Doctoral Degrees (Obstetrics and Gynaecology)

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Browsing Doctoral Degrees (Obstetrics and Gynaecology) by Subject "Embryo markers"

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    The evaluation of different embryo markers and their subsequent effect on embryo development, implantation and pregnancy outcome in an in-vitro fertilization program
    (Stellenbosch : Stellenbosch University, 2012-12) Kotze, Dirk Jacobus; Kruger, T. F.; Sher, G.; Stellenbosch University. Faculty of Medicine and Health Sciences. Dept. of Obstetrics and Gynaecology.
    ENGLISH ABSTRACT: CHAPTER 1 In this chapter the aim is to outline the different chapters under section A. Against this background, we will conduct a literature review of relevant studies performed, and evaluate their comments regarding identifying embryo markers which can be utilized to improve overall ART outcome. We will evaluate the embryo marker sHLA-G in detail, using a prospective randomized study as well as a retrospective multi-centre study. The role of the morphology and genetic profile of an oocyte, zygote and embryo and subsequent blastocyst formation will be evaluated with the help of WGA/CGH. The work will then be summarized and conclusions will be made as well as possible suggestions for future directions will be indicated. In section B the methodology of the studies explaining the role of the candidate is illustrated. CHAPTER 2 In this chapter the impact of the oocyte/zygote and the embryo on implantation/pregnancy rate was discussed. The morphologic characteristics of the oocyte, the cumulus–oocyte-complex (COC), the zona pellucida, the perivitelline space, cytoplasm and meiotic spindle and the polar body and its appearance were discussed in detail. The morphologic characteristics of embryo fragmentation and its effect on embryo development, ploidy and blastocyst formation were also studied. Embryo markers to predict pregnancy outcome were researched based on the international literature. The pronuclear morphology and early cleavage were highlighted as non-invasive embryo markers to predict outcome. A non-invasive biochemical marker, soluble Human Leucocyte-Antigen-G (sHLA-G), that is expressed by developing embryos was researched. The value of blastocyst transfer and the improved ongoing pregnancy rate compared to cleavage stage embryos were highlighted based on a recent meta-analysis. A detailed discussion on sHLA-G as well as Array-CGH and the future of these tests followed. CHAPTER 3 In this chapter the aim was to compare pregnancy and implantation rates when embryos are selected based on a single Day 3 (D 3) morphology score vs. a GES score plus sHLA-G expression. This was a prospective randomized study (n=214) undergoing fresh ICSI cycles. Embryos were selected for transfer based on either Day 3 morphology score (Group A) or GES-scoring plus sHLA-G expression (Group B). The following results were reported: Clinical [35/107 (33%) vs. 52/107 (49%)] and ongoing pregnancy [20/107 (19%) vs. 52/107 (49%)] rates were significantly different between Group A and Group B (p<0.05). Implantation rates were not significantly different between Group A [52/353 (15%)] and Group B [73/417 (18%)] (p<0.05). The number of pregnancies lost during the first trimester was nearly 12 times higher in Group A [25/52 (48%)]. It was concluded that the miscarriage rate was significantly lower in Group B than Group A and the pregnancy results were superior when embryos were selected based on GES plus sHLA-G expression. CHAPTER 4 Several studies have reported an association between the presence of soluble human leukocyte antigen-G (sHLA-G) in human embryo culture supernatants (ES) with implantation and pregnancy outcome in vitro. However, the actual presence role during implantation and effect on implantation and pregnancy outcome are still controversial. A retrospective multi-centre study was performed on 2040 ICSI patients in six different centers. All embryos were individually cultured and a chemiluminescence enzyme-linked immunosorbent assay (ELISA) was used to detect the presence of sHLA-G in culture medium surrounding embryos. In all centers, a positive sHLA-G result was associated with an increase in odds of multiple clinical implantations (OR: 1.48, 95% CI: 1.07 to 2.05, p-value: 0.0170), and an increased odds of multiple on-going pregnancies (OR: 1.66, 95% CI: 1.10, 2.51, p-value: 0.0170). Data from this multi-centre study emphasize that sHLA-G expression is a valuable non-invasive embryo marker to assist in improving pregnancy outcome with the theoretical potential to reduce multiple pregnancies. A combination of sHLA-G expression and extended embryo culture to the blastocyst stage might provide future tools by which to select single embryos for transfer and reduce the risk of multiple gestational, without compromising their pregnancy rates. CHAPTER 5 In this chapter the ploidy status of first and second polar bodies and Day 3 blastomere, embryo morphology and biochemical (sHLA-G) characteristics were correlated with blastocyst development and subsequent pregnancy outcome. All oocytes/zygotes and embryos were individually cultured to the blastocyst stage. PB-I, PB-II and blastomeres underwent whole genome amplification (WGA) and comparative genome hybridization (CGH) and complete karyotyping. Each embryo‟s culture medium supernatant was collected and analyzed for sHLA-G expression on Day 2. The following results were reported: Fifty seven mature (MII) donor oocytes were obtained, 33/57 (57.9%) were aneuploid, 21/57 (36.8%) were euploid and 3/57 (5%) were “inconclusive”. No correlation was found between CGH status of PB-I, PB-II and the GES-score. Furthermore, no correlation was established between PB-I CGH results and blastocyst morphology grade. There was a significant correlation between PB-I CGH and blastomere CGH results. Euploid and aneuploid PB-I developed into 58% and 67% blastocysts, respectively. Kappa statistics (>0.7) revealed a positive correlation between the ploidy of PB-I, PB-II and the blastomeres. It was concluded that following ICSI and sequential genetic karyotyping of the oocyte/zygote and subsequent blastomeres, the majority of oocytes fertilized and subsequent zygotes developed into blastocysts, despite their ploidy status. We therefore conclude that blastocyst development is not associated with ploidy. CHAPTER 6 Identifying a developmentally competent embryo to transfer that has the highest probability to develop into a live baby has been an issue of debate and continues research. The aim of this chapter is to discuss the morphological, biochemical and genetic features of an embryo that has been shown to be predictive of implantation and pregnancy outcome in ART using most current evidence. A literature search was performed looking at the correlation between pronuclear morphology, early cleavage, cleavage stage embryos, blastocyst development, the presence of sHLA-G, CGH, embryo development and implantation/pregnancy rates in ART. Based on the available literature, a combination of observations could assist the scientist with embryo selection. The pronuclear stage morphology, the early embryo division, cleavage embryo stage and quality of the day 3 embryos provides limited guidance. However, choosing a blastocyst with a positive sHLA-G result on Day 5 is the optimal combination to make the final selection before embryo transfer or freezing. This non-invasive approach should improve pregnancy outcome and reduce multiple pregnancy rates. As far as the use of the more invasive technology such as aCGH is concerned, more research on pregnancy outcome is needed. CHAPTER 7 A combination of observations for embryo selection, starting with oocyte grading, pronuclear stage morphology, early zygote cleaving and cleavage-stage embryo morphology/quality on Day-3, however, ultimately using extended embryo culture and choosing a blastocyst on Day 5 with positive sHLA-G values available, will assist the scientist in making the final decision before selecting an embryo for transfer or cryopreservation. The use of aCGH (for chromosomal analysis) is invasive and is still considered experimental. Finally we conclude that despite all the above mentioned parameters to select an embryo for transfer that will develop into a live baby, more extensive research and international corroboration is needed in order to improve and standardize embryo selection criteria.