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Preimplantation Genetic Diagnosis for Single Gene Disorders
Published in Carlos Simón, Carmen Rubio, Handbook of Genetic Diagnostic Technologies in Reproductive Medicine, 2022
Ana Cervero, Jose Antonio Martínez-Conejero, Lucía Sanz-Salvador, Claudia Gil-Sanchís, Maribel Sánchez-Piris, Laura Iñiguez Quiles
The presence of DNA in the blastocoel fluid and spent embryo culture media is a potential alternative for a non-invasive genetic assessment of preimplantation embryos (57). Several studies have been published using both types of samples and the spent culture media seems to be a better source of embryonic DNA (58). However, the amplification rates are lower and ADO rates are still higher than those observed using conventional cellular biopsy and consequently, the risk of misdiagnosis is elevated (59). Linkage analysis strategies may help to overcome these technical problems, but the potential presence of maternal DNA contamination could be a significant issue. Therefore, although there have been published promising results, more work needs to be done to define optimal methods for the collection and analysis of this extra-embryonic DNA before the clinical application can be a reality.
Preimplantation Genetic Testing for Structural Rearrangements
Published in Darren K. Griffin, Gary L. Harton, Preimplantation Genetic Testing, 2020
Recent studies have shown that blastocoel fluid and spent blastocyst medium could potentially serve as alternative sources for template DNA sampling for the diagnosis of both numerical and segmental aneuploidy of the early embryo [46–48]. For blastocoel fluid sampling alone, the concordance rate between the diagnosis results of trophectoderm samples from the same embryo is high (97.1%). The amplification rate (82%) is not ideal, however, and it may still require an invasive biopsy for a significant proportion of the embryos to have a confirmatory diagnosis of the segregational status of the chromosomes [47]. Assessment of spent culture medium sampling for PGT-SR has been successfully applied on a patient with a male carrier of a balanced translocation [46,XY,t(14;15)(q22;q24)] which resulted in the live birth of a chromosomally normal and a healthy baby [49]. Recently, a combination of both blastocoel fluid and spent culture medium made an improvement in the accuracy of the diagnosis. Jiao et al. [50] first systematically performed PGT-SR on mixed blastocoel fluid and spent blastocyst medium, trophectoderm (TE), and whole embryo samples while making significant modifications to the pre-existing multiple annealing and looping-based amplification cycle (MALBAC) techniques used for DNA amplification [50]. Using this combination, 41 blastocysts donated by 22 couples known to carry a chromosome rearrangement were successfully quantified by segmental abnormalities with high resolution (∼1 Mb), and in none of the samples was maternal contamination detected [50].
Regulation of Reproduction by Dopamine
Published in Nira Ben-Jonathan, Dopamine, 2020
After fertilization, the zygote undergoes several mitotic divisions and forms a solid ball, the morula, which travels down the oviduct toward the uterus (Figure 10.15). Upon reaching the 20–30 cell stage, the morula starts a process of differentiation and cavitation to form a blastocyst. The blastocyst, still surrounded by the ZP, is composed of three regions: (1) an inner cell mass, which will develop into the embryo; (2) a surrounding outer layer called the trophoblast, which will develop into the placenta; and (3) a fluid-filled cavity called the blastocoele. DA-producing pluripotent cells isolated from the inner cell mass of preimplantation monkey blastocysts provided evidence for a very early expression of DA, even before implantation [84]. Yet, what functions are fulfilled by this DA in implantation and/or in growth of the embryo, remain to be determined.
