FAS AND BC1-2 AND GRANULOSA CELL APOPTOSIS

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Candidate. The candidate has been trained in obstetrics and gynecology and has completed a subspecialty fellowship in Reproductive Endocrinology. He has been actively engaged in basic research on human granulosa cell physiology for over 4 years and has experience with a number of the techniques to be used in this proposal. He has published five first-author papers, and another has been submitted (Appendix). Research plan. During embryonic and fetal development, the number of oocytes in the human ovary increases until mid-gestation, after which it steadily declines until the menopause. Oocytes which are not surrounded by granulosa cells nearly i ovarian development undergo atresia, while follicles also undergo atresia at every stage of development from the primordial to the tertiary antral stage. The vast majority of oocytes are lost through atresia; only a minority mature to ovulation. In non-human models, atresia of antral follicles has been associated with granulosa cell apoptosis, or programmed cell death. Activation of apoptosis can be regulated by both extracellular (cytokine, growth facto) and intracellular mechanisms. The extracellular factors which regulate non- human granulosa cell apoptosis are being reported. The proposed studies seek to identify the intracellular pathways that signal the human granulosa cell to undergo apoptosis, as well as the source and identity of th extracellular effectors that activate the apoptotic signal through these pathways. A better understanding of the process of granulosa cell apoptosis may aid in the design of new therapies to retard the loss of oocytes that accompanies both normal aging and therapies for malignancy. The specific aims of this proposal are to investigate the roles of the intracellular apoptosis-regulating molecules Fas/APO-1 and Bcl-2 in the human ovary, as well as the roles of the cytokines interferon-gamma (IFN- gamma) and tumor necrosis factor-alpha (TNF-alpha) in transmitting the apoptotic signal. These studies may also help to determine whether the apoptotic signal originates within ovarian parenchymal cells or from leukocytes. It is hypothesized that in human granulosa cells, expression and activation of Fas promotes apoptosis, while Bcl-2 expression in excess of its antagonist Bax prevents apoptosis. It is further hypothesized that IFN-gamma induces Fas expression, while TNF-alpha may induce apoptosis through its own receptor. Environment. The Reproductive Endocrinology Center at the university of California, San Francisco is supported by NIH Grant #HD11979. Included in the Center are Morphology, Molecular Biology, and Radioimmunoassay Cores, the facilities of which are available to the investigator. The Primary Sponsor's laboratory and these Cores contain all of the necessary equipment for the studies proposed.