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  • About
  • The Global ETD Search service is a free service for researchers to find electronic theses and dissertations. This service is provided by the Networked Digital Library of Theses and Dissertations.
    Our metadata is collected from universities around the world. If you manage a university/consortium/country archive and want to be added, details can be found on the NDLTD website.
41

Estudo da associação entre polimorfismos do gene do receptor de vitamina D (VDR) e do SNP-71 A/G do gene 17 beta- hidroxiesteróide desidrogenase tipo 5 (HSD17B5) e variáveis clínicas, hormonais e metabólicas em pacientes com pubarca precoce e controles

Santos, Betânia Rodrigues dos January 2011 (has links)
A pubarca precoce (PP) é definida como o desenvolvimento de pêlos pubianos antes dos 8 anos de idade em meninas e 9 anos de idade em meninos. Embora a PP não interfira diretamente com os eventos da puberdade, algumas evidências sugerem que estas meninas tenham maior risco para desenvolver, mais tarde, a Síndrome dos Ovários Policísticos (PCOS). A 17ß-hidroxiesteróide desidrogenase tipo 5 (17ßHSD5) é a principal responsável pela conversão de androstenediona em testosterona. Variações no gene que codifica para essa enzima, em especial os polimorfismos de nucleotídeo único (SNPs), podem estar relacionados com hiperandrogenismo e PCOS. A vitamina D, além dos efeitos sobre metabolismo ósseo, parece modular outras ações extra-esqueléticas, incluindo secreção e sensibilidade tecidual à insulina. A Vitamina D vem sendo associada com resistência insulínica e variantes do gene do receptor da vitamina D (VDR), vem sendo estudadas em populações de risco como no diabetes. No entanto, pouco se sabe sobre o envolvimento destes polimorfismos na PP. Os objetivos do presente trabalho foram: Avaliar os níveis de 25-hidroxivitamina D; determinar a frequência dos polimorfismos FokI, BsmI, ApaI e TaqI do gene do VDR e do SNP-71AG do gene da 17ßHSD5; verificar se existe associação entre esses polimorfismos com variáveis antropométricas, metabólicas e hormonais em uma amostra de pacientes com PP e controles do sul do Brasil. Foram arroladas 36 meninas com PP e 197 controles saudáveis. As genotipagens foram realizadas por PCR em tempo real para os SNPs -71AG, BsmI e FokI e por PCR-RFLP para os SNPs ApaI e TaqI. O SNP -71 AG do gene da 17ßHSD5 apresentou distribuição genotípica de 52,4% AA, 39,1% AG e 8,6% GG, sendo a frequência dos alelos A:G de 0,72:0,28. Analisando os dois grupos, verificamos uma maior freqüência do alelo variante (G) no grupo de meninas com PP quando comparadas aos controles (0,37 e 0,26, respectivamente), no entanto sem diferença estatística (p=0,054); não foram verificadas associações do polimorfismo com os dados clínicos e hormonais. As meninas com PP apresentaram níveis séricos de 25(OH)D inferiores aos das meninas controles (18,08±8,32 versus 21,27±7,03; p=0,032). Na análise dos polimorfismos, observou-se que o genótipo polimórfico GG do SNP ApaI TG, apresentou uma frequência maior em PP (30,6%) do que nas controles (16,2%) (Odds Ratio: 2,269; 95% Intervalo de Confiança: 1,015 – 5,076; p=0,042). Este genótipo foi também associado com níveis mais baixos de estradiol (35,30 (14,80 – 50,48) versus 12,22 (6,49 – 23,69); p=0,030) e testosterona total (0,52 (0,39 – 0,84) versus 0,20 (0,11 – 0,47); p=0,009) nas meninas com PP, mas não foi associado com os níveis de 25(OH)D. Por outro lado, verificou-se associação entre a presença dos polimorfismos TaqI TC (genótipo TC+CC) e BsmI GA (genótipo GA+AA) e níveis séricos de 25(OH)D mais elevados no grupo de meninas saudáveis (19,86±7,16 versus 22,55±6,69, p=0,007; 19,53±6,94 versus 22,88±6,76, p=0,001, respectivamente). Em conclusão, os dados deste estudo indicam que: 1) houve maior freqüência do alelo variante G SNP -71 AG do gene da 17βHSD5, com uma associação limítrofe desse alelo com o diagnóstico clínico de PP; 2) o polimorfismo ApaI TG associou-se com PP e parece estar modulando os processos esteroidogênicos nas meninas com PP; 3) houve uma interação entre os polimorfismos TaqI TC e BsmI GA e concentrações circulantes de vitamina D em meninas do sul do Brasil. / Precocious pubarche (PP) is usually defined as the development of pubic hair before the age of 8 in girls and age of 9 in boys. Although the PP does not interfere directly in puberty events, some evidence suggests that these girls have higher risk for the development of Polycystic Ovary Syndrome (PCOS) at later ages. The Type 5 17β-Hydroxysteroid Dehydrogenase (17ßHSD5) is the principal responsible for the conversion of androstenedione to testosterone. Variations in the gene encoding for this enzyme, especially Single Nucleotide polymorphisms (SNPs), may be related with hyperandrogenism, and PCOS. Besides the effects on bone metabolism, vitamin D appears to modulate other extra-skeletal actions, including secretions and tissue sensitivity to insulin. Vitamin D has been associated with insulin resistance and variants in the vitamin D receptor (VDR) gene, have been studied in populations at risk of Diabetes. However, little is known about these polymorphisms in the PP. The aims of this work were: to evaluate the levels of the 25-hydroxyvitamin D; to determine the polymorphisms FokI, BsmI, ApaI and TaqI in VDR gene and SNP -71AG in 17ßHSD5 gene frequencies; to asses if exist association between this SNPs and anthropometric, metabolic and hormonal characteristics in patients with PP and controls of the southern Brazil. Were enrolled 36 girls with PP and 197 healthy controls. Genotypic analyzes were evaluated by Real Time for the SNPs -71AG, BsmI and FokI and by PCR-RFLP for the ApaI e TaqI polymorphisms. Genotype frequency for SNP -71 AG of the 17ßHSD5 gene was 52.4% AA, 39.1% AG and 8.6% GG, A:G allelic frequency was 0.72:0.28. Analyzing both groups, higher frequency of the variant allele (G) in patient PP than controls (0.37 e 0.26, respectively) was found but without statistical difference (p=0.054); there were no associations between this polymorphism and clinical and hormonal features. PP girls have serum levels of 25(OH)D lower than those from control group (18.08±8.32 versus 21.27±7.03; p=0.032). The polymorphism analyze was observed that genotype GG of the SNP ApaI TG showed a higher frequency in PP (30.6%) than controls (16.2%) (Odds Ratio: 2.269; 95% confidence interval: 1.015 – 5.076; p=0.042). The same genotype was associated with lower estradiol (35.30 (14.80 – 50.48) versus 12.22 (6.49 – 23.69); p=0.030) and total testosterone levels (0.52 (0.39 – 0.84) versus 0.20 (0.11 – 0.47); p=0.009), in girls with PP. There were no association between this polymorphism and serum 25(OH)D. On the other hand, there was association between the presence of the polymorphisms TaqI TC (TC + CC genotype) and BsmI GA (GA + AA genotype) and higher serum 25(OH)D in the group of healthy girls (19.86 ± 7.16 versus 6.69 ± 22:55 , p = 0.007; 19:53 ± 6.94 versus 22.88 ± 6.76, p = 0.001, respectively). In conclusion, data from this study indicate that: 1) there was a higher frequency of the variant allele G of the SNP -71 AG of the 17ßHSD5 gene, with a borderline association of this allele with the clinical diagnosis of PP; 2) ApaI TG polymorphism is associated with PP and seems to modulate the processes steroidogenesis in girls with PP; 3) there was an interaction between the polymorphisms TaqI TC and BsmI GA and vitamin D concentrations in girls from southern Brazil.
42

