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Objective: The present study evaluated the efficiency of embryo development and pregnancy of somatic cell nuclear transfer (SCNT) embryos using different source-matured oocytes in Camelus dromedarius.Methods: Camelus dromedarius embryos were produced by SCNT using in vivo- and in vitro- matured oocytes. In vitro embryo developmental capacity of reconstructed embryos was evaluated. To confirm the efficiency of pregnancy and live birth rates, a total of 72 blastocysts using in vitro- matured oocytes transferred into 45 surrogates and 95 blastocysts using in vivo- matured oocytes were transferred into 62 surrogates by transvaginal method.Results: The collected oocytes derived from ovum pick up showed higher maturation potential into metaphase II oocytes than oocytes from the slaughterhouse. The competence of cleavage, and blastocyst were also significantly higher in in vivo- matured oocytes than in vitro- matured oocytes. After embryo transfer, 11 pregnant and 10 live births were confirmed in in vivo- matured oocytes group, and 2 pregnant and 1 live birth were confirmed in in vitro- matured oocytes group. Furthermore, blastocysts produced by in vivo-matured oocytes resulted in significantly higher early pregnancy and live birth rates than in vitromatured oocytes.Conclusion: In this study, SCNT embryos using in vivo- and in vitro-matured camel oocytes were successfully developed, and pregnancy was established in recipient camels. We also confirmed that in vivo-matured oocytes improved the development of embryos and the pregnancy capacity using the blastocyst embryo transfer method.

Background The evaluation of the DNA damage generated in cumulus cells after mature cumulus-oocyte complexes vitrification can be considered as an indicator of oocyte quality since these cells play important roles in oocyte developmental competence. Therefore, the aim of this study was to determine if matured cumulus-oocyte complexes exposure to cryoprotectants (CPAs) or vitrification affects oocytes and cumulus cells viability, but also if DNA damage is generated in cumulus cells, affecting fertilization and embryo development. Results The DNA damage in cumulus cells was measured using the alkaline comet assay and expressed as Comet Tail Length (CTL) and Olive Tail Moment (OTM). Results demonstrate that oocyte exposure to CPAs or vitrification reduced oocyte (75.5 ± 3.69%, Toxicity; 66.7 ± 4.57%, Vitrification) and cumulus cells viability (32.7 ± 5.85%, Toxicity; 7.7 ± 2.21%, Vitrification) compared to control (95.5 ± 4.04%, oocytes; 89 ± 4.24%, cumulus cells). Also, significantly higher DNA damage expressed as OTM was generated in the cumulus cells after exposure to CPAs and vitrification (39 ± 17.41, 33.6 ± 16.69, respectively) compared to control (7.4 ± 4.22). In addition, fertilization and embryo development rates also decreased after exposure to CPAs (35.3 ± 16.65%, 22.6 ± 3.05%, respectively) and vitrification (32.3 ± 9.29%, 20 ± 1%, respectively). It was also found that fertilization and embryo development rates in granulose-intact oocytes were significantly higher compared to denuded oocytes in the control groups. However, a decline in embryo development to the blastocyst stage was observed after CPAs exposure (1.66 ± 0.57%) or vitrification (2 ± 1%) compared to control (22.3 ± 2.51%). This could be attributed to the reduction in both cell types viability, and the generation of DNA damage in the cumulus cells. Conclusion This study demonstrates that oocyte exposure to CPAs or vitrification reduced viability in oocytes and cumulus cells, and generated DNA damage in the cumulus cells, affecting fertilization and embryo development rates. These findings will allow to understand some of the mechanisms of oocyte damage after vitrification that compromise their developmental capacity, as well as the search for new vitrification strategies to increase fertilization and embryo development rates by preserving the integrity of the cumulus cells.

