Research
Research
Oocyte quality impacts early embryonic survival, the establishment and maintenance of pregnancy, fetal development, and even adult disease. As a faculty member in the Department of Animal Sciences at the University of Illinois, I have developed a competitive and innovative research program focusing on oocyte physiology. Quality, or developmental competence, is acquired during folliculogenesis as the oocyte grows, and during the period of oocyte maturation. Although meiosis, or nuclear maturation, may be completed successfully, there are a variety of other processes occurring within the cytoplasm of the oocyte that are required for complete developmental competence following fertilization. However, the cellular mechanisms that impart oocyte developmental competence are entirely unclear. My long range goal is to understand the mechanisms of nuclear and cytoplasmic maturation in mammalian oocytes in relation to developmental potential. In pursuit of this goal, I have pursued novel ways to think about the control of developmental competence in oocytes, using pigs and mice as model species.
One aspect of my research involves the investigation of metabolic pathways in the oocyte and developing zygote. I have demonstrated that metabolic events occurring in the oocyte are related to the ultimate developmental ability of the resulting zygote after fertilization. Importantly, I have shown that the metabolism of in vivo matured porcine oocytes is far more active than those matured in vitro, providing both a baseline of normal metabolic activity and knowledge of possible deficiencies. I have also demonstrated that glutathione, but surprisingly not ATP, or energy charge, was deficient in those oocytes matured in the laboratory. Interestingly, the pentose phosphate pathway (PPP), which is much more active in porcine oocytes than those of other domestic species, is related to glutathione production with the cell. Results point to this pathway as being key to successful maturation of porcine oocytes. Chemical manipulation of this pathway has recently shown this to be true as an increase in glucose metabolism via the PPP in the oocyte resulted in increased embryonic development after fertilization. Interestingly, oocyte meiotic arrest can be induced by chemically blocking the pentose phosphate pathway. This arrest can be overcome by addition of end products and cofactors of the pathway. Work in my laboratory has definitively demonstrated the intimate relationship between glucose metabolism via the pentose phosphate pathway and meiotic progression. This is the first time that a metabolic pathway has been demonstrated to control both meiosis and developmental competence in a mammalian species. My current goal in this area is to elucidate specific molecules related to glucose metabolism that play critical roles in the control mechanisms of resumption of meiosis and acquisition of developmental potential.
An additional aspect of my research, and the current focus of my research program, is to identify molecular markers in oocytes that are correlated with developmental competence. I hypothesize that a differential pattern of mRNA expression exists in cumulus-oocyte complexes with high and low developmental potential that reflects functional differences within these two classes of oocytes. My laboratory is now using microarray technology to analyze transcription in mouse and pig oocytes with different developmental competencies, thereby discovering target molecules that may be involved in the mechanism of developmental competence. A major project in the laboratory at this time is comparing gene expression in mature and immature oocytes derived from mice of varying ages with varying oocyte quality. This work is funded by an NIH R03 grant, entitled "Molecular Signature of Oocyte Developmental Competence," of which I am PI. We are also investigating mRNA expression in mature oocytes derived from gilts and sows. Gilt oocytes are not competent to support embryonic development until after the third estrous cycle, so a comparison of these two groups of oocytes will establish transcripts that are present in competent sow oocytes but not in incompetent oocytes from gilts. Because the transcriptome of oocytes with high and low developmental competencies will be compared, target molecules will be identified that are critical for oocyte viability and support of subsequent development. These target genes can then be further studied to elucidate critical developmental mechanisms in oocytes. I am complementing my genomics work with proteomic technology. We are examining the proteins present in follicular fluid of sows and gilts to establish those correlated with increased development of the enclosed oocyte. We are also investigating proteins in the ovaries of young versus aged mice to elucidate proteins involved in loss of fertility with age. Experiments on maternal aging of the oocyte have immediate application to human infertility. Overall, these series of studies will result in the discovery of mechanisms involved in and markers for oocyte developmental competence.
When differentially expressed gene transcripts and proteins related to oocyte quality have been identified, effective strategies can be developed to alter specific pathways critical to developmental potential of oocytes. This research will enable the development of assays to test for oocyte developmental competence, strategies to enhance oocyte maturation in vitro, including improved media formulation for in vitro oocyte maturation in humans and livestock species, and perhaps the identification and treatment of specific types of infertility by RNA or protein injection into oocytes. The successful application of reproductive biotechnologies to livestock species could impact both agricultural food production, preservation of genetic diversity, and biomedical research. Transgenic technology has the potential for improving food animal performance characteristics and disease resistance, as well as the production of human pharmaceutical proteins, models for biomedical research and a source of organs for xenotransplantation. Success of these technologies is intimately linked to the developmental potential of the oocyte.
A recent direction of my research program involves characterizing the Ossabaw mini-pig as a complete animal model for polycystic ovarian disease in women. Polycystic Ovary Syndrome (PCOS) is not only a gynecological condition affecting 5-10% of reproductive aged women, but a comprehensive syndrome with a variety of associated metabolic disorders including insulin resistance, diabetes, dyslipidemia, atherosclerosis and cardiovascular disease. Gynecologically, there is an association between PCOS and hyperandrogenemia, hirsutism, infertility, recurrent pregnancy loss, gestational diabetes and endometrial carcinoma. The reproductive and metabolic morbidities of PCOS make it an important women's health issue. Unfortunately, the etiology and pathogenesis of the syndrome are unclear, and there is no effective cure. Currently, no comparative animal model of PCOS exists that embodies the complexity of the syndrome. Our study will attempt to characterize and validate the female Ossabaw pig as a model of PCOS that comprises both its metabolic and reproductive aspects. Our preliminary findings suggest that female Ossabaw swine naturally develop many of the characteristics of MS when fed in excess, and that they exhibit one of the primary features of PCOS, namely hyperandrogenemia. This work is being carried out in collaboration with the Indiana University School of Medicine, and is funded by an Indidan University-Purdue University CBR2 pilot grant entitled "The Ossabaw Pig as a Model of Polycystic Ovary Syndrome," of which I am PI. In addition, I have 2 pending NIH submissions to further characterize this model and elucidate the mechanisms of PCOS.
I am also actively involved in research to apply assisted reproductive technologies to exotic species, and germplasm preservation. In a collaborative effort with the USDA germplasm banking program, we evaluated the longevity and usefulness of cryopreserved boar semen as a safeguard against loss of genes for production traits and biodiversity. It was discovered that cryopreserved semen can be stored for at least 20 years and then successfully used to create embryos in an in vitro fertilization program. In collaboration with The Saint Louis Zoo, The Cincinnati Zoo, Omaha's Henry Doorly Zoo and the Wildlife Biological Resource Center in South Africa, I am developing and applying novel techniques for the in vitro production of embryos from sperm and oocytes, as well as cryopreservation of these gametes and embryos, in antelope and exotic bovid species. My current study of in vitro embryo production in South African antelope species is the most thorough investigation of this topic ever undertaken in these species.