOA-H-6R) allele when compared to the Indian-derived subjects. The results suggest that the Chinese-derived rhesus macaques possess a higher frequency of alleles associated with novelty-seeking, impulsivity, and aggressiveness compared to their Indian-derived peers and that those genotypically-mediated traits may have beneficial to both humans and rhesus macaques as they spread into novel and unfamiliar environments.

No description available.

En damm används i huvudsak för att magasinera vatten i energiutvinningssyfte. Dammen rör sig fram och tillbaka i ett säsongsmönster mestadels beroende på skillnader i utomhustemperatur och vattentemperaturen i magasinet. Det nordiska klimatet innebär risk för isläggning i magasinet, för vilken lasten är relativt outforskad. Denna rapport syftar till ett med multipla linjära regressionsmodeller samt dynamiska regressionsmodeller avgöra vilka variabler som förklarar en specifik svensk dammkonstruktions rörelse. Dammens rörelse mäts genom att mäta dammens förflyttning kontra berggrunden med data från dammens inverterade pendlar. Av särskilt intresse är att avgöra islastens påverkan på rörelsen. Resultaten visar att multipla linjära regressions-modeller inte fullständigt lyckas modellera dammens rörelse, då de har problem med autokorrelerade residualer. Detta hanteras med hjälp av autoregressiva regressionsmodeller där de initiala förklarande variablerna inkluderas, kallat dynamisk regression. Denna rapports resultat visar att de autoregressiva parametrarna fungerar mycket väl för att förklara pendlarna, men att även tid, temperatur, det hydrostatiska trycket samt istjocklek är användbara förklarande variabler. Istjockleken visar signifikant påverkan på 5 % signifikansnivå på två av de undersökta pendlarna, vilket är ett noterbart resultat. Författarna menar att rapportens resultat indikerar att det finns anledning att fortsätta forska kring islastens påverkan på dammkonstruktioner. / A dam is a structure mainly used for storing water and generating electricity. The structure of a dam moves in a season-based pattern, mainly because of the difference in temperature between the air on outside of the dam and the water on the inside. Due to the Nordic climate, occurrences of icing on the water in the basin is fairly frequent. The effects of ice on the structural load of the dam are relatively unexplored and are the subject to this bachelor’s thesis. The goal of this project is to evaluate which predictors are significant to the movement of the dam with multiple linear regression models and dynamic regressions. The movement is measured by inverted pendulums that register the dam’s movement compared to the foundation. It is of particular interest to determine if the ice load influences the movement of the dam. The multiple regression models used to explain the dam’s movement were all discarded due to autocorrelation in the residuals. This falsifies the models, since autocorrelation means that they don’t meet the needed assumptions. To counteract the autocorrelation, dynamic models with autoregressive terms were fitted. These models showed no problem with autocorrelation. The result from the dynamic models were successful and managed to significantly explain the movement of the dam. The autoregressive terms proved to be efficient explanatory variables. The dynamic regression models also show that the time, temperature, hydrostatic pressure and ice thickness variables are also useful explanatory variables. The ice thickness shows a significant effect at the 5 % significance level on two of the investigated pendulums. The report's results indicate that there is reason to continue research on the ice load impact on dam constructions.

