Effective method for in-vitro culture of cryopreserved human ovarian tissue.

It is widely accepted that it is possible to successfully cryopreserve human ovarian tissue by direct plunging into liquid nitrogen using permeable cryoprotectants only, without disaccharides. This study aimed to search for and test a new method for in-vitro culture of vitrified tissue. Ovarian biopsies were obtained during operative laparoscopy. Pieces of ovarian tissue were vitrified and warmed. After warming, tissue pieces were randomly distributed into three groups for further culture: in 2 ml of culture medium which was regularly renewed (group 1), in 30 ml of culture medium without agitation (group 2) and in 30 ml of culture medium with agitation (group 3). During the 2-week and 6-week culture, the growth of follicles within the vitrified-warmed ovarian tissue pieces was investigated. After 2 weeks of culture, mean numbers of non-degenerated follicles per mm(2) of tissue were 1.5, 1.7 and 4.5 for groups 1, 2 and 3 respectively (groups 1 and 2 versus group 3, P < 0.05). Agitation during culture of ovarian tissue is beneficial, and can be used as a prognostic tool for future warming and autotransplantation of ovarian tissue.