Estudo do efeito bioquímico e estrutural de novas mutações em enzimas da esteroidogênese / Study of the biochemistry and structural effects of rare mutations in steroidogenesis enzymes

Lusa, Ana Leticia Gori, 1983- 05 March 2013 (has links)
Orientadores: Maricilda Palandi de Mello, Fernanda Caroline Soardi / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-23T09:56:23Z (GMT). No. of bitstreams: 1 Lusa_AnaLeticiaGori_D.pdf: 4582728 bytes, checksum: a14d0d0453e90b4a59cb02532b21d83b (MD5) Previous issue date: 2013 / Resumo: A Hiperplasia congênita da Adrenal (HCA) é uma das doenças mais comuns com herança autossômica recessiva. Uma das causas é a deficiência da 21-hidroxilase que surge por mutações no gene CYP21A2. O desequilíbrio na síntese de cortisol causa um estímulo excessivo das glândulas adrenais pelo hormônio adrenocorticotrópico (ACTH) e, consequentemente, à hiperplasia da glândula e ao aumento na síntese de andrógenos. O presente trabalho teve por objetivo clonar e expressar variantes do gene CYP21A2 e, avaliar as variantes enzimáticas da 21-hidroxilase quanto à atividade residual, comparando suas formas nativa e mutantes. Foi também realizado o estudo in silico para se comparar as estruturas das enzimas 21-hidroxilase e 3?-hidroxiesteróide desidrogenase tipo 2 (HSD3B2) avaliando o impacto das alterações provocadas por mutações sobre a estrutura da proteína nativa. Para a enzima 21-hidroxilase, as mutações analisadas quanto à atividade foram: p.Ser113Phe, p.Pro267Leu, p.Val358Ile, p.Arg426Cis e, as combinações p.Ile172Asn+p.Val358Ile e p.Val281Leu+p.Arg426Cis. Nos testes de atividade enzimática, as enzimas com as mutações p.Val358Ile e as combinações p.Ile172Asn+p.Val358Ile e p.Val281Leu+p.Arg426Cis apresentaram valores de atividade residual compatíveis com a forma perdedora de sal. Por outro lado, a enzima com a mutação p.Pro267Leu apresentou atividade residual maior que 50%, compatível com mutações da forma não clássica. As mutações p.Ser113Phe e p.Arg426Cis, por sua vez, apresentaram atividade residual entre 2 e 7% que, em relação aos controles utilizados, foram correspondentes à forma clássica virilizante simples. Os modelos estruturais gerados para as enzimas 21-hidroxilase portadoras da deleção p.(Gln389_Ala391del) e das mutações individuais ou combinadas acima citadas foram comparados ao da enzima nativa. Estudo similar foi realizado com enzimas HSD3B2 nativa e portadoras de duas mutações, p.Pro222Gln e p.Pro222Thr, relacionadas à forma perdedora de sal da deficiência de HSD3B2. Esse estudo demonstrou boa correlação das mutações encontradas nos pacientes com os seus fenótipos tanto no que diz respeito à atividade enzimática quanto às alterações estruturais. Assim, ficou demonstrado que este tipo de abordagem é eficaz para a avaliação dos aspectos funcionais de mutações inéditas / Abstract: Congenital adrenal hyperplasia (CAH) is one of the most common hereditary autosomal recessive diseases. 21-hydroxylase deficiency is one important cause of the disease and results from mutations in CYP21A2 gene. The unbalance in cortisol synthesis leads to excessive stimulation of adrenal glands by adrenocorticotropic hormone (ACTH), consequently, to adrenal hyperplasia and excessive androgen synthesis. The present study aimed to obtain and express CYP21A2 clones carrying sequence variations and to evaluate 21-hydroxylase enzymatic residual activities of variant enzymes in comparision to the wild-type. In silico studies were also performed to compare the structure of 21-hydroxylase and 3?-hydroxysteroid dehydrogenase type 2 (HSDB2) evaluating the impact of mutations upon the native structure of the enzymes. Residual activity analyses for 21-hydroxylase enzyme comprised the mutations p.Ser113Phe, p.Pro267Leu, p.Val358Ile and the combined mutations p.Ile172Asn+p.Val358Ile and p.Val281Leu+p.Arg426Cys. The assays indicated that p.Val358Ile and p.Ile172Asn+p.Val358Ile and p.Val281Leu+p.Arg426Cys reduced CYP21A2 activity to levels corresponding to the salt wasting form. Whereas the enzyme bearing p.Pro267Leu mutation showed residual activity higher than 50% indicating a nonclassic mutation. Conversely, p.Ser113Phe and p.Arg426Cys mutations showed residual activities that varied between 2% and 7% that, compared to controls, corresponded to simple virilizing form. Structural models generated for 21-hydroxylase enzyme with p.(Gln389_Ala391del) and each individual or combined mutations cited above have been compared to the native enzime. Similar study was performed with HSD3B2 enzyme in its native and mutated formas with p.Pro222Gln e p.Pro222Thr that are related to the salt wasting form of HSD3B2 deficiency. This study demonstrated a good genotype-phenotype correlation in patients either for residual activities or for structural changes. Therefore, it has been demonstrated that such analyses are effective to evaluate functional aspects for novel mutations / Doutorado / Genetica Animal e Evolução / Doutora em Genética e Biologia Molecular
43

