Unravelling The Role Of Androgens In Polycystic Ovary Syndrome

Abstract

Polycystic Ovary Syndrome (PCOS) is a multifaceted hormonal disorder which affects 5-15% of reproductive-aged women worldwide. While classically recognised as an ovarian disorder, PCOS is associated with a variety of reproductive, endocrine and metabolic features including ovulatory dysfunction, infertility, hyperandrogenism, obesity, and an increased risk of type 2 diabetes mellitus and cardiovascular disease. The most consistently present of these is hyperandrogenism – supraphysiological levels of androgens such as testosterone (T) and dihydrotestosterone (DHT). Typically thought of as male steroid hormones, androgens have been shown to play important role in the maintenance of normal female reproductive function. Despite the high prevalence of hyperandrogenism amongst patients, the role of androgens in the etiology and pathogenesis of PCOS has yet to be determined. The aim of this work was to unravel the association between excess androgen exposure and the development and advancement of the PCOS phenotype in mouse model of androgen-induced PCOS. The first study contained within this work (Chapter Three) provides the first comprehensive characterization of a range of reproductive, endocrine and metabolic traits associated with PCOS in four distinct mouse models: a model of prenatal androgenisation utilising the potent non-aromatizable androgen DHT administered during days 16-18 of gestation, and three diverse models of postnatal androgen exposure employing a long-term treatment with either DHT, the proandrogen dehydroepiandrosterone (DHEA), or letrozole (an aromatase inhibitor) for 90 days beginning at 3 weeks of age. Prenatal androgenisation produced some reproductive and endocrine traits, but failed to induce the metabolic abnormalities seen in PCOS. DHEA treatment did not reproduce any features associated with PCOS while treatment with letrozole produced few PCOS-like characteristics and some aberrant changes not typical of the syndrome. Additionally, letrozole treatment did not reproduce any metabolic attributes of PCOS. On the other hand, postnatal exposure to excess androgen, by way of DHT treatment, produced a breadth of reproductive, endocrine and metabolic traits that mimic those seen in human PCOS. This study revealed that a treatment regime of long-term postnatal exposure to DHT reproduced the strongest PCOS-like phenotype in our mice and provides a robust animal model in which to study the pathogenesis of PCOS. The second study (Chapter Four) aimed to explore the involvement of genomic androgen receptor (AR)-mediated actions in the development of these PCOS traits. As a prenatally androgenised mouse model of PCOS had previously been reported to exhibit impaired neuroendocrine hypothalamic feedback of the hypothalamic-pituitary-gonadal (HPG) axis, we took this opportunity to utilise mice from our own prenatal model to investigate the neuroendocrine regulation of the HPG axis in PCOS in addition to the effects of AR inactivation on the PCOS phenotype. PCOS was induced in wild-type (WT) and androgen receptor knockout (ARKO) mice using DHT administered on days 16-18 of gestation. A subset of these mice were also exposed to 17β-estradiol for 7 days prior to collection, via a subdermal implant, to investigate the impaired estradiol negative feedback on the hypothalamus. WT mice with DHT-induced PCOS displayed several reproductive abnormalities including aberrant cycling and ovulatory dysfunction in addition to adipocyte hypertrophy and hepatic steatosis. However, diestrus serum luteinising hormone and follicle stimulating hormone, and estradiol-induced negative feedback as well as hypothalamic expression of several neuropeptides were unaffected by DHT treatment in WT mice. Mice both homozygous and heterozygous for the global inactivation of the AR (ARKO), did not display any PCOS traits when exposed to excess androgens during prenatal life. This study showed the importance of AR signalling in the development of PCOS and revealed that even AR haplosufficiency is adequate to prevent induction of PCOS by prenatal hyperandrogenism. Finally, the third study (Chapter Five) aimed to shed further light on the AR-mediated androgen actions in PCOS with a focus on identifying the tissue-specific targets of these actions. Employing our postnatal model of PCOS induction, this study investigates the effects of: 1) global loss of AR signalling (ARKO), 2) neuronal knockout (NeurARKO) and 3) granulosa cell-specific AR inactivation (GCARKO) on the development of the PCOS phenotype induced by exposure to exogenous DHT. As in our prenatal model, ARKO mice were fully protected from all DHT-induced features of PCOS. Neuron-specific AR signaling was required for the development of a variety of reproductive and metabolic traits including classic polycystic ovaries, dysfunctional ovulation, obesity and dyslipidemia. In contrast, loss of AR signalling in granulosa cells did not impede the pathogenesis of PCOS-like features in GCARKO mice. To further examine the role of extra-ovarian AR signalling in PCOS, reciprocal ovary transplants were carried out in WT and ARKO mice. Results from ARKO hosts with transplanted WT ovaries revealed that excess androgen exposure requires functional extra-ovarian, and not intra-ovarian, AR signalling in order to produce features of PCOS. This study provides strong evidence that neuroendocrine genomic AR signaling is an important mediator in the development of PCOS. The studies contained within this thesis are the first to provide a comprehensive analysis of a mouse model of PCOS encompassing a breadth of reproductive, endocrine and metabolic features. This work has identified the optimal model in which to study this complex, multifactorial condition which affects a significant number of women worldwide. Additionally, our results have shown that the effects of androgens on the pathogenesis of PCOS are mediated via the androgen receptor in a dose-dependent manner such that two functional copies are required for DHT to reproduce features of PCOS in the mouse. Finally, in a crucial study to investigate the locus of androgen actions we have revealed the previously overlooked importance of extra-ovarian neuroendocrine androgen action in the origins and progression of PCOS, despite it being thought of primarily as an ovarian disorder. Overall, these studies have provided valuable insights into both the role of androgens in Polycystic Ovary Syndrome and potential new targets for the development of mechanism-based treatments of this disorder.