O-11: Prenatal Oogenesis: Selecting the Qualityand Quantity of Oocytes in the OvarianReserve

Background: The purpose of this research programme is to improve understanding of the molecular and cellular processes that lead to selection of oocytes before birth. Vastly more oocytes are produced prenatally than can be utilised in reproductive life. Around 70%, of oocytes formed are eliminated before birth and never contribute to the ovarian reserve. After birth, the number of oocytes continues to decline throughout life to the menopause. Which oocytes are selected for deletion and why? This question remains enigmatic. With such powerful selection, one might expect that only the most perfect oocytes, with the highest capacity for development, will survive. However, this is clearly not the case because many oocytes that reach ovulation and fertilisation are abnormal and human fertility is often suboptimal. Therefore, our research aims to understand the criteria by which oocytes are selected for contribution to the ovarian reserve. A number of approaches and papers spanning over a decade have helped to elucidate some of the answers.Materials and Methods: Human fetal ovaries are collected by collaboration with organisations undertaking termination of pregnancy in the second trimester. There is no contact between researchers and patients. Patients are given information about the study by a member of their healthcare team and invited to participate, without any effect upon their treatment, regardless of their decision. If patients consent, ovaries are removed from fetuses as soon as possible after delivery and taken to the laboratory. In view of the limited supply of human material for research, mouse fetal ovaries are also utilised, from timed matings, to ensure precise dating of pregnancies. On occasion, specific mouse genotypes are also used, for example in our work on p53 knockout mice. Ovaries are examined by a range of methods, specifically including (1) immunocytochemistry, to unequivocally identify the stages of meiotic prophase I or particular molecular configurations such as recombination foci, (2) FISH, to identify individual chromosomes, (3) tissue culture, with a range of environmental conditions and supplements, to identify which oocytes survive and to optimise conditions for their study, (4) molecular markers of apoptosis, to identify oocytes that have been selected for removal from the viable pool.Results: Our results have shown that oocyte selection for apoptosis includes, but is not restricted to those with genetic abnormalities, since apoptotic markers do not overlap 100% with degenerating oocytes. We haveshown that oocytes can survive in vitro under a range of conditions, but that progression beyond the pachytene stage of meiotic prophase I is rarely observed. We have shown that the timing of entry into meiotic prophase I, and the timing of progression through it, may affect likelihood of survival, and that p53 status in the mouse mother or the fetus may affect oocyte formation.Conclusion: Apoptotic markers and cytogenetic abnormalities are both indicative of oocyte lack of viability, however, these are not coincident and environmental factors also contribute to oocyte survival or selection for removal from the pool.