Program

Location

Edwards Hall Lobby

Document Type

Poster Presentation (1 hour)

Description

Background: There is considerable evidence of bidirectional communication between the oocyte and the surrounding granulosa cells (GC) within the maturing human ovarian follicle. Study of GC gene expression could provide insight into predicting oocyte quality and embryo outcomes during in vitro fertilization (IVF). This study aims at establishing a model for future research with primary human GCs. Methods: Oocytes and GCs from a single follicle were harvested from IVF patients using Ultrasound-guided transvaginal retrieval. Oocytes were stripped of the surrounding GC which were isolated using gradient-based differential centrifugation and then frozen and stored at -80oC. GC samples were tracked to the IVF outcome of their associated oocyte including embryo fragmentation score, blastomere symmetry and embryo cell number. Total RNA extractions were performed only on GCs tracked with a known embryo outcome. RNA extracts were quantified and qualified and then converted into cDNA for future studies. Results: Embryos were successfully generated from 131 oocytes out of the total 290 follicles sampled. Of the 131 samples, which had both GC samples and embryo outcomes, only 31 GC samples yielded sufficient quantity and quality of total RNA (A260/A280 ratios between 1.8-2.0 and a minimum 3ug) for focused gene expression PCR array studies. Initial qPCR , run using a GAPDH housekeeping gene and CYP11A1 target gene results have shown presence of good quality cDNA . Conclusion: Investigation of the associations between GC gene expression and oocyte/embryo outcomes is dependent on obtaining sufficient amounts of high quality GC-RNA necessary for RT-PCR-Array analysis. We have shown here that, during follicle sampling of standard clinical IVF cycles, the expected efficiency of obtaining both embryos and sufficient extracted RNA is very low; <10% of sampled follicles.

Start Date

20-4-2013 10:30 AM

Comments

Math, Sciences and Engineering Poster Session

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Apr 20th, 10:30 AM

Predicting Oocyte Quality and Embryo Outcomes for IVF

Edwards Hall Lobby

Background: There is considerable evidence of bidirectional communication between the oocyte and the surrounding granulosa cells (GC) within the maturing human ovarian follicle. Study of GC gene expression could provide insight into predicting oocyte quality and embryo outcomes during in vitro fertilization (IVF). This study aims at establishing a model for future research with primary human GCs. Methods: Oocytes and GCs from a single follicle were harvested from IVF patients using Ultrasound-guided transvaginal retrieval. Oocytes were stripped of the surrounding GC which were isolated using gradient-based differential centrifugation and then frozen and stored at -80oC. GC samples were tracked to the IVF outcome of their associated oocyte including embryo fragmentation score, blastomere symmetry and embryo cell number. Total RNA extractions were performed only on GCs tracked with a known embryo outcome. RNA extracts were quantified and qualified and then converted into cDNA for future studies. Results: Embryos were successfully generated from 131 oocytes out of the total 290 follicles sampled. Of the 131 samples, which had both GC samples and embryo outcomes, only 31 GC samples yielded sufficient quantity and quality of total RNA (A260/A280 ratios between 1.8-2.0 and a minimum 3ug) for focused gene expression PCR array studies. Initial qPCR , run using a GAPDH housekeeping gene and CYP11A1 target gene results have shown presence of good quality cDNA . Conclusion: Investigation of the associations between GC gene expression and oocyte/embryo outcomes is dependent on obtaining sufficient amounts of high quality GC-RNA necessary for RT-PCR-Array analysis. We have shown here that, during follicle sampling of standard clinical IVF cycles, the expected efficiency of obtaining both embryos and sufficient extracted RNA is very low; <10% of sampled follicles.