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INTRODUÇÃO: A obesidade na infância e adolescência representa uma epidemia global e figura como um problema de saúde pública proeminente de prevalência crescente. A obesidade frequentemente está associada à compulsão alimentar periódica (CAP) e componentes genéticos participam de sua etiologia multifatorial. Polimorfismos de nucleotídeo único (SNPs) no gene da leptina (LEP) e do receptor da leptina (LEPR) podem modificar a expressão da leptina e de suas vias de sinalização e, consequentemente, alterar a regulação do apetite e da saciedade, contribuindo assim para a etiopatogenia e manutenção da CAP. O objetivo deste trabalho foi investigar a influência dos polimorfismos rs7799039 (G > A) no gene LEP e rs1137100 (A > G), rs1137101 (A > G) e rs8179183 (G > C) no gene LEPR sobre a CAP em crianças e adolescentes obesos, além de caracterizar a população quanto à CAP e verificar a associação dos SNPs com o risco cardiometabólico (RCM) e a obesidade. MÉTODOS: Estudo transversal que incluiu 465 crianças e adolescentes obesos com idade entre 7 e 19 anos avaliados quanto a variáveis antropométricas e metabólicas. Os fatores de RCM consistiram de hipertensão arterial sistêmica, glicemia de jejum alterada, HDL-colesterol baixo e hipertrigliceridemia. A CAP foi avaliada por meio da Escala de Compulsão Alimentar Periódica (ECAP). Para investigar o efeito dos SNPs no risco para a obesidade foi incluído um grupo controle composto por 135 crianças e adolescentes eutróficos. A genotipagem foi realizada por PCR em tempo real e para análise dos SNPs, adotou-se o modelo dominante. Foi calculado o desequilíbrio de ligação entre os SNPs e estimada as frequências dos haplótipos. As comparações entre os grupos foram realizadas estratificadamente por gênero e estádio puberal. Para avaliar a magnitude do risco dos SNPs sobre a CAP e a obesidade foi realizada regressão logística ajustada para variáveis de confusão (idade, Z-IMC e estádio puberal). RESULTADOS: As crianças e adolescentes obesos (12,5 ± 2,9 anos; 52,7% meninas) classificados com CAP apresentaram maior adiposidade e a frequência da CAP foi mais elevada no gênero feminino (OR= 2,146; IC 95% 1,461-3,152; p < 0,001). A frequência do alelo A do rs7799039 foi mais elevada no grupo de obesos (OR= 1,530; IC 95% 1,022-2,292; p= 0,039) e o alelo associou-se ao maior nível de leptina e colesterol total em meninas e à maior glicemia em meninos (p < 0,05). No rs1137100 e o rs1137101, a presença do alelo G em meninas conferiu risco para a hipertrigliceridemia (OR= 1,926; IC 95% 1,010-3,673; p= 0,047 e OR= 2,039; IC 95% 1,057-3,931; p= 0,033, respectivamente). O alelo C do rs8179183 relacionou-se, em meninas, à relação cintura-estatura e glicemia mais elevadas e, em meninos, ao maior percentil de pressão arterial diastólica, glicemia, colesterol total e LDL-colesterol (p <0,05). CONCLUSÃO: Os polimorfismos não foram associados à compulsão alimentar periódica. A CAP foi relacionada ao pior grau de adiposidade e o maior risco foi observado no gênero feminino. O SNP rs7799039 no gene LEP conferiu risco para obesidade, enquanto o rs1137100, rs1137101 e rs8179183 no gene LEPR relacionaram-se ao pior perfil cardiometabólico em crianças e adolescentes obesos / INTRODUCTION: Obesity during childhood and adolescence represents a global epidemic and consists in a prominent public health issue of increasing prevalence. Obesity is frequently associated with binge eating (BE) and genetic factors participate of its multifactorial etiology. Single nucleotide polymorphisms (SNPs) in the leptin (LEP) and leptin receptor (LEPR) genes may modify the leptin expression and its signaling pathways and, consequently, alter appetite and satiety regulation, thus contributing to the etiopathogeny and maintenance of BE. The aim of this study was to investigate the influence of polymorphisms rs7799039 (G > A) in the LEP gene and rs1137100 (A > G), rs1137101 (A > G) and rs8179183 (G > C) in the LEPR gene on BE in obese children and adolescents, besides characterize the population regarding to BE and examine the association of SNPs with cardiometabolic risk (CMR) and obesity. METHODS: Cross-sectional study in which 465 obese children and adolescents aged from 7 to 19 years were enrolled and had anthropometric and metabolic variables assessed. The CMR factors consisted of systemic hypertension, impaired fasting glucose, low HDL-cholesterol levels and hypertriglyceridemia. The BE was evaluated through the Binge Eating Scale (BES). To investigate the effect of SNPs on obesity risk, a control group of 135 