Die Bedeutung partieller 21-Hydroxylase- und 3beta-Hydroxysteroiddehydrogenasedefizienzen für die Ätiopathogenese von Fertilitätsstörungen

Ghanaati, Zahra 12 March 2001 (has links)
Ziel der Untersuchungen war, zur Klärung der Ursachen einer während der letzten Jahrzehnte erhöhten Frequenz sowohl von PCOS als auch von IO beizutragen. Es war zu ermitteln, ob hormonelle Verschiebungen bei den Patienten nachweisbar und diese durch genetische und epigenetische Faktoren erklärbar sind. Ausgehend von dem Postulat, daß verminderte 21-OH- und 3beta-HSD-Aktivitäten als prädisponierende Faktoren von PCOS und IO angesehen werden, waren hormonanalytische Untersuchungen zur Ermittlung partieller 21-OH- bzw. 3beta-HSD-Defizienzen durchgeführt worden. Den eigenen Erfahrungen und Darstellungen der internationalen Literatur entsprechend befaßt sich ein Teil der Methodik mit der Entwicklung einer neuen, der üblichen 17alfa-OHP-Messung überlegenen Methode zur Ermittlung von 21-OH-Defizienzen durch 21-DOF-Bestimmung nach ACTH-Test im Blutplasma. Wir erhielten bei vier von 21 PCOS-Patientinnen und drei von acht Patienten mit IO erhöhte 21-DOF-, 21-DOF/F- bzw. 17alfa-OHP-Werte nach ACTH-Test, die auf partielle 21-OH-Defizienzen hinweisen. Zusätzlich wurden bei 12 PCOS-Patientinnen erhöhte basale DHEAS- oder DHEAS/F-Werte gefunden, die als Hinweise auf partielle 3beta-HSD-Defizienzen oder 17,20-Lyase-Hyperaktivität gedeutet wurden. In der Stichprobe der IO waren DHEAS oder DHEAS/F-Werte bei vier Patienten erhöht. Da bei vier der 12 Patientinnen mit PCOS und zwei von vier Patienten mit IO genetisch und endokrinologisch gleichzeitig eine partielle 21-OH-Defizienz nachgewiesen wurde, kann bei diesen Patienten eine partielle 3beta-HSD-Defizienz weitgehend ausgeschlossen werden. Es wurden molekulargenetische Untersuchungen für die 14 häufigsten Mutationen in CYP21 bei Cohorten mit AGS, PCOS und IO durchgeführt. Die Untersuchung der AGS-Patienten sollte dazu dienen, ein effizientes und schnelles System der Mutationssuche für diagnostische Zwecke zu etablieren. Es wurden die häufigsten, phänotypisch wirksamen Mutationen in CYP21 bei der Mehrzahl dieser Patientengruppe im homozygoten bzw. compound heterozygoten Zustand gefunden und eine deutliche Genotyp-Phänotyp-Korrelation festgestellt. Auch bei Patientinnen mit PCOS sowie bei IO, bei denen partielle 21-OH-Defizienzen nachweisbar waren, wurden Mutationen in CYP21 gefunden. Die hierbei heterozygot vorliegenden Mutationen waren dieselben, die homozygot oder compound heterozygot bei schweren Formen des AGS gefunden wurden. Es ergab sich eine Korrelation molekulargenetischer und hormonanalytischer Befunde bei AGS, PCOS sowie IO. Allerdings konnten bei der Mehrzahl der Fälle mit PCOS und mit IO weder Mutationen noch hormonelle Auffälligkeiten hinsichtlich partieller 21-OH-Defizienzen gefunden werden. Die jedoch bei vielen Patientinnen gefundenen erhöhten DHEAS- und DHEAS/F-Werte stimmen mit Untersuchungen überein, die parallel starke Zunahmen der Häufigkeit der Hemmung des Enzyms 3ß-HSD bzw. der Aktivierung der 17,20-Lyase bei PCOS-Patientinnen und der Prävalenz des PCOS selbst bei nach 1955 geborenen Frauen und von Spermatogenesestörungen bei nach 1960 geborenen Männern fanden. Die Ursache hierfür wird in der Beeinflussung der adrenalen und gonadalen Steroidhormonsynthese vor allem durch das Umweltteratogen DDT und seine Metaboliten gesehen. Weiterhin wurde der Umweltfaktor Streß diskutiert. Für die Ätiopathogenese der untersuchten Fertilitätsstörungen werden materno-fetale Mechanismen postuliert, worauf unsere sowohl molekulargenetischen als auch hormonanalytischen Befunde hinweisen. Insgesamt bestätigen die Ergebnisse unserer Arbeit die These, daß Leben auf der Interaktion von Genen und Umweltfaktoren beruht und daß Hormone dabei als Mediatoren wirken. In gen- oder umweltbedingten unphysiologischen Konzentrationen können sie während kritischer Entwicklungsphasen des neuroendokrinen Systems als Teratogene wirken und zu lebenslangen Reproduktionsstörungen führen. / This paper describes a mutational and hormonal screening in a cohort of 21 patients ultrasonically diagnosed with PCO. Our data show single heterozygous base pair CYP21 mutations in 4 patients. The four women with PCOS and CYP21 mutations also displayed clear signs of partial 21-hydroxylase deficiency through a significant rise in 21DOF or 17alfa-OHP plasma levels after ACTH stimulation. Azziz et al. have reported several heterozygous mutations in hyperandrogenic women with LO-CAH. Other studies report several heterozygous point mutations in hyperandrogenic woman who, however, were not examined for polycystic ovaries.The correlation between the hormone profiles and genetic screening results found with our patients underscores the latter s usefulness with PCOS patients. In contrast to the hormone profile, genetic screening is not influenced by external factors. The frequency of heterozygous CYP21 mutations is higher (19%) than in the normal population (5-8%), suggesting a link with PCOS in some cases. The ratio of LH/FSH was significantly raised in 43% of the cases. Most importantly, basal plasma DHEA-S levels and DHEA-S/F ratios were clearly increased, higher than the means +2SD in controls. This suggests a partial 3beta-hydroxysteroid dehydrogenase deficiency or 17,20 lyase hyperactivity. Other authors, however, were not able to find mutations in the corresponding genes. This could be explained by the fact that the DDT metabolite o,p DDD is a strong inhibitor of 3beta-HSD, and that DDT and its metabolites may be able to activate the 17,20 lyase, a cytochrome P450 enzyme. Furthermore, DDT has some oestrogen activity, and its perinatal administration can produce a PCOS-like syndrome in rats. Very significantly, there has not only been an approximately fourfold increased prevalence of PCO in women borne since 1955 in eastern Germany, following a massive prenatal exposure to DDT, but also a notable shift in the hormone profiles of those affected. A predominance of 3beta-HSD deficiencies and 17,20 lyase hyperactivity (70%) vs. 21-hydroxylase deficiency (23%) has emerged, in contrast with 21-hydroxylase deficiencies in 70% vs. 3beta-HSD deficiencies or 17,20 lyase hyperactivity in 14% for those born earlier than 1955. Similar results were obtained in this study for women with PCOS born since 1955, suggesting that the prenatal exposure of high amounts of DDT and its metabolites indeed appear to be responsible - at least in part - for the major increase in PCO and PCOS.
44

