Thesis (Ph. D.)--University of Virginia, 2007. / Includes bibliographical references. Also available online through Digital Dissertations.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 5 uppsatser.

Tez (Uzmanlık) - Süleyman Demirel Üniversitesi, Tıp Fakültesi, İç Hastalıkları Anabilim Dalı, 2003. / Kaynakça var.

Diss. (sammanfattning) Umeå : Univ., 2007. / Härtill 5 uppsatser.

Thesis (PH. D.)--Michigan State University. Dept. of Psychology, 2008. / Title from PDF t.p. (viewed on Sept. 8, 2009) Includes bibliographical references (p. 157-178). Also issued in print.

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L’hypertrophie bénigne de la prostate (HBP) est une pathologie qui affecte 50% des hommes dès l’âge de 60 ans et qui conduit à des troubles de la miction. L’HBP se caractérise par une hypertrophie exclusive ou composite de plusieurs compartiments tissulaires de la prostate que sont l’épithélium, le stroma et les fibres musculaires qui définissent respectivement les composantes glandulaire, fibreuse et musculaire de cette pathologie. Il a récemment été montré que les souris dépourvues en récepteurs nucléaires LXR (Liver‐X‐receptor) α (souris lxrα‐/‐) développent une hypertrophie de la prostate dont les signes histologiques évoquent une HBP de type fibreuse. Par ailleurs, un des traitements de l’HBP, vise à éteindre la signalisation androgénique en inhibant la conversion de la testostérone en son métabolite actif, la dihydrotestostérone (DHT). Le phénotype d’hypertrophie de la prostate pourrait donc également s’expliquer par une altération de la signalisation androgénique dans les souris lxrα‐/‐. Dans ce contexte, notre projet de recherche a été centré sur l’étude du rôle des LXR dans l’apparition de l’HBP dans sa composante glandulaire et l’analyse des relations moléculaires associant les signalisations dépendantes de LXRα et du récepteur des androgènes (AR) au sein de la prostate. Le phénotype d’HBP observé dans les souris lxrα‐/‐ résulte d’altérations importantes de l’homéostasie de l’épithélium qui miment la composante glandulaire : 1) une activité sécrétoire accrue ; 2) une altération des processus de sécrétion associée à une altération de l’expression des gènes codant des protéines du transport vésiculaire ; 3) une réponse altérée de certains gènes androgéno‐dépendants associée à une hypersensibilité aux androgènes ; 4) des modifications du réseau paracrine reliant le stroma et l’épithélium. Au final, ces travaux définissent LXRα comme un acteur clé de l’homéostasie prostatique et ouvrent des pistes intéressantes pour la compréhension de l’étiologie de l’HBP chez l’homme. Ces résultats montrent qu’il est possible de moduler la réponse androgénique de la prostate en ciblant LXRα. Ainsi, à plus long terme, l’activation pharmacologique de LXRα constitue une piste potentielle dans le traitement de l’HBP. / Benign prostate hyperplasia (BPH) is a very common prostatic disorder that affects 50% of men after 60 years. In BPH, prostate enlargement causes urinary disorders. BPH is characterized by a hypertrophy of several tissue compartments such as the epithelium, stroma and/or muscle fibers. Hence, three main forms of BPH have been described : glandular, fibrous and muscular form. It has been recently shown that LXR (Liver‐X‐receptor) α (lxrα‐/‐) mice develop a prostate enlargement with histological signs of fibrous BPH. Inhibition of testosterone conversion into DHT is one the most effective pharmacological treatment of BPH. Thus, the lxrα‐/‐ prostate phenotype could be in part due to an alteration of androgen signaling. In this context, the aim of this work was to study the role of LXR in glandular BPH development and to understand the relationships between LXRα and the androgen receptor (AR) dependent signaling pathway in prostate. The prostate enlargement observed in lxrα‐/‐ mice results from major alterations in epithelium homeostasis mimicking the glandular alteration of BPH : 1) increase of secretory activity ; 2) alteration of the secretory process associated with altered expression of vesicular transport protein encoding genes ; 3) a disruption in the response of androgen‐dependent genes associated with androgen hypersensitivity ; 4) changes in the paracrine network between stroma and epithelium. Finally, this work defines LXRα as a key player in prostate homeostasis and opens interesting way to the understanding of BPH etiology. These results show that targeting LXRα modulate the prostate androgenic response. Thus, pharmacological activation of LXRα could constitute a new option for the treatment of BPH.