eutrophic children and adolescents was enrolled. Genotyping was performed by real-time PCR and for the SNPs analysis, the dominant model was adopted. The linkage disequilibrium between SNPs was calculated and the haplotype frequencies were estimated. Comparisons between groups were performed stratified by gender and pubertal stage. To assess the risk magnitude for the SNPs on BE and obesity, logistic regression adjusted for confounding variables (age, Z-BMI and pubertal stage) was performed. RESULTS: Obese children and adolescents (12.5 ± 2.9 years, 52.7% girls) classified with BE showed greater adiposity and BE frequency was higher among females (OR= 2.146; 95% CI 1.461-3.152; p < 0.001). The observed frequency of A allele of rs7799039 was a higher in the obese group (OR= 1.530; 95% CI 1.022-2.292; p= 0.039) and the allele was associated with higher leptin and total cholesterol levels in girls and higher glucose levels in boys (p < 0.05). For the rs1137100 and rs1137101, the presence of the G allele among girls, conferred risk for hypertriglyceridemia (OR= 1.926; 95% CI 1.010-3.673; p= 0.047 and OR= 2.039; 95% CI 1.057-3.931; p= 0.033, respectively). The C allele of rs8179183 was associated, among girls, with a higher waist-to-height ratio and glucose levels and, among boys, with greater diastolic blood pressure percentile, glucose, total cholesterol and LDL-cholesterol levels (p < 0.05). CONCLUSION: Polymorphisms were not associated with binge eating. BE was related with a more severe adiposity and an increased risk was observed among females. The SNP rs7799039 in the LEP gene contributed to the risk of obesity, whereas the rs1137100, rs1137101 and rs8179183 in LEPR gene were related to a worse cardiometabolic profile in obese children and adolescents

Thesis (PhD (Biomedical Sciences. Medical Physiology))--University of Stellenbosch, 2009. / ENGLISH ABSTRACT: In recent years there has been an increase in obesity and diabetes mellitus (DM). These conditions have for a long time been associated with infertility. Obesity is characterized by high levels of circulating leptin and cytokines as well as insulin resistance. Type I DM is associated with low or no insulin whereas, Type II DM is characterised by insulin resistance. As the prevalence of obesity and DM continues to rise, it is likely that the incidence of infertility associated with these pathological conditions will likewise increase. The effects of insulin and leptin on male reproductive function have been reported on the endocrine and spermatogenesis level, but their effects on cellular level of human ejaculated spermatozoa are yet to be elucidated. This study presents data on the role of insulin and leptin on human ejaculated spermatozoa and their interaction with cytokines and nitric oxide. In the first part of the study, we established the suitable concentrations of glucose, insulin and leptin that could be administered to human spermatozoa in vitro. Glucose concentration of 5.6 mM was chosen as the suitable concentration to be administered to human spermatozoa because it has previously been reported in the literature; furthermore, it is within the range of the physiological glucose levels found in the blood of fasting humans. Insulin and leptin concentrations of 10 μIU and 10 nmol were chosen respectively because they gave much improved sperm function and this was within the range of insulin and leptin levels previously measured in human ejaculated spermatozoa. This was followed by investigating the signalling pathway of insulin and its beneficial effects on human spermatozoa function. Endogenous insulin secretion from human ejaculated spermatozoa was blocked by nifedipine and its receptor tyrosine phosphorylation effects were inhibited by erbstatin while phosphatidylinositol 3-kinase (PI3K) phosphorylation activity was inhibited by wortmannin. Exogenous insulin administration significantly increased human sperm motility parameters as well as the sperm ability to acrosome react. The inhibition of endogenous insulin release from spermatozoa as well as the inhibition of the insulin receptor substrate (IRS) tyrosine phosphorylation significantly decreased motility parameters and the ability of spermatozoa to acrosome react. The study also investigated the effects of insulin and leptin on human sperm motility, viability, acrosome reaction and nitric oxide (NO) production. Both insulin and leptin significantly increased sperm motility parameters, acrosome reaction and NO production. The NO production induced by insulin and leptin was via PI3K signalling as evidenced by a reduction in NO levels