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Vajta G.,James Cook University | Rienzi L.,Genera Center For Reproductive Medicine | Bavister B.D.,Wayne State University
Reproductive BioMedicine Online

Sporadic reports published during the previous decade have documented pregnancies achieved with transfer of zona-free human embryos. Although the overall efficiency seems to be good and some authors have suggested systematic application for special infertility problems, there have been only a few attempts to compare the benefits of zona-free embryo culture and transfer with the traditional approach using zona-intact embryos. So far, the majority of instances in which zona-free culture has been applied have occurred accidentally. This review summarizes the known functions of the zona pellucida, analyses natural and artificial situations where its function is compromised, including zona hardening and difficult hatching that seem to be related to in-vitro embryo culture, and discusses possible methods and timing for artificial zona removal. With the availability of in-vitro systems capable of replacing important functions of the zona pellucida, routine use of zona-free culture for the whole in-vitro period, after or even before fertilization, is a realistic possibility with potential additional benefits. Based on the increasing amount of animal studies, a systematic comparison is suggested that may eventually diminish the handicaps of the in-vitro situation and lead to simplification of manipulations as well as higher success rates after embryo transfer. © 2010, Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved. Source

Vajta G.,Cairns Fertility Center | Rienzi L.,Genera Center For Reproductive Medicine | Cobo A.,University of Valencia | Yovich J.,Cairns Fertility Center | Yovich J.,Medical Center
Reproductive BioMedicine Online

Culture of preimplantation-stage embryos has always been a key element of laboratory embryology and has contributed substantially to the success of many assisted reproduction procedures. During the past decade, its importance has increased as extended in-vitro embryo culture and single blastocyst transfer have become indispensable parts of the approach to decreasing the chance of multiple pregnancy while preserving the overall efficiency of the treatment. However, in spite of the scientific and commercial challenge stimulating research worldwide to optimize embryo culture conditions, a consensus is missing even in the basic principles, including composition and exchange of media, the required physical and biological environment and even the temperature of incubation. This review attempts to summarize the controversies, demonstrate the fragility of some widely accepted dogmas and generate an open-minded debate towards rapid and efficient optimization. New approaches expanding the traditional frames of mammalian embryo culture are also discussed. Although some researchers suppose that the efficiency of the presently applied in-vitro culture systems have already approached the biological limits, authors are confident that substantial improvement may be achieved that may expand considerably the possibilities of future assisted reproduction in humans. © 2010, Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved. Source

Vajta G.,BGI Shenzhen | Vajta G.,Central Queensland University | Rienzi L.,Genera Center For Reproductive Medicine | Ubaldi F.M.,Genera Center For Reproductive Medicine
Reproductive BioMedicine Online

Vitrification is now the dominant approach for cryopreservation of human oocytes and embryos; however, serious disagreement persists, particularly about biosafety issues. Techniques are categorized as either 'open' or 'closed' according to occurrence of direct contact between the medium and liquid nitrogen during cryopreservation. Advocates of closed systems emphasize the potential danger of disease transmission mediated through liquid nitrogen, and praise the safety of their approach; those who use the open systems refer to the lack of evidence of disease transmission and regard their systems as more consistent and efficient. The purpose of this review is to clarify whether open and closed systems are really open and closed; if closed systems are safe and free of any danger of contamination; if closed systems are equally efficient as open ones for cryopreservation of human embryos and oocytes by considering overall outcome; and finally, if ethical and legal concerns are sound when risks and benefits are considered in a broader sense. On the basis of these answers, implementation of rational measures to lower the theoretical danger of disease transmission are proposed while maintaining the achievements in cryopreservation that have contributed substantially to the advancement in assisted reproduction techniques during the past decade. © 2014 Reproductive Healthcare Ltd. All rights reserved. Source

Rienzi L.,Genera Center For Reproductive Medicine | Vajta G.,BGI Shenzhen | Ubaldi F.,Genera Center For Reproductive Medicine

During the past decades, improvements in culture of preimplantation embryos have contributed substantially in the success of human assisted reproductive techniques. However, most efforts were focused on optimization of media and gas components, while the established physical conditions and applied devices have remained essentially unchanged. Very recently, however, intensive research has been started to provide a more appropriate environment for the embryos and to replace the rather primitive and inappropriate devices with more sophisticated and practical instruments. Success has been reported with simple or sophisticated tools (microwells or microchannels) that allow accumulation of autocrine factors and establishment of a proper microenvironment for embryos cultured individually or in groups. The microchannel system may also offer certain level of automation and increased standardization of culture parameters. Continuous monitoring of individual embryos by optical or biochemical methods may help to determine the optimal day of transfer, and selection of the embryo with highest developmental competence for transfer. This advancement may eventually lead to adjustment of the culture environment to each individual embryo according to its actual needs. Connection of these techniques to additional radical approaches as automated ICSI or an ultimate assisted hatching with full removal of the zona pellucida after or even before fertilization may result in devices with high reliability and consistency, to increase the overall efficiency and decrease the work-intensity, and to eliminate the existing technological gap between laboratory embryology work and most other fields of biomedical sciences. © 2011 Elsevier Ltd. All rights reserved. Source

Fiorentino F.,GENOMA Molecular Genetics Laboratory | Spizzichino L.,GENOMA Molecular Genetics Laboratory | Bono S.,GENOMA Molecular Genetics Laboratory | Biricik A.,GENOMA Molecular Genetics Laboratory | And 6 more authors.
Human Reproduction

Background: Fluorescence in situ hybridization (FISH) is the most widely used method for detecting unbalanced chromosome rearrangements in preimplantation embryos but it is known to have several technical limitations. We describe the clinical application of a molecular-based assay, array comparative genomic hybridization (array-CGH), to simultaneously screen for unbalanced translocation derivatives and aneuploidy of all 24 chromosomes. Methods: Cell biopsy was carried out on cleavage-stage embryos (Day 3). Single cells were first lysed and DNA amplified by whole-genome amplification (WGA). WGA products were then processed by array-CGH using 24sure arrays, BlueGnome. Balanced/normal euploid embryos were then selected for transfer on Day 5 of the same cycle. Results: Twenty-eight consecutive cycles of preimplantation genetic diagnosis were carried out for 24 couples carrying 18 different balanced translocations. Overall, 187/200 (93.5) embryos were successfully diagnosed. Embryos suitable for transfer were identified in 17 cycles (60.7), with transfer of 22 embryos (mean 1.3 ± 0.5). Twelve couples achieved a clinical pregnancy (70.6 per embryo transfer), with a total of 14 embryos implanted (63.6 per transferred embryo). Three patients delivered three healthy babies, during writing, the other pregnancies (two twins and seven singletons) are ongoing beyond 20 weeks of gestation. Conclusions: The data obtained demonstrate that array-CGH can detect chromosome imbalances in embryos, also providing the added benefit of simultaneous aneuploidy screening of all 24 chromosomes. Array-CGH has the potential to overcome several inherent limitations of FISH-based tests, providing improvements in terms of test performance, automation, sensitivity and reliability. © 2011 The Author. Source

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