La huntingtine (HTT) est une protéine d’échafaudage participant à des fonctions indispensables au bon fonctionnement cellulaire. Elle est codée par le gène HTT qui présente une répétition polymorphique de triplet CAG. Une répétition excédant 35 CAG dans la HTT est à l’origine de la maladie de Huntington, une maladie neurodégénérative héréditaire sévère. Ainsi, bien que d’expression ubiquitaire, la HTT est principalement étudiée dans le système nerveux. Par exemple, ses implications dans le tissu mammaire, en condition normale et pathologique, sont inconnues. Nous avons observé que la forme mutante de la HTT accélère le développement de cancer du sein et en accentue la sévérité et que la forme sauvage est impliquée dans le développement normal de la glande mammaire. Mon projet principal de thèse était de caractériser le rôle de la HTT, de sa phosphorylation à la sérine 421 (S421-P-HTT) ainsi que du polymorphisme des répétitions CAG dans les cancers du sein.En utilisant des modèles cellulaires et murins et par des études d’expression chez des patientes atteintes d’un cancer du sein, j’ai observé que l’expression de la HTT et de la S421-P-HTT corrèlent avec le stade de différenciation tumorale. Au niveau moléculaire, la HTT régule, par sa phosphorylation à la S421, l’expression et la localisation d’une des protéines des jonctions serrées, ZO1 et module ainsi l’adhésion intercellulaire. ZO1 colocalise avec la S421-P-HTT aux jonctions intercellulaires et forme un complexe avec la HTT. La perte d’expression de HTT est pro-Métastatique chez la souris et est moindre dans les cancers du sein métastatiques. De plus, les niveaux d’expression de HTT et de ZO1 sont diminués en parallèle dans les carcinomes humains de bas grades.J’ai également montré que le polymorphisme CAG présent dans la HTT sauvage joue un « double emploi » : tandis que de longues répétitions protègent de l’apparition de cancers, elles accentuent sa sévérité lorsque la maladie se développe. Dans le sous-Type HER2 spécifiquement, la longueur de la répétition CAG est un facteur pronostic indépendant du développement de métastases.Ainsi, ces travaux ont permis de mettre en évidence un rôle clé pour la HTT au cours de la progression tumorale mammaire, et devraient conduire à une meilleure compréhension des mécanismes moléculaires impliqués dans le développement de métastases dans le cancer du sein. / Huntingtin (HTT) is a scaffold protein involved in numerous cellular mechanisms essentials for appropriate physiological functions. HTT is encoded by HTT gene which carries a polymorphic repetition of CAG triplet. When the CAG repetition exceeds 35, it leads to Huntington’s disease, a hereditary severe neurodegenerative disorder. While HTT expression is ubiquitous, it is mainly studied in nervous system. For example, HTT roles in breast physiology and cancer are unknown. We demonstrated that mutant HTT accelerates breast tumor and metastasis development and that wild-Type HTT is involved in normal mammary gland development. My main project was to characterize the roles of HTT and of its phosphorylation at S421 (S421-P-HTT) and that of the polymorphic CAG length in mammary carcinomas.First, leaning on cellular and murine models as well as on expression studies in breast cancer patients, I observed that HTT and S421-P-HTT expression correlates with tumoral differentiation stage. At the molecular level, HTT regulates through its phosphorylation at S421, the expression and localization of ZO1, a marker of intercellular junction and therefore modulates intercellular adhesion. ZO1 colocalizes with S421-P-HTT specifically at tight junctions and forms a complex with HTT. Loss of HTT is itself pro-Metastatic in mice and is decreased in metastatic human breast cancer. Moreover, HTT and ZO1 are concomitantly downregulated in low-Grade human carcinomas.On the other hand, the polymorphism of CAG repetitions in HTT has a dual-Purpose: while long repetitions protect against cancer development, it increases its severity once cancer is developed. In HER2 subtype specifically, HTT appears as an independent prognostic factor of metastasis development.Thus, these studies point out a key function of HTT outside the brain during mammary carcinoma progression and should lead to a better understanding of molecular mechanisms involved in metastasis development.

No description available.

