Dry biobanking has many advantages over the current paradigm of storing cryopreserved cells under liquid nitrogen. During drying, however, the cells become damaged. The highly condensed spermatozoa DNA has been shown in many desiccation studies to generally maintain its integrity. Using ram freeze-dried epididymal spermatozoa as a model, Palazzese et al. were the first to evaluate both single- and double-strand DNA breaks (SSBs and DSBs, respectively), showing that drying causes minimal DSBs but extensive SSBs (Palazzese L et al., 2018. DNA fragmentation in epididymal freeze-dried ram spermatozoa impairs embryo development, pp. 393–400). Furthermore, the authors also demonstrated that spermatozoa capable of directing embryo development to the blastocyst stage in vitro originated from rams with the least DNA damage Overall, the impact of sperm DNA damage on embryonic development depends on a balance between the extent of sperm DNA fragmentation, fragmentation type, and the oocyte’s repair capacity.
Cover Story: Oog1, an oocyte-specific gene, encodes the protein belonging to the leucine-rich repeat (LRR) superfamily. LRR is a motif involved in protein-protein interactions. Complete knockout of Oog1 is challenging because five copies of the Oog1 gene are present on chromosomes 4 and 12. Honda et al. generated Oog1 RNA interference (RNAi)-transgenic mice to investigate the effects of Oog1 knockdown on gene expression in the oocytes (Honda et al. Oocyte-specific gene Oog1 suppresses the expression of spermatogenesis-specific genes in oocytes, pp. 297–301). The abundance of spermatogenesis-specific transcripts was elevated in the Oog1 knockdown ovaries. In addition, a few abnormal oocytes were observed in 6-month-old Oog1 knockdown mouse ovaries. These findings suggested that OOG1 suppresses the expression of spermatogenesis-specific genes in the oocytes and plays important roles during oogenesis.
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Based on the results obtained for the studies of in vitro development of cloned embryos, epigenetic modifications have been widely used for cloning farm animals. However, such studies remain few in canids because of the lack of optimal in vitro oocyte maturation, embryo culture, and superovulation system. Kim et al. investigated whether a histone deacetylase inhibitor used in dog to pig interspecies somatic cell nuclear transfer (iSCNT), which improves nuclear reprogramming, could be used in dog cloning (Kim et al.: Suberoylanilide hydroxamic acid during in vitro culture improves development of dog-pig interspecies cloned embryos but not dog cloned embryos. p. 277–282). Porcine oocytes supported reprogramming of nuclei from dog fibroblasts up to early developmental stage of iSCNT embryos, and treating the embryos with suberoylanilide hydroxamic acid (SAHA) increased their developmental competence. However, unfortunately, SAHA treatment for dog to pig iSCNT embryos was not sufficient to improve their in vivo development because three and one clones were successfully produced from the control and SAHA treated groups, respectively.
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Recently, the effects of macromolecular crowding on cultured cells have gathered increasing interest. Crowded culture medium prepared by the addition of polyvinylpyrrolidone (PVP; Mw 360,000) positively influences the viability of oocytes during in vitro growth. Mizumachi et al. found that crowding affects a wide range of factors, including oocyte viability, complex morphology, and intimate conjunction of oocytes with cumulus/granulosa cells across the zona pellucida (Mizumachi et al. Macromolecular crowded conditions strengthen contacts between mouse oocytes and companion granulosa cells during in vitro growth, pp. 153-160). Confocal laser scanning microscopy indicated a higher number of transzonal processes (TZPs) reaching the oocyte from cumulus cells in 2% PVP medium than in 0% PVP medium.
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Whether cumulus cells are required for the successful vitrification of mature oocytes in cattle is controversial. Ishii et al. re-evaluated the effects of the presence of multi-layered cumulus cells (MCCs) on the vitrification of mature bovine oocytes (Ishii et al., Embryogenesis of vitrified mature bovine oocytes is improved in the presence of multi-layered cumulus cells. pp. 95–99). In the presence of MCCs, there was no difference between the embryonic development of fresh and vitrified mature bovine oocytes. These results suggest that MCCs protect mature oocytes from freezing, and promote their survival and development after in vitro fertilization. Ishii et al. successfully produced calves using a small number of vitrified mature oocytes with MCCs collected from the ovaries of individual cows post-slaughter.
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Okuyama et al. reported a transvaginal endoscopy-based technique for conducting ovarian examination in sows in a standing position. Sows were sedated in pig stalls, and their vaginal walls were punctured. Subsequently, a urethroscope was inserted into their abdomen, and an examination was conducted after their ovaries had moved towards the urethroscope camera via rectal palpation (Okuyama MW, et al. A transvaginal endoscopy-based technique for performing ovarian examinations in sows. pp. 617–622). This less invasive procedure without the use of general anesthesia may allow repeated ovarian examinations and increase our understanding of the ovarian dynamics in pigs.