Variation de l’expression et de l’activité des 11β-hydroxystéroïde déshydrogénases rénales, cardiaques et placentaires au cours de la gestation de la rate

Barrette, Mathieu 08 1900 (has links)
L’activation du système rénine-angiotensine-aldostérone peut entraîner le développement d’une hypertension artérielle et de la fibrose cardiaque. Toutefois, au cours de la grossesse, malgré une hausse substantielle des niveaux d’aldostérone, ces effets délétères ne sont pas observés. L’aldostérone exerce ses effets via les récepteurs des minéralocorticoïdes, les MR, qui peuvent également lier le cortisol avec une affinité similaire. La régulation des niveaux locaux de ce glucocorticoïde par les 11β-hydroxystéroïde déshydrogénases (11β-HSD) est donc essentielle pour éviter une stimulation inappropriée des MR. Nous suggérons que, durant la grossesse, ces enzymes sont impliquées dans la protection de la mère et du foetus contre les niveaux élevés d’aldostérone et de cortisol. Notre hypothèse de travail est que les mécanismes d’adaptation qui prennent place au cours de la grossesse nécessitent des changements d’expression (ARNm et protéine) et d’activité des 11β-HSD spécifiques selon le tissu. Des rates Sprague-Dawley ont été sacrifiées aux jours 14, 17, 19 et 22 de gestation (terme = jour 23) et leurs organes ont été collectés. Dans le rein, les niveaux protéiques des 11β-HSD sont diminués en fin de gestation. Dans le placenta, on observe une importante chute de l’expression génique et protéique de la 11β-HSD1 au jour 17 tandis que la 11β-HSD2 y est augmentée. L’expression et l’activité de la 11β-HSD2 sont par la suite diminuées jusqu’à terme. Aucune différence significative n’est retrouvée dans le ventricule gauche cardiaque. En conclusion, nos résultats démontrent que la gestation est accompagnée d’importants changements dans le placenta, possiblement pour assurer un développement foetal adéquat, tandis que le rein et le coeur sont peu ou pas affectés. Des études plus approfondies sur l’expression des MR dans ces tissus nous aideront à mieux comprendre l’implication des 11β-HSD au fil de la gestation. / The activation of the renin-angiotensin-aldosterone system can lead to hypertension and cardiac fibrosis. However, despite a substantial elevation of aldosterone during pregnancy, those adverse effects are not observed. Aldosterone acts via the mineralocorticoid receptors (MR) which can also bind cortisol with a similar affinity. Regulation of the local levels of this glucocorticoid by the 11β-hydroxysteroid dehydrogenases (11β-HSD) is thus crucial to avoid overstimulation of MRs. We believe that these enzymes are involved in the maternal and fetal protections against the high levels of aldosterone and cortisol observed during pregnancy. We propose that the adaptative mechanisms occurring during normal pregnancy involve tissue-specific changes in the expression (mRNA and protein) and activity of both 11β-HSDs. Pregnant Sprague-Dawley rats were sacrified on day 14, 17, 19 or 22 of gestation (term = day 23) and their organs were collected. In the kidney, our results have shown that 11β-HSDs protein levels decrease in late gestation. In the placenta, a dramatic decrease of 11β-HSD1 mRNA and protein expressions is observed on day 17 while 11β-HSD2 levels are increased. Expression and activity of the 11β-HSD2 are then decreased up to day 22. No significant differences were detected in the left cardiac ventricle. In conclusion, our results demonstrate that gestation is associated with important modifications in the placenta, possibly to ensure a normal fetal growth, while expression in the kidney and the heart is barely changed. More studies on MR expression in those tissues will be required to better characterise the function of the 11β-HSDs throughout pregnancy.
45

Vliv stresu na expresi 11β-hydroxysteroiddehydrogenasy v mozku laboratorního potkana / Effect of stress on expression of 11β-hydroxysteroid dehydrogenase in rat brain

