GROWTH FACTORS IN FOLLICULAR DEVELOPMENT

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Malfunction of granulosa cells results in follicular atresia, oocyte degeneration, sterility and impairment of a variety of physiological processes that depend on ovarian steroids. The purpose of the present proposal is to provide insight into the intricate cellular and molecular mechanisms involved in gonadotropin regulated folliculogenesis in general and granulosa cell function in particular. Because conventional granulosa cell cultures result in the loss of follicular structure and alteration in cell functions, we have developed an alternative approach to study follicular functions by use of isolated intact preantral follicles from the hamster ovary and a long-term culture system for their in vitro development. This technique opens an entirely new dimension to study directly the physiology and biochemistry of proliferating follicular cells in their native, follicular microenvironment. We have determined that follicular cells in these cultures synthesize cAMP and steroids, proliferate in response to gonadotropins; express gonadotropin receptors as well as unique follicular proteins. We have observed that mitogenic action of FSH is transduced via EGF. Radioimmunoassay data, Western blotting and immunoprecipitation studies indicated that the hamster ovary has both EGF, TGF beta1 and beta2-like proteins, and EGF receptors. However, the cellular and molecular changes during development of the follicles and the roles of growth factors in these processes remain to be determined. I hypothesize that gonadotropin and steroid regulation of follicular cell proliferation and differentiation depend upon critical modulation of the activities of ovarian growth factors (e.g., EGF and TGFs-beta) and their receptors. Disruption of this process results in malfunctions in granulosa cells and sterility. The present study will focus on one important question: (1) Do gonadotropins regulate folliculogenesis by modulating ovarian EGF and TGFs-beta (beta1 and beta2) activities? The study will involve (1) immunohistochemical analyses of growth factor expression and their hormonal control during in vivo and in vitro follicular development; (2) biochemical analyses of growth factor effects on follicular growth and steroidogenesis, and (3) immunohistochemical and biochemical assessment of EGF- receptors during hormonal control of follicular development. Identification of the pathway for gonadotropin action in intact ovarian follicles will significantly add to our knowledge of the fundamental mechanisms of folliculogenesis.