Embryo Cleavage Kinetics Research Articles

Are Early Embryo Cleavage Kinetics Affected by Energy Substrates in Different Culture Media?

This study aimed to investigate the influence of different culture media on early embryonic cleavage kinetics using time-lapse analysis and to determine the possible relationships between energy substrates in culture media and the cleavage kinetics. A total of 10 021 embryos from 1310 couples were cultured in time-lapse incubators. Embryos cultured in Vitrolife media were allocated to group I, and those in COOK media to group II. Embryo cleavage time points up to the 8-cell stage (t2-t8) were observed after pronuclei fading. The baseline demographic features, in vitro fertilization indications, ovarian stimulation protocol, oocyte-cumulus complexes, fertilization rate, together with pregnancy and perinatal outcomes were similar (P>0.05) between groups I and II. According to the time-lapse analysis, all embryos in group I showed significantly faster cleavage speed than those in group II (P<0.05). Furthermore, there was better synchrony in division (s3) and a longer length of the third cell cycle duration (cc3) in group II. Interestingly, implanted embryos in group II showed faster cleavage speed than those in group I, especially at t4 and t7. The glucose contents and multiple major amino acids were similar between the two groups. Lactic and pyruvic acid contents were generally higher in group I than those in group II. Because different commercial culture media may influence cleavage kinetics of embryos, it is essential for embryologists to take culture media into consideration in selecting a potential embryo when using a time-lapse system before implantation.

Stress Management during the Intracytoplasmic Sperm Injection Cycle May Slow Down First Embryo Cleavage and Accelerate Embryo Compaction: A Pilot Randomized Controlled Trial

Introduction: A firm consensus on the effectiveness of psychological interventions during infertility treatment has not been reached yet in terms of mental health and pregnancy rates. Moreover, the influence of these interventions on embryo cleavage kinetics has not been investigated. Objective: The aim of this work was to study whether stress management in couples undergoing an intracytoplasmic sperm injection (ICSI) cycle influences stress levels, mitochondrial DNA (mtDNA) levels in granulosa cells, and cleavage-stage embryos. Methods: Infertile couples were randomized into a treatment as usual (TAU) group (n = 30) and stress management program (SMP) group (n = 29) at the beginning of an ICSI cycle. Couples in the SMP group attended education and relaxation sessions at each visit to the clinic for folliculometry. The perceived stress scale (PSS) was used to assess stress levels at the beginning and end of the cycle. Moreover, mtDNA levels of granulosa cells and embryo morphokinetics were evaluated. Results: Post-intervention, women in the SMP group had significantly lower PSS scores than their initial PSS (p < 0.001; effect size, ES = 0.5) and than the final PSS of the TAU group (p = 0.02; ES = 0.09). Additionally, mtDNA levels were significantly lower in luteal granulosa cells of the SMP group than the TAU group (p = 0.02). An earlier time of pronuclei appearance (p = 0.03) and time to 2 cells (p = 0.015) and a faster time to full compaction (p = 0.045) were detected in the embryos of the SMP group compared with the TAU group. Conclusion(s): The implemented program may reduce stress levels, retard first embryo cleavage, and accelerate embryo compaction. Further studies with an active control group are needed to confirm these results.

Relief of endoplasmic reticulum stress enhances DNA damage repair and improves development of pre-implantation embryos.

Early-cleaving embryos are known to have better capacity to reach the blastocyst stage and produce better quality embryos compared to late-cleaving embryos. To investigate the significance of endoplasmic reticulum (ER) stress on early embryo cleavage kinetics and development, porcine embryos produced in vitro were separated into early- and late-cleaving groups and then cultured in the absence or presence of the ER stress inhibitor tauroursodeoxycholic acid (TUDCA). Developing embryos were collected at days 3 to 7 of culture for assessment of ER stress status, incidence of DNA double-strand breaks (DSBs), development and total cell number. In the absence of TUDCA treatment, late-cleaving embryos exhibited ER stress, higher incidence of DNA DSBs, as well as reductions in development to the blastocyst stage and total embryo cell numbers. Treatment of late-cleaving embryos with TUDCA mitigated these effects and markedly improved embryo quality and development. These results demonstrate the importance of stress coping responses in early developing embryos, and that reduction of ER stress is a potential means to improve embryo quality and developmental competence.

A prospective randomized comparison of early embryo cleavage kinetics between two media culture systems.

