Abnormal cleavage is involved in the self-correction of bovine preimplantation embryos.

Chromosome instability leading to aneuploidy during early cleavage is well known in humans and cattle. Partial compaction (PC), which occurs only in some blastomeres, is suggested as a self-correction mechanism through which human embryos avoid aneuploid mosaicism. Partially compacted embryos show abnormal cleavages more frequently during early development; however, the mechanism by which blastomeres are excluded has not been elucidated. Here, we confirmed PC in approximately half of the tested bovine embryos, similar to that in human embryos. DNA sequencing of single-cell and intact embryos revealed that the morulae that excluded some blastomeres had euploidy, but many of the excluded blastomeres had aneuploidy. Time-lapse imaging of zygotes without the zona pellucida revealed that the excluded blastomeres underwent reverse and direct cleavages, which are abnormal cleavages, more frequently than the blastomeres involved in compaction. These results suggest the potential role of abnormal cleavage in the self-correction mechanism during the development of mammalian preimplantation embryos.

Effect of Age and Morphology on Live Birth Rate After Cleavage Stage Embryo Transfer.

Accurate knowledge of the live birth rate for cleavage stage embryos is essential to determine an appropriate number of embryos to transfer at once. Results from previous studies lack details needed for practical use. This is a mathematical analysis and model building study of day 3 cleavage stage embryo transfers. A total of 996 embryos were transferred in 274 fresh and 83 frozen embryo transfers. Embryo morphology was divided into 4 groups based on number of cells and fragmentation percentage. Each embryo transfer was modeled as an equation equating the sum of the live birth rates of the transferred embryos to the number of live births that resulted. The least squares solution to the system of embryo transfer equations was determined using linear algebra. This analysis was repeated for ages 35 to 42 years old at oocyte retrieval. The best fit live birth rates per embryo in the age group centered on 35 years old were 29%, 13%, 10%, and 9% for embryos in the 8-cell with ≤ 5% fragmentation, 8-cell with > 5% fragmentation, 9-12 cell, and 6-7 cell groups, respectively. Cleavage stage embryos with fewer than 6 cells on day 3 had very low best fit live birth rates close to 0% at age 39 years and were excluded from the primary analysis to prevent overfitting. These live birth rates can be used with a simple embryo transfer model to predict rates of single and multiple gestation prior to a planned cleavage stage embryo transfer.

Laser assisted blastomere extrusion biopsy of in vitro produced cattle embryos-A potential high throughput, minimally invasive approach for sampling pre-morula and morula stage embryos.

Whilst adoption of in vitro production (IVP) of cattle embryos and subsequent biopsy for genetic evaluation is increasing, biopsy techniques primarily used were developed to sample in vivo-produced blastocysts. This study was conducted to develop a laser-assisted blastomere extrusion approach for rapid and minimal-invasive biopsy of IVP cattle embryos at pre-morula to morula stages of development (Day 5 or 6 post-fertilisation). Embryo development into blastocysts was not compromised when ≤3 cells were collected by blastomere extrusion on Day 5 (44.4 ± 4.4 % and 34.3 ± 4.6 %) or Day 6 (58.0 ± 4.3 % and 57.5 ± 5.3 %) post-fertilisation compared with non-biopsied control embryos. Similarly, capacity to withstand cryopreservation was not different between embryos biopsied at Day 5 and 6 post-fertilisation and control-embryos (58.8 ± 6.0 %, 63.5 ± 5.6 %, and 56.0 ± 4.8 %, respectively). When more cells were collected from embryos at Day 6 post-fertilisation (≥8 compared to ≤3 cells), subsequent embryo development was not different (63.6 ± 6.1 % and 73.1 ± 6.2 %, respectively) nor was the capacity to withstand cryopreservation (67.9 ± 9.0 % and 62.5 ± 8.7 %, respectively). For biopsies on Day 6 post-fertilization, 95 % of samples produced a PCR product; however, when compared to the whole embryo PCR results, approximately 11 % of biopsy-samples classified as being from a male embryo were from female embryos (false positive), indicating DNA contamination between samples. In conclusion, results of this study indicate laser-assisted blastomere extrusion is a time efficient and minimally invasive approach to biopsy IVP morula and pre-morula cattle embryos to facilitate genetic analysis.