Kuželová, Andrea January 2013 (has links)
This thesis examines the influence of stress on the activity of hippocampal CA1 area. The main task was to determine whether the stress load affects the changes of the local metabolism of glucocorticoids, and whether the levels of corticosteroid receptors in the CA1 hippocampus are modulated in response to stress. In order to answer these questions, the experiments were carried out using three different rat strains - Fisher, Lewis and Wistar which differ in their activities of hypothalamic-pituitary-adrenal axis. Our results demonstrate that stress has no effect on expression of MR mRNA. Conversely, stress reduces the levels of GR mRNA in CA1 area of the dorsal hippocampus. Moreover, we confirmed that the Lewis and Wistar rats didn't change metabolism of glucocorticoids after stress response. By the Fisher rats increased levels of 11β-HSD1 mRNA expression and therefore increased the metabolism of corticosterone.
46

Vztah metabolismu kortikosteroidů a ontogeneze ke stresové odpovědi / Relationship between corticosteroid metabolism, ontogenesis and stress response

Makal, Jakub January 2013 (has links)
Stress is a widespread phenomenon in the western society of these days. It is a risky factor for health and well-being of the majority of people. Based on these facts, it is the main subject for the field of "stress physiology" research, which aims to study processes occurring during stress response and tries to elucidate mechanisms leading to stress-induced health impairment. The first aim of this thesis was to describe effects of psycho-social stress on organism. The second aim was to find out if can stress applied in juvenile age affect the stress response in adulthood. If so, how is the role of glucocorticoid-metabolism enzyme 11β-HSD1 in this influence? To answer these questions, two different animal models inducing stress response in the laboratory rat were used. The first one is the model of mild social stress based on the resident-intruder paradigm. Our results show efficancy of this model. Fisher 344 male rats treated under this model for seven consecutive days show highly elevated plasma corticosterone concentrations and elevated expression of the glucocorticoid receptor gene in the pituitary. Behavioral analysis demonstrates a decreased social behavioral profile of the intruders, suggesting submisive social position of these animals in the resident-intruder paradigm. The second model used is...
47

Análise de fatores genéticos associados ao desenvolvimento da síndrome metabólica durante a terapia com glicocorticoide em pacientes portadores da deficiência da 21-hidroxilase / Analysis of genetic factors associated with the development of the metabolic syndrome during therapy with glucocorticoids in patients with 21hydroxylase deficiency