Objective:To investigate whether early embryo cleavage kinetics were affected by type of culture media.Methods:In this prospective sibling-split study, 620 oocytes from 37 patients were randomly allocated into two groups: Cook group and Vitrolife group. Oocytes/embryos in Cook group, would be cultured with Cook sequential culture medium, while oocytes/embryos in Vitrolife group, would be cultured with Vitrolife sequential culture medium. Time-lapse imaging technology was used to calculate exact timing of early embryo cleavage events which included time to 2PN breakdown, cleavage to 2-, 3-, 4-, 5- cell and the time duration in the 2-,3-cell stage. Then these timing of early embryo cleavage events were compared between Cook group and Vitrolife group. Moreover, fertilization rate, cleavage rate, high quality embryo rate, usable blastocyst rate, pregnancy rate and implantation rate of these two groups were also analyzed.Results:The results showed there were no differences in all timing of early embryo cleavage events between the two groups. In addition, the two groups were similar in fertilization rate (Cook 71.0% vs. Vitrolife 71.3%, P>0.05), cleavage rate (Cook 98.1% vs. Vitrolife 98.2%, P>0.05), high quality embryo rate (Cook 52.1% vs. Vitrolife 52.7%, P>0.05), usable blastocyst rate (Cook 29.7% vs. Vitrolife 28.0%, P>0.05), pregnancy rate (Cook 46.7% VS. Vitrolife 50.0%, P>0.05) and implantation rate (Cook 30.3% VS. Vitrolife 29.0%, P>0.05).Conclusions:Morphokinetics used for embryo selection are not affected by the two different culture media.

The Incidence of DNA Double-Strand Breaks Is Higher in Late-Cleaving and Less Developmentally Competent Porcine Embryos.

Studies in different species, including human, mice, bovine, and swine, demonstrated that early-cleaving embryos have higher capacity to develop to the blastocyst stage and produce better quality embryos with superior capacity to establish pregnancy than late-cleaving embryos. It has also been shown that experimentally induced DNA damage delays embryo cleavage kinetics and reduces blastocyst formation. To gain additional insights into the effects of genome damage on embryo cleavage kinetics and development, the present study compared the occurrence of DNA double-strand breaks (DSBs) with the expression profile of genes involved in DNA repair and cell cycle control between early- and late-cleaving embryos. Porcine oocytes matured in vitro were activated, and then early-cleaving (before 24 h) and late-cleaving (between 24 and 48 h) embryos were identified and cultured separately. Developing embryos, on Days 3, 5, and 7, were used to evaluate the total cell number and presence of DSBs (by counting the number of immunofluorescent foci for phosphorylated histone H2A.x [H2AX139ph] and RAD51 proteins) and to quantify transcripts of genes involved in DNA repair and cell cycle control by quantitative RT-PCR. Early-cleaving embryos had fewer DSBs, lower transcript levels for genes encoding DNA repair and cell cycle checkpoint proteins, and more cells than late-cleaving embryos. Interestingly, at the blastocyst stage, embryos that developed from early- and late-cleaving groups had similar number of DSBs as well as transcript levels of genes induced by DNA damage. This indicates that only embryos with less DNA damage and/or superior capacity for DNA repair are able to progress to the blastocyst stage. Collectively, findings in this study revealed a negative correlation between the occurrence of DSBs and embryo cleavage kinetics and embryo developmental capacity to the blastocyst stage.

Impact of PCOS on early embryo cleavage kinetics

This study investigated whether polycystic ovary syndrome (PCOS) affected early embryo development assessed by time-lapse analysis of embryo kinetics from fertilization to the blastocyst stage. This was a prospective cohort study of two pronuclei (2PN) embryos from 25 hyperandrogenic PCOS patients (110 2PN embryos), 26 normoandrogenic PCOS patients (140 2PN embryos) and 20 healthy, regularly cycling women (controls, 97 2PN embryos). Patients underwent the same baseline evaluation and the same ovarian stimulation from April 2010 to February 2013. Oocytes were fertilized by intracytoplasmic sperm injection and incubated in an EmbryoScope with pictures taken every 20min in seven focal planes. Time to 2PN breakdown, first cleavage and cleavage to 3, 4, 5, 6, 7 and 8 cells, morula and blastocyst (t2, t3, t4, t5, t6, t7, t8, tM, tB) were annotated. Differences in embryo kinetics between groups were assessed by mixed modelling. Compared with controls, embryos from hyperandrogenic PCOS patients were significantly delayed at 2PN breakdown, t2, t3, t4 and t7 but not at t5, t6, t8, tM or tB. Embryos from hyperandrogenic PCOS women had developed slower from fertilization to the 8-cell stage compared with embryos from controls.We investigated whether polycystic ovary syndrome (PCOS) affected early embryo development assessed by time-lapse analysis of embryo kinetics from fertilization to the blastocyst stage. This was a prospective cohort study of two pronuclei (2PN) embryos from 25 hyperandrogenic PCOS patients (110 2PN embryos), 26 normoandrogenic PCOS patients (140 2PN embryos) and 20 healthy, regularly cycling women (controls, 97 2PN embryos). Patients underwent the same baseline evaluation and the same ovarian stimulation from April 2010 to February 2013. Oocytes were fertilized by intracytoplasmic sperm injection and incubated in an EmbryoScope with pictures taken every 20min in seven focal planes. Time to 2PN breakdown, first cleavage, cleavage to 3, 4, 5, 6, 7 and 8 cells, morula and blastocyst (t2, t3, t4, t5, t6, t7, t8, tM, tB) were annotated. Differences in embryo kinetics between groups were assessed by mixed modelling. With mixed modelling, the ‘patient factor’ is adjusted for, as each patient contributed several embryos for analysis. Compared with embryos from controls, embryos from hyperandrogenic PCOS patients were significantly delayed at 2PN breakdown (P=0.0128), t2 (P=0.0061), t3 (P=0.0171), t4 (P=0.0017), t7 (P=0.04) and but not at t5, t6, t8, tM or tB. We concluded that embryos from hyperandrogenic PCOS women developed slower from fertilization to the 7-cell stage compared with embryos from controls. The clinical impact of the embryo delay was unknown, since this study was not powered to detect differences in implantation and pregnancy rates between groups.

Embryo cleavage kinetics under different oxygen tensions predict embryonic developmental potential in the mouse

Embryo cleavage kinetics under different oxygen tensions predict embryonic developmental potential in the mouse

From ultrastructural flagellar sperm defects to the health of babies conceived by ICSI

The objective of this retrospective study was to describe a population of patients displaying impaired sperm motility due to ultrastructural flagellar defects and to analyse the intracytoplasmic sperm injection (ICSI) results and neonatal outcomes in this population. The fertilization rate, embryo quality, clinical pregnancy rate, implantation rate, birth rate and perinatal health of babies were determined. Patients ( n = 20) were divided into seven categories according to ultrastructural flagellar abnormalities. The type of flagellar abnormality significantly affected the fertilization rate ( P < 0.025). Two types of flagellar abnormalities showed slower early embryo cleavage kinetics ( P < 0.001) when axonemal central structures and periaxonemmal columns were abnormal or absent. Of 53 ICSI attempts, 14 resulted in clinical pregnancies (26.4% per cycle) after fresh and frozen embryo transfer. Three (21.4%) of these pregnancies ended in miscarriages and, in the remaining, 12 infants were born (7.2% of transferred embryos). The outcomes differed according to the ultrastructural defect. This study demonstrates that a high proportion of patients could father a child (45.0%). However, flagellar abnormalities appear to influence ICSI results and fetal development.

Qualité et sélection des embryons

Embryo quality is routinely evaluated based on morphological and kinematic criteria, but it should also be founded on clinical and biological data from the ovulation induction treatments and the data from the SPZ test, with, if necessary, their genetic analysis. Zygote quality evaluation is based on the zygote score or the P0 profiles. Embryo cleavage kinetics is evaluated at D1 (early cleavage 25-27 h post-insemination), at D2 (four cells), and at D3 (eight cells), and their selection, particularly for eSET, is for the most part based on the cell number and regularity, the percentage of fragmentation (< or = 20 %) and the location of the fragments and the absence of blastomere polynucleation. Imaging software (Fertimorph) now facilitates the evaluation of embryo quality.

Assisted Reproductive Technology affects developmental kinetics, H19 Imprinting Control Region methylation and H19gene expression in individual mouse embryos

BackgroundIn the last few years, an increase in imprinting anomalies has been reported in children born from Assisted Reproductive Technology (ART). Various clinical and experimental studies also suggest alterations of embryo development after ART. Therefore, there is a need for studying early epigenetic anomalies which could result from ART manipulations, especially on single embryos. In this study, we evaluated the impact of superovulation, in vitro fertilization (IVF) and embryo culture conditions on proper genomic imprinting and blastocyst development in single mouse embryos.In this study, different experimental groups were established to obtain embryos from superovulated and non-superovulated females, either from in vivo or in vitro fertilized oocytes, themselves grown in vitro or not. The embryos were cultured either in M16 medium or in G1.2/G2.2 sequential medium. The methylation status of H19 Imprinting Control Region (ICR) and H19 promoter was assessed, as well as the gene expression level of H19, in individual blastocysts. In parallel, we have evaluated embryo cleavage kinetics and recorded morphological data.ResultsWe show that:1. The culture medium influences early embryo development with faster cleavage kinetics for culture in G1.2/G2.2 medium compared to M16 medium.2. Epigenetic alterations of the H19 ICR and H19 PP are influenced by the fertilization method since methylation anomalies were observed only in the in vitro fertilized subgroup, however to different degrees according to the culture medium.3. Superovulation clearly disrupted H19 gene expression in individual blastocysts. Moreover, when embryos were cultured in vitro after either in vivo or in vitro fertilization, the percentage of blastocysts which expressed H19 was higher in G1.2/G2.2 medium compared to M16.ConclusionCompared to previous reports utilizing pools of embryos, our study enables us to emphasize a high individual variability of blastocysts in the H19 ICR and H19 promoter methylation and H19 gene expression, with a striking effect of each manipulation associated to ART practices. Our results suggest that H19 could be used as a sensor of the epigenetic disturbance of the utilized techniques.