High three pro-nuclei (3PN) zygotes proportion associated with normal embryo multinucleation at the two-cell stage: two cases report.

Objective: Blastomere multinucleation at the two-cell stage (MN2) is a common nuclear abnormality observed in early human embryos and known to decrease the implantation rate. The aim of this study is to explore whether or not there is a link between high 3PN zygotes proportion and MN2 incidence.Methods: For embryo culture in the conventional incubator, the evaluation of nuclear status of two-cell stage is usually not performed. Therefore, the MN2 phenomenon is easily ignored. The time-lapse monitoring system (TLS) offers a promising new method to evaluate embryo development. We reported two cases who had single 2PN zygote and more than four 3PN zygotes in the conventional in-vitro fertilization (c-IVF) cycle.Results: We observed the MN2 incidence in the single normal embryo by TLS which suggested that it might be resulted from high 3PN zygotes proportion incidence. No available embryo was obtained in the first c-IVF cycle and the intracytoplasmic sperm injection (ICSI) treatment was performed in the second cycle. In subsequent ICSI cycles, we observed no 3PN zygotes incidence and transferred two day 3 embryos for the patients. Finally, the two couples successfully obtained healthy babies.Conclusions: High 3PN zygotes proportion might be associated with the MN2 incidence.

Live birth following a single oocyte fertilized with ICSI and embryo biopsy at blastocyst stage: a case report.

In this report, we present a case of a couple who obtained a live birth with a single oocyte fertilized by ICSI. Two oocytes were collected at 35.5 hours (h) post trigger and both were at metaphase II when sperm injection was performed (38 h). At 18 h post injection, one oocyte was fertilized, developed to the blastocyst stage and underwent to trophectoderm biopsy for preimplantation genetic testing on day 5. Following biopsy, the blastocyst was vitrified and 4 h after warming procedure, the euploid blastocyst was replaced to the uterine cavity. Healthy live birth was obtained after 39 weeks of gestation.

The effect of short-term disturbance of day 3 embryo culture on the development and implantation.

This study is carried out to estimate the effect on embryo development and implantation potential performed by inevitably occurring short-term 3-day embryo culture condition disorders due to light microscopy applied for morphology parameter assay. One thousand two hundred and fifty-three IVF program results were analyzed to measure the pregnancy rate on embryo transfer and the percentage of embryos reached the blastocyst stage. In 58% of cases, on the 3rd day the quality of embryos was not evaluated (n = 730), while in 42% of cases the evaluation was done on day 3 (n = 523). Embryo development estimation on day 3 affects the pregnancy rate and implantation potential especially in patients of older age group. Additional embryo analysis also affects the number of blastocysts obtained. The pregnancy rate for 3rd and 5th day transfer does not differ. Additional analysis of the 3rd day embryos reduces the PR in patients of the older age group. Embryo culture condition variation has a considerable impact on obtained blastocyst number (i.e blastocyst formation) especially in patients of the older age group.

Developmental Potential and Clinical Value of Embryos with Abnormal Cleavage Rate.

