Cryopreservation of whole ovine ovaries with pedicles as a model for human: parameters of perfusion with simultaneous saturations by cryoprotectants.

BACKGROUND: The aim of this research was to study the effectiveness of perfusion of intact ovine ovaries with different rates of perfusion and time-period elapsed between extraction of these ovaries and the beginning of perfusion. METHODS: Ovaries were perfused through the arteria ovarica (ovarian arteries) with culture medium supplemented with 5% bovine calf serum, 6% dimethyl sulfoxide, 6% ethylene glycol, 0.15M sucrose, Indian ink, and 100 IU/mL heparin at room temperature (22 degrees C). In the first cycle of experiments, ovaries (n = 96) were perfused for 60 minutes just after extraction of ovaries at the following rates of perfusion (mL/hour): 150, 100, 75, 50, 25, 12.5, and 6.3. In the second cycle of experiments, ovaries (n = 26) were perfused at a rate of 25 mL/hour for 60 minutes after extraction of ovaries and their storage at room temperature for 2, 3, 4, and 5 hours, for groups 1, 2, 3, and, 4, respectively. Successful perfusion of blood vessels was detected visible by a blue coloration of the ovarian tissues. RESULTS: The first cycle of experiments showed that the optimal perfusion rates were 50 mL/hour and 25 mL/hour. In the second cycle of experiments, good perfusion of ovaries was established at a perfusion rate of 25 mL/hour for the ovaries of groups when 2 and 3 hours had elapsed after extraction. CONCLUSIONS: Effective perfusion of ovine intact ovaries with vascular pedicle was established using freezing medium at room temperature at the rate of perfusion of 25 mL/hour or 50 mL/hour. The ovaries must be perfused not later than 3 hours after the death of animals.