Moreira, Ricardo Paranhos Pires 05 June 2014 (has links)
Introdução: A deficiência da 21-hidroxilase (21-OHD) é um frequente erro herdado do metabolismo que resulta no comprometimento da síntese do cortisol e/ou aldosterona e aumento da produção de andrógenos. A doença é caracterizada por uma diversidade fenotípica, variando desde virilização pré-natal da genitália externa de fetos femininos e pós-natal em ambos os sexos, com ou sem perda de sal, até quadros assintomáticos. Em seu tratamento é necessária reposição com glicocorticoide para se evitar a insuficiência adrenocortical e os sinais de virilização. Um fino ajuste na dose diária do glicocorticoide é essencial para se evitar sub ou supertratamento, com o objetivo de preservar o potencial de estatura final e fertilidade. Entretanto, tem sido observada maior frequência de obesidade e outras comorbidades metabólicas nestes pacientes; porém, a prevalência destas complicações ainda não é conhecida, bem como se estariam associadas à exposição ao glicocorticoide e/ou com fatores genéticos. Objetivos: avaliar a frequência de obesidade e de síndrome metabólica (SM) em pacientes com 21OHD; caracterizar a distribuição alélica dos polimorfismos dos genes do receptor de glicocorticoide (NR3C1) e da enzima 11beta-hidroxiesteróide desidrogenase tipo I (HSD11B1), e correlacionar a distribuição destes polimorfismos com a presença das complicações metabólicas. Métodos: Foram selecionados 109 pacientes (60 PS/49 VS), sendo 41 crianças e adolescentes (idade média 11,4 ± 3,9 anos) e 68 adultos (idade média 28,4 ± 9 anos) em tratamento com glicocorticoide e com adequado controle hormonal. Pacientes com a forma PS também receberam fludrocortisona. Adequado controle foi caracterizado por concentração normal de atividade plasmática de renina e de andrógenos de acordo com o sexo e idade nos últimos 2 anos. A obesidade nos adultos foi definida pelo IMC >= 30 kg/m² e em crianças e adolescentes pelo IMC acima do percentil 95. Síndrome metabólica foi definida segundo o critério do National Cholesterol Education Program em adultos e crianças. História familiar de hipertensão arterial, diabetes, dislipidemia, obesidade e/ou doença cardiovascular também foi avaliada. Foram mensuradas glicemia, lipoproteínas, triglicérides, colesterol total e insulina. Os alelos BclI, A3669G, ER22/23EK e N363S do gene NR3C1 e o alelo 4436InsA do gene HSD11B1 foram genotipados e as análises de associação com os fenótipos foram realizadas por meio dos testes Chi-quadrado, t-studant e análise de regressão. As análises de correlação foram feitas utilizando o teste de correlação de Pearson. Resultados: Obesidade foi observada em 31,7% das crianças e 23,5% dos adultos. Síndrome metabólica foi observada em 14,6% das crianças e 7,3% dos adultos. A prevalência dos componentes da SM foi maior no grupo dos obesos quando comparada a de pacientes não obesos (crianças e adultos). Não houve correlação significante entre o IMC, sexo, forma clínica da 21-OHD, duração da terapia e dose de GC. História familiar positiva para obesidade, hipertensão, dislipidemia e doença cardiovascular foi mais frequente nos pacientes obesos quando comparada a de pacientes não obesos, em adultos e crianças. Os polimorfismos BclI, A3669G e 4436InsA foram identificados em 23,2%, 9,7% e 14,6% dos alelos das crianças, respectivamente, e nos adultos em 26,4%, 9,6% e 18,4% dos alelos, respectivamente. A variante A3669G foi associada à maiores concentrações de LDL-c em crianças quando comparada aos carreadores do alelo selvagem. Os pacientes adultos carreadores do polimorfismo BclI apresentaram maior IMC, circunferência abdominal e PAS quando comparados aos carreadores do alelo selvagem. Não observamos diferenças estatisticamente significantes no perfil metabólico entre pacientes carreadores e não carreadores do polimorfismo 4436InsA (adultos e crianças). Conclusão: observamos que pacientes 21-OHD possuem maior prevalência de obesidade, e o grupo pediátrico maior prevalência de SM em relação à população de referência, sendo ambas independentes da dose de glicocorticoide e do tempo do tratamento. A presença de perfil metabólico adverso esteve associada à obesidade e à predisposição genética, tais como história familiar e variantes genéticas do receptor de glicocorticoide / Introduction: Congenital adrenal hyperplasia due to 21-hydroxylase deficiency (21OHD) is a common autosomal recessive disorder that leads to decreased glucocorticoid secretion, with or without mineralocorticoid deficiency, and increased androgen production. The disease is characterized by phenotypic variability, including a severe form with prenatal virilization of the external genitalia in female fetuses and postnatal virilization in both sexes, with or without salt loss. Current therapy aims to provide adequate glucocorticoid (GC) replacement and to suppress the abnormal androgen secretion; mineralocorticoid replacement aims to control the renal salt balance to avoid adrenal crisis. Nevertheless, these therapeutic goals are difficult to achieve in practice due to the complexity of replicating the physiologic cortisol circadian rhythm. Increased prevalence of obesity, insulin resistance, hypertension and adverse lipid profile have been observed among CAH patients under GC therapy; however, the extent of its prevalence and also whether it is associated with the GC dose or with genetic factors are not known. Objectives: to evaluate the obesity and metabolic syndrome (SM) frequencies in 21-OHD patients; to characterize the allelic distribution of the NR3C1 and HSD11B1 polymorphisms, and to correlate with the metabolic profile. Methods: One hundred and nine patients (60SW/49SV) were selected, 41 being children and adolescents (mean age 11.4 ± 3.9 yrs) and 68 adults (mean age 28.4 ± 9 yrs) all of whom received GC treatment and had adequate hormonal control. SW patients also received fludrocortisone. Adequate hormonal control was characterized by normal plasmatic rennin activity and androgen levels according to age and sex for at least two years. Blood fasting was used to obtain glucose, lipoproteins, triglycerides, total cholesterol and insulin levels. Obesity in the adult group was defined by BMI >= 30 kg/m², and in the young group by BMI > 95th percentile. Metabolic syndrome was defined by the NCEP ATPIII criteria. Family history of the hypertension, diabetes, dyslipidemia, obesity and/or cardiovascular disease was also evaluated. The BclI, A3669G, ER22/23EK and N363S alleles of the NR3C1 gene and 4436InsA of the HSD11B1 gene were genotyped and association analyses with phenotype were carried out with Chi-square, t-test and regression analysis. Correlation analyses were performed by Pearson correlation test. Results: obesity was observed in 31.7% of children and 23.5% of adults. SM was observed in 14.6% of young and 7.3% of adult patients. SM prevalence was higher in the obese group than the nonobese group (children and adults). There was no significant correlation between GC dose and BMI, sex, clinical form or treatment duration. Prevalence of family history of obesity, hypertension, dyslipidemia and cardiovascular disease was higher in the obese than in non-obese patients (children and adults). The BclI, A3669G and 4436InsA polymorphisms were found in 23.2%, 9.7% and 14.6% of the alleles in children, respectively and in 26.4%, 9.6% and 18.4% of the alleles in adults. The A3669G variant was associated to increased LDL-c levels in comparison with noncarriers in the young group. The BclI adult carriers presented higher BMI, abdominal circumference and systolic blood pressure in comparison with noncarriers. Statistically significant differences were not observed in the metabolic profile between carriers and non-carriers of the 4436InsA polymorphism (children and adults). Conclusion: in the present study, which analyzed the clinical and metabolic profile of 21-OHD patients, high obesity prevalence, independent of GC dose and treatment duration, was observed. Adverse metabolic profile was mainly associated with obesity and genetic predisposition, such as family history and NR3C1 polymorphisms
48

Variation de l’expression et de l’activité des 11β-hydroxystéroïde déshydrogénases rénales, cardiaques et placentaires au cours de la gestation de la rate

Barrette, Mathieu 08 1900 (has links)
L’activation du système rénine-angiotensine-aldostérone peut entraîner le développement d’une hypertension artérielle et de la fibrose cardiaque. Toutefois, au cours de la grossesse, malgré une hausse substantielle des niveaux d’aldostérone, ces effets délétères ne sont pas observés. L’aldostérone exerce ses effets via les récepteurs des minéralocorticoïdes, les MR, qui peuvent également lier le cortisol avec une affinité similaire. La régulation des niveaux locaux de ce glucocorticoïde par les 11β-hydroxystéroïde déshydrogénases (11β-HSD) est donc essentielle pour éviter une stimulation inappropriée des MR. Nous suggérons que, durant la grossesse, ces enzymes sont impliquées dans la protection de la mère et du foetus contre les niveaux élevés d’aldostérone et de cortisol. Notre hypothèse de travail est que les mécanismes d’adaptation qui prennent place au cours de la grossesse nécessitent des changements d’expression (ARNm et protéine) et d’activité des 11β-HSD spécifiques selon le tissu. Des rates Sprague-Dawley ont été sacrifiées aux jours 14, 17, 19 et 22 de gestation (terme = jour 23) et leurs organes ont été collectés. Dans le rein, les niveaux protéiques des 11β-HSD sont diminués en fin de gestation. Dans le placenta, on observe une importante chute de l’expression génique et protéique de la 11β-HSD1 au jour 17 tandis que la 11β-HSD2 y est augmentée. L’expression et l’activité de la 11β-HSD2 sont par la suite diminuées jusqu’à terme. Aucune différence significative n’est retrouvée dans le ventricule gauche cardiaque. En conclusion, nos résultats démontrent que la gestation est accompagnée d’importants changements dans le placenta, possiblement pour assurer un développement foetal adéquat, tandis que le rein et le coeur sont peu ou pas affectés. Des études plus approfondies sur l’expression des MR dans ces tissus nous aideront à mieux comprendre l’implication des 11β-HSD au fil de la gestation. / The activation of the renin-angiotensin-aldosterone system can lead to hypertension and cardiac fibrosis. However, despite a substantial elevation of aldosterone during pregnancy, those adverse effects are not observed. Aldosterone acts via the mineralocorticoid receptors (MR) which can also bind cortisol with a similar affinity. Regulation of the local levels of this glucocorticoid by the 11β-hydroxysteroid dehydrogenases (11β-HSD) is thus crucial to avoid overstimulation of MRs. We believe that these enzymes are involved in the maternal and fetal protections against the high levels of aldosterone and cortisol observed during pregnancy. We propose that the adaptative mechanisms occurring during normal pregnancy involve tissue-specific changes in the expression (mRNA and protein) and activity of both 11β-HSDs. Pregnant Sprague-Dawley rats were sacrified on day 14, 17, 19 or 22 of gestation (term = day 23) and their organs were collected. In the kidney, our results have shown that 11β-HSDs protein levels decrease in late gestation. In the placenta, a dramatic decrease of 11β-HSD1 mRNA and protein expressions is observed on day 17 while 11β-HSD2 levels are increased. Expression and activity of the 11β-HSD2 are then decreased up to day 22. No significant differences were detected in the left cardiac ventricle. In conclusion, our results demonstrate that gestation is associated with important modifications in the placenta, possibly to ensure a normal fetal growth, while expression in the kidney and the heart is barely changed. More studies on MR expression in those tissues will be required to better characterise the function of the 11β-HSDs throughout pregnancy.
49

