Vitrification of Porcine Oocytes and Zygotes in Microdrops on a Solid Metal Surface or Liquid Nitrogen

Somfai, T.; Kikuchi, Kazuhiro

doi:10.1007/978-1-0716-0783-1_21

Cited by 13 publications

(14 citation statements)

References 16 publications

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“…Vitrification of COCs was performed according to our previous reports [21,22] either before IVM or after IVF according to the experimental design (detailed below). The basic medium (BM) for vitrification and warming was a modified NCSU-37 [23] without glucose, but supplemented with 20 mM HEPES, 50 μM β-mercaptoethanol, 0.17 mM sodium pyruvate, 2.73 mM sodium lactate and 4 mg/ml BSA.…”

Section: Vitrification/warming Of Cocsmentioning

confidence: 99%

Vitrification of porcine immature oocytes and zygotes results in different levels of DNA damage which reflects developmental competence to the blastocyst stage

et al. 2023

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The present study investigated the effects of vitrification of porcine oocytes either at the immature Germinal Vesicle (GV) stage before in vitro maturation (GV-stage oocytes) or at the pronuclear stage after in vitro maturation and fertilization (zygotes) on DNA integrity in relevance with their subsequent embryo development. Vitrification at the GV stage but not at the pronuclear stage significantly increased the abundance of double-strand breaks (DSBs) in the DNA measured by the relative fluorescence after γH2AX immunostaining. Treatment of GV-stage oocytes with cryoprotectant agents alone had no effect on DSB levels. When oocytes were vitrified at the GV stage and subjected to in vitro maturation and fertilization (Day 0) and embryo culture, significantly increased DSB levels were detected in subsequent cleavage-stage embryos which were associated with low cell numbers on Day 2, the upregulation of the RAD51 gene at the 4–8 cell stage (measured by RT-qPCR) and reduced developmental ability to the blastocyst stage when compared with the non-vitrified control. However, total cell numbers and percentages of apoptotic cells (measured by TUNEL) in resultant blastocysts were not different from those of the non-vitrified control. On the other hand, vitrification of zygotes had no effect on DSB levels and the expression of DNA-repair genes in resultant embryos, and their development did not differ from that of the non-vitrified control. These results indicate that during vitrification GV-stage oocytes are more susceptible to DNA damages than zygotes, which affects their subsequent development to the blastocyst stage.

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Section: Vitrification/warming Of Cocsmentioning

confidence: 99%

Vitrification of porcine immature oocytes and zygotes results in different levels of DNA damage which reflects developmental competence to the blastocyst stage

et al. 2023

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“…Since the lipid removal procedure by micromanipulation is very labor intensive, a similar strategy was developed that separated the lipids through centrifugation but did not remove them from within the zona pellucida as a high-throughput method of cryopreservation for IVP embryos (Spate et al, 2013). An alternative method uses solid surface vitrification, whereby 50 oocytes or zygotes are moved through a series of equilibration and vitrification media and placed onto a cooling surface, such as aluminum foil, sitting on top of liquid nitrogen before being transferred to cryovials (Somfai and Kikuchi, 2021).…”

Section: Cryopreservationmentioning

confidence: 99%

Production of Pigs From Porcine Embryos Generated in vitro

Chen

Redel

et al. 2022

Front. Anim. Sci.

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Generating porcine embryos in vitro is a critical process for creating genetically modified pigs as agricultural and biomedical models; however, these embryo technologies have been scarcely applied by the swine industry. Currently, the primary issue with in vitro-produced porcine embryos is low pregnancy rate after transfer and small litter size, which may be exasperated by micromanipulation procedures. Thus, in this review, we discuss improvements that have been made to the in vitro porcine embryo production system to increase the number of live piglets per pregnancy as well as abnormalities in the embryos and piglets that may arise from in vitro culture and manipulation techniques. Furthermore, we examine areas related to embryo production and transfer where improvements are warranted that will have direct applications for increasing pregnancy rate after transfer and the number of live born piglets per litter.

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“…At the end of IVM, the COCs were treated with 0.1% (w/v) hyaluronidase in the collection medium for 2 min and then denuded by gentle pipetting using a fine glass pipette in the collection medium without hyaluronidase. To assess live/dead status, the denuded oocytes were evaluated under a stereomicroscope as described previously (Somfai & Kikuchi, 2021). Those with membrane damage and a brownish, faded cytoplasm were considered as dead oocytes.…”

Section: Ivm and The Assessment Of The Live/dead Status And Nuclear M...mentioning

confidence: 99%

Dibutyryl‐cAMP and roscovitine differently affect premature meiotic resumption and embryo development of vitrified immature porcine oocytes

Nguyen

Somfai

Hirao

et al. 2022

Animal Science Journal

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Vitrification and warming can trigger premature meiosis in immature porcine oocytes.Our aim was to compare the efficacies of two meiotic inhibitors, dibutyryl-cAMP and roscovitine for the meiosis synchronization during in vitro maturation (IVM) of porcine oocytes vitrified at the germinal vesicle (GV) stage. We first compared the efficacy of 1 mM dibutyryl-cAMP and 25 μM roscovitine on meiotic arrest during the first 22 h of IVM. Dibutyryl-cAMP could maintain the GV stage in 83.5% of oocytes; however, roscovitine was even more effective (96.6%), whereas only 17.4% of the oocytes remained at the GV stage without these additives. Temporal meiotic arrest for 22 h by roscovitine did not reduce the percentage of oocytes reaching the Metaphase II stage during subsequent IVM. However, after parthenogenetic stimulation or in vitro fertilization, subsequent embryo development to the blastocyst stage was compromised after roscovitine treatment, whereas dibutyryl-cAMP improved the percentage of blastocyst development. In conclusion, dibutyryl-cAMP could derogate but not completely prevent premature meiosis in vitrified oocytes, whereas roscovitine could more efficiently prevent it. However, for embryo production, the use of roscovitine was disadvantageous, whereas the use of dibutyryl-cAMP was beneficial.

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Vitrification of Porcine Oocytes and Zygotes in Microdrops on a Solid Metal Surface or Liquid Nitrogen

Cited by 13 publications

References 16 publications

Vitrification of porcine immature oocytes and zygotes results in different levels of DNA damage which reflects developmental competence to the blastocyst stage

Vitrification of porcine immature oocytes and zygotes results in different levels of DNA damage which reflects developmental competence to the blastocyst stage

Production of Pigs From Porcine Embryos Generated in vitro

Dibutyryl‐cAMP and roscovitine differently affect premature meiotic resumption and embryo development of vitrified immature porcine oocytes

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