In vitro maturation (IVM) of transvaginally aspirated immature oocytes is an effective and safe assisted reproductive treatment for predicted or high responder patients. Currently, immature oocytes are also being collected from the contralateral ovary during laparoscopy/laparotomy and even ex vivo from the excised ovary or the spent media during ovarian tissue preparation prior to ovarian cortex cryopreservation. The first live births from in vitro-matured ovarian tissue oocytes (OTO-IVM) were reported after monophasic OTO-IVM, showing the ability to achieve mature OTO-IVM oocytes. However, fertilisations rates and further embryological developmental capacity appeared impaired. The introduction of a biphasic IVM, also called capacitation (CAPA)-IVM, has been a significant improvement of the oocytes maturation protocol. However, evidence on OTO-IVM is still scarce and validation of the first results is of utmost importance to confirm reproducibility, including the follow-up of OTO-IVM children. Differences between IVM and OTO-IVM should be well understood to provide realistic expectations to patients.

Introduction: Priming with human chorionic gonadotrophin (hCG) before oocyte retrieval is a common strategy in oocyte in vitro maturation (IVM) treatment. Although this approach may have a beneficial effect on the number of oocytes retrieved and on oocyte maturation rates, hCG may disrupt important signaling processes at the oocyte-somatic cell interface during oocyte maturation in vitro, and there is no strong evidence that hCG triggering leads to higher pregnancy rates in patients with polycystic ovary syndrome (PCOS). In these patients, an important number of cumulus-oocyte complexes (COCs) obtained during oocyte retrieval are aspirated from small antral follicles with a mean diameter between 4 and 6 mm. We investigated whether oocytes derived from these small antral follicles can produce embryos that implant after maturation in an IVM system without hCG priming. Material and Methods: Between January and December 2010, 96 IVM-treatment cycles were performed at UZ Brussel in patients with PCOS. Patients were administered an average cumulative dose of 510 ± 190 IU uFSH or HPHMG daily before oocyte retrieval. When endometrial thickness was at least 5 mm, the oocyte retrieval was scheduled on the following day. No hCG trigger was given. In 33 of these cycles, all follicles were between 4 and 6 mm diameter and no dominant follicle was present at the moment of oocyte retrieval. COCs were retrieved and cultured in IVM culture medium suite (ORIGIO) for 38 hrs at 5% CO2 and 20% O2. After IVM, oocytes were denuded and ICSI was performed at one time point (one hour after denuding). Maturation rate, in vitro production of embryos and clinical outcomes after embryo transfer were evaluated. Results: The mean age of the patients was 28.5 ± 3.9 years. All COCs were retrieved from follicles with a diameter between 4 and 6 mm. In total, 587 COCs (with an average of 17.8 ± 10.2 COCs / cycle) were obtained, from which 287 (48.9%) were at metaphase II (average 8.7 ± 5.3 COCs / cycle) after 38 hours of IVM. The mean fertilisation rate was 63% (average 5.6 ± 3.3 / cycle). On day 3 after ICSI, 50% of embryos (average 2.9 ± 2.3 / cycle) had a good morphological quality (more than 6 cells and classified as grade 1 or 2) and were eligible for transfer. In five cycles (15%), no embryo transfer was performed due to poor embryo quality. In 28 cycles, an embryo transfer was performed on day 3 after ICSI, with an average of 1.29 ± 0.46 embryos per transfer. Four ongoing pregnancies (clinical pregnancy rate of 14.2%) were established and two healthy babies were born. Conclusion: PCOS patients represent a heterogeneous group. In a subset of these patients, follicular development is arrested at the small antral follicular stage, and follicles do not grow beyond 6 mm diameter despite uFSH or HP-HMG priming in IVM treatment cycles. In an IVM system without hCG-priming before oocyte retrieval, the oocytes derived from these small follicles can complete meiosis in vitro, be fertilised, and develop into healthy offspring. Further research is needed to optimise the clinical and laboratory protocol of IVM treatment, tailored to the specific follicular constellation of the ovary in patients with PCOS.