The use of morphokinetics as a predictor of embryo implantation
TL;DR: A multivariable model is proposed based on the findings to classify embryos according to their probability of implantation and it is proposed that the image acquisition and time-lapse analysis system makes it possible to determine exact timing of embryo cleavages in a clinical setting.
Abstract: background: Time-lapse observation presents an opportunity for optimizing embryo selection based on morphological grading as well as providing novel kinetic parameters, which may further improve accurate selection of viable embryos. The objective of this retrospective study was to identify the morphokinetic parameters specific to embryos that were capable of implanting. In order to compare a large number of embryos, with minimal variation in culture conditions, we have used an automatic embryo monitoring system. methods: Using a tri-gas IVF incubator with a built-in camera designed to automatically acquire images at defined time points, we have simultaneously monitored up to 72 individual embryos without removing the embryos from the controlled environment. Images were acquired every 15 min in five different focal planes for at least 64 h for each embryo. We have monitored the development of transferred embryos from 285 couples undergoing their first ICSI cycle. The total number of transferred embryos was 522, of which 247 either failed to implant or fully implanted, with full implantation meaning that all transferred embryos in a treatment implanted. results: A detailed retrospective analysis of cleavage times, blastomere size and multinucleation was made for the 247 transferred embryos with either failed or full implantation. We found that several parameters were significantly correlated with subsequent implantation (e.g. time of first and subsequent cleavages as well as the time between cleavages). The most predictive parameters were: (i) time of division to 5 cells, t5 (48.8–56.6 h after ICSI); (ii) time between division to 3 cells and subsequent division to 4 cells, s2 (≤0.76 h) and (iii) duration of cell cycle two, i.e. time between division to 2 cells and division to 3 cells, cc2 (≤11.9 h). We also observed aberrant behavior such as multinucleation at the 4 cell stage, uneven blastomere size at the 2 cell stage and abrupt cell division to three or more cells, which appeared to largely preclude implantation. conclusions: The image acquisition and time-lapse analysis system makes it possible to determine exact timing of embryo cleavages in a clinical setting. We propose a multivariable model based on our findings to classify embryos according to their probability of implantation. The efficacy of this classification will be evaluated in a prospective randomized study that ultimately will determine if implantation rates can be improved by time-lapse analysis.
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TL;DR: No significant differences were observed in first or second cell-cycle length, synchrony of the second or third cell cycles, duration of blastulation, multinucleation at the 2-cell stage and irregular division patterns between euploid and aneuploid embryos.
Abstract: This study determined whether morphokinetic variables between aneuploid and euploid embryos differ as a potential aid to select euploid embryos for transfer. Following insemination, EmbryoScope time-lapse images from 98 blastocysts were collected and analysed blinded to ploidy. The morphokinetic variables were retrospectively compared with ploidy, which was determined fol- lowing trophectoderm biopsy and analysis by array comparative genomic hybridization or single-nucleotide polymorphic array. Multiple aneuploid embryos were delayed at the initiation of compaction (tSC; median 85.1 hours post insemination (hpi); P = 0.02) and the time to reach full blastocyst stage (tB; median 110.9 hpi, P = 0.01) compared with euploid embryos (tSC median 79.7 hpi, tB median 105.9 hpi). Embryos having single or multiple aneuploidy (median 103.4 hpi, P = 0.004 and 101.9 hpi, P = 0.006, respec- tively) had delayed initiation of blastulation compared with euploid embryos (median 95.1 hpi). No significant differences were observed in first or second cell-cycle length, synchrony of the second or third cell cycles, duration of blastulation, multinucleation at the 2-cell stage and irregular division patterns between euploid and aneuploid embryos. This non-invasive model for ploidy clas- sification may be used to avoid selecting embryos with high risk of aneuploidy while selecting those with reduced risk. RBMOnline
292 citations
TL;DR: Analysis of retrospective data indicated that culturing and selecting embryos by TMS significantly improved the relative probability of clinical pregnancy and was attributed to a combination of stable culture conditions and the use of morphokinetic parameters for embryo selection.
