Vitrification of mouse ovaries using ethylene glycol and DMSO as cryoprotectants: histopathological evaluation

Ten, 4to 6-week-old BALB/c mice were randomly assigned to either control (non-vitrified, n = 5) or treatment (vitrified, n = 5) groups. Ovaries in the vitrified group were frozen sequentially by immersion into two vitrification solutions VS1: 10% ethylene glycol (EG) + 10% DMSO in holding medium (TCM-199 + 20% FBS) and VS2: 20% EG + 20% DMSO in holding medium. After thawing at 37 °C in 1.0 M sucrose, vitrified as well as non-vitrified ovaries were serially sectioned and examined histopathologically. The proportion of atretic follicles between non-vitrified and vitrified samples was significantly different (36.5 vs. 78.9%, P<0.001). No statistical difference due to vitrification was observed for the percentage of small follicles between the two experimental groups. In contrast, the rate of atresia for the growing and antral follicles in the vitrified ovaries was statistically higher than in the non-vitrified group (70.1 vs. 30.6%, P<0.001). Although many antral follicles were atretic following vitrification, sufficient follicles, especially small class, survived. Therefore, vitrification using EG and DMSO is an efficient procedure for cryopreservation of ovaries.