when PI3K activity was inhibited by wortmannin. To investigate whether insulin and leptin could improve motility parameters of asthernozoospermic and teratozoospermic spermatozoa, the spermatozoa were separated into two fractions by means of a double density gradient technique. The gradient system was able to separate spermatozoa into high morphologically abnormal and less motile spermatozoa similar to that of asthernozoospermic and teratozoospermic patients as well as a more motile fraction. Insulin and leptin significantly increased the motility parameters of spermatozoa from the immature and less motile fraction. The fourth part of the study was aimed at investigating the effects of the cytokines, tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6), on human sperm motility, viability, acrosome reaction and NO production. The study shows that TNF-α and IL-6 significantly reduced motility parameters and acrosome reaction in a dose4 and time-dependent manner. These cytokines were also shown to significantly increase NO production from human spermatozoa. The decreased motility parameters induced by these cytokines could be attributed to their ability to induce excessive NO production. It is not yet clear how they inhibit spermatozoa to undergo the acrosome reaction. The fifth part of the study was to investigate the expression and localization of glucose transporter 8 (GLUT8) in human spermatozoa. This study shows that GLUT8 is constitutively expressed and located in the midpiece region of the human spermatozoa. The study also showed that stimulating spermatozoa with insulin led to an increase in GLUT8 expression as well as translocation to the acrosomal region. In the last part of the study we wanted to investigate why the increase in NO generation by spermatozoa due to insulin and leptin stimulation is accompanied with increased sperm function whereas NO increased due to TNF-α and IL-6 stimulation is accompanied with decreased sperm function. We observed that TNF-α and IL-6 not only increased NO production but also ROS production. This study speculates that the decrease in sperm motility and acrosome reaction when TNF-α and IL-6 were administered was due to the concomitant high increase in NO and ROS they induced. In conclusion, this study has established in vitro beneficial effects of insulin and leptin in normozoospermic and asthernozoospermic human sperm function. These hormones influence sperm function via the PI3K signalling pathway in two ways. Firstly, by increasing GLUT8 expression and translocation thereby possibly increasing glucose uptake and metabolism and secondly, by increasing NO production. The study has also established that TNF-α and IL-6 have detrimental effects on human spermatozoa in a dose and time dependent manner. These effects are mediated via their ability to stimulate both NO and ROS production in human spermatozoa. This study reports that GLUT8 is expressed in the midpiece region of human spermatozoa and that insulin stimulation upgrades its expression and leads to its translocation to the acrosomal region. / AFRIKAANSE OPSOMMING: Oor die afgelope jare was daar `n toename in obesiteit en diabetes mellitus (DM). Hierdie toestande word reeds vir ’n geruime tyd geassosieer met onvrugbaarheid. Obesiteit word gekenmerk deur verhoogde sirkulerende vlakke van leptiene en sitokiene sowel as insulien weerstandigheid. Tipe I DM word geassosieer met lae of geen insulien terwyl Tipe II DM gekenmerk word deur insulien weerstandigheid. Soos wat die voorkoms van obesiteit en DM toeneem, is dit waarskynlik dat die insidensie van onvrugbaarheid wat met hierdie patologiese toestande geassosieer word, gevolglik ook sal toeneem. Die effek van insulien en leptien op die manlike voortplantingsfunksie is alreeds aangetoon op endokriene en spermatogenese vlak, maar hul effekte op sellulêre vlak van menslike geëjakuleerde spermatosoë is nog onduidelik. Die studie vertoon data oor die rol van insulien en leptien op die menslike geëjakuleerde spermatosoë en hul interaksie met sitokiene en stikstofoksied (NO). In die eerste gedeelte van die studie, het ons ’n toepaslike konsentrasie van insulien en leptien bepaal wat aan menslike spermatosoë in vitro toegedien kan word. Glukose konsentrasies van 5,6 mM is bepaal as die gepaste konsentrasie om aan menslike spermatosoë toe te dien, omdat dit beter resultate tot gevolg het; verder is dit vergelykbaar met fisiologiese glukose vlakke in die bloed van `n vastende persoon. Insulien en leptien konsentrasies is op 10 μIU en 10 nm onderskeidelik vasgestel, aangesien dit tot beter resultate gelei