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Authors
Supervisors
Van Langendonckt, Anne
 ;
Donnez, Jacques
Abstract
Recent advances in cancer therapy have resulted in an increased number of long-term cancer survivors. Unfortunately, aggressive chemotherapy, ionizing radiotherapy and bone marrow transplantation can severely affect the ovarian follicular store and subsequently lead to a loss of fertility and premature menopause. The different options available for fertility preservation in cancer patients are embryo cryopreservation, oocyte cryopreservation and ovarian tissue cryopreservation. The choice depends on various parameters: the type and timing of chemotherapy, the type of cancer, the patient's age and the partner status. Most female cancer patients of reproductive age do not have the option of utilizing established assisted reproductive technologies as embryo cryopreservation to safeguard their fertility. Indeed, in many cancers, chemotherapy is initiated soon after diagnosis. A promising alternative to prevent fertility loss in these patients is the cryopreservation and transplantation of ovarian tissue. With the latest advances in cryobiology, ovarian tissue cryopreservation is rapidly becoming a more widely offered technique by many medical centers around the world. The indications now extend beyond cancer, as gonadotoxic chemotherapy is being used in a number of benign systemic diseases as well. Moreover, patients undergoing oophorectomy for benign ovarian conditions or for prophylaxis potentially may benefit from ovarian cryopreservation. Recent advances in orthotopic transplantation of cryopreserved ovarian tissue signal a promising future for ovarian tissue cryobanking. However, there are still many unresolved issues related to these technologies. Safeguarding patients from cancer cell reintroduction as well as minimizing ischemic tissue damage are critical issues for the successful clinical application of ovarian tissue cryobanking. Our objective is to offer young patients at risk of premature ovarian failure after gonadotoxic treatment, realistic and safe fertility preservation options. Our study focused on developments of cryopreservation and transplantation of ovarian tissue. In order to propose the most suitable transplantation procedure in each clinical situation (type of cancer, patient s age, risk of transmission), three options were investigated: 1) Cryopreservation and grafting of ovarian cortical fragments, the clinical application of which has been demonstrated. We reported the first live birth after orthotopic transplantation of cryopreserved ovarian tissue. To date, four patients have had their ovarian tissue reimplanted for malignant disease and one for benign disease. These results prove that autotransplantation of cryopreserved ovarian tissue is clinically applicable. However, this technique can only be proposed to selected patients at present. Further research is needed to minimize follicular loss after transplantation and develop new options for patients with cancer at risk of ovarian metastasis. 2) Cryopreservation of entire ovary with its vascular pedicle, reducing follicular loss due to ischemia. We showed that freezing an entire human ovary was possible, by perfusing the ovarian vessels with a cryoprotectant solution before freezing it with a passive cooling device. The protocol we applied ensured a high survival rate of follicles, small vessels and stromal cells after thawing, as well as a well-preserved ultrastructure and no signs of apoptosis. This was the first time that entire human ovaries had been cryopreserved in liquid nitrogen using an accessible cryopreservation protocol, and that survival of oocytes and granulosa cells could be proved. 3) Isolation and transplantation of ovarian follicles, in order to avoid transmission of malignant cells through the graft. We set up a protocol to digest human ovarian cortex using the Liberase enzyme blend to isolate primordial and primary follicles from ovarian cortical tissue. This optimized protocol yielded good quality isolated follicles (good morphology and integrity, and high viability), which is an absolute prerequisite for the further successful processing of these follicles, either for culture or transplantation. The Ficoll density gradient method we developed allows us to maximize the recovery of isolated human ovarian follicles and minimize the manipulation time, while maintaining high follicular viability. Our results demonstrate, for the first time, that isolated human primordial follicles are able to survive and grow after grafting. As this approach has successfully restored fertility to mice, our optimization of follicle isolation and recovery protocols now allows us to consider its development for humans. Our experimental study has highlighted certain areas that need to be addressed in order to continue to progress in this field. Further studies investigating whole ovary cryopreservation and vascular patency after transplantation, follicle isolation and follicle transplantation or culture may lead us to convert these techniques from research laboratory to clinical practice. ...
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Dolmans, M.-M. (2006). Human ovarian tissue cryopreservation and transplantation. https://hdl.handle.net/2078.5/112175