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Preservation of caprine preantral follicle viability after cryopreservation in sucrose and ethylene glycol

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Abstract

Caprine preantral follicles within ovarian fragments were cryopreserved in the absence or presence of 0.5 M sucrose with or without 1 M dimethyl sulfoxide and/or 1 M ethylene glycol (EG). After being thawed, they were washed in minimum essential medium with or without 0.3 M sucrose. Histological analysis of follicle integrity immediately after cryopreservation showed consistent beneficial effects of including sucrose in the three cryoprotectant solutions analyzed when tissue was thawed without sucrose (53.9±14.8–82.4±3.2% normal vs 27.6±1.6–36.6±6.5%, P<0.05). However, in further studies, the addition of sucrose to the thaw solutions proved detrimental or of no benefit. An analysis of the cryopreserved material with calcein-AM and ethidium homodimer (markers for living and dead cells, respectively) gave comparable results to those obtained by histology. Follicles cryopreserved in EG, EG plus sucrose, or sucrose alone were cultured in vitro for 24 h following warming. During this culture period, viability fell most rapidly in material cryopreserved in sucrose alone and was no longer correlated with either the viability or integrity estimates made immediately after warming. By contrast, the viability of follicles cryopreserved in EG with sucrose and then cultured for 24 h was not significantly different from the cultured non-frozen controls. These results indicate that cryopreservation in 1 M EG plus 0.5 M sucrose combined with thawing without sucrose is effective for caprine ovarian tissue.

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Acknowledgements

The authors thank Peter Ursem, Arend Rijneveld, Ineke Daemen, Frans van Kooi, and the technicians from the Department of Pathobiology for their help.

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Authors and Affiliations

  1. Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands

    R. R. Santos & R. van den Hurk

  2. Department of Equine Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands

    T. Tharasanit

  3. Laboratory of Manipulation of Oocytes Enclosed in Preantral Follicles - LAMOFOPA, Faculty of Veterinary, State University of Ceará, Fortaleza, CE, Brazil

    R. R. Santos & J. R. Figueiredo

  4. Department of Biochemistry and Cell Biology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands

    T. van Haeften

  5. P.O. Box 80151, NL-3584 TD, Yalelaan 7, Utrecht, The Netherlands

    R. van den Hurk

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  1. R. R. Santos
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  2. T. Tharasanit
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  3. J. R. Figueiredo
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  4. T. van Haeften
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  5. R. van den Hurk
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Corresponding author

Correspondence to R. van den Hurk.

Additional information

This work was supported by CAPES/Brazil. Regiane Rodrigues dos Santos is a recipient of a grant from FUNCAP of Brazil.

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Santos, R.R., Tharasanit, T., Figueiredo, J.R. et al. Preservation of caprine preantral follicle viability after cryopreservation in sucrose and ethylene glycol. Cell Tissue Res 325, 523–531 (2006). https://doi.org/10.1007/s00441-006-0193-5

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  • DOI: https://doi.org/10.1007/s00441-006-0193-5

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