het, en omdat dit vergelykbaar was met insulien en leptien vlakke wat reeds voorheen in menslike geëjakuleerde spermatosoë gemeet is. Dit was gevolg deur `n ondersoek na die insulien seintransduksie pad en sy voordelige effekte op menslike spermatosoë funksie. Endogene insulien afskeiding deur menslike geëjakuleerde spermatosoë was deur nifedipien geïnhibeer en sy reseptor tirosien fosforilasie effekte was deur erbstatin geïnhibeer terwyl fosfatidielinositol 3-kinase (PI3K) fosforilasie deur wortmannin geïnhibeer is. Eksogene insulien toediening het menslike sperm-motiliteit parameters betekenisvol laat toeneem asook die vermoë van sperme om die akrosoomreaksie te ondergaan. Die inhibisie van endogene insulien afskeiding deur spermatosoë sowel as die inhibisie van die insulien reseptor substraat (IRS) tirosien fosforilasie het die motiliteit parameters en die akrosoomreaksievermoë van spermatosoë verlaag. Die studie het ook die effekte van insulien en leptien op menslike sperm-motiliteit, -lewensvatbaarheid, -akrosoomreaksie en -NO produksie nagevors. Beide insulien en leptien het sperm-motiliteit parameters, -akrosoomreaksie en -NO produksie betekenisvol verhoog. NO produksie is deur insulien en leptien via PI3K seintransduksie geïnduseer, soos bewys deur die verlaging waargeneem in NO vlakke toe PI3K aktiwiteit deur wortmannin geïnhibeer was. Om vas te stel of insulien en leptien die motiliteit parameters van asthenozoospermiese en teratozoospermiese spermatosoë kon verbeter, het ons spermatosoë in twee fraksies met ’n dubbel digtheid gradiënt geskei. Die gradiënt sisteem was daartoe instaat om die spermatosoë in ’n onvolwasse, (morfologies abnormaal en minder motiel - soortgelyk aan dié van asthenozoospermiese en teratozoospermiese pasiënte), sowel as ’n volwasse meer motiele fraksie te skei. Insulien en leptien het die motiliteit parameters van spermatosoë van die onvolwasse en minder motiele fraksie verhoog. Die vierde gedeelte van die studie was daarop gemik om die effekte van die sitokiene tumor nekrose faktor alfa (TNF-α) en interleukin-6 (IL-6) op menslike sperm-motiliteit, -lewensvatbaarheid, -akrosoomreaksie en -NO produksie, te ondersoek. Die studie het getoon dat TNF-α en IL-6 motiliteit parameters en akrosoomreaksie in ’n tyd- en dosis-afhanklike wyse betekenisvol verlaag het. Hierdie sitokiene was ook in staat om NO produksie in menslike spermatosoë te verhoog. Die verlaging in motiliteit parameters wat deur hierdie sitokiene geïnduseer is, kan toegeskryf word aan hul vermoë om die produksie van oormatige hoeveelhede NO te stimuleer. Dit is nog nie duidelik hoe hulle die akrosoomreaksie in spermatosoë kan inhibeer nie. Die vyfde gedeelte van die studie het dit ten doel gehad om die uitdrukking en lokalisering van die glukose transporter 8 (GLUT8) in menslike spermatosoë te ondersoek. Hierdie studie kon aantoon dat GLUT8 konstitutief uitgedruk is en in die middelstuk van die menslike spermatosoë voorkom. Die studie bewys ook dat stimulering van die spermatosoë met insulien tot `n toename in GLUT8 uitdrukking sowel as translokasie na die akrosomale area, lei. In die finale gedeelte van die studie wou ons ondersoek waarom die toename in NO produksie in spermatosoë (as gevolg van insulien en leptien stimulasie) deur `n toename in spermfunksie gekenmerk word, terwyl die toename in NO produksie (as gevolg van TNF-α en IL-6 stimulasie) deur ’n afname in spermfunksie gekenmerk word. Ons het waargeneem dat TNF-α en IL-6 nie alleen NO produksie nie, maar ook reaktiewe suurstof spesies (ROS) produksie verhoog het. Ons vermoed dat die afname in sperm motiliteit en akrosoomreaksie met TNF-α en IL-6 toediening, die gevolg van die gelyktydige verhoging in NO en ROS was. In gevolgtrekking kan ons sê dat hierdie studie die voordelige in vitro effekte van insulien en leptien op asthenozoospermiese en teratozoospermiese menslike spermfunksie aangetoon het. Hierdie hormone beïnvloed spermfunksie via die PI3K seintransduksie pad op twee maniere. Eerstens, deur `n toename in GLUT8 uitdrukking en translokasie, met die gevolg dat glukose opname en metabolisme moontlik verhoog is, en tweedens, deur die toename in NO produksie. Die studie het ook vasgestel dat TNF-α en IL-6 nadelige effekte op menslike spermatosoë in `n dosis- en tyd-afhanklike wyse het. Hierdie effekte vind plaas a.g.v. hul vermoë om beide NO en ROS produksie in menslike spermatosoë te induseer. Die studie toon aan dat GLUT8 uitdrukking in die middelstuk van die menslike spermatosoon voorkom en dat insulien stimulasie GLUT8 uitdrukking opreguleer en tot translokasie na die akrosomale area lei.