277 citations
TL;DR: It is demonstrated that precise cell cycle parameter timing is observed in all euploid embryos to the four-cell stage, whereas only 30% of aneuploids embryos exhibit parameter values within normal timing windows, suggesting that cell cycle and fragmentation parameters of individual blastomeres are diagnostic of ploidy, amenable to automated tracking algorithms, and likely of clinical relevance in reducing transfer of embryos prone to miscarriage.
Abstract: Previous studies have demonstrated that aneuploidy in human embryos is surprisingly frequent with 50–80% of cleavage-stage human embryos carrying an abnormal chromosome number. Here we combine non-invasive time-lapse imaging with karyotypic reconstruction of all blastomeres in four-cell human embryos to address the hypothesis that blastomere behaviour may reflect ploidy during the first two cleavage divisions. We demonstrate that precise cell cycle parameter timing is observed in all euploid embryos to the four-cell stage, whereas only 30% of aneuploid embryos exhibit parameter values within normal timing windows. Further, we observe that the generation of human embryonic aneuploidy is complex with contribution from chromosome-containing fragments/micronuclei that frequently emerge and may persist or become reabsorbed during interphase. These findings suggest that cell cycle and fragmentation parameters of individual blastomeres are diagnostic of ploidy, amenable to automated tracking algorithms, and likely of clinical relevance in reducing transfer of embryos prone to miscarriage.
273 citations
TL;DR: Embryos with DC2-3 had a statistically significantly lower implantation rate than embryos with a normal cleavage pattern, suggesting that rejection of these embryos for transfer could improve the implantation rates.
270 citations
TL;DR: Experienced embryologists using Eeva in combination with D3 morphology significantly improved their ability to identify embryos that would reach the usable blastocyst stage, and adjunctive use of morphology plus Eeva also reduced interindividual variability in embryo selection.
261 citations
References
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TL;DR: It is found that success in progression to the blastocyst stage can be predicted with >93% sensitivity and specificity by measuring three dynamic, noninvasive imaging parameters by day 2 after fertilization, before embryonic genome activation (EGA).
Abstract: We report studies of preimplantation human embryo development that correlate time-lapse image analysis and gene expression profiling. By examining a large set of zygotes from in vitro fertilization (IVF), we find that success in progression to the blastocyst stage can be predicted with >93% sensitivity and specificity by measuring three dynamic, noninvasive imaging parameters by day 2 after fertilization, before embryonic genome activation (EGA). These parameters can be reliably monitored by automated image analysis, confirming that successful development follows a set of carefully orchestrated and predictable events. Moreover, we show that imaging phenotypes reflect molecular programs of the embryo and of individual blastomeres. Single-cell gene expression analysis reveals that blastomeres develop cell autonomously, with some cells advancing to EGA and others arresting. These studies indicate that success and failure in human embryo development is largely determined before EGA. Our methods and algorithms may provide an approach for early diagnosis of embryo potential in assisted reproduction.
714 citations
"The use of morphokinetics as a pred..." refers background or result in this paper
...In a more recent study, Wong et al. (2010) found that development of human embryos to the blastocyst stage was correlated with: (i) the duration of the first cytoplasmic cleavage from 1 cell to 2 cells; (ii) time between division to 2 cells and subsequent division to 3 cells and (iii) time between…...
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...One promising predictive factor is the precise timing of key events in early embryo development (Payne et al., 1997; Lemmen et al., 2008; Mio and Maeda, 2008; Nakahara et al., 2010; Wong et al., 2010)....
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...They studied supernumerary IVF embryos that had been cryopreserved at the zygote stage; these embryos were frozen with a procedure that significantly affects the outcome after thawing and could not be directly comparable with fresh embryos (Wong et al., 2010)....
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...…analysis presented in this study is based on a larger set of 247 exclusively transferred embryos, all of which were evaluated based on their ability to implant and form a gestational sac, which largely confirms the findings of the previous studies (Lemmen et al., 2008; Wong et al., 2010)....
