Extracellular matrix improves survival of both stored and fresh human primordial and primary ovarian follicles in long-term culture

• Published in: Human reproduction (Oxford) Vol. 12; no. 5; pp. 1032 - 1036
• Main Authors: Hovatta, O, Silye, R, Abir, R, Krausz, T, Winston, R M
• Format: Journal Article
• Language: English
• Published: Oxford Oxford University Press 01.05.1997
• Subjects: Adult; Biological and medical sciences; Biopsy; Birth control; Cryopreservation; Culture Techniques; Extracellular Matrix - physiology; Female; Follicular Atresia - physiology; Freezing; Gynecology. Andrology. Obstetrics; Humans; Medical sciences; Ovarian Follicle - cytology; Ovarian Follicle - physiology; Ovariectomy; Ovary - cytology; Ovary - pathology; Sterility. Assisted procreation; Time Factors; Human; Culture medium; Ovary; Cryopreservation; Female genital system; Assisted procreation; Tissue culture; Female; Extracellular matrix; Ovarian follicle; In vitro; Optimization
• ISSN: 0268-1161; 1460-2350
• DOI: 10.1093/humrep/12.5.1032

Ovarian cortical tissue was obtained during gynaecological operations by biopsy or after oophorectomy from 20 women aged 25-42 years. It was placed in organ culture, either fresh or following thawing after cryopreservation, for 1-4 months. The tissue was cut in slices 0.1-0.3 mm in diameter and transferred to 12 mm inserts in 24-well culture plates. These slices were cultured for 4-21 days in either alpha minimum essential medium (alpha-MEM) or Earle's balanced salt solution with added pyruvate. Both media were supplemented with 10% human serum, insulin, gonadotrophins and antibiotics. Half of the inserts were precoated with extracellular matrix (Matrigel). Histological samples revealed that there were viable, non-atretic, primordial, primary and secondary follicles in all the cultures. Mitoses were seen in the granulosa cells of the secondary follicles. Although the proportion of atretic follicles increased during culture, non-atretic follicles were still present after 21 days. After 4-11 days the proportion of viable follicles was significantly higher when cultured in Earle's solution supplemented with pyruvate, than when cultured in MEM (77 versus 38%, P < 0.001). In cultures with extracellular matrix the proportion of viable follicles was significantly higher after 10-15 days than it was without matrix (85 versus 19%, P < 0.001). Culture after thawing frozen ovarian tissue did not affect the density or the proportion of the viable follicles. Two-thirds of follicles in cryopreserved tissue were viable after 10-15 days in culture. The results indicate that it is possible to culture human primary and primordial follicles in vitro, and follicles in cryopreserved tissue are viable.