To investigate the developmental potential and clinical value of embryos with abnormal cleavage rate, a retrospective analysis was performed on 66 635 2-prokaryotic (2PN) and 1-pronuclear (1PN) embryos. The embryos were given conventionally in vitro fertilization (IVF) treatment and continuously cultured on the day 3 (D3) at the Reproductive Medicine Center, Tongji Medical College, Huazhong University of Science and Technology from January 2016 to December 2017. The embryos were separated into the day-2 (D2) undivided group with 106 cases, the arrested development group with 3482 cases, the blastomere reduction group with 541 cases, and the control group with 62 506 cases, respectively. The blastocyst utilization rates of these three abnormal groups were 2.83%, 10.86% and 6.84%, respectively, which were significantly different from that in control group (39.46%). Furthermore, 2 cases of anabiosis and 1 case of live birth were found in D2 undivided group. In arrested development group, there were 55 cases of anabiosis, 11 cases of clinical pregnancy in single-embryo transplantation (including 6 cases of live birth), and 25 cases of clinical pregnancy in combination with one normal embryo transplantation (including 23 cases of live births and 15 cases of dizygotic twins under B-ultrasound). There were 13 case of anabiosis in blastomere reduction group: there was 1 case of single embryo transplantation and clinical pregnancy was obtained; there were also 6 cases of clinical pregnancy in combination with one single normal embryo transplantation (including 5 cases of live births and 2 cases of dizygotic twins under B-ultrasound). In conclusion, embryos with abnormal cleavage rate still have the potential to continue to develop, and have certain blastocyst utilization rate and live birth.

Are cleavage anomalies, multinucleation, or specific cell cycle kinetics observed with time-lapse imaging predictive of embryo developmental capacity or ploidy?

OBJECTIVE: To determine whether cleavage anomalies, multinucleation, and specific cellular kinetic parameters available from time-lapse imaging are predictive of developmental capacity or blastocyst chromosomal status. DESIGN: Retrospective analysis of prospectively collected data. SETTING: Single academic center. PATIENT(S): A total of 1,478 zygotes from patients with blastocysts biopsied for preimplantation genetic screening were cultured in the EmbryoScope. INTERVENTION(S): Trophectoderm biopsy. MAIN OUTCOME MEASURE(S): Embryo dysmorphisms, developmental kinetics, and euploidy. RESULT(S): Of the 767 biopsied blastocysts, 41.6% (95% confidence interval [CI], 38%-45%) were diagnosed as euploid. Individual dysmorphisms such as multinucleation, reverse cleavage, irregular chaotic division, or direct uneven cleavage were not associated with aneuploidy. Direct uneven cleavage and irregular chaotic division embryos did, however, exhibit lower developmental potential. The presence of two or more dysmorphisms was associated with an overall lower euploidy rate, 27.6% (95% CI 19%-39%). Early embryo kinetics were predictive of blastocyst development but not ploidy status. In contrast, chromosomal status correlated significantly with start time of blastulation (tSB), expansion (tEB), and the tEB-tSB interval. A lower euploidy rate, 36.6% (95% CI 33%-42%) was observed with tSB ≥ 96.2 hours, compared with 48.2% with tSB < 96.2 (95% CI 42%-54%). A drop in euploidy rate to 30% (95% CI 25%-37%) was observed in blastocysts with delayed expansion (tEB > 116). The proportion of euploid blastocysts was increased with tEB-tSB intervals of ≤13 hours. A logistic regression model to enhance the probability of selecting a euploid blastocyst was constructed. CONCLUSION(S): Morphokinetics may aid in selection of euploid embryos from a cohort of day 5/6 blastocysts.

Production of calves by the transfer of cryopreserved bovine elongating conceptuses and possible application for preimplantation genomic selection.

Preimplantation genomic selection based on single nucleotide polymorphism (SNP) genotypes is expected to accelerate genetic improvement in cattle. However, genome-wide genotyping at the early embryonic stage has several limitations, such as the technical difficulty of embryonic biopsy and low accuracy of genotyping resulting from a limited number of biopsied cells. After hatching from the zona pellucida, the morphology of the bovine embryo changes from spherical to filamentous, in a process known as elongation. The bovine nonsurgical elongating conceptus transfer technique was recently developed and applied for sexing without requiring specialized skills for biopsy. In order to develop a bovine preimplantation genomic selection system combined with the elongating conceptus transfer technique, we examined the accuracy of genotyping by SNP chip analysis using the DNA from elongating conceptuses (Experiment 1) and optimal cryopreservation methods for elongating conceptuses (Experiment 2). In Experiment 1, the call rates of SNP chip analysis following whole genome amplification in biopsied cells from two elongating conceptuses were 95.14% and 99.32%, which were sufficient for estimating genomic breeding value. In Experiment 2, the rates of dead cells in elongating conceptuses cryopreserved by slow freezing were comparable to those in fresh elongating conceptuses. In addition, we obtained healthy calves by the transfer of elongating conceptuses cryopreserved by slow freezing. Our findings indicate that the elongating conceptus transfer technology enables preimplantation genomic selection in cattle based on SNP chip analysis. Further studies on the optimization of cryopreservation methods for elongating conceptuses are required for practical application of the selection system.

