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Quality and functionality of human ovarian tissue after cryopreservation using an original slow freezing procedure

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

Abstract

Purpose

To evaluate the efficiency of an original slow freezing protocol on the quality and function of human ovarian cortex.

Methods

Human ovarian tissues were cryopreserved using a freezing medium supplemented with propanediol and raffinose as cryoprotectants and antioxidants (L-glutamine, taurine). Samples were then frozen using a faster cooling rate than the usual one. Viability and morphology of follicles, DNA fragmentation in follicles and stroma as well as histology of the vascular endothelium were analyzed before and after freezing/thawing. Moreover, a functional analysis was performed based on the evaluation of follicular growth and development in thawed ovarian tissues that were cultured in vitro.

Results

Our freezing/thawing protocol allows preservation of a high proportion of viable follicles and the preservation of the different follicle developmental stages (p > 0.05 versus fresh control). 70.5 ± 5.2 % of follicles retained an intact morphology after cryopreservation (p = 0.04). Stroma cells but not follicles exhibited a slight increase of DNA fragmentation after thawing (p < 0.05). Microvessel endothelium within thawed tissues appeared to be preserved. Granulosa cells showed signs of proliferation in follicles cultured for 12 days. Secretion of 17β-oestradiol significantly increased during in vitro culture.

Conclusions

This protocol leads to good preservation of ovarian integrity and functionality post-thawing and thus appears as a suitable technique of ovarian tissue cryopreservation in clinical settings. Further research could be extended to optimize conditions of in vitro culture.

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Acknowledgments

We would like to thank all the surgical team in the department of gynecology, CHU Estaing, Clermont-Ferrand (France) for their help to recruit patients. We are grateful to Dr Wassim Essamet for his expertise regarding the histology, Emmanuel Bourgeois and Christine Artonne for their precious technical assistance and Mrs Elizabeth Petit for language revision of the manuscript. Thanks are also due to the women who donated tissue for this research.

This work is supported by an industrial PhD fellowship (Convention Industrielle de Formation par la Recherche, CIFRE) with the Centre International de Chirurgie Endoscopique (CICE), France (Grant No: 176/2009).

Conflicts of interest

The authors declare that there are no conflicts of interest.

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Authors and Affiliations

  1. Laboratoire de Biologie de la Reproduction, Université Clermont 1, UFR Médecine, E.A. 975, 28 Place Henri Dunant, 63001, Clermont-Ferrand Cedex 1, France

    Sandra Sanfilippo, Laurent Janny & Florence Brugnon

  2. Centre International de Chirurgie Endoscopique (CICE), 63001, Clermont-Ferrand Cedex 1, France

    Sandra Sanfilippo, Michel Canis & Jean-Luc Pouly

  3. Pôle Gynécologie obstétrique et Reproduction humaine, CHU Clermont-Ferrand, CHU Estaing, 63003, Clermont-Ferrand Cedex 1, France

    Michel Canis, Jean-Luc Pouly, Laurent Janny & Florence Brugnon

  4. Follicle Biology Laboratory, Vrije Universiteit Brussel, 1090, Jette, Belgium

    Sergio Romero & Johan Smitz

  5. Inserm U1107 NEURO-DOL, Laboratoire de Physiologie, Université Clermont 1, UFR Pharmacie, 63001, Clermont-Ferrand Cedex 1, France

    Benoît Sion

  6. Anatomie Pathologique, CHU Clermont-Ferrand, CHU Estaing, 63003, Clermont-Ferrand Cedex1, France

    Pierre Déchelotte

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  1. Sandra Sanfilippo
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  2. Michel Canis
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  3. Sergio Romero
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  4. Benoît Sion
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  5. Pierre Déchelotte
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  6. Jean-Luc Pouly
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  7. Laurent Janny
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  8. Johan Smitz
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  9. Florence Brugnon
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Corresponding author

Correspondence to Sandra Sanfilippo.

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Capsule An original slow freezing protocol able to protect both quality and functionality of human ovarian tissue, and easily applicable in a clinical setting.

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Sanfilippo, S., Canis, M., Romero, S. et al. Quality and functionality of human ovarian tissue after cryopreservation using an original slow freezing procedure. J Assist Reprod Genet 30, 25–34 (2013). https://doi.org/10.1007/s10815-012-9917-5

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  • DOI: https://doi.org/10.1007/s10815-012-9917-5

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