132 BLASTOCYST PRODUCTION FROM BOVINE OVARIAN CORTEX FRAGMENTS XENOGRAFTED UNDER THE BACK SKIN OF MICE

Abstract

Chemo- or radiotherapy negatively affects the fertility of female patients undergoing oncological treatments. Ovaries are sensitive to such treatments, resulting in an increasing number of premature ovarian failures. Graft techniques are a promising alternative to preserve the fertility of such patients. So far, 35 birthed from human ovarian cortex autografts were reported in the literature; however, in this approach there is a risk of neoplastic reincidence. The aim of the present study was to evaluate the feasibility of ectopic ovarian cortex xenograft (using the bovine model) under the back skin of immunodeficient mice. Female SCID mice (~60 days, n = 38) were anesthetized with ketamine/xilazine and were placed on ventral decubitus. Ovarian cortex fragments from 8 cows (1.5 mm³; n = 152) were grafted through incisions made in the dorsal region (4 grafts per mouse). Ten days after ovarian fragments xenograft, the recipients were killed and the xenografts were harvested. The mice and grafts were weighed before and after the transplant. From the xenografts recovered, 88 were either routinely processed for histology (n = 26), to evaluate the progression of folliculogenesis, or sliced (n = 62) to recover the cumulus‐oocyte complexes, which were morphologically classified and used for in vitro embryo production, using standard procedures (in vitro maturation, fertilization, and embryo culture). The remaining grafts recovered (64) were stored in liquid nitrogen for future studies. Differences between means were compared using Student’s t-test. There was no difference between the body weight of recipient mice before and after xenograft (20.5 ± 0.4 v. 21 ± 0.8 g, respectively; P > 0.05). On the other hand, the grafts increased weight (11.6 ± 3.4 v. 14.8 ± 5.2 mg before and after transplant; P < 0.01). Histological analysis of the slices showed primordial, primary, multilaminar, antral, and atretic follicles, indicating the progression of folliculogenesis and neo-angiogenesis in the grafts. Twenty-four viable cumulus‐oocyte complexes were recovered from ovarian xenografts, from which 2 blastocysts were produced in vitro 8 days later (8.3% blastocyst rate). In summary, this study showed that ovarian xenografts were (i) healthily maintained under the back skin of immunodeficient mice, (ii) responsive to murine gonadotrophins, and (iii) able to produce viable cumulus-oocyte complexes that, (iv) by in vitro fertilization, can originate blastocysts. In general, our findings show the feasibility of the ovary xenograft as an alternative technique to fertility preservation in oncogenic patients, avoiding the risk of neoplastic re-incidence.

Study approved by Animal Experimentation Ethics Committee/FUSJ-009/15. Financial support was received from Fapemig and CNPq.