Viability of primordial follicles derived from cryopreserved ovine ovarian cortex tissue

Abstract

Objective: To develop an efficient technique to preserve primordial follicles from cryopreserved ovarian tissue.Design: Frozen-thawed and fresh preantral follicles were mechanically isolated for viability testing, and their morphology was histologically analyzed.Setting: Laboratory of Animal Reproduction, Faculty of Veterinary Medicine and Zootechny, University State of São Paulo, Brazil.Animal(s): Lambs 12-24 months of age.Intervention(s): Ovarian cortical fragments were prepared for cryoprotectant toxicity testing, freezing and thawing procedures, and in vitro culture.Main Outcome Measure(s): Histologic structure and follicular viability.Result(s): on day zero, no morphologic differences were observed between follicles isolated from fresh tissue and those treated with the cryopreservatives ethylene glycol (EG) and dimethyl sulfoxide (DMSO) and subjected to freezing. Even so, frozen follicles treated with DMSO + EG showed dark staining, indicating degeneration. on day zero, the follicular viability was similar between the control group (78.9%) and those treated with EG (77%) and frozen with EG (75%). After 10 days in culture, a reduced percentage of follicles was considered viable in all groups. This decrease was accentuated in those treated with DMSO (37.5% and 35.2% in those exposed to and frozen with DMSO, respectively) and DMSO + EG (33.9% and 30% in those exposed to and frozen with DMSO + EG, respectively) as compared with the control group (45%) and EG-treated groups (40.1% and 40% for those exposed to and frozen with EG, respectively).Conclusion(s): Ethylene glycol seems to be the best cryoprotectant for the cryopreservation of ovine ovarian tissue. (Fertil Steril (R) 2009;91:1976-83. (C)2009 by American Society for Reproductive Medicine.)