This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Progress has been made in understanding the regulation and role of apoptosis in female germ cells (Nat Rev Mol Cell Biol 2001, 2:838-848), providing insight into the basis of ovarian failure under both normal and pathological conditions. In the course of conducting out studies of oocyte death, we noticed that the incidence of apoptosis in oocytes harvested from different inbred strains of mice and cultured for 24 hours under identical conditions was dramatically different, ranging from around 5% (apoptosis-resistant) to over 90% (apoptosis-prone). Using conventional electron microscopic evaluation in oocytes of different inbred strain of mice we observed electron-dense mitochondria with enlarged vesicles, but intact membranes or more dense mitochondria with smaller vesicles, but disrupted membranes. The oocytes with abnormal mitochondria exhibited an elevated incidence of apoptosis (93 2%) after 24 h of culture. To further explore the consequences of these latter observations, mitochondria were isolated from ovarian somatic cells of mice with either the apoptosis-prone or apoptosis-resistant oocytes. These pools of purified mitochondria were then microinjected into oocytes of the opposite strain to determine their influence, if any, on apoptosis. Introduction of approximately 5,000 apoptosis-prone mitochondria into apoptosis-resistant oocytes increased the incidence of culture-induced apoptosis nearly 5-fold. On the other hand, introduction of approximately 5,000 apoptosis-resistant mitochondria into apoptosis-prone oocytes reduced the incidence of culture-induced apoptosis by approximately 50%. These data suggest underlying mitochondrial phenotype manifested by ultrastructural and biochemical properties (not discussed here) of mitochondria and susceptibility to apoptosis in the female germ line. We are collaborating with Drs. Perkins and Ellisman to further explore this phenotype using EM tomography, which we is revealing finer structural changes associated with cell death in the oocytes prone to death.
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