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...It is thus unclear if embryos with the suggested morphokinetic cleavage pattern would have implanted (Wong et al., 2010)....
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TL;DR: Preimplantation genetic screening did not increase but instead significantly reduced the rates of ongoing pregnancies and live births after IVF in women of advanced maternal age.
Abstract: BACKGROUND Pregnancy rates in women of advanced maternal age undergoing in vitro fertilization (IVF) are disappointingly low. It has been suggested that the use of preimplantation genetic screening of cleavage-stage embryos for aneuploidies may improve the ef- fectiveness of IVF in these women. METHODS We conducted a multicenter, randomized, double-blind, controlled trial comparing three cycles of IVF with and without preimplantation genetic screening in women 35 through 41 years of age. The primary outcome measure was ongoing pregnancy at 12 weeks of gestation. The secondary outcome measures were biochemical preg- nancy, clinical pregnancy, miscarriage, and live birth. RESULTS Four hundred eight women (206 assigned to preimplantation genetic screening and 202 assigned to the control group) underwent 836 cycles of IVF (434 cycles with and 402 cycles without preimplantation genetic screening). The ongoing-pregnancy rate was significantly lower in the women assigned to preimplantation genetic screen- ing (52 of 206 women (25%)) than in those not assigned to preimplantation genetic screening (74 of 202 women (37%); rate ratio, 0.69; 95% confidence interval (CI), 0.51 to 0.93). The women assigned to preimplantation genetic screening also had a significantly lower live-birth rate (49 of 206 women (24%) vs. 71 of 202 women (35%); rate ratio, 0.68; 95% CI, 0.50 to 0.92). CONCLUSIONS Preimplantation genetic screening did not increase but instead significantly reduced the rates of ongoing pregnancies and live births after IVF in women of advanced maternal age. (Current Controlled Trials number, ISRCTN76355836.)
640 citations
TL;DR: This controlled-randomized, clinical trial confirmed the effectiveness of oocyte cryo-storage in an ovum donation programme, failing to demonstrate the superiority of using fresh oocytes with respect to the use of vitrified egg-banked ones in terms of OPR, but instead confirmed the non-inferiority of Vitrified oocytes.
Abstract: background: An efficient oocyte cryopreservation method is mandatory to establish a successful egg-banking programme. Although there are increasing reports showing good clinical outcomes after oocyte cryopreservation, there is still a lack of large controlled studies evaluating the effectiveness of oocyte cryo-banking. In this study, we aimed to compare the outcome of vitrified-banked oocytes with the gold standard procedure of employing fresh oocytes. methods: A randomized, prospective, triple-blind, single-centre, parallel-group controlled-clinical trial (NCT00785993), including 600 recipients (a ¼ 0.05 and power of 80% for sample-size calculation) selected among 1032 eligible patients from November 2008 to September 2009, was designed to compare the outcome of vitrified-banked oocytes with the gold standard procedure of employing fresh oocytes. The study was designed to establish the superiority of the ongoing pregnancy rate (OPR) of fresh oocytes over that of vitrified oocytes, by performing a likelihood ratio test in a logistic regression analysis expressed as odds ratio (OR) with 95% confidence interval (CI). A limit of 0.66 for OR of vitrified versus fresh groups was defined to set up a possible conversion from superiority to non-inferiority. Randomization was performed 1:1 based on a computer randomization list in vitrification (n ¼ 300) or fresh groups (n ¼ 300). The primary end-point was the OPR per randomized patient i.e. intention-to-treat population (ITT). Secondary end-points were clinical pregnancy (CPR), implantation (IR) and fertilization rates, respectively. Additionally, embryo developmental characteristics were recorded. results: There were no differences in donor ovarian stimulation parameters, demographic baseline characteristics for donors and recipients, ovum donation indications or male factor distribution between groups (NS). The OPR per ITT was 43.7 and 41.7% in the vitrification and fresh groups, respectively. The OR of OPR was 0.921 in favour of the vitrification group. Nevertheless, the 95% CI was 0.667–1.274, thus the superiority of fresh group