Leptin is a protein hormone primarily secreted by the subcutaneous adipose tissue and is responsible for regulating the energy balance. Individuals with more adipose tissue have much higher leptin levels and often suffer from a condition known as leptin resistance. Various factors such as gender, age, ethnicity, diet and physical activity influence the body's leptin concentration. Males have lower leptin levels than females of the same age and body fat content. Black people tend to have higher leptin concentrations than white people because of a more subcutaneous adipose tissue distribution. Physical activity serve as a main manipulator of the body's energy balance and influence leptin concentration through various adaptations associated with a more favorable body composition such as, an increase in lean tissue and a decrease in body fat. The object of this study was firstly to investigate the association between body composition - with special emphasis on adipose tissue and leptin concentration in black adolescent boys and girls of the North-West Province of South Africa. Secondly, the study aimed to determine the influence of physical fitness components and leptin concentration in the same population. In this study, 148 girls and 114 boys supplied overnight fasting blood samples. Anthropometric data: length (m), weight (kg), skin folds (mm) and circumferences of the waist (cm) and hip (cm) were measured. The percentage body fat were also measured in the BOD-POD. Finally a battery of physical fitness tests were done which included: the maximum number of sit-ups, push-ups and the bleep-test for indirect V&-max. Spearman Rank Order correlations determined that there should be adjusted for age and Tanner stage. Partial correlations were done with body composition variables, [BMI (body mass index), SSF (sum of 6 skin folds), body fat percentage], and physical fitness variables, bush-ups, sit-ups and indirect V02-max]. In both genders a strong positive relationship occurred between all the above mentioned body composition variables and leptin. In boys the strongest correlation (I= 0.8420) was found between SSF and leptin levels. In girls the strongest correlation (r = 0.7489) was found between BMI and leptin concentration. In both genders, body fat percentage indicated the lowest correlation, although it was still significant. In both genders the indirect V02-max values indicated a significant negative relationship with serum leptin concentration, although it was weak, it was the highest in boys (r = - 0314). In girls the indirect V02-max values (r = -0.235) and the maximum amount of push-ups (r = -0205) indicated significant, but weak correlations. According to the results of this study it is clear that serum leptin concentration has a strong positive relationship with body fat, more accurate, with the subcutaneous adipose tissue. It was also indicated that baseline physical fitness in these black adolescents from the selected school in Potchefstroom, North-west Province (South Africa) are statistically significant negatively correlated with leptin levels, although it was not a strong correlation. / Thesis (M.Sc. (Human Movement Science))--North-West University, Potchefstroom Campus, 2006.

La présente thèse est liée à l'étude des indices externes au réseau neuronal et comment ceux-ci impactent le développement du système nerveux central. Spécifiquement, notre objectif était d'explorer l'effet de la leptine, une hormone sécrétée par les adipocytes, sur la plasticité développementale GABAergique. Nous avons utilisé des tranches aigues d'hippocampe de rat nouveau-né pour montrer que la leptine induit une potentialisation de la fréquence de l'activité miniature GABAergique, nécessitant une augmentation postsynaptique de calcium et l'activation de voies de signalisation spécifiques. Nous avons confirmé cet effet sur des cultures de neurones hippocampiques, sur lesquelles nous avons commencé à développer une méthode pour mesurer le corrélat morphologique de la plasticité fonctionnelle des synapses GABAergiques en culture. Cette approche suggère que la plasticité GABAergique induite par la leptine pourrait survenir à densité constante de récepteurs GABAA membranaires. La leptine induit donc une potentialisation de l'activité GABAergique dans les neurones hippocampiques en développement. Enfin, nous avons trouvé que les neurones pyramidaux de CA3 reçoivent une activité miniature GABAergique réduite chez des souris ob/ob ne produisant pas de leptine, suggérant que la leptine contribue au développement de la circuiterie GABAergique in vivo. Dans l'ensemble, les études que nous présentons apportent un éclairage nouveau sur le développement d'aires cérébrales dites de « haut niveau », dont nous avons observé qu'elles intègrent des signaux dits de « bas niveau », c'est-à-dire en provenance de la périphérie afin de modeler leur développement. / The present dissertation tackles the larger question of how external cues impact the development of the central nervous system. Our specific aim was to explore the effect of leptin, an adipocyte-derived hormone, on GABAergic plasticity in the developing rodent hippocampus. We used acute hippocampal slices of newborn rats to show that leptin induces a long lasting potentiation of the frequency of miniature GABAergic activity. Using pharmacological tools we found that this event requires a postsynaptic increase in intracellular calcium as well as specific postsynaptic signaling pathways. To address the mechanistic action of leptin we confirmed the leptin-induced plasticity on hippocampal cultures and began to develop a method to measure the morphological correlate of GABAergic synapses in culture. Applying this method suggested that the leptin-induced GABAergic plasticity might occur with a constant density of postsynaptic GABAA receptor puncta. Taken together, these data show that leptin induces a potentiation of GABAergic activity in developing hippocampal neurons, perhaps by recruiting clusters of GABAA receptors expressed at the membrane to form newly functional GABAergic synapses. In addition we found that CA3 pyramidal neurons of leptin-deficient ob/ob mice exhibit lower miniature GABAergic activity compared to wild type littermates, which suggests that leptin contributes to the development of the hippocampal GABAergic circuitry in vivo. Overall, these studies shed a new light on the development of admittedly "higher-level" cerebral regions which were found here to integrate "lower-level", peripheral signals to shape their development.

