Vitrification of human ovarian tissue: a practical and relevant alternative to slow freezing

Reproductive Biology and Endocrinology

Florence Brugnon³  Laurent Janny³  Claude Darcha¹  Benoît Sion⁵  Johan Smitz²  Michel Canis⁴  Sandra Sanfilippo⁴ 

[1] CHU Clermont-Ferrand, CHU Gabriel Montpied, Anatomie et cytologie pathologiques, Centre de biologie, Clermont-Ferrand, France;Follicle biology laboratory, Vrije Universiteit Brussel, Jette, B - 1090, Belgium;Faculté de médecine, CNRS-UMR 6293/INSERM U1103, Laboratoire de génétique, reproduction et développement, Université d’Auvergne, Clermont-Ferrand, France;CHU Clermont-Ferrand, CHU Estaing, Pôle gynécologie obstétrique et reproduction humaine - Assistance médicale à la procréation - CECOS, Clermont-Ferrand, France;Laboratoire de physiologie, Université Clermont 1, UFR Pharmacie, Inserm U1107 NEURO-DOL, Clermont-Ferrand, France

关键词: DNA fragmentation;    Follicle morphology;    Conventional slow-freezing;    Vitrification;    Human ovarian tissue;    Cryopreservation;   

Others  :  1216369
DOI  :  10.1186/s12958-015-0065-5

 received in 2015-01-16, accepted in 2015-06-15,  发布年份 2015

【 摘 要 】

Background

Cryopreservation of ovarian tissue can be used to preserve the fertility of patients who are about to receive treatment(s) that could compromise their future ovarian function. Here we evaluate the effectiveness of a vitrification protocol by carrying out a systematic comparison with a conventional slow-freezing method on human ovarian tissue.

Methods

Human ovarian samples (mean age 28.0 ± 1.1 years) were processed in parallel for each cryopreservation procedure: vitrification and slow-freezing. Following warming/thawing, histological observations and a TUNEL assay in ovarian follicles were performed and compared to unfrozen control.

Results

Both cryopreservation protocols gave comparable histological outcomes. Percentage of intact follicles was 83.6 % following vitrification in a 1.5 M 1,2-propanediol (PrOH), 1.5 M ethylene glycol (EG) and 0.5 M raffinose solution, 80.7 % after slow-freezing in 1.5 M PrOH and 0.025 M raffinose, and 99.6 % in fresh tissue. Follicle density was unchanged by vitrification (0.6 follicles/mm2) or slow-freezing (0.5 follicles/mm2) compared to fresh tissue (0.7 follicles/mm2). Percentage of follicles with DNA fragmentation was not statistically different in vitrified (20.8 %) or slow-frozen (31.3 %) tissues compared to the unfrozen control (35.0 %). There was no difference in proportion of stroma cells with DNA fragmentation in vitrified (6.4 %) and slow-frozen (3.7 %) tissues compared to unfrozen tissue (4.2 %).

Conclusions

This vitrification protocol enables good preservation of ovarian quality post-warming. The evaluation of endocrine function after vitrification need to be perform in a higher cohort to evaluate if this protocol may offer a relevant alternative to conventional slow-freezing for the cryopreservation of human ovarian tissue.

【 授权许可】

   
2015 Sanfilippo et al.

【 图 表 】

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