PGE2 Supplementation of Oocyte Culture Media Improves the Developmental and Cryotolerance Performance of Bovine Blastocysts Derived From a Serum-Free in vitro Production System, Mirroring the Inner Cell Mass Transcriptome

Abstract

The culture media used throughout the in vitro production (IVP) of bovine embryos remain complex. The serum added to culture media in order to improve embryo development negatively impacts the cryotolerance of blastocysts. Periconceptional prostaglandin E2 (PGE2) signaling is known to exert prosurvival effects on in vitro-generated blastocysts. The purpose of the present study was to evaluate the effects on developmental and cryotolerance performance of a serum-free (SF) IVP system that included defined oocyte culture media supplemented or not with PGE2, versus serum-containing (SC) IVP. RNA-sequencing analysis was used to examine the gene expression of ICM derived under the different IVP conditions. We assessed the degree of cryotolerance of grade-I blastocysts during a three-day post-thaw culture by measuring survival and hatching rates, counting trophectoderm and inner cell mass (ICM) blastomere numbers. We also determined the proportion of ICM cells expressing octamer-binding transcription factor 4 protein (OCT4/POU5F1). We showed that grade-I blastocyst development rates under SF + PGE2 conditions were similar to those obtained under SC conditions, although the cleavage rate remained significantly lower. SC IVP conditions induced changes to ICM gene expression relative to several metabolic processes, catabolic activities, cell death and apoptosis. These alterations were associated with significantly higher levels of ICM cell death at day 7 post-fertilization, and lower survival and hatching rates after thawing. SF IVP conditions supplemented or not with PGE2 induced changes to ICM gene expression related to DNA replication, metabolism and double-strand break repair processes, and were associated with significantly larger ICM cell populations after thawing. SF + PGE2 IVP induced changes to ICM gene expression related to epigenetic regulation and were associated with a significantly higher proportion of ICM cells expressing OCT4. For the first time, our study thus offers a comprehensive analysis of the ICM transcriptome regulated by IVP culture conditions in terms of the cellular changes revealed during culture for three days after thawing.

OCT4/POU5F1 is indispensable for the lineage differentiation of the inner cell mass in bovine embryos

“…SOX2 was expressed pan‐ICM at day 7 and restricted to EPI by day 9, resulting in a decreased percentage of SOX2‐positive cells. Together with NANOG, SOX2 cells were still present in the embryonic disk at day 12 and it has been shown previously that OCT4 is present in this lineage until day 17 11,12 . A total of 28 day 7 blastocysts were stained against SOX17, where this transcription factor was not present in eight embryos and only faint and restricted to the ICM in the remainder.…”

“…Together with NANOG, SOX2 cells were still present in the embryonic disk at day 12 and it has been shown previously that OCT4 is present in this lineage until day 17. 11,12 A total of 28 day 7 blastocysts were stained against SOX17, where this transcription factor was not present in eight embryos and only faint and restricted to the ICM in the remainder. By day 9, the HB began to form an inner lining of the blastocoel cavity consisting of the visceral and parietal HB, 13,14 which were both marked by SOX17 until day 12.…”

Epidermal Growth Factor (EGF) and Basic Fibroblast Growth Factor (FGF2) Profiles in Follicular Fluid, Maturation Media and Embryo Culture of Bali Cattle

“…In addition, it is suspected that embryonic development abilities are related to the existing growth factors (Prasad et al, 2018). Research results Charpigny et al (2021) reports several growth factors have been shown to affect the rate of embryonic development, the proportion of embryos that progress to the blastocyst stage, the number of blastocyst cells, metabolism and apoptosis. After re-culture for 48 hours or 96 hours post insemination (Table 2) Embryos of stage 4 cells, 8 cells,…”

Scoping review of primary studies evaluating Bovine in vitro oocyte maturation and embryo development

“…(9) The production of embryos involves crucial steps such as oocyte maturation (meiotic, cytoplasmic, and molecular), fertilization, and embryo development, which significantly affect embryo survival. (10) Therefore, the culture system used during the oocyte maturation and embryo culture stages has a significant impact on the development and quality of the blastocysts produced. (11) Despite the importance of culture conditions during the different stages of embryo production, there is still no consensus on the optimal medium for Bovine oocyte maturation or the optimal culture conditions and characteristics after fertilization for embryo development.…”