Obesity and associated metabolic disorders, called as "metabolic syndrome", currently represent a major social and economical problem of public health. From the energy balance point of view, long-lasting energy surplus leads eventually to massive accumulation of energy stores resulting in various adverse effects on metabolism and health. General goal of the thesis was to examine these metabolic disorders at cellular and whole-body level using suitable mouse models. The main focus was on the most metabolically active tissue, namely skeletal muscle, liver and adipose tissue and on the regulatory roles of AMP-activated protein kinase (AMPK) and leptin in the energy metabolism. The whole thesis is based on four published studies. Two studies were focused on skeletal muscle. In the first study, we proved the involvement of leptin and AMPK in the metabolic response to high-fat diet-feeding. We described a mechanism of muscle non- shivering thermogenesis based on enhanced lipid catabolism, which contributes to the genetically-determined resistance of inbred A/J mice to obesity. Such mechanism was not operating in obesity-prone C57BL/6 mice. In the second study, performed using C57BL/6 mice, we have described beneficial effect of combination treatment using n-3 polyunsaturated fatty acids (n-3 PUFA) of...

Nos últimos anos o gene da leptina, vem sendo objeto de intensa investigação em bovinos pela sua associação com características econômicas, tais como, metabolismo energético, eficiência e consumo alimentar, deposição de gordura e características reprodutivas. Estudos anteriores apontam que esse gene possa influenciar o início da puberdade tanto em animais como em humanos. Tendo em vista a importância desse gene para a pecuária nacional, os objetivos do presente estudo foram determinar a relação entre expressão do gene da leptina e puberdade em novilhas Nelore; identificar polimorfismos no promotor e no gene da leptina; avaliar os efeitos desses polimorfismos com a puberdade, expressão do gene e diferença esperada na progênie (DEP) para probabilidade de prenhez de novilha aos 14 meses (PP14). Amostras de tecido adiposo subcutâneo foram coletadas de 28 novilhas Nelore com idade em torno de 25 meses, sendo 14 pré-púberes e 14 púberes. Análises foram feitas por RT-PCR quantitativa, utilizando cDNA sintetizado a partir de RNA total extraído das amostras de tecido adiposo. A expressão da leptina foi 2,1 vezes maior (P=0,0337) no tecido adiposo das novilhas púberes, sugerindo que esse gene possa estar influenciando o início da puberdade nestes animais. Amostras de DNA foram extraídas para identificar polimorfismos no gene e no promotor da leptina que poderiam estar associados com a puberdade e a expressão do gene no tecido adiposo subcutâneo. Foram identificados 37 polimorfismos, dos quais 20 não haviam sido reportados previamente. Nenhum dos 37 polimorfismos encontrados ou dos 10 haplótipos formados a partir dos polimorfismos, foram associados (P0,05) com a maior expressão do gene ou com a puberdade nestas novilhas. Aparte disso, o DNA de 96 fêmeas Nelore, sendo 48 com alta DEP para PP14 e 48 com baixa DEP para PP14 pertencentes a outro rebanho, foi extraído a partir de sangue periférico. Uma região de 715 pb do gene da leptina foi seqüenciada nestas 96 amostras. Nenhuns dos 11 polimorfismos identificados apresentaram associação (P0,05) com os valores de alta ou baixa DEP para PP14, entretanto a substituição do haplótipo 1, o mais freqüente, pelo haplótipo 2 revelou uma associação sugestiva (P=0,0923) com a característica, indicando que a substituição possa estar contribuindo para a variação da DEP para PP14. Fêmeas com o haplótipo 2 em média tiveram efeito de -9,51 a DEP para PP14 em relação aquelas com o haplótipo mais freqüente. Este estudo confirma a associação da expressão do gene da leptina com a puberdade, descreve novos polimorfismos identificados neste gene e apresenta resultados iniciais da associação dos polimorfismos com expressão e puberdade em Nelore. / In recent years, the leptin gene has been subject of intense research in cattle, possibly because of its association with economic characteristics, such as, energy metabolism, feed efficiency, feed intake, fat deposition and reproductive characteristics. Previous studies suggest that this gene might be influencing the onset of puberty in both animals and humans. Given the importance of this gene for the national livestock, the goals of this study was to determine the relationship between leptin gene expression and puberty in Nelore heifers; identified polymorphisms in the promoter and leptin gene; evaluate the effect of these polymorphisms on puberty, gene expression and expected progeny difference (EPD) for probability of pregnancy of heifers at 14 months (PP14). Samples of subcutaneous adipose tissue were collected from 28 heifers aged around 25 months, 14 prepubertal and 14 pubertal. Leptin gene expression was determined by quantitative RT-PCR using cDNA synthesized from total RNA extracted from adipose tissue samples. The expression of leptin was 2.1 times higher (P=0,0337) in subcutaneous adipose tissue of pubertal heifers, suggesting that this gene might be influencing the onset of puberty in these animals. DNA from these heifers were extracted to identify polymorphisms in the promoter and leptin gene that could be associated with puberty and gene expression in subcutaneous adipose tissue of these heifers. We identified a total of 37 polymorphisms, 20 of which have not been reported previously. None of the 37 polymorphisms found or the 10 haplotypes formed from the polymorphisms were associated (P0.05) with higher expression of the gene or with puberty in these heifers. Apart from this, DNA of 96 Nelore, being 48 with high EPD for PP14 and 48 with low EPD for PP14 belonging to another herd, was extracted from peripheral blood. A region of 715 bp in the leptin gene was sequenced in these 96 samples. None of the 11 polymorphisms identified were associated (P0.05) with high or low values of EPD for PP14, however replacing the haplotype 1, the most frequent, for haplotype 2 showed a suggestive association (P=0,0923) with the trait, indicating that the replacement can contribute to the variation of EPD for PP14. Females with haplotype 2 had an average effect of -9.51 on EPD for PP14 compared to those with the most frequent haplotype. This study confirms the association of leptin expression on puberty, describes new polymorphisms identified in this gene and presents initial results of the association of polymorphisms with expression and puberty in Nelore.

