Original delayed-start ovarian stimulation protocol with a gonadotropin-releasing hormone antagonist, medroxyprogesterone acetate, and high-dose gonadotropin for poor responders and patients with poor-quality embryos

IntroductionThe delayed-start gonadotropin-releasing hormone antagonist protocol seems effective for patients who are poor ovarian responders, but there are insufficient data on whether it is also effective for patients with poor-quality embryos and low rates of good blastocyst formation. Specifically, the effectiveness of delayed-start gonadotropin-releasing hormone antagonists with progesterone has not been adequately investigated. Therefore, we compared the efficacy of the original delayed-start gonadotropin-releasing hormone antagonist protocol using medroxyprogesterone acetate (MPA) and high-dose gonadotropin in patients with poor ovarian response.MethodsOverall, 156 patients with recurrent assisted reproductive technology failure who underwent the original protocol were included. They received cetrorelix acetate (3 mg) and MPA (10 mg) on cycle day 3, and high-dose gonadotropin was initiated on day 11. When the leading follicle reached 14 mm, ganirelix acetate (0.25 mg) was administered until the trigger day. The number of oocytes retrieved, metaphase II (MII) oocytes, two pronuclear (2PN) zygotes, and good blastocysts and live birth rates were compared between the previous (Cycle A) and original (Cycle B) cycles in three groups (Group A, all patients; Group B, poor responders; and Group C, patients with poor-quality embryos).ResultsIn Group A (n=156), the number of MII oocytes (3.6 ± 3.3 versus 4.5 ± 3.6), 2PN zygotes (2.8 ± 2.9 versus 3.8 ± 3.1), good blastocysts (0.5 ± 0.9 versus 1.2 ± 1.6), and live birth rates (0.6 versus 24.4) significantly increased in Cycle B. Similar results were obtained in Group B (n=83; 2PN zygotes [1.7 ± 1.7 versus 2.3 ± 1.8], good blastocysts [0.4 ± 0.7 versus 0.9 ± 1.3], live birth rates [0 versus 18.1]) and Group C (n=73; MII oocytes [5.1 ± 3.8 versus 6.6 ± 4.0], 2PN zygotes [4.0 ± 3.4 versus 5.4 ± 3.4], good blastocysts [0.7 ± 1.1 versus 1.6 ± 1.9], and live birth rates [1.4 versus 31.5]).ConclusionThis original protocol increased the number of MII oocytes retrieved, 2PN zygotes, good blastocysts, and live birth rates in both poor responders and in patients with poor-quality embryos

Synchrony of the first division as an index of the blastocyst formation rate during embryonic development

Abstract Purpose To devise an uninvasive selection system for human embryos with high developmental potential after a single oocyte retrieval cycle by comparing the in vitro and in vivo effectiveness of first division synchrony against subsequent embryonic developmental stages. Methods The effects of using assisted reproductive technology on 948 embryos that were produced in 137 cycles were examined by dividing the embryos into “early cleavage” (first division within 25.90 hours) and “late cleavage” (first division at or after 25.90 hours) groups and comparing the blastocysts and good‐quality blastocyst formation rates between the two groups. These two groups were each divided further into “high synchrony” (first division synchrony within 3.96 hours) and “low synchrony” (first division synchrony at or after 3.96 hours) groups. The blastocysts, good‐quality blastocyst formation rates, and pregnancy rates were compared among these four groups. Results Both the blastocysts and good‐quality blastocyst formation rates were significantly higher in the early‐cleavage groups than in the late‐cleavage groups. The blastocyst formation rate of the latter was also significantly increased in the high‐synchrony, compared with the low‐synchrony, group. Conclusion First division synchrony in a single oocyte retrieval cycle could be a useful assessment of the blastocyst formation rate that enables the selection of viable embryos at an early stage of culture

A novel trophectoderm biopsy technique for all blastocyst stages

Abstract Purpose This study was conducted to assess the effectiveness of a new trophectoderm (TE) biopsy method that does not require prior opening of the zona pellucida at the blastocyst stage. Methods TE biopsy was conducted using a modified extrusion method for embryos during the cleavage stage. In this method, culture medium was injected into the perivitelline space to help extrude TE cells from the zona pellucida before TE biopsy. Results Our extrusion method preserves the embryo culture environment until immediately before biopsy because it does not require opening of the zona pellucida prior to TE biopsy. Furthermore, this method does not require a waiting time for blastocyst hatching after laser irradiation, thereby minimizing damage to the embryos and maintaining the time schedule of culture operations. Conclusions TE biopsy using this novel extrusion method may be useful in various applications, including the collection of TE cells for next‐generation sequencing analysis

The effects of differences in trophectoderm biopsy techniques and the number of cells collected for biopsy on next‐generation sequencing results

Abstract Purpose To examine how differences in trophectoderm biopsy techniques affect the frequency of mosaic embryos and sequencing results. Methods We examined differences in next‐generation sequencing (NGS) analysis results among operators or according to biopsy technique. Additionally, we determined the cut‐off for the number of collected cells to predict the occurrence of mosaicism. We collected cells according to the cut‐off value and examined whether there was a difference in the NGS analysis results between the pulling and flicking methods. Results There was no difference in the NGS analysis results among the operators. Regarding re‐biopsy, changes in the mosaic were observed in all specimens. The cut‐off value for the number of collected cells was five, and when more than five cells were collected, there was no difference in the NGS analysis results between the two methods. Conclusions We demonstrated that if trophectoderm biopsy techniques and NGS are stable, the cell collection location has a greater effect on NGS results than the biopsy technique

Production of Cloned Miniature Pigs Expressing High Levels of Human Apolipoprotein(a) in Plasma

<div><p>High lipoprotein(a) [Lp(a)] levels are a major risk factor for the development of atherosclerosis. However, because apolipoprotein(a) [apo(a)], the unique component of Lp(a), is found only in primates and humans, the study of human Lp(a) has been hampered due to the lack of appropriate animal models. Using somatic cell nuclear transfer (SCNT) techniques, we produced transgenic miniature pigs expressing human apo(a) in the plasma. First, we placed the hemagglutinin (HA)-tagged cDNA of human apo(a) under the control of the β-actin promoter and cytomegalovirus enhancer, and then introduced this construct into kidney epithelial cells. Immunostaining of cells with anti-HA antibody allowed identification of cells stably expressing apo(a); one of the positive clones was used to provide donor cells for SCNT, yielding blastocysts that expressed apo(a). Immunohistochemical analysis of tissue sections and RT-PCR analysis of total RNA from organs of cloned piglet revealed that apo(a) is expressed in various tissues/organs including heart, liver, kidney, and intestine. More importantly, a transgenic line exhibited a high level (>400 mg/dL) of Lp(a) in plasma, and the transgenic apo(a) gene was transmitted to the offspring. Thus, we generated a human apo(a)–transgenic miniature pig that can be used as a model system to study advanced atherosclerosis related to human disease. The anatomical and physiological similarities between the swine and human cardiovascular systems will make this pig model a valuable source of information on the role of apo(a) in the formation of atherosclerosis, as well as the mechanisms underlying vascular health and disease.</p></div