Steroid hormone regulation of fetal mouse oocyte development

The factors governing maintenance of the non-renewable reservoir of primordial follicles in female mammals remain largely elusive. During the early stages of fetal development, oocytes grow in nests or clusters known as germ cell cysts. Cysts break down into individual oocytes in the perinatal period and become enclosed by somatic pregranulosa cells to form primordial follicles. Steroid hormones have been shown by numerous studies to be one of the important factors which critically govern the process of cyst breakdown and follicle formation. There has been earlier evidence from this laboratory to indicate that exogenous exposure of neonatal mice ovaries to estradiol (E2), progesterone (P4) or E2 mimicking chemicals known as xenoestrogens such as Diethylstilbestrol (DES) and Bisphenol-A (BPA) delay cyst breakdown and follicle formation. The overall goal of this dissertation project is centered on the pivotal question: "What is the source of steroid hormone signaling and its role in meiotic progression during fetal oocyte development in mice?" One of the two objectives of this dissertation was to identify the sources of the steroid hormones (maternal circulation or fetal ovaries) which regulate fetal oogenesis. Our studies showed prominent expression of both mRNA and protein in the fetal ovaries for aromatase and 3-beta-hydroxysteroid dehydrogenase (3betaHSD), cardinal steroidogenic enzymes required for E2 and P4 synthesis respectively. The mRNA levels for both aromatase and 3betaHSD in the fetal ovaries detected by qPCR were found to decrease prior to cyst breakdown. These results align to our previous model that high levels of steroid hormones keep oocytes in cysts during fetal development and the drop in hormone levels is required to trigger cyst breakdown. To analyze the functional significance of this local steroid action, we used aromatase and 3betaHSD inhibitors (letrozole and trilostane respectively) in organ culture to block hormone production by fetal ovaries. We find that the total number of oocytes was reduced in treated ovaries compared to controls. The second objective of the dissertation was to examine the relation between two temporal events in mice oogenesis: progression to the diplotene stage and primordial follicle formation. We performed a thorough quantitative analysis by nuclear morphological observations of diplotene versus prediplotene nuclei of hematoxylin and eosin stained serial sections of ovaries at different ages. Interestingly, we observed that primordial follicle formation occurs irrespective of the meiotic stage of the oocyte nuclei. Thus oocytes in follicles were found both at diplotene and pre-diplotene stages. We also wanted to understand the role of steroid hormone signaling in meiotic progression of oocytes. Our data indicate that exogenous treatment of P4 and not E2 decrease the number of follicles containing oocytes at diplotene. Such insights from the murine research models significantly contribute to our knowledge of the meiotic defects caused due to E2 or P4 exposure during fetal oogenesis in the case of human pregnancies (which often results from exposure to environmental estrogens or xenoestrogens). Aneuploidy is one of the prevalent causes for genetic disorders in humans and it arises from anomalies in the chromosome content of the gametes (sperms and ova). Any disruption in the normal meiotic events during fetal gametogenesis may be amplified along the way to give rise to aneuploid gametes. Meiotic studies in model organisms are therefore indispensable to our understanding of human aneuploidy. In summary, this dissertation project has focused on the critical role of steroid hormone regulation of fetal mouse oocyte development and its role on meiotic progression, thus contributing to our understanding of early ovarian differentiation.