Establishment of a novel method for cryopreservation and thawing of the mouse ovary

Abstract

During cryopreservation of ovarian tissue, the conditions of freezing and thawing are big factors controlling the survival rate of oocytes obtained. However, the conditions and procedures as they pertain to ovarian follicles and oocytes have not been established. Thus, we tried to determine the appropriate freeze-thaw times using the vitrification method with ethylene glycol and DMSO as cryoprotective agents and dd Y female mouse ovaries. The maturity rate from GV to the metaphase-II (MII) stage was 62.8% with ethylene glycol and 69.3% using DMSO, while the controls (GV oocytes obtained from a fresh ovary) showed a maturation rate of 83.6% (46/55). MII oocytes obtained by culturing GV oocytes in vitro showed a 64.3% (18/28) fertility rate via in vitro fertilization and a developmental rate into a 2 cell stage embryo of 35.7% (10/28) and into a 4-cell stage, 7.1% (2/28). However, development beyond the 8 cell stage embryo was not observed. A significant difference was not recognized between control (fresh) and ovarian tissues that had been frozen/thawed with respect to their ability to produce hormones. It is concluded that the vitrification method was effective for both freezing ovarian tissues and preserving its functional ability (maturation and capacitation).

The use of a metal container for vitrification of mouse ovaries, as a clinical grade model for human ovarian tissue cryopreservation, after different times and temperatures of transport

“…Vitrification, as a choice of cryopreservation, has shown promising outcomes in terms of follicle and stromal preservation in rodents and other species including humans [1,2,4,10,13,16,17,19,21]. Despite these promising results, little attention has been paid to how the tissue should be handled before cryopreservation, to guarantee the best possible post-storage outcome.…”

The effect of leukemia inhibitory factor and coculture on the in vitro maturation and ultrastructure of vitrified and nonvitrified isolated mouse preantral follicles

“…Vitrification is a simple and suitable alternative technique to preserve mammalian ovaries for long-time storage (27,28). During the vitrification procedure, no ice crystals are formed and it allows rapid freezing of the samples; thus the tissues maintain their structure and morphology with minimal damage (29)(30)(31).…”

Preservation of female fertility in humans and animal species

“…Histological evaluations and assessment of the viability of oocytes grown in vitro from vitrified ovarian tissues have been published for a range of species including, for example, mouse (Tokieda et al, 2002;Migishima et al, 2003), sheep (Bordes et al, 2005) cow (Kagawa et al, 2009); goat (Santos et al, 2007, Carvalho et al, 2013; camel (Madboly et al, 2017) dog (Ishijima et al, 2006), nonhuman primate (Santana et al, 2012) and human (Suzuki et al, 2015). Collectively these results suggest that it is possible to maintain tissue integrity and oocyte health following super cooling and warming.…”