Candidate gene expression patterns in rabbit preimplantation embryos developed in vivo and in vitro

Abstract

Purpose The levels and timing of expression of genes like BCLXL, HDAC1 and pluripotency marker genes namely, OCT4, SOX2, NANOG and KLF4 are known to influence preimplantation embryo development. Despite this information, precise understanding of their influence during preimplantation embryo development is lacking. The present study attempts to compare the expression of these genes in the in vivo and in vitro developed preimplantation embryos. Methods The in vivo and in vitro developed rabbit embryos collected at distinct developmental stages namely, pronuclear, 2 cell, 4 cell, 8 cell, 16 cell, Morula and blastocyst were compared at the transcriptional and translational levels using Real Time PCR and immunocytochemical studies respectively. Results The study establishes the altered levels of candidate genes at the transcriptional level and translational level with reference to the zygotic genome activation (ZGA) phase of embryo development in the in vivo and in vitro developed embryos. The expression of OCT4, KLF4, NANOG and SOX2 genes were higher in the in vitro developed embryos whereas and HDAC1 was lower. BCLXL expression had its peak at ZGA in in vivo developed embryos. Protein expression of all the candidate genes was observed in the embryos. BCLXL, KLF4 and NANOG exhibited diffused localisation whereas HDAC1, OCT4, and SOX2 exhibited nuclear localisation. Conclusions This study leads to conclude that BCLXL peak expression at the ZGA phase may be a requirement for embryo development. Further expression of all the candidate genes was influenced by ZGA phase of development at the transcript level, but not at the protein level.

Cloning and characterization of rabbit POU5F1, SOX2, KLF4, C-MYC and NANOG pluripotency-associated genes

“…Particularly, the expression of POU5F1 could be followed in developing embryos of POU5F1 promoter-EGFP transgenic rabbits (Yin et al, 2013). In another study, the expression level of POU5F1, SOX2, NANOG and KLF4 was measured by quantitative PCR during rabbit preimplantation development (Henderson et al, 2014). In agreement with our results, it was shown that these genes are expressed during this period, and were suggested to be key regulators of rabbit preimplantation development.…”

Extracellular vesicles and melatonin benefit embryonic develop by regulating reactive oxygen species and 5‐methylcytosine

“…During the preimplantation stage of embryos, paternal and maternal genomes undergo dramatic reprogramming changes, particularly embryonic genome activation (EGA) . EGA of rabbit embryos generally occurs at the 8‐cell stage and is characterized by strict temporal and stage‐specific gene regulation . In particular, DNA methylation, an important epigenetic modification, plays a crucial role in regulating gene expression and other biological processes .…”

“…36 EGA of rabbit embryos generally occurs at the 8-cell stage and is characterized by strict temporal and stage-specific gene regulation. 32,37 In particular, DNA methylation, an important epigenetic modification, plays a crucial role in regulating gene expression and other biological processes. 38 DNA methylation states are altered by in vitro culture conditions of preimplantation stage rabbit embryos.…”

The Oviductal Extracellular Vesicles’ RNA Cargo Regulates the Bovine Embryonic Transcriptome

“…One of the highly connected downregulated genes was NANOG, well-known as a pluripotency-sustaining factor in embryonic stem cells [63]. In rabbits, lower NANOG expression has been found in in vivo embryos at the 16 cell, morula, and blastocyst stages compared to the same stages in in vitro produced embryos [64]. Since NANOG expression is restricted to the embryoblast (EB) and is repressed in trophectoderm (TE) cells [65], this lower expression in the total blastocyst could derive from a different ratio of the number of EB and TE cells at a given stage in in-vitro-produced and in vivo embryos.…”