Embryo biopsy and development: the known and the unknown.

Preimplantation genetic diagnosis (PGD) has been introduced in clinical practice as a tool for selecting 'healthy' embryos before their transfer in utero. PGD protocols include biopsy of cleaving embryos (blastomere biopsy (BB)) or blastocysts (trophectoderm biopsy (TB)), followed by genetic analysis to select 'healthy' embryos for transfer in utero. Currently, TB is replacing the use of BB in the clinical practice. However, based on the European Society of Human Reproduction and Embryology Preimplantation Genetic Diagnosis Consortium reports, BB has been used in >87% of PGD cycles for more than 10 years. An exhaustive evaluation of embryo biopsy (both BB and TB) risks and safety is still missing. The few epidemiological studies available are quite controversial and/or are limited to normalcy at birth or early childhood. On the other hand, studies on animals have shown that BB can be a risk factor for impaired development, during both pre- and postnatal life, while little is known on TB. Thus, there is an urgent need of focused researches on BB, as it has contributed to give birth to children for more than 10 years, and on TB, as its application is significantly growing in clinical practice. In this context, the aim of this review is to provide a complete overview of the current knowledge on the short-, medium- and long-term effects of embryo biopsy in the mouse model.

A single trophectoderm biopsy at blastocyst stage is mathematically unable to determine embryo ploidy accurately enough for clinical use.

BACKGROUND: It has become increasingly apparent that the trophectoderm (TE) at blastocyst stage is much more mosaic than has been appreciated. Whether preimplantation genetic screening (PGS), utilizing a single TE biopsy (TEB), can reliably determine embryo ploidy has, therefore, increasingly been questioned in parallel. METHODS: We for that reason here established 2 mathematical models to assess probabilities of false-negative and false-positive results of an on average 6-cell biopsy from an approximately 300-cell TE. This study was a collaborative effort between investigators at The Center for Human Reproduction in New York City and the Center for Studies in Physics and Biology and the Brivanlou Laboratory of Stem Cell Biology and Molecular Embryology, the latter two both at Rockefeller University in New York City. RESULTS: Both models revealed that even under best case scenario, assuming even distribution of mosaicism in TE (since mosaicism is usually clonal, a highly unlikely scenario), a biopsy of at least 27 TE cells would be required to reach minimal diagnostic predictability from a single TEB. CONCLUSIONS: As currently performed, a single TEB is, therefore, mathematically incapable of reliably determining whether an embryo can be transferred or should be discarded. Since a single TEB, as currently performed, apparently is not representative of the complete TE, this study, thus, raises additional concern about the clinical utilization of PGS.

Clinical outcome of cycles with oocyte degeneration after intracytoplasmic sperm injection.

There are variant rates of oocyte degeneration after intracytoplasmic sperm injection (ICSI) among different patients. Oocyte degeneration after ICSI may reflect the cohort of oocyte quality and subsequent embryo development capacity and clinical outcome. This retrospective study analyzed 255 cycles with at least one degenerated oocyte after ICSI (degeneration group) and 243 cycles with no degenerated oocytes after ICSI (control group). Basic characteristics like female age, body mass index, duration of infertility, hormone (FSH, LH, E2) levels on day 3 of menses, and primary infertility patient rate were similar between the two groups (p > 0.05). Total dose of gonadotropin and length of stimulation were also similar between the two groups (p > 0.05), but the degeneration group exhibited a more exuberant response to ovarian stimulation as reflected by more oocytes retrieved (p < 0.05). The number of 2PN embryos available and high quality embryos were similar between the two groups (p > 0.05), but the high quality embryo rate, early cleavage embryo rate, and available embryo rate were all statistically lower than the control group (p < 0.05). Embryo developmental kinetics seemed to be disturbed and embryo fragmentation rate increased in the degeneration group (p < 0.05). However, there was no statistical difference in the distribution of graded embryos transferred, and there were no statistical differences in the pregnancy rate, implantation rate, and abortion rate between the two groups (p > 0.05). We deduce that the presence of oocyte degeneration after ICSI may be associated with decreased embryo quality with embryo development kinetics disturbed. However, the clinical outcomes may not be affected if the premise that sufficient high quality degeneration group embryos are available for transfer.

