Extended ex vivo culture of fresh and cryopreserved whole sheep ovaries

We describe an original perfusion system for the culture of whole ovine ovaries for up to 4 days. A total of 33 ovaries were divided into six groups: control (n≤6), not perfused and fixed; Groups SM72 and SM72-FSH (n≤6 each), perfused with a simple medium for 72h with or without FSH; Groups CM96 and CM96-FSH (n≤6 each), perfused with a complex medium for 96h with or without FSH; Group CM96-FSH-cryo, (n≤3) cryopreserved and perfused for 96h with Group CM96-FSH medium. Depending on the medium used, morphological parameters of cultured ovaries differed from fresh organs after 72 (SM72, SM72-FSH) or 96 (CM96, CM96-FSH) h of perfusion. Oestradiol and progesterone were secreted in all groups but FSH had an effect only on Group CM96-FSH, stimulating continued oestradiol secretion 10 times higher than in all other groups. Morphological parameters and hormone secretion of cryopreserved ovaries were not different from fresh controls. This method enables the culture of whole ovaries for up to 4 days, the time required in vivo for 0.5-mm follicles to grow to 2.2mm and then for these follicles to reach the ovulatory size of 4mm or more. It could be used as a research tool or to complement current techniques for preserving female fertility.