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Journal of assisted reproduction and genetics2023; 40(11); 2565-2576; doi: 10.1007/s10815-023-02935-4

Use of confocal microscopy and intracytoplasmic sperm injection (ICSI) to assess viability of equine oocytes from young and old mares after vitrification.

Abstract: The impact of vitrification on oocyte developmental competence as a function of donor age remains an important issue in assisted reproductive technologies (ARTs). Methods: Equine germinal vesicle (GV) or metaphase II (M(II) oocytes were vitrified using the Cryotop® method. Spindle organization and chromosome alignment were evaluated from confocal imaging data sets of in vivo (IVO) or in vitro (IVM) matured oocytes subjected to vitrification or not. Intracytoplasmic sperm injection (ICSI) from the same groups was used to assess developmental potential. Results: An increase in chromosome misalignment was observed in spindles from older mares when compared to those of younger mares (P < 0.05). When MII oocytes subjected to vitrification were examined following warming, there was no difference in the percentage of oocytes displaying chromosome misalignment. Next, GV oocytes, collected from the ovaries of younger and older mares, were compared between fresh IVM and IVM following vitrification and warming. For nonvitrified samples, an age difference was again noted for spindle organization and chromosome alignment, with a higher (P < 0.05) percentage of normal bipolar meiotic spindles with aligned chromosomes observed in nonvitrified oocytes from young versus older mares. Vitrification led to a reduction of spindle length (P < 0.05) for oocytes from old mares, whether vitrified at GV or MII stages, whereas this effect was not observed in oocytes from young mares except those vitrified at GV and subjected to IVM. Oocyte developmental potential after vitrification was evaluated after ICSI of vitrified and warmed MII or GV oocytes from young mares. From 25 MII oocytes, 18 oocytes were injected with sperm, and six blastocysts were produced, which, upon transfer to mares' uteri, resulted in four pregnancies. Immature (GV) oocytes collected from live mares were also vitrified, warmed, and matured in vitro before ICSI. In this group, nonvitrified, control, and vitrified oocytes did not differ (P > 0.05) with respect to the incidence of maturation to MII, cleavage after ICSI, or blastocyst development. Conclusions: These findings demonstrate an effect of maternal age in an equine model at the level of meiotic spindle integrity and chromosome positioning that is influenced by both the meiotic stage at which oocytes are vitrified and whether meiotic maturation occurred in vivo or in vitro.
© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.
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Publication Date: 2023-09-19 PubMed ID: 37725179PubMed Central: PMC10643763DOI: 10.1007/s10815-023-02935-4Google Scholar: Lookup
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Summary

This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.

The research article focuses on understanding the effect of vitrification on the viability of equine oocytes – female reproductive cells – taken from both young and old mares. Using techniques of confocal microscopy and intracytoplasmic sperm injection (ICSI), the study found that maternal age had an impact on the integrity of meiotic spindles and chromosome positioning in the oocytes.

Objective of the Study

  • The primary objective of this study was to explore the impact of vitrification, a rapid freezing technique used in assisted reproductive technologies (ARTs), on the competency of equine oocytes derived from mares of different age groups. The research also aimed to compare the spindle organization and chromosome alignment of both in vivo (occurred naturally) and in vitro (occurred artificially) matured oocytes.

Methodology

  • The researchers used equine germinal vesicle (GV) or metaphase II (M(II) oocytes and subjected them to vitrification using the Cryotop® method.
  • Spindle organization and chromosome alignment were evaluated using confocal imaging, comparing in vivo (IVO) or in vitro (IVM) matured oocytes.
  • Intracytoplasmic sperm injection (ICSI) was performed to gauge the developmental potential of these oocytes.

Findings

  • The analyses showed a significant increase in chromosome misalignment in older mares compared to younger ones.
  • However, when MII oocytes subjected to vitrification were examined after warming, no difference was observed in chromosome misalignment, regardless of the age of the donor mares.
  • Vitrification caused a reduction of spindle length in oocytes from older mares, whether vitrified at GV or MII stages. This effect, however, was not observed in the oocytes from young mares.

Conclusion

  • The results suggest that maternal age has considerable effects on spindle integrity and chromosome positioning in equine oocytes. This impact is influenced both by the meiotic stage at which oocytes are vitrified and whether meiotic maturation occurred in vivo or in vitro.

Cite This Article

APA
Maclellan LJ, Albertini DF, Stokes JE, Carnevale EM. (2023). Use of confocal microscopy and intracytoplasmic sperm injection (ICSI) to assess viability of equine oocytes from young and old mares after vitrification. J Assist Reprod Genet, 40(11), 2565-2576. https://doi.org/10.1007/s10815-023-02935-4

Publication

Journal of assisted reproduction and genetics
ISSN: 1573-7330
NlmUniqueID: 9206495
Country: Netherlands
Language: English
Volume: 40
Issue: 11
Pages: 2565-2576

Researcher Affiliations

Maclellan, Lisa J
  • Department of Biomedical Sciences, Colorado State University, 1601 Campus Delivery, Fort Collins, Colorado, 80523, USA.
  • Seven Creeks Equine Reproduction, Euroa, 3666, Australia.
Albertini, David F
  • Bedford Research Foundation, Bedford, MA, 01730, USA.
Stokes, Joanne E
  • Department of Biomedical Sciences, Colorado State University, 1601 Campus Delivery, Fort Collins, Colorado, 80523, USA.
Carnevale, Elaine M
  • Department of Biomedical Sciences, Colorado State University, 1601 Campus Delivery, Fort Collins, Colorado, 80523, USA. Elaine.Carnevale@colostate.edu.

MeSH Terms

  • Animals
  • Horses
  • Male
  • Female
  • Vitrification
  • Sperm Injections, Intracytoplasmic / veterinary
  • Cryopreservation / veterinary
  • Cryopreservation / methods
  • Semen
  • Oocytes
  • Microscopy, Confocal

Conflict of Interest Statement

The authors declare no competing interests.

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