Direct Unequal Cleavages: Embryo Developmental Competence, Genetic Constitution and Clinical Outcome.

OBJECTIVE: To investigate the prevalence, developmental potential, chromosomal constitution and clinical outcome of embryos with direct unequal cleavages (DUC). DESIGN: A retrospective observational study. SETTING: Academic Institution. PARTICIPANT: 21,261 embryos from 3,155 cycles cultured in EmbryoScope®. RESULTS: The total incidence of DUCs per embryo occupying the first three cleavages were 26.1%. Depending of the cell stage, DUC rate was 9.8% at first cleavage (DUC-1), 9.1% at second cleavage (DUC-2), and 3.7% at third cleavage (DUC-3) with 3.6% of embryos exhibiting multiple DUCs (DUC-Plus). The occurrence of DUCs was not correlated with female gamete age or source. The incidence of DUC-1 was significantly higher in embryos fertilized by epididymal and testicular sperm (13.6% and 11.4%, respectively) compared to ejaculated sperm (9.1%, all p<0.05). The total incidences of DUCs were strongly correlated with the onset of blastomere multinucleation (MNB) during the first three divisions. In MNB embryos, DUCs incidence are two to three times more likely to develop when compared to non-MNB embryos (OR = 3.11, 95% CI [2.64, 3.67] at 1-cell stage, OR = 2.64, 95% CI [2.39, 2.91] at 2-cell stage and OR = 2.51, 95% CI [1.84, 3.43] at 4-cell stage). The blastocyst formation rates gradually decreased from 61.0% in non-DUC to 40.2% in DUC-3, 18.8% in DUC-2, 8.2% in DUC-1 and 5.6% in multiple DUC embryos (DUC-Plus). The known implantation rates (FH) for day 3 (D3) transfers were 12.42% (n = 3172) in Non-DUC embryos, 6.3% (n = 127) in DUC-3, and 2.7% (n = 260) in DUC-2 embryos. No live births resulted from either DUC-1 (n = 225) or DUC-Plus (n = 100) embryo transfers. For blastocyst transfers, lower implantation rates (33.3%) but similar live birth (LB) rates (40%) were observed if DUC blastocysts were transferred. Comparatively rates in Non-DUC blastocyst were 45.2% and 34.8%, respectively. The euploid rate gradually increased from DUC-1, -2, -3 to Non-DUC (13.3%, 19.5%, 33.3%, 45.6%, p<0.001) for D3 biopsied embryos. Interestingly, the trend of decreased euploidy disappeared in DUC D5/6 biopsied embryos and similar rates were exemplified in DUC (D5 56.3%, D6 35.6%) vs. non-DUC (D5 51.4%, D6 33.8%) embryos. CONCLUSION: Blastocyst formation, implantation potential and euploid rate were significantly reduced in DUC embryos. DUC embryos should be deselected for D3 transfers, but should be culture to blastocyst stage for possible ET.

Cleavage-stage or blastocyst transfer: what are the benefits and harms?

