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Effect of embryo morphology and morphometrics on implantation of vitrified day 3 embryos after warming: a retrospective cohort study.

ABSTRACT: Characteristics routinely used to evaluate embryo quality after thawing include number of blastomeres survived and presence of mitosis resumption after overnight culture. It is unknown to which extent symmetry and fragmentation affect implantation after warming and whether application of stricter criteria either before vitrification or after warming would improve implantation rate (IR) of vitrified/warmed embryos. This study aimed to find new parameters to improve selection criteria for vitrification and for transfer after warming.Firstly, