Coculture of monkey ovarian tissue increases survival after vitrification and slow-rate freezing.
OBJECTIVE
To assess whether coculture of monkey ovarian tissue after low-temperature storage enhances follicular viability. To assess a novel method of vitrifying ovarian tissue.


METHODS
Prospective in vitro study.


METHODS
University-affiliated national research center.


METHODS
Ovaries from 15 cynomolgus or rhesus macaques (1-11 years).


METHODS
Vitrification using a containerless liquid nitrogen emersion system that involves dropping thin cortical pieces suspended in cyroprotectant directly into liquid nitrogen with outcome compared with slow-rate-controlled freezing. Before analysis, some of the thawed tissue was cocultured on mitotically inactivated mouse fetal fibroblast monolayers supplemented with FSH, insulin, transferrin, and selenium.


METHODS
Percentage of oocytes viable using live-dead fluorescent staining with carboxyfluorescein diacetate succinimidyl ester and propidium iodide.


RESULTS
Postthaw survival rates were 70.4% +/- 4.8% of 1,705 follicles after vitrification and 67.3% +/- 1.9% of 1,895 follicles after slow-rate freeze in six trials with each method. Coculture of the thawed tissue increased the viability, respectively, to 89% +/- 2.1% of 2,833 follicles previously vitrified and to 90.3% +/- 1.9% of 2,109 follicles after a slow-rate freeze (P<.01). Primordial follicles (30- to 50-microm diameter) were the vast majority of surviving follicles after thaw and coculture. Follicular viability in control fresh tissue (eight trials) was 76.0% +/- 4.1%, suggesting negligible loss in follicular viability after cryopreservation.


CONCLUSIONS
Coculture of thawed ovarian tissue on mouse fetal fibroblasts and FSH increases the percentage of viable follicles. A novel method of vitrifying ovarian tissue is as effective as slow-rate freezing. These approaches may improve graft survival and function when used to treat chemotherapy-induced sterility.
Yeoman
Richard R
RR
Division of Reproductive Sciences, Oregon National Primate Research Center, Oregon Health and Sciences University, 505 185th Avenue, Beaverton, OR 97006, USA. yeomanr@ohsu.edu
Wolf
Don P
DP
Lee
David M
DM
D000818:Animals; D002470:Cell Survival; D018920:Coculture Techniques; D015925:Cryopreservation / Q000379:methods*; D005260:Female; D005347:Fibroblasts / Q000166:cytology; D008252:Macaca fascicularis; D008253:Macaca mulatta; D051379:Mice; D010053:Ovary / Q000166:cytology* / Q000637:transplantation