ET is a critical step in an assisted reproduction cycle. Over the past decade there has been an increasing trend to extending culture from cleavage-stage to blastocyst transfer. There has also been a trend to single ET and reporting the success of an assisted reproductive cycle as a cumulative live-birth rate after using both fresh and frozen embryos. There is low evidence that fresh blastocyst transfer is associated with improved live-birth rates compared with fresh cleavage-stage embryos. However, in the few studies that report cumulative pregnancy rates after fresh and frozen transfers, no significant difference was found. Cleavage-stage transfer is associated with greater numbers of embryos available for freezing, and blastocyst transfer is associated with increased number of cycles with no embryos to transfer. Further well-designed studies are warranted to evaluate the outcomes for blastocyst transfer including cumulative live-birth rate after fresh and frozen transfers, time to live birth, costs of the different transfer strategies, and perinatal mortality and severe perinatal morbidity.

Karyomapping identifies second polar body DNA persisting to the blastocyst stage: implications for embryo biopsy.

Blastocyst biopsy is now widely used for both preimplantation genetic screening (PGS) and preimplantation genetic diagnosis (PGD). Although this approach yields good results, variable embryo quality and rates of development remain a challenge. Here, a case is reported in which a blastocyst was biopsied for PGS by array comparative genomic hybridization on day 6 after insemination, having hatched completely. In addition to a small trophectoderm sample, excluded cell fragments from the subzonal space from this embryo were also sampled. Unexpectedly, the array comparative genomic hybridization results from the fragments and trophectoderm sample were non-concordant: 47,XX,+19 and 46,XY, respectively. DNA fingerprinting by short tandem repeat and amelogenin analysis confirmed the sex chromosome difference but seemed to show that the two samples were related but non-identical. Genome-wide single nucleotide polymorphism genotyping and karyomapping identified that the origin of the DNA amplified from the fragments was that of the second polar body corresponding to the oocyte from which the biopsied embryo developed. The fact that polar body DNA can persist to the blastocyst stage provides evidence that excluded cell fragments should not be used for diagnostic purposes and should be avoided when performing embryo biopsies as there is a risk of diagnostic errors.

Morphokinetics as a predictor of self-correction to diploidy in tripronucleated intracytoplasmic sperm injection-derived human embryos.

OBJECTIVE: To describe, in morphokinetic terms, a tripronucleated embryo (TPN) population according to ploidy and to explore the value of such variables for predicting ploidy. DESIGN: Experimental. SETTING: In vitro fertilization laboratory. PATIENT(S): Seventy-nine TPN embryos obtained after intracytoplasmic sperm injection (TPN-ICSI) were cultured in a time-lapse incubator for 6 days. INTERVENTION(S): Ploidy determinations were carried out for 35 TPN-ICSI at the cleavage and/or blastocyst stage. Their morphokinetics were then retrospectively compared. MAIN OUTCOME MEASURE(S): Direct (cleavage time from 2- to 8-cell stages) and indirect (cell cycle duration and blastomere synchrony at cleavage) morphokinetic variables; ploidy determination by FISH; in vitro development to the blastocyst stage. RESULT(S): TPN-ICSI cleaved later than bipronucleated control embryos (BPN). Diploid TPN displayed morphokinetic behavior closer to BPN than triploid TPN regarding almost all of the direct and indirect morphokinetic variables measured. Variable t5 was found to be a predictable variable of ploidy in TPN. CONCLUSION(S): TPN-ICSI are not homogeneous in ploidy, cleavage, or morphokinetic terms. Diploid, but nontriploid, TPN are morphokinetically similar to diploid BPN. The ploidy of TPN can be predicted by variable t5.

Can repeated IVF-ICSI-cycles be avoided by using blastocysts developing from poor-quality cleavage stage embryos?

In many clinics, good-quality embryos are selected for embryo transfer and cryopreservation at the cleavage stage, and poor-quality embryos are discarded. The aim of this retrospective study was to examine how many repeated IVF cycles could be avoided by culturing the cleavage stage poor-quality embryos to blastocyst stage and transferring them after vitrification and warming (604 IVF and intracytoplasmic sperm injection [IVF-ICSI] cycles were included). Poor-quality cleavage stage embryos not eligible for transfer or cryopreservation were cultured until day 5 or 6, and those developing to the blastocyst stage were vitrified. The rate of vitrified blastocysts and clinical pregnancy and delivery rate of the warmed blastocysts was evaluated. The effect of the extended culture on the cumulative delivery rate, and the number of avoided new treatment cycles was calculated. The surplus blastocysts resulted in clinical pregnancy, spontaneous abortion and delivery rates of 24.6%, 27.3% and 17.2% respectively. The use of surplus blastocysts raised cumulative delivery rate from 43% to 47% and 53 repeated new cycles were avoided. This study shows that the cumulative delivery rate can be increased, and repeated IVF-ICSI treatments avoided by using blastocysts developing from poor-quality cleavage stage embryos, which otherwise would have been discarded.