Androgens are well documented as important regulators of male health, primarily in the maintenance and development of male sexual characteristics. However, a decline in circulating androgens has also been associated with co-morbidities such as obesity, cardiac disease and metabolic syndrome. Previous research has focussed upon the body wide impact of adrenal androgens, however whilst androgen receptor (AR) is abundantly expressed in the adrenal cortex of both rodents and humans, surprisingly little is known about androgen action on the adrenal cortex itself. This gap in our understanding is at least in part due to the perceived lack of suitable animal models. Rodents have largely been overlooked as a model system as their adrenals are unable to produce androgens due to lack of 17α Hydroxylase and 17, 20 lyse activity and they therefore do not have a zona reticularis. However, historical studies using castrated mice showed that removal of androgens leads to the redevelopment of an additional cortex zone known as the transient X-zone. The foetal adrenal is thought to give rise the adult adrenal cortex in human and rodents. These foetal cells are maintained for a period postnatally and regress differently depending on species and sex. In the human this zone is known as the ‘foetal zone’, and the rodent homologue termed the ‘X-zone’. The mechanisms underpinning the regression of the X-zone and its purpose and maintenance postnatally still aren’t clearly understood. To provide a comprehensive overview of androgen signalling in the adrenal cortex, multiple mouse models were utilised. First, Cre/loxP technology was used to ablate AR specifically from the adrenal cortex. Further androgen manipulation was achieved through castration (removal of androgens) and human chorionic gonadotropin (hCG) treatment (increased androgens). The initial study investigates the impacts on the male mouse adrenal. Histology analysis revealed the presence of an X-zone in all experimental cohorts following loss of AR or circulating androgens, confirmed by 20- α-hydroxysteroid dehydrogenase (20 alpha-HSD) expression. These data demonstrate that androgens signalling via AR is required for X-zone regression during puberty. However, interrogation of morphology of hCG treated cohorts revealed no phenotypic changes compared to controls, this demonstrates that hyper stimulation with androgens does not negatively impact the adrenal cortex or influence X-zone morphology. Differences in X-zone morphology and 20 alpha-HSD localization prompted cortex measurements which revealed significant differences in X-zone depth and cell density depending on ablation of AR, circulating androgens or both. This suggests that androgens and androgen receptor are working together and also independently to regulate the adrenal cortex. This result was strengthened through analysis of steroid enzyme genes and cortex markers, which revealed that normal AKR1B7 expression was absent following loss of androgens but not androgen receptor. A final part of this study examined the impacts long term androgen receptor ablation and long term castration in ageing animals. A final part of this study examined the impacts long term androgen receptor ablation and long term castration in ageing animals. These results demonstrate that following prolonged loss of androgens that there is no major disruption to the adrenal cortex. Morphology analysis and X-zone measurements revealed that X-zone regression was occurring in mice with long term castration, characterized by a reduction in size and pockets of vacuolization throughout the X-zone. This phenotype is also observed in ageing females with X-zone regression via vacuolization. These data suggest that following prolonged loss of androgens, the male adrenal is feminized and behaves as such. In contrast, AR ablation only, results in an enlarged adrenal with large spindle cell lesions and X-zone expansion confirmed by X-zone measurements. Initial experiments have demonstrated that androgens can work independently of AR to regulate the adrenal cortex. Together these data suggests that AR is required to control the appropriate action of circulating androgens in the adrenal cortex, with loss of AR resulting in off target signalling from circulating androgens in the adrenal leading to spindle cell hyperplasia, X-zone expansion and X-zone mislocation. A second set of studies were carried out to determine the role of androgen signalling in the female adrenal, specifically, if loss of AR leads to the absence of normal X-zone regression during pregnancy. To answer this question the same selective AR ablation model was used. Analysis of litters comparing observed and expected genetic distribution revealed significantly fewer females being born carrying complete ablation of adrenal AR. Morphology analysis of these mice revealed severe cortex disruption and spindle cell hyperplasia similar to that observed in mutant males. Investigation of adrenals following pregnancy revealed that X-zone regression still occurred despite loss of AR. This result shows that X-zone regression in the female is under different regulation compared to male adrenal and occurs via an androgen-independent signalling mechanism. However, loss of AR still leads to anatomical dysregulation of the adrenal cortex. AR ablation revealed changes in glucocorticoid receptor (GR) expression in the adrenal cortex. To dissect this relationship further a final study was conducted, attempting to ablate GR from the adrenal cortex also using the Cyp11a1 Cre. Initial observations of these mice revealed excessive hair loss through barbering, curved spines and stressed behaviour when monitored in the cage under normal conditions. Immunohistochemistry was used to confirm GR ablation in the adrenal cortex, however, to our surprise, GR expressing cells were not steroidogenic and thus were not targeted by the Cre recombinase. Despite no GR ablation in the adrenal, morphology analysis revealed severe disruption to the adrenal cortex. The Cyp11a1 Cre not only targets the adrenal but is expressed in the hindbrain. To determine if GR ablation in the hindbrain explains the phenotype, we next used PCR analysis interrogating hindbrain genomic DNA to determine if there was recombination of GR. Results confirmed GR recombination in the hindbrain. Due to the observation of stressed behaviour and adrenal cortex disruption, we wanted to determine if this was a result of hyperactivity of the adrenal cortex. Serum corticosterone was analysed and was elevated in these animals. These data revealed that GR ablation in the hindbrain results in adrenal cortex disruption and an elevated stress response, potentially highlighting a new model to investigate stress disorders and their impact on the hypothalamic-pituitary-adrenal axis. Together this data defines new roles for AR signalling in the adrenal cortex and the role of the hindbrain GR signalling in regulating adrenal morphology and function.

