To efficiently produce high-quality bovine calves by transferring embryos obtained from in vitro fertilization (IVF), it is important to evaluate their ability to conceive and their ability to develop into normal litters. In this study, we aimed to clarify the effects of timing and morphology of blastomere cleavage on the gene expression status of bovine IVF embryos. Bovine IVF embryos were classified in four categories, which were divided according to the time of the first blastomere cleavage and the presence or absence of direct cleavage. In addition, the gene expression profiles of these embryos were examined. The timing and morphology of the blastomere cleavage was involved in pre-implantation development and gene expression status of bovine IVF embryos. Our results indicate the possibility of multiple evaluations for bovine IVF embryos and the selection of the most suitable embryos for embryo transfer.

The conventional ovum pick-up method requires oocytes to be transported from local farms to the laboratory, where they undergo nuclear maturation. However, atmospheric conditions for oocyte transportation differ from those for normal oocyte maturation in vitro. In this study, we examined the effects of conventional and modified oocyte transport conditions on oocyte quality and subsequent embryonic development. Cumulus-oocyte complexes were collected from slaughterhouse-derived bovine ovaries and cultured in few drops of medium on plastic plates in a CO₂-incubator (Control), in plastic tubes containing medium (C-T) in air, or in tubes containing gellan gum and medium (MC-T) in air. C-T conditions reduced mitochondrial functionality (mitochondrial membrane potential and adenosine triphosphate), lipid content, and DNA methylation but increased mitochondrial DNA copy number and phosphorylated AMP-activated protein kinase (P-AMPK) levels compared to those in control oocytes. Furthermore, RNA sequencing analysis of blastocysts derived from these oocytes revealed that C-T conditions affected mitophagy- and AMPK-signaling-related genes. However, MC-T conditions attenuated these C-T-associated changes. In conclusion, conventional C-T conditions affect oocyte metabolism and alter embryo quality, whereas the use of gellan gum as a substrate ameliorates such adverse effects. The oocyte transportation system is inadequate for embryonic production and can induce epigenetic changes. Modifying these conditions with gellan gum is a useful counter-measure.

Golden hamsters (Mesocricetus auratus) have been extensively used in biomedical research. With the advent of genome-editing technology, it is now possible to generate gene-knockout hamsters, providing unique research models that cannot be achieved with mice or rats. Therefore, the development of cryopreservation techniques for hamster embryos is in high demand. In this study, we present a simplified vitrification protocol for hamster embryo preservation. In vivo-derived 8-cell or morula embryos (Day 3) were vitrified using Cryotop in modified HECM-3 medium containing ethylene glycol, DMSO, and sucrose. After warming, the embryos were transferred into the uteri of Day 3-pregnant females with a different coat color. The results showed that 21–26% of the transferred embryos developed to the term. The experiments were conducted in a conventional laboratory setting, avoiding direct light exposure. Given the reproducibility of our vitrification protocol, it has broad applicability in laboratories that use hamsters.