Prevalence, consequence, and significance of reverse cleavage by human embryos viewed with the use of the Embryoscope time-lapse video system.

OBJECTIVE: To investigate the prevalence and potential causes of reverse cleavage (RC) by human early-cleavage embryos and its associations with embryonic development and implantation after transfer. DESIGN: Clinical retrospective cohort study. SETTING: Private fertility treatment center. PATIENT(S): A total of 126 consecutive in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI) treatment cycles, with 353 IVF and 436 ICSI embryos cultured in the Embryoscope until day 3. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Embryo assessment on day 3, incidence of abnormal division, embryo morphokinetic parameters, and fetal heart beat. RESULT(S): RC, referring to either blastomere fusion or failed cytokinesis, occurred up to three times per individual embryo in 27.4% of embryos during the first three cleavage cycles. A higher incidence was associated with GnRH antagonist cycles compared with agonist cycles (odds ratio [OR] 1.683), or with ICSI compared with IVF (OR 1.600). After ICSI, sperm progressive motility was associated with RC (area under the receiver operating characteristic curve: 0.573). Compared with RC-negative embryos, a lower proportion of RC-positive embryos reached 6-cell stage or beyond by day 3 (47.7% vs. 71.7%), and were more likely to have multinucleation at the 4-cell stage (10.1% vs. 5.0%). Embryos showing RC had significantly poorer performance in both conventional grading and morphokinetic parameters, and they implanted less (0/22 vs. 29/131) than those not showing RC. CONCLUSION(S): RC significantly compromised embryo development, culminating in poor implantation potential. For each embryo, it can occur on more than one occasion at any stage during the first 3 days of culture. It is associated with factors affecting both oocyte and sperm.

Changes in sex ratio from fertilization to birth in assisted-reproductive-treatment cycles.

BACKGROUND: In Western gender-neutral countries, the sex ratio at birth is estimated to be approximately 1.06. This ratio is lower than the estimated sex ratio at fertilization which ranges from 1.07 to 1.70 depending on the figures of sex ratio at birth and differential embryo/fetal mortality rates taken into account to perform these estimations. Likewise, little is known about the sex ratio at implantation in natural and assisted-reproduction-treatment (ART) cycles. In this bioessay, we aim to estimate the sex ratio at fertilization and implantation using data from embryos generated by standard in-vitro fertilization (IVF) or intracytoplasmic sperm injection (ICSI) in preimplantation genetic diagnosis cycles. Thereafter, we compare sex ratios at implantation and birth in cleavage- and blastocyst-stage-transfer cycles to propose molecular mechanisms accounting for differences in post-implantation male and female mortality and thereby variations in sex ratios at birth in ART cycles. METHODS: A literature review based on publications up to December 2013 identified by PubMed database searches. RESULTS: Sex ratio at both fertilization and implantation is estimated to be between 1.29 and 1.50 in IVF cycles and 1.07 in ICSI cycles. Compared with the estimated sex ratio at implantation, sex ratio at birth is lower in IVF cycles (1.03 after cleavage-stage transfer and 1.25 after blastocyst-stage transfer) but similar and close to unity in ICSI cycles (0.95 after cleavage-stage transfer and 1.04 after blastocyst-stage transfer). CONCLUSIONS: In-vitro-culture-induced precocious X-chromosome inactivation together with ICSI-induced decrease in number of trophectoderm cells in female blastocysts may account for preferential female mortality at early post-implantation stages and thereby variations in sex ratios at birth in ART cycles.