Abstract

Polycystic ovary syndrome (PCOS) is the most frequent cause of hyperandrogenism in women of reproductive age. PCOS is associated with metabolic derangements such as IR and diabetes mellitus 2 with an important medical burden in terms of public health. It has been observed that an excess of androgens during gestation induces PCOS-like reproductive and metabolic traits (including hyperandrogenism, oligomenorrhea, polyfollicular ovaries, obesity, IR and impaired insulin secretion) in female mammals, suggesting that the intrauterine environment may play a role in the etiology of PCOS. We were the first to demonstrate that pregnant women with PCOS exhibit elevated androgen and insulin levels, and proposed that PCOS mothers could provide an altered maternal environment for both female and male fetuses. In addition, we have developed an animal model of prenatal androgenization to further test this hypothesis. Our prenatal testosterone (T) exposure model is the closest model for PCOS, since it resembles the increasing amount of T that characterizes ongoing pregnancy in PCOS mothers. We have observed that the offspring of T treated sheep mothers have several reproductive and metabolic disruptions resembling those observed in the offspring of PCOS mothers. In addition, the animal model offers mainly an androgenized millieu to the fetus in comparison to the PCOS mother who offers a hyperandrogenic and hyperinsulinemic intrauterine environment. Therefore the sheep model lets us dissect the effect of androgens from other stimuli such as insulin, and provides tissue samples for studies in the offspring. The underlying mechanism of the fetal origins of PCOS has not been established. Epigenetic changes in key genes programmed in utero may be a possibility. Thus, site-specific epigenetic studies in selected target tissues are needed in order to verify this hypothesis. In the present proposal we aim to study epigenetic changes in the offspring of PCOS mothers (methylation of peripheral blood leukocyte DNA) and T treated sheep mothers (methylation of peripheral blood leukocyte DNA and methylation, gene and protein expression in target tissues). We will explore possible candidate genes associated with metabolic and reproductive dysfunction based on the phenotypic changes observed in our human and experimental models. Therefore our hypothesis is: The altered intrauterine environment modifies the expression of key genes trough epigenetic changes leading to metabolic and reproductive abnormalities in the offspring of hyperandrogenic mothers. To test this hypothesis, our design will evaluate 2 stages of development. For the human model a) early infancy and b) peripuberty in females and males born to PCOS and control mothers; and for the animal model a) fetal period and b) peripuberty in the offspring of T-treated and control sheep mothers. These two stages are key to know whether metabolic and reproductive epigenetic changes arise early in development and/or if they are expressed later when T levels rise due to pubertal development. We will accomplish the following general goals: a. To demonstrate that the metabolic and reproductive derangements observed on the offspring born to PCOS and T-treated sheep mothers occur through epigenetic changes of specific genes induced by an altered intrauterine environment. b. To establish if the findings of the human model correlate with the methylation pattern and expression of these specific genes in metabolic and reproductive tissues in the ovine model. Specific goals: 1. To evaluate the methylation patterns of ADIPOQ, ADIPOR1, ADIPOR2, LEP, LEPR, AMH and AR genes in peripheral blood leukocytes of the offspring born to PCOS women during early infancy. 2. To determine the methylation patterns of ADIPOQ, ADIPOR1, ADIPOR2,LEP, LEPR, AMH and AR genes in peripheral blood leukocytes of the offspring born to PCOS women during the pre and late pubertal periods. 3. To determine the methylation patterns of ADIPOQ, ADIPOR1, ADIPOR2,LEP, LEPR, AMH and AR genes in peripheral blood leukocytes in fetal and in pubertal offspring born to T-treated sheep mothers. 4. To assess the methylation patterns of the adiponectin, leptin, and their receptor genes in the main metabolic tissues (adipose tissue, liver and muscle) in fetal and in peripubertal offspring born to T-treated sheep mothers. 5. To assess the methylation patterns of the AR and AMH in the main reproductive tissues (ovary and testis) in fetal and in peripubertal offspring born to T-treated sheep mothers. 6. To evaluate gene and protein expression of these candidate genes in metabolic and reproductive tissues in fetal and in pubertal offspring born to T-treated sheep mothers. The results from the present proposal will help to clarify the mechanisms by which the altered intrauterine environment modifies the expression of key genes, trough epigenetic changes, leading to metabolic and reproductive abnormalities in the offspring of hyperandrogenic mothers. Moreover, it may suggest early preventive strategies focused in the adequate management of pregnant PCOS women in order to avoid the perpetuation of this syndrome.