with respect to OPR was not proven (P ¼ 0.744). Non-inferiority of the vitrified group compared with the fresh group was shown with a margin of 0.667, which was above the pre-established non-inferiority limit of 0.66. CPR per cycle (50.2 versus 49.8%; P ¼ 0.933) or per embryo-transfer (55.4 versus 55.6% ; P ¼ 0.974), and IR (39.9 versus 40.9%; P ¼ 0.745) were similar for patients receiving either vitrified or fresh oocytes. The proportion of top-quality embryos obtained either by inseminated oocyte (30.8 versus 30.8% for Day-2; and 36.1 versus 37.7% for Day-3, respectively) or by cleaved embryos (43.6 versus 43.8% for Day-2 and 58.4 versus 60.7% for Day-3, respectively) was similar between groups (NS). conclusions: This controlled-randomized, clinical trial confirmed the effectiveness of oocyte cryo-storage in an ovum donation programme, failing to demonstrate the superiority of using fresh oocytes with respect to the use of vitrified egg-banked ones in terms of OPR. Instead, the non-inferiority of vitrified oocytes was confirmed. These findings involve highly relevant issues that may open a new range of possibilities in ART. Clinical Trials identifier: www.clinicaltrials.gov: NCT 00785993.
455 citations
"The use of morphokinetics as a pred..." refers methods in this paper
...Supernumerary embryos were frozen for potential future transfers using IVI standard vitrification technique (Cobo et al., 2010)....
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TL;DR: The results of all treatments including patient's responses during the follicular and luteal phases, oocyte recovery, fertilization, cleavage, replacement, implantation, abortion, and birth and the effect of factors such as replacing two or more embryos, maternal age, and previous obstetric history are described in detail.
Abstract: The program for in vitro fertilization at Bourn Hall began in October 1980. Various types of infertility have been treated during this time using the natural menstrual cycle or stimulation of follicular growth with antiestrogens and gonadotrophins. Follicular growth and maturation are assayed by urinary estrogens and LH, monitored regularly during the later follicular stage. Many patients had an endogenous LH surge; others needed an injection of HCG to induce ovulation. All oocytes were recovered by laparoscopy. Wide variations occurred in the time interval between the start of the LH surge and oocyte recovery and between oocyte recovery and insemination. Embryos taken between the one- and the eight-cell stage were replaced into their mother, no standard procedure being adopted for all patients. The results of all treatments including patient's responses during the follicular and luteal phases, oocyte recovery, fertilization, cleavage, replacement, implantation, abortion, and birth and the effect of factors such as replacing two or more embryos, maternal age, and previous obstetric history are described in detail. The incidence of implantation after embryo replacement improved from 16.5% initially to 30% currently. More than 118 babies have been born, and many pregnancies are continuing.
429 citations
"The use of morphokinetics as a pred..." refers methods in this paper
...…on August 9, 2011 doi:10.1093/humrep/der256 D ow nloaded from https://academ ic.oup.com /hum rep/article-abstract/26/10/2658/611030 by guest on 07 M arch 2019 of inspection 25–27 h post ICSI, and its impact on pregnancy rate in humans was first published by the Edwards group (Edwards et al., 1984)....
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...of inspection 25–27 h post ICSI, and its impact on pregnancy rate in humans was first published by the Edwards group (Edwards et al., 1984)....
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TL;DR: Single blastocyst transfer is an effective method of eliminating multiple births while maintaining high pregnancy rates in this selected group of patients.
416 citations
"The use of morphokinetics as a pred..." refers background in this paper
...Extended culture has been advocated as a way to increase implantation rate and improve reproductive outcomes (Mercader et al., 2003; Gardner et al., 2004)....
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...Given the uncertainties associated with evaluation at Day 3, some clinics have turned to extended culture regimens to improve the assessment of embryo implantation potential (Milki et al., 2000; Gardner et al., 2004)....
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