The goal of this proposal is to explore a novel role of oxygen on regulating primordial follicle quiescence in the primate ovary with long term clinical impacts of controlling ovarian aging and improving fertility preservation methods for young cancer patients. Women are born with a finite number of eggs/oocytes (~ 1 million), sequestered in ?quiescent primordial follicles? that comprise of a resting oocyte surrounded by pre-granulosa cells. This pool of follicles, also known as the ovarian reserve, serves as the source of growing follicles and fertilizable oocytes. Primordial follicles can stay quiescent for decades; however, ~75% of the pool is depleted before puberty. After puberty, ~1000 primordial follicles are activated, an irreversible process, during each menstrual cycle, but only 1 matures and is capable of releasing a fertilizable oocyte, while the rest degenerate. This natural decline of the ovarian reserve is accelerated by various stresses and disease states that cause premature ovarian aging (e.g. cancer therapy, Turner's syndrome, endometriosis, etc). One could control ovarian aging or prevent premature ovarian failure by delaying the activation of primordial follicles. However, mechanisms that govern primordial follicle quiescence are not understood, especially in primates. Despite studies characterizing the relationship between angiogenesis and hypoxia-inducible factors, very little information exists about the actual O2 levels and their roles in the ovary. The PIs' preliminary data suggest that primordial follicles exist in a state of physiological hypoxia, and that inappropriate introduction of oxygen to tissue or culture medium results in follicle activation. In order to test this hypothesis, the PIs propose experiments to measure the physiological concentration of oxygen in the ovarian cortex, explore effects of hypoxia/reoxygenation (in vivo) and determine the concentration of oxygen required to maintain follicles in the quiescent state (in vitro). The rhesus macaque will be used as the model system with the strongest parallels to human ovarian biology. Follicle activation will be measured at the morphological and molecular level (i.e. markers of cellular proliferation and oxidative stress), with additional characterization of hypoxic signaling. Successful implementation of this proposal will determine specific roles of oxygen in maintaining the quiescent state of the primordial follicle. The findings will support future in-depth, mechanistic studies to identify factors that regulate follicle activation in response to oxygen concentration. Ultimately, these studies will lead to the development of innovative technologies that can advance progress in reproductive medicine, such as: 1) Methods to suppress inappropriate follicle loss during ovarian tissue cryopreservation and transplantation in young female cancer patients, 2) Improved interventions for ovarian torsion, which occur during ovarian stimulation protocols, 3) Novel techniques for in vitro maturation of follicles from the primordial to mature stage, and 4) Increase the number of gonadotropin-dependent follicles in the growing pool, increasing the number of fertilizable eggs and improving the odds of pregnancy for women using assisted reproductive technology.
Women are born with limited numbers (~1 million) of quiescent eggs (primordial oocytes) in the ovary; however, the majority become atretic during resting or growing stages; only 25% remains at puberty, and only ~500 oocytes will mature and become fertilizable throughout a women?s lifetime. In order to address this poorly understood phenomena, we propose using the clinically relevant rhesus macaque model to investigate the novel idea that oxygen acts as a modulator for primordial oocyte quiescence and growth. If proven, results from this proposal may offer strategies to delay ovarian aging or prevent premature ovarian failure (i.e. following cancer treatment) by controlling the activation of primordial follicles.
