Morphological and ultrastructural evaluation of cultured frozen-thawed human fetal ovarian tissue.

Objective: To investigate a defined culture condition for the culture of frozen-thawed human ovarian tissue.

Design: Prospective laboratory study.

Setting: Reproductive biology laboratories in university hospitals.

Patient(s): Fetal ovarian tissue from elective termination of pregnancy.

Intervention(s): Culture of frozen-thawed fetal ovarian tissue for up to 63 days.

Main Outcome Measure(s): Morphology, morphometry, and survival of follicles in relation to culture times.

Result(s): The proportion of primordial, early primary, and primary follicles in frozen-thawed (day 0) ovarian tissue was 77.5%, 21.7%, and 0.8%, respectively. Pronounced degeneration was found in all cell types, and < or =36% of the follicles had signs of atresia at days 7-14, but this figure improved with culture time to <20% of the total follicular population. After 7-14 and 21-35 days of culture, the relative proportion of the follicles in the different classes remained nearly stable. Morphometric examination of healthy follicles showed a significant increase in both follicle and oocyte diameter compared with control. A few follicles had developed to the early secondary stage. Ultrastructural analysis demonstrated well-preserved morphological integrity of healthy primordial and early primary follicles. Immunohistochemical localization of proliferating cell nuclear antigen was positive in proliferating follicular cells at days 7-14 and 21-35 of culture.

Conclusion(s): The present culture condition leads to good survival and progressive follicular growth and differentiation that is comparable to the physiological pattern of early folliculogenesis.