Ovarian function, including follicle development, ovulation, and corpus luteum formation/degradation, is dependent upon tissue remodeling events, many of which are associated with a family of Zn+dependent endopeptidases, the matrix metalloproteinases (MMPs). Extreme remodeling of ovarian tissue is observed in photoperiodic species where seasonal changes in day length can either inhibit or stimulate hypothalamic/ pituitary secretion of GnRH/gonadotropins, leading to atrophy or resumption of ovarian function. Photo- responsive individuals are therefore excellent models for basic ovarian function, because modifications of day length naturally induce and then reverse ovarian atrophy. In photoperiodic Siberian hamsters, ovarian remodeling prompted by changes in day length is associated with differential expression of MMPs. Indeed, photostimulated return to ovarian function (recrudescence) can be impeded following in vivo administration of a broad-spectrum MMP inhibitor, GM6001. While this suggests an important role for these proteases, the action of MMPs during recrudescence and the process of how the quiescent ovary can resume cycling following weeks of atrophy is unknown. This proposal hypothesizes that 1) inhibition of recrudescence in response to GM6001 occurs because cleavage of MMP substrates that normally mediate return to ovarian function is impeded, 2) that the GM6001-treated ovary fails to return to function because key ovarian processes such as angiogenesis, granulosa cell proliferation, and steroidogenesis are dependent on MMP activity, and 3) that because gelatinases (MMPs-2/-9) are key players in ovarian cyclicity, and gelatinase activity is significantly down regulated following GM6001 administration, much of the remodeling can be attributed to gelatinases. Initial experiments will use a hypothesis-driven proteomics approach to identify substrates cleaved by MMPs during recrudescence by comparing the proteome of GM6001-treated vs. control ovaries. Examination of both mRNA and protein for key markers of angiogenesis (e.g., CD34, VEGF-R1), proliferation of granulosa cells (PCNA), and steroidogenesis (e.g., Cyp19, 32HSD) will reveal a mechanism of MMP action during recrudescence, and define a role for MMPs in these processes as GM6001 treated tissue is compared to controls. Finally, a gelatinase-specific inhibitor (SB-3CT) will be administered in vivo during photostimulated recrudescence to provide direct evidence of gelatinase action and function in the recovering ovary. Together, these studies should provide novel insight into the cellular and molecular regulation of recrudescence of ovarian function. In addition, these data will provide the first evidence of the targets of MMP action as ovarian cyclicity returns. Understanding the role that MMPs play in mammalian ovarian function by using a photoperiodic model helps to elucidate critical clinical questions of how to shut down (contraception) and restart (assisted reproduction, premature menopause) ovarian activity with non-hormonal mechanisms.
Ovarian recrudescence is a natural process stimulated by changes in photoperiod (the number of hours of light per day) whereby the atrophied and non-functional ovary returns to a fully functional organ in 2-8 weeks. While the hormonal initiation of recrudescence has been thoroughly examined, the cellular changes that occur at the level of the ovary remain unknown. Understanding the cellular/molecular pathways of mammalian ovarian recrudescence, such as matrix metalloproteinase (MMP) action, is novel and timely, as new non-hormonal methods of contraception and assisted reproduction techniques are clinically sought. Our studies will provide potential non-hormonal targets for clinically- geared studies focused on """"""""turning on"""""""" and """"""""turning off"""""""" ovarian function.