Influence of post-thaw culture duration on pregnancy outcomes in frozen blastocyst transfer cycles
Published in Systems Biology in Reproductive Medicine, 2023
Hui Ji, Shanren Cao, Hui Ding, Li Dong, Chun Zhao, Junqiang Zhang, Jing Lu, Xiuling Li, Xiufeng Ling
The selection criteria for thawed embryos before transfer must be carefully planned and rigorous to maximize the pregnancy outcomes. Normally, embryos are transferred after a short culture duration, that is, after thawing for 1–6 h (Ahlstrom et al. 2013; Ferreux et al. 2018; Tubbing et al. 2018). In contrast, some fertility centers use a long post-thaw culture interval (16–24 h) (Kang et al. 2013; Haas et al. 2016; Yang et al. 2016). One advantage of having the different protocols is the flexibility of laboratory workflow; embryos can be thawed in the afternoon one day before transfer on a working day or in the morning of a transfer day during weekends and holidays (Fang et al. 2016). Moreover, the uterine cavity is an optimal incubator, and in vitro embryo culture conditions cannot replicate the fallopian tube and uterine environments in vivo. As such, some blastocysts fail to develop in vitro but can show high implantation rates in vivo (Haas et al. 2018). Therefore, a short post-thaw culture protocol is more feasible. However, several publications have suggested that a short culture period might be insufficient to evaluate the developmental potential of blastocysts and that a long culture period could increase the degree of blastocoel expansion and provide more valuable information required for the selection of embryos (Guerif et al. 2003; Du et al. 2016; Minasi et al. 2016; Herbemont et al. 2018).
The age-related required number of zygotes estimated from prior clinical studies of preimplantation genetic testing for aneuploidy (PGT-A)
Published in Systems Biology in Reproductive Medicine, 2023
Tasuku Mariya, Takeshi Sugimoto, Takema Kato, Toshiaki Endo, Hiroki Kurahashi
Table 2 summarizes the prior studies used for our current calculations. Since the reported finding of Gardner and colleagues that transferring blastocyst stage embryos can improve the pregnancy rate (Gardner et al. 1998), many studies have further investigated this phenomenon. However, few subsequent reports have confirmed the development rate of zygotes to blastocysts by age group. The investigation by Kato et al. (2012) was relatively large-scale with detailed age grouping but was a single-center study using a minimal ovarian stimulation method. Warshaviak et al. (2019) described the development rate from a zygote to the 8-cell stage in two age groups. Although the relatively older report by Janny and Menezo (1996) satisfied the inclusion criteria for our current analyses, we used the most recent large-scale report by Sainte-Rose et al. (2021) to estimate the rate of zygotes that will develop a useful blastocyst. Useful blastocysts were defined morphologically, based on the expansion of the blastocoel cavity (B1–B6) and the number and cohesiveness of the inner cell mass (ICM) and trophectoderm (TE) cells (Gardner et al. 1998). Sainte-Rose et al. grouped blastocysts morphologically as ‘good’ (B3–B6, AA/AB/BA/BB) or ‘average’ (B3–B6, AC/CA/BC/CB) with regard to useful blastocysts in their investigation (Sainte-Rose et al. 2021).
Preimplantation genetic diagnosis (PGD) and genetic testing for aneuploidy (PGT-A): status and future challenges
Published in Gynecological Endocrinology, 2020
Romualdo Sciorio, Luca Tramontano, James Catt
In the late 1990s, it became standard practice to extend embryo culture up to the blastocyst stage [30]. At this point, the embryo is already differentiated into two distinct cell types: the inner-cell mass (ICM) that will develop and form the fetus, and the trophectoderm (TE) cells which will become the placenta. As mentioned earlier, in recent times, many IVF units have left cleavage stage biopsy, and have turned to TEB as this biopsy offers some advantages. First, 5-10 cells can be biopsied from the blastocyst and this makes the genetic diagnosis more reliable and less prone to errors [31,32]. Second, TEB is believed to have a smaller detrimental effect on embryo viability compared to biopsy on day-3. Finally, TE biopsy reduced the risk of mosaicism [25]. To perform TEB, it is required to make a hole in the ZP either on day-3 or on day-5, and to wait for the beginning of TE herniation, which represents the optimal time to start the biopsy. Recently, the biopsy of blastocoel fluid has been proposed by Palini et al. [33] as new source of embryonic genetic material (Figure 2). With the introduction of vitrification as a highly efficient cryopreservation method, it is possible to combine blastocyst biopsy with vitrification and genetic screening, followed by warming and transfer of euploid embryos in a subsequent cycle [34,35].