Análise de fatores genéticos associados ao desenvolvimento da síndrome metabólica durante a terapia com glicocorticoide em pacientes portadores da deficiência da 21-hidroxilase / Analysis of genetic factors associated with the development of the metabolic syndrome during therapy with glucocorticoids in patients with 21hydroxylase deficiency

Ricardo Paranhos Pires Moreira 05 June 2014 (has links)
Introdução: A deficiência da 21-hidroxilase (21-OHD) é um frequente erro herdado do metabolismo que resulta no comprometimento da síntese do cortisol e/ou aldosterona e aumento da produção de andrógenos. A doença é caracterizada por uma diversidade fenotípica, variando desde virilização pré-natal da genitália externa de fetos femininos e pós-natal em ambos os sexos, com ou sem perda de sal, até quadros assintomáticos. Em seu tratamento é necessária reposição com glicocorticoide para se evitar a insuficiência adrenocortical e os sinais de virilização. Um fino ajuste na dose diária do glicocorticoide é essencial para se evitar sub ou supertratamento, com o objetivo de preservar o potencial de estatura final e fertilidade. Entretanto, tem sido observada maior frequência de obesidade e outras comorbidades metabólicas nestes pacientes; porém, a prevalência destas complicações ainda não é conhecida, bem como se estariam associadas à exposição ao glicocorticoide e/ou com fatores genéticos. Objetivos: avaliar a frequência de obesidade e de síndrome metabólica (SM) em pacientes com 21OHD; caracterizar a distribuição alélica dos polimorfismos dos genes do receptor de glicocorticoide (NR3C1) e da enzima 11beta-hidroxiesteróide desidrogenase tipo I (HSD11B1), e correlacionar a distribuição destes polimorfismos com a presença das complicações metabólicas. Métodos: Foram selecionados 109 pacientes (60 PS/49 VS), sendo 41 crianças e adolescentes (idade média 11,4 ± 3,9 anos) e 68 adultos (idade média 28,4 ± 9 anos) em tratamento com glicocorticoide e com adequado controle hormonal. Pacientes com a forma PS também receberam fludrocortisona. Adequado controle foi caracterizado por concentração normal de atividade plasmática de renina e de andrógenos de acordo com o sexo e idade nos últimos 2 anos. A obesidade nos adultos foi definida pelo IMC >= 30 kg/m² e em crianças e adolescentes pelo IMC acima do percentil 95. Síndrome metabólica foi definida segundo o critério do National Cholesterol Education Program em adultos e crianças. História familiar de hipertensão arterial, diabetes, dislipidemia, obesidade e/ou doença cardiovascular também foi avaliada. Foram mensuradas glicemia, lipoproteínas, triglicérides, colesterol total e insulina. Os alelos BclI, A3669G, ER22/23EK e N363S do gene NR3C1 e o alelo 4436InsA do gene HSD11B1 foram genotipados e as análises de associação com os fenótipos foram realizadas por meio dos testes Chi-quadrado, t-studant e análise de regressão. As análises de correlação foram feitas utilizando o teste de correlação de Pearson. Resultados: Obesidade foi observada em 31,7% das crianças e 23,5% dos adultos. Síndrome metabólica foi observada em 14,6% das crianças e 7,3% dos adultos. A prevalência dos componentes da SM foi maior no grupo dos obesos quando comparada a de pacientes não obesos (crianças e adultos). Não houve correlação significante entre o IMC, sexo, forma clínica da 21-OHD, duração da terapia e dose de GC. História familiar positiva para obesidade, hipertensão, dislipidemia e doença cardiovascular foi mais frequente nos pacientes obesos quando comparada a de pacientes não obesos, em adultos e crianças. Os polimorfismos BclI, A3669G e 4436InsA foram identificados em 23,2%, 9,7% e 14,6% dos alelos das crianças, respectivamente, e nos adultos em 26,4%, 9,6% e 18,4% dos alelos, respectivamente. A variante A3669G foi associada à maiores concentrações de LDL-c em crianças quando comparada aos carreadores do alelo selvagem. Os pacientes adultos carreadores do polimorfismo BclI apresentaram maior IMC, circunferência abdominal e PAS quando comparados aos carreadores do alelo selvagem. Não observamos diferenças estatisticamente significantes no perfil metabólico entre pacientes carreadores e não carreadores do polimorfismo 4436InsA (adultos e crianças). Conclusão: observamos que pacientes 21-OHD possuem maior prevalência de obesidade, e o grupo pediátrico maior prevalência de SM em relação à população de referência, sendo ambas independentes da dose de glicocorticoide e do tempo do tratamento. A presença de perfil metabólico adverso esteve associada à obesidade e à predisposição genética, tais como história familiar e variantes genéticas do receptor de glicocorticoide / Introduction: Congenital adrenal hyperplasia due to 21-hydroxylase deficiency (21OHD) is a common autosomal recessive disorder that leads to decreased glucocorticoid secretion, with or without mineralocorticoid deficiency, and increased androgen production. The disease is characterized by phenotypic variability, including a severe form with prenatal virilization of the external genitalia in female fetuses and postnatal virilization in both sexes, with or without salt loss. Current therapy aims to provide adequate glucocorticoid (GC) replacement and to suppress the abnormal androgen secretion; mineralocorticoid replacement aims to control the renal salt balance to avoid adrenal crisis. Nevertheless, these therapeutic goals are difficult to achieve in practice due to the complexity of replicating the physiologic cortisol circadian rhythm. Increased prevalence of obesity, insulin resistance, hypertension and adverse lipid profile have been observed among CAH patients under GC therapy; however, the extent of its prevalence and also whether it is associated with the GC dose or with genetic factors are not known. Objectives: to evaluate the obesity and metabolic syndrome (SM) frequencies in 21-OHD patients; to characterize the allelic distribution of the NR3C1 and HSD11B1 polymorphisms, and to correlate with the metabolic profile. Methods: One hundred and nine patients (60SW/49SV) were selected, 41 being children and adolescents (mean age 11.4 ± 3.9 yrs) and 68 adults (mean age 28.4 ± 9 yrs) all of whom received GC treatment and had adequate hormonal control. SW patients also received fludrocortisone. Adequate hormonal control was characterized by normal plasmatic rennin activity and androgen levels according to age and sex for at least two years. Blood fasting was used to obtain glucose, lipoproteins, triglycerides, total cholesterol and insulin levels. Obesity in the adult group was defined by BMI >= 30 kg/m², and in the young group by BMI > 95th percentile. Metabolic syndrome was defined by the NCEP ATPIII criteria. Family history of the hypertension, diabetes, dyslipidemia, obesity and/or cardiovascular disease was also evaluated. The BclI, A3669G, ER22/23EK and N363S alleles of the NR3C1 gene and 4436InsA of the HSD11B1 gene were genotyped and association analyses with phenotype were carried out with Chi-square, t-test and regression analysis. Correlation analyses were performed by Pearson correlation test. Results: obesity was observed in 31.7% of children and 23.5% of adults. SM was observed in 14.6% of young and 7.3% of adult patients. SM prevalence was higher in the obese group than the nonobese group (children and adults). There was no significant correlation between GC dose and BMI, sex, clinical form or treatment duration. Prevalence of family history of obesity, hypertension, dyslipidemia and cardiovascular disease was higher in the obese than in non-obese patients (children and adults). The BclI, A3669G and 4436InsA polymorphisms were found in 23.2%, 9.7% and 14.6% of the alleles in children, respectively and in 26.4%, 9.6% and 18.4% of the alleles in adults. The A3669G variant was associated to increased LDL-c levels in comparison with noncarriers in the young group. The BclI adult carriers presented higher BMI, abdominal circumference and systolic blood pressure in comparison with noncarriers. Statistically significant differences were not observed in the metabolic profile between carriers and non-carriers of the 4436InsA polymorphism (children and adults). Conclusion: in the present study, which analyzed the clinical and metabolic profile of 21-OHD patients, high obesity prevalence, independent of GC dose and treatment duration, was observed. Adverse metabolic profile was mainly associated with obesity and genetic predisposition, such as family history and NR3C1 polymorphisms
50