O tecido adiposo é considerado não somente uma fonte de energia estocável, mas principalmente um órgão endócrino que secreta várias citoquinas, as quais podem contribuir para o desenvolvimento de doenças relacionadas à obesidade, incluindo o diabetes mellitus e a doença vascular aterosclerótica. Dentre esse pool de moléculas, a adiponectina (Arcp30, AdipoQ, apM1, ou GBP28), uma nova proteína semelhante ao colágeno, foi descoberta como uma citoquina específica do adipócito. Neste estudo, realizamos a determinação dos níveis séricos da adiponectina em uma amostra de pacientes hospitalizados com SCA e, posteriormente, avaliamos a associação com os eventos cardiovasculares no seguimento clínico. Método: avaliamos 114 pacientes com SCA de ambos os sexos neste estudo de corte transversa com seguimento clínico mediano de 18 meses. Realizamos análise de regressão multivariada de Cox para identificar associação independente entre adiponectina e o risco subsequente de óbito CV, IAM não-fatal, AVE não-fatal, e rehospitalização com revascularização. Também comparamos os diversos biomarcadores metabólicos, inflamatórios, de coagulação e de necrose miocárdica por quartis de adiponectina. Resultados: Adiponectina não correlacionou-se de modo independente com o risco CV, tanto na análise univariada quanto nos modelos de Cox. Das variáveis metabólicas, a única com valor preditivo para os desfechos primário e co-primário foi a glicemia de jejum, com OR ajustado=1,06 (IC95% 1,01-1,11), p=0,016, e OR ajustado=1,10 (IC95% 1,03-1,17), p=0,003, ambos para incrementos de 10 na glicemia de jejum. Conclusões: Adiponectina não se mostrou variável independente de risco cardiovascular nesta amostra de pacientes com SCA. A glicemia de jejum foi a única variável metabólica com valor preditivo independente para os desfechos cardiovasculares. / Background: The adipose tissue is considered not only an energy resource stock, but mainly an endocrine organ that secretes several cytokines, which may contribute to the development of diseases related to obesity, including diabetes mellitus and the atherosclerotic vascular diseases. Within this pool of molecules, adiponectin (Arcp30, AdipoQ, apM1, or GBP28), a novel protein similar to collagen, was discovered as an adipocyte-specific cytokine. In this study, we determined the serum levels of adiponectin in a sample of patients hospitalized with acute coronary syndromes (ACS), and subsequently we evaluated the association with the cardiovascular events during the follow-up phase. Methods: we evaluated 114 patients with ACS of both genders in this cross-sectional study with a median follow-up of 18 months. We performed a proportional multiple regression analysis of Cox to identify independent association between adiponectina and risk of cardiovascular death, non-fatal acute MI, non-fatal CVA, and rehospitalization requiring revascularization. We also compared the various biomarkers of metabolism, inflammation, coagulation, and of myocardial necrosis divided by quartiles of adiponectin. Results: Adiponectin did not correlate independently with CV risk, either on univariate analysis or on the multiple regression models of Cox. Among the metabolic biomarkers, the only variable with predictive value for the primary and co-primary endpoints was fasting glucose, with an adjusted OR=1,06 (95%CI 1,01-1,11), p=0,016, and adjusted OR=1,10 (95%CI 1,03-1,17), p=0,003, both for increments of 10. Conclusions: Adiponectin was not an independent risk factor for cardiovascular risk in this sample of ACS patients. Fasting glucose was the only metabolic biomarker with a significant and independent predictive value for cardiovascular outcomes.

