Extended embryo culture and ploidy of morphologically normal embryos assessed by next gen sequencing PGS: a single center retrospective study

Abstract

To analyze embryo culture metrics with regard to absolute measures (chromosome ploidy by NGS-PGS, age of oocyte) and trends (embryo age, embryo grade, egg donor recipient). A single - center, retrospective, observational study. Oocyte retrieval was performed 34 hours after trigger under ultrasound guidance. After retrieval, the oocytes were denuded mechanically by pipetting with cumulase (Origio, enzyme activity 40-120 U/ml). ICSI was performed on all metaphase II oocytes. Fertilized oocytes were cultured in microdroplets of culture media (Vitrolife) overlayed with Ovoil (Vitrolife), gassed with a blood gas mixture (WestAir Gases, 5% oxygen, 7.2% carbon dioxide, balanced nitrogen), until the day of assisted hatching (day 3 of embryo culture). Assisted hatching was performed and embryos were subsequently transferred to fresh culture medium. Embryos were assessed for blastocyst development, biopsy and cryopreservation criteria on days 5, 6, and 7. Blastocysts that met the criteria, either good (distinct inner cell mass (ICM)/numerous trophectoderm (TE) cells) or fair (small ICM/some TE cells) were biopsied and cryopreserved. All genetic analysis was performed by Ovation Fertility Genetics (Las Vegas) using Next Gen Sequencing. 2557 blastocysts from 257 cycles were analyzed in the present study. A total of 1179 on Day 5, 1289 on day 6, and 89 blastocysts at day 7 were biopsied and analyzed by NGS -PGS. On day 5 of culture 64% of embryos tested were euploid, 34% aneuploid. On day 6 of culture 49% of embryos were euploid and 49% aneuploid, and day 7 of culture, 46% of embryos tested were euploid and 52% were aneuploid. The chi-square statistic is 58.1 and is significant at p < .01, indicating that ploidy and length of time in culture are dependent variables. Our optimized culture system routinely yields blastocysts that are good and fair quality on day 7, which did not meet our biopsy and cryopreservation criteria on day 5 or 6 of culture. In the present study, we retrospectively analyzed PGS data from day 5, 6, and 7 blastocysts derived from IVF patients in our clinic during 2017- 2018, and analyzed trends in embryo development and ploidy. Our results (46% euploid blastson day 7) agree with the small, but growing body of evidence that suggests that extended embryo culture to day 7 is clinically important for patients who do not have day 5 and/or day 6 blastocysts. Additionally, we believe for patients with day 5 and/or day 6 blastocysts, day 7 culture may still be necessary, considering the time and costs of IVF cycles, particularly when constructing an IVF cycle with donor eggs.