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Can frozen-thawed human ovary withstand refreezing-rethawing in the form of cortical strips?

  • Fertility Preservation
  • Published:
Journal of Assisted Reproduction and Genetics Aims and scope Submit manuscript

Abstract

Purpose

The aim of this study was to elucidate whether ovarian tissue is able to withstand a double freezing-thawing procedure.

Methods

Human ovarian cortical biopsies from 4 thawed whole ovaries were divided into 4 experimental subgroups: (a) frozen-thawed non-grafted group, (b) frozen-thawed xenografted group, (c) refrozen-rethawed non-grafted group, and (d) refrozen-rethawed xenografted group. Xenografting was performed using 8 severe combined immunodeficient mice for a total duration of 21 days. The following analyses were conducted: classic hematoxylin and eosin staining, Ki67 immunolabeling, transmission electron microscopy, Masson’s green trichrome, and double CD34 immunostaining.

Results

Morphologically normal preantral follicles were detected in all groups. We observed a dramatic decline of more than 65% in early preantral follicle survival rates after grafting of both frozen-thawed (p < 0.0001) and refrozen-rethawed (p < 0.0001) ovarian tissue. However, mean follicle densities remained comparable between the frozen-thawed and refrozen-rethawed non-grafted groups, as well as both grafted groups. Equivalent proportions of proliferating early preantral follicles were identified in frozen-thawed and refrozen-rethawed samples, whether the tissue was grafted or not. Furthermore, we did not observe any significant difference in atretic follicle rates between any of the four groups, and the ultrastructural quality of follicles appeared unaffected by the refreezing procedure. Similar proportions of fibrosis were noted in the frozen-thawed and refrozen-rethawed groups, irrespective of grafting. Finally, no significant differences were witnessed in terms of vascularization.

Conclusion

We were able to demonstrate, for the first time, that refrozen-rethawed ovarian tissue has the same functional characteristics as frozen-thawed ovarian tissue.

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Acknowledgments

The authors thank Mira Hryniuk for reviewing the English language of the article and Dolores Gonzalez, Olivier Van Kerk, Sarah Storder, and Alberte Lefèvre for their technical assistance. The authors also extend their thanks to Guillaume Courtoy for his assistance with Visiopharm® software.

Funding

This study was supported by grants from the Fonds National de la Recherche Scientifique de Belgique (Télévie grant 7.4602.18F awarded to C.H., F.R.S.-FNRS/FRIA FC29657 awarded to L.C., Télévie grant 7.4590.16 to R.M., 5/4/150/5 grant to M.M.D., EOS grant 30443682 and FNRS-PDR Convention T.0077.14), the Fonds Spéciaux de Recherche, the Foundation Against Cancer (grant 2018-042 awarded to A.C.), and donations from the Ferrero family and Philippe de Spoelberch.

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Authors and Affiliations

  1. Pôle de Recherche en Gynécologie, Institut de Recherche Expérimentale et Clinique, Université Catholique de Louvain, Avenue Mounier 52, bte. B1.52.02, 1200, Brussels, Belgium

    Camille Hossay, Alessandra Camboni, Luciana Cacciottola, Thu Y. T. Nguyen, Rossella Masciangelo & Marie-Madeleine Dolmans

  2. Anatomopathology Department, Cliniques Universitaires Saint-Luc, Avenue Hippocrate 10, 1200, Brussels, Belgium

    Alessandra Camboni

  3. Society for Research into Infertility, Avenue Grandchamp 143, 1150, Brussels, Belgium

    Jacques Donnez

  4. Gynecology Department, Cliniques Universitaires Saint-Luc, Avenue Hippocrate 10, 1200, Brussels, Belgium

    Marie-Madeleine Dolmans

Authors
  1. Camille Hossay
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  2. Alessandra Camboni
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  3. Luciana Cacciottola
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  4. Thu Y. T. Nguyen
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  5. Rossella Masciangelo
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  6. Jacques Donnez
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  7. Marie-Madeleine Dolmans
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Contributions

C.H.: Conception and design of the study, experimental procedures, analysis of results, statistical analysis, and article preparation. A.C.: Experimental procedures, analysis of results, and discussion contribution. L.C.: Experimental procedures and analysis of results and discussion contribution. T.Y.T.N.: Experimental procedures. R.M.: Experimental procedures and analysis of results. J.D.: Data evaluation, discussion contribution, and article revision. M.M.D.: Conception of the study, data evaluation, discussion contribution, and article revision.

Corresponding author

Correspondence to Marie-Madeleine Dolmans.

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The authors declare that they have no competing interests.

Ethics approval

The Institutional Review Board of the Université Catholique de Louvain (reference 2012/23MAR/125) approved the use of human ovarian tissue for this study after obtaining written informed consent from all subjects. Animal welfare guidelines were followed, and the protocol was approved by the Committee on Animal Research of the Université Catholique de Louvain (reference 2018/UCL/MD/40).

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Hossay, C., Camboni, A., Cacciottola, L. et al. Can frozen-thawed human ovary withstand refreezing-rethawing in the form of cortical strips?. J Assist Reprod Genet 37, 3077–3087 (2020). https://doi.org/10.1007/s10815-020-01960-x

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