La cellule principale de l’os sous-chondral est l’ostéoblaste qui joue un rôle central dans la production qualitative et quantitative de la matrice ostéoïde. Plusieurs études montrent que le collagène de type I et la phosphatase alcaline sont des marqueurs précoces de la différenciation des ostéoblastes tandis que l’ostéocalcine (OCN) et la minéralisation sont des marqueurs des stades tardifs de cette différenciation. L’os sous-chondral est le site actif de nombreux changements morphologiques qui peuvent être différents au cours de l’arthrose (OA) mais qui sont partie prenante du processus pathologique. Ces changements consistent en une formation de la matrice osseuse importante associée une inhibition de la minéralisation. Ces phénomènes peuvent être reliés aux changements phénotypiques des ostéoblastes. La galectine-3 (gal-3) est un facteur inflammatoire qui a été détecté dans le tissu synovial et dans le liquide synovial lors d’inflammation d’OA. Des études antérieures ont montré que la gal-3 participait à la destruction du cartilage et inhibait fortement la production d’OCN par les ostéoblastes OA. Ces faits ont permis de suggérer que la gal-3 pouvait participer soit à l’initiation soit à la progression de l’arthrose. Peu d’études ont globalement été réalisées sur le rôle de la galectine-3 dans l’arthrose et encore moins sur le rôle de gal-3 sur les altérations de l’os sous-chondral.Dans ce contexte, ce travail de thèse a consisté à caractériser les ostéoblastes arthrosiques, puis investigué la modulation du phénotype des ostéoblastes arthrosiques par gal-3 en et enfin identifier les mécanismes cellulaires impliqués. D’une part, nous avons identifié deux populations ostéoblastes OA grâce à l’expression d’OCN. Dans les conditions basales, ces deux populations expriment de façon différentielle le TGF-ß1, Wnt5b et DKK2, ce qui suggère une différenciation et un phénotype hétérogènes des ostéoblastes chez les patients OA. D’autre part, nous confirmons le rôle délétère de la gal-3 dans l’articulation lors d’inflammation puisqu’elle stimule la production de collagénase 1 impliquée dans la dégradation osseuse. De plus, elle accentue la perturbation phénotypique des ostéoblastes qui produisent plus de leptine lors d’épisodes hypoxiques. Bien que plusieurs ligands membranaires puissent médier les effets de gal-3, 4F2hc semble jouer un rôle récurrent / Osteoblasts are the main cells in subchondral bone (SCB), which are responsible for the bone matrix production. Their differentiation can be evaluated by type I collagen and alkaline phosphatase in the early stage and by osteocalcin (OCN) and mineralization in the late stage. Alterations of SCB are essential episodes of osteoarthritis (OA) and are represented by a significant bone formation accompanied with abnormal hypomineralization. These changes in SCB are related to phenotypic modifications of osteoblasts. Galectin-3 (Gal-3) is an inflammatory factor markedly detected in the synovial tissue and synovial fluid during OA inflammation. Previous studies have demonstrated that gal-3 was deleterious for cartilage and inhibited the production of OCN in OA osteoblasts. These findings suggest that gal-3 can participate in either the initiation or progression of osteoarthritis. So far, a few studies have been conducted to explore the role of Gal-3 in OA and particularly related to SCB. In this context, the thesis has consisted to characterize OA osteoblasts, to investigate the modulation of the OA osteoblast phenotype by gal-3 and finally to identify the involved cellular mechanisms. We have identified two populations of OA osteoblasts according to the OCN expression. Under basal conditions, these two populations express TGF-ß1, Wnt5b and DKK2 differentially, suggesting various differentiation and heterogeneous phenotype of osteoblasts in OA patients. Moreover, we confirmed the deleterious role of gal-3 in the joint during inflammation since it stimulates the production of collagenase 1 involved in bone degradation. In addition, it emphasizes the disruption of phenotypic osteoblasts by producing more leptin during hypoxic episodes. Although several membrane ligands can mediate the effects of gal-3, 4F2hc seems to play a prominent role

Made available in DSpace on 2016-03-22T17:27:02Z (GMT). No. of bitstreams: 1 gisele.pdf: 962432 bytes, checksum: ec65fd91558d80cca72d2f5de776a6d7 (MD5) Previous issue date: 2013-02-27 / Major depression disorder is associated with elevated risk of numerous metabolic disturbances. Leptin resistance or deficiency have been suggested in mood regulation. Here, we investigate whether leptin levels are associated with current depression in a nested population-based study cross-sectional paired. We evaluate 256 subjects, 128 control and 128 individuals diagnosed with current depression by a structured diagnostic interview Mini International Neuropsychiatric Interview and were drugnaïve for antidepressive. Individuals were categorized according to body mass index (BMI) in normal weight (&#8805;18.5 and <25.0), overweight (&#8805;25.0 and <29.9) or obese (&#8805;30.0). Higher leptin levels was associated with female sex (p=0.001), body mass index (p=0.093), and decreased in physically active subjects (p=0.001). In gender and BMI stratified sample, lower leptin levels were associated with current depression only in normal-weight women related to control group [12.57(5.34-24.94) ng/mL vs 23.35 (11.70-53.58) ng/mL, respectively; p=0.016]. Linear regression analysis reveals that leptin is an independent factor for current depression (&#946;=-0.239; p=0.045) after adjustment for physical activity. Our results shows that current depression in women are associated with reduced levels of serum leptin, in a gender and BMI dependent manner / N