Made available in DSpace on 2015-12-06T22:54:32Z (GMT). No. of bitstreams: 0 Previous issue date: 2009 / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / Neste estudo, 151 isolados brasileiros de Sporothrix schenckii foram caracterizados quanto as suas características fenotípicas e genotípicas. Quanto às características fenotípicas, a maioria dos isolados apresentou conídios ovais, sendo que os de origem animal são maiores que os de origem humana (animal: 2,96 ± 1,07 versus forma linfocutânea: 2,37 ± 0,43; forma fixa: 2,33 ± 0,53). Os isolados provenientes da região Norte do Brasil são os mais termotolerantes e os isolados do Nordeste são os mais termossensíveis. Os isolados de origem animal são mais tolerantes à temperatura quando comparados com isolados obtidos de pacientes. Os isolados brasileiros apresentaram grande variabilidade genética quando avaliados por RAPD. Por Southern Blot, utilizando a enzima APA I para o estudo do polimorfismo das regiões ITS, três genótipos foram obtidos. Não houve correlação entre o perfil obtido por RAPD ou Southern Blot com a origem clínica ou geográfica dos isolados. A secreção de proteínas/glicoproteínas é variável entre os isolados, sofrendo influência direta do meio de cultura. Os isolados brasileiros avaliados são produtores das enzimas DNAse e urease, 15,78% produzem gelatinase, 26,31% produzem proteinase e 21,05% produzem caseinase. Todos os isolados avaliados toleram pressão osmótica de 16,6% de glicerol, 89,47% toleram 20% e todos são inibidos a 23% e 28,5 % de glicerol. Todos os isolados avaliados toleram pressão salina de 6% e 8%; 42,10% toleram pressão de 9% e 10% de sal e todos são inibidos a 12% de sal. Todos os isolados avaliados crescem na faixa de pH de 2,2 a 12,5. A patogenia e virulência são variáveis entre os isolados, podendo estar relacionada ao genótipo do isolado. Diferentes proteínas são reconhecidas pelos soros dos camundongos infectados, sendo a molécula de 60 kDa, a mais reconhecida neste sistema. Em relação à esporotricose humana, as moléculas de 70 kDa e 38 kDa são comumente reconhecidas por soros de pacientes com esporotricose fixa e linfocutânea. O antígeno bruto do isolado Ss 118 e a fração SsCBF são eficientes para diagnóstico da esporotricose felina através da técnica de ELISA. A fração SsCBF é capaz de elicitar resposta imune do tipo HTT em camundongos experimentalmente infectados. Em geral, os resultados indicam que os isolados brasileiros são heterogêneos, apresentando características genéticas e fenotípicas individuais, independentes de sua origem. Somente a área conidial e a tolerância a temperatura foram características fenotípicas relacionadas à origem. / In this study, we analyzed 151 Brazilian S. schenckii isolates by phenotyping and genotyping aspects. About phenotyping aspects, most of isolates showed oval conidia and the mean conidial area of S. schenckii animal isolates was greater than clinical isolates (animal: 2.96 ± 1.07 versus lymphocutaneous form: 2.37 ± 0.43; fixed form: 2.33 ± 0.53). Isolates from the Northeast region exhibited the lowest thermotolerance and the Northern isolates exhibited the highest thermotolerance. Animal isolates are better thermotolerants than clinical isolates. The Brazilian isolates showed great genetic diversity when analyzed by RAPD. By Southern blot using the endonuclease APA I to study polymorphism of ITS regions, three genotypes were obtained. There was no association between RAPD and Southern profiles with clinical forms origin or geographic origin. The protein/glycoprotein secretion was variable among isolates and was directly influenced by the medium composition. All Brazilian isolates studied produced DNAse and urease exoenzymes, 15.78% produced gelatinase, 26.31% produced proteinase and 21.05% produced caseinase. All analyzed isolates grew at 16,6% of glycerol, 89,47% grew at 20% of glycerol and all of them were inhibited at 23% and 28,5 % of glycerol. All analyzed isolates grew at 6% and 8% of NaCl, 42.10% grew at 9% and 10% of NaCl and all of them were inhibited at 12% of NaCl. All of them were able to grow at pH 2.2 to 12.5. The pathogenicity and virulence were variable among isolates and it can be related to the genotype. Different proteins were recognized by sera of infected mice and the 60 kDa molecule was the most one recognized in murine system. In relation to human sporotrichosis, 70 kDa and 38 kDa molecules were common recognized by sera from patients with fixed and lymphocutaneous sporotrichosis. The crude exoantigen of Ss 118 isolate and the SsCBF fraction are able to be used in the diagnosis of feline sporotrichosis by ELISA. The SsCBF fraction is able to elicit DTH immune response in infected mice. In general, our results indicate that Brazilian isolates are heterogeneous, which present individual phenotyping and genotyping characteristics, independent of origin. Only the conidial area and thermotolerance were related to origin. / BV UNIFESP: Teses e dissertações