Regulation of the 11beta-hydroxysteroid dehydrogenase type 2 promoter by steroid hormones in breast cancer cells. Convergence of progesterone receptor binding to DNA and JAK/START pathway activation

Subtil Rodriguez, Alicia 27 June 2007 (has links)
El gen humano 11-HSD2 es un modelo para investigar la contribución de los efectos de los receptores de esteroides en células de cáncer de mama. El análisis del promotor mostró que la región distal está implicada en la mayor parte de la activación dependiente de hormona. En respuesta a hormona, STAT5A se recluta a la región distal y PR a las regiones distal y proximal del promotor. El reclutamiento de PR se debe a dos mecanismos diferentes, la unión directa de PR a la región proximal, y la implicación vía JAK/STAT en el reclutamiento a la región distal. La inducción del gen 11-HSD2 por hormonas disminuye parcialmente por inhibidores de MAPK y PI3K/Akt y totalmente por inhibidores de JAK/STAT. Así, los efectos citoplasmáticos del PR están implicados en la inducción del gen progesterona. La forma activa de la ARN-polimerasa II es reclutada por la inducción con hormonas a la región distal del promotor 11-HSD2 y la región distal tiene respuesta a hormonas por sí misma, indicando que la inducción del gen por hormonas empieza antes del sitio de inicio de transcripción descrito previamente. / The human 11-HSD2 gene is a model to investigate the contribution of steroid hormone receptors effects on a progesterone responsive promoter in breast cancer cells. Deletion analysis of the 11-HSD2 promoter showed that the distal region is involved in most of the hormone-dependent activation. ChIP showed hormone-dependent STAT5A-recruitment to the distal region and PR-recruitment to the distal and proximal promoter regions. Results suggest two different mechanisms of hormone-induced PR-recruitment, since cells stably expressing PR containing a mutated DNA-binding domain have affected hormone-dependent PR-recruitment to proximal promoter, and JAK/STAT pathway inhibition blocks PR-recruitment to distal promoter. Hormone-stimulated 11-HSD2 gene-expression was partially decreased by MAPK and PI3K/AKT pathway inhibitors and totally blocked by JAK/STAT pathways inhibitors, indicating that cytoplasmic PR effects involvement in progestin-induced 11-HSD2 expression. Importantly, upon hormone induction active RNA-polymerase II is recruited from the 11-HSD2 distal promoter region and the distal minimal promoter has hormone-responsiveness by itself, suggesting that progesterone-dependent 11-HSD2 expression starts upstream the previously characterized transcription start site.

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