Hormonal Characteristics of Women Receiving Ovarian Tissue Transplantation with or without Endogenous Ovarian Activity

Ovarian tissue cryopreservation (OTC) and transplantation of frozen/thawed ovarian tissue (OTT) are used for fertility preservation in girls and women. Here, we evaluated the hormonal characteristics of women with or without postmenopausal levels of FSH at the time of OTT to study differences and conditions that best support the initiation of ovarian function. A total of 74 women undergoing OTT (n = 51 with menopausal levels of FSH; n = 23 with premenopausal levels) were followed by measurements of FSH, LH, AMH, and oestradiol. Concentrations of FSH and LH returned to premenopausal levels after 20 weeks on average, with a concomitant increase in oestradiol. Despite resumption of ovarian activity, AMH concentrations were in most instances below the detection limit in the menopausal group, suggesting a low ovarian reserve. Despite a higher age in the premenopausal group, they more often experienced an AMH increase than the menopausal group, suggesting that conditions in the premenopausal ovary better sustain follicle survival, perhaps due to the higher concentrations of oestradiol. Collectively, this study highlights the need for improving follicle survival after OTT. Age and the amount of tissue transplanted are important factors that influence the ability to regain ovarian activity and levels of FSH may need to be downregulated and oestradiol increased prior to OTT.

Follicle Rescue From Prepubertal Ovaries After Recent Treatment With Cyclophosphamide—An Experimental Culture System Using Mice to Achieve Mature Oocytes for Fertility Preservation

Ovarian tissue cryopreservation is the only feasible method for fertility preservation in prepubertal girls that will undergo gonadotoxic chemotherapy. To date, the only clinical use of cryopreserved tissue is by a later tissue retransplantation to the patient. Clinical challenges in fertility preservation of very young patients with cancer include time constraints that do not allow to retrieve the tissue for cryopreservation before starting chemotherapy and the preclusion of future ovarian tissue transplantation due to the risk of reintroduction of malignant cells in patients with systemic diseases. To overcome these two challenges, we investigated using an experimental model the feasibility of retrieving secondary follicles from ovaries of prepubertal mice after cyclophosphamide (CPA) treatment in increasing doses of 50, 75, and 100 mg/kg. The follicles were thereafter cultured and matured in vitro. The main outcomes included the efficiency of the method in terms of obtained matured oocytes and the safety of these potentially fertility preservative procedures in terms of analyses of oocyte competence regarding normality of the spindle and chromosome configurations. Our findings demonstrated that it was feasible to isolate and culture secondary follicles and to obtain mature oocytes from prepubertal mice ovaries recently treated with CPA. The efficiency of this method was highly demonstrated in the 100 mg/kg CPA group, with near 90% follicle survival rate after 12 days’ culture, similarly to control. Around 80% of the follicles met the criteria to put into maturation, and more than 40% of them achieved metaphase II, with normal spindle and chromosome configurations observed. Suboptimal results were obtained in the 50 and 75 mg/kg CPA groups. These paradoxical findings towards CPA dose might probably reflect a more difficult selection of damaged growing follicles from ovaries recently treated with lower doses of CPA and a hampered ability to identify and discard those with reduced viability for the culture.

Equine ovarian tissue xenografting: Impacts of cooling, vitrification, and VEGF

Ovarian tissue transplantation methods using cooled and cryopreserved samples have been attractive options for fertility preservation in animal models and humans. The aim of this study was to evaluate the impact of previous exposure to cooling, cryopreservation, and VEGF exposure on the overall efficiency of equine ovarian tissue after heterotopic xenotransplantation in mice. The end points evaluated were follicular morphology and development, follicular and stromal cell densities, angiogenesis (i.e., density of new and mature blood vessels), collagen types I and III fiber densities, and total fibrosis. Ovaries of adult mares were harvested after ovariectomy and ovarian fragments were xenografted in the intraperitoneal wall of BALB nude mice. Ten types of treatments involving different combinations of cooling, cryopreservation, and xenografting procedures, and VEGF exposure were compared. The novel aspect of this study was the use of equine ovarian tissue xenotransplantation in mice, challenging the fragments with different combinations of treatments. The main findings were (i) cooling but not cryopreservation was effective in preserving the follicular morphology, (ii) a greater percentage of developing follicles but lower follicular and stromal cell densities were observed after ovarian tissue engraftment, (iii) exposure to VEGF increased new and mature vessels in cryopreserved transplanted tissue, and (iv) an appropriate balance in the collagen types I and III fiber ratio in cooling-transplanted tissue was observed after exposure to VEGF. This study contributes to advancing knowledge in preservation of ovarian tissue after cooling-cryopreservation and transplantation aiming to be applied to genetically superior/valuable horses, livestock, endangered animals, and, possibly, humans.