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dc.contributor.advisorRussell, Darryl Lyndonen
dc.contributor.advisorLane, Michelle Thereseen
dc.contributor.authorGebhardt, Kathryn Michelleen
dc.description.abstractCurrent infertility treatments are confounded by an inability to identify oocytes and embryos with the highest developmental potential to generate and sustain a pregnancy resulting in a live birth, reducing the efficiency of treatment cycles and resulting in low pregnancy success rates. Embryos have varying capacity to form a successful pregnancy and embryo developmental potential is particularly reliant on nuclear and cytoplasmic qualities of the oocyte from which it is derived. A biochemical marker of oocyte and therefore embryo developmental potential would improve pregnancy success rates following assisted reproductive technologies by optimising oocyte and embryo selection techniques. The communication between an oocyte and its surrounding cumulus cells is essential for growth, maturation and metabolic activity, and strengthens the rationale to utilise cumulus cells to assess oocyte quality and predict treatment outcomes and health parameters for women undergoing assisted reproduction. The potential for cumulus cell gene expression to predict clinical embryo grade and pregnancy success was investigated in cumulus masses from single human oocytes which were fertilised and cultured individually. To make direct correlations between cumulus cell gene expression and treatment outcomes patients underwent single embryo culture and transfer. Gene expression was analysed in cumulus cells from independent oocytes that yielded a successful term pregnancy compared to those for which treatment failed and pregnancy was not established. Patient matched cumulus cell pairs were utilised to investigate a potential correlation between cumulus gene expression and clinical embryo grade. Cumulus cell gene expression was assessed using both a microarray platform for non-biased genome wide gene expression analyses and real-time RT-PCR assays focused on genes with known important functions related to oocyte maturation. Real time RT-PCR analyses identified cumulus expressed genes which significantly correlated with pregnancy success following single embryo transfer. Specifically, cumulus cell PTGS2, VCAN and GAS5 mRNA expression significantly (p < 0.02) correlated with establishment of a pregnancy resulting in a live birth, while PTX3 mRNA expression showed a trend towards significance (p = 0.066). Additionally, cumulus cell levels of VCAN, GREM1 and PFKP showed a significant correlation with birth weight in the patients who achieved pregnancy, indicating their role as potential predictors of health outcomes for babies born from assisted reproduction. No significant differences were seen for other genes analysed in relation to pregnancy outcome or when gene expression was correlated with clinical embryo grade. The use of a microarray platform led to the identification of new genes, never before identified in the COC as markers of human oocyte quality and pregnancy success. The characterisation of GAS5 and PEPSINOGEN transcripts in both human and murine follicular cells furthered the rationale for their potential as markers of oocyte quality and provided an understanding of the pattern of expression and hormonal regulation within human and murine ovarian cells. The expression of the GAS5 transcript was confirmed by real time RT-PCR analysis and shown to be significantly correlated with live birth following initial identification by microarray experiments. The findings demonstrate that expression of VCAN, PTGS2, GAS5 and PTX3 represent molecular markers of oocyte quality. The molecular markers identified in this study provide a unique tool to assess the relative potential of individual oocytes to achieve successful pregnancy from a pool of oocytes generated by one patient. In conjunction with embryo selection techniques of visual assessment and developmental milestones in culture the present biomarkers provide information to differentiate between embryos with similar appearance as viable or non-viable, to improve ART efficiency while decreasing multiple gestations and even extend to predict birth weight and hence general health expectancy for babies conceived following assisted reproduction.en
dc.subjectoocyte; cumulus cells; pregnancy; infertility; gene expressionen
dc.titleIdentification of molecular markers of pregnancy success for assisted reproduction.en
dc.contributor.schoolSchool of Paediatrics and Reproductive Healthen
dc.provenanceCopyright material removed from digital thesis. See print copy in University of Adelaide Library for full text.en
dc.description.dissertationThesis (Ph.D.) -- University of Adelaide, School of Paediatrics and Reproductive Health, 2010en
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