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Orientador: Luis Carlos Spolidorio / Banca: Ticiana Sidorenko de Oliveira Capone / Banca: Thallita Pereira Queiroz / Resumo: Os mecanismos de ação dos andrógenos sobre homeostase e regulação das células que participam do turnover ósseo em fêmeas ainda são pouco compreendidos. Este trabalho teve como principal objetivo avaliar a participação de andrógenos na diferenciação e atividade de osteoclastos in vitro. Células totais de medula óssea de fêmur e tíbia de camundongos fêmeas foram utilizadas como fonte de células precursoras de osteoclastos, sendo cultivadas utilizando-se α-MEM suplementado e em presença de RANK-L (30ng/mL) e M-CSF (50ng/mL). As células foram tratadas com testosterona (T) diidrotestosterona (DHT) e antagonistas de receptores de hormônios sexuais, como flutamida (FLU) e fulvestranto (FUL). O anastrozol (ANA) foi usado para inibição da enzima aromatase e o etanol (0,01%) foi utilizado como controle. Após cinco dias, as células foram fixadas, coradas com TRAP e contadas, considerando-se células TRAP-positivas com 3 ou mais núcleos. Para o ensaio de atividade, foram utilizadas placas revestidas com fosfato de cálcio inorgânico e a área de reabsorção foi calculada com o auxílio de software. O estágio de diferenciação osteoclástica foi avaliado por RT-qPCR e a modulação da expressão de receptores para hormônios sexuais foi avaliada por Western Blot. Os andrógenos (T e DHT) não exerceram efeitos sobre a diferenciação e atividade de osteoclastos (ANOVA; p>0,05). Por outro lado, os tratamentos com ANA, FLU e FUL, associados ou não a T, regularam positivamente a diferenciação e atividade d... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The action mechanisms of androgens on homeostasis and the regulation of cells that participate in bone turnover in females are still poorly understood. This study had as main objective to evaluate the participation of androgens in the differentiation and activity of in vitro osteoclasts. Total bone marrow cells from femur and tibia of female mice were used as a source of precursor cells of osteoclasts, they were cultivated using supplemented α-MEM and in the presence of RANK-L (30ng/mL) and M-CSF (50ng/mL). The cells were treated with testosterone (T), dihydrotestosterone (DHT) and antagonists of sexual hormone receptors such as flutamide (FLU) and fulvestrant (FUL). Anastrozole (ANA) was used for inhibiting the aromatase enzyme and ethanol (0.01%) was used as a control. After five days, the cells were fixed, colored with TRAP and counted, considering TRAP-positive cells those ones containing 3 or more nuclei. For the activity assay, were used plaques covered with inorganic calcium phosphate and the area of reabsorption was calculated with the assistance of a software. The osteoclast differentiation stage was evaluated by RT-qPCR and the modulation of the expression of receptors for sexual hormones was assessed by Western Blotting. The androgens (T and DHT) did not exert effects in differentiation and activity of osteoclasts (ANOVA, p> 0.05). On the other hand, the treatments with ANA, FLU and FUL, associated or not to T, positively regulated the differentiation and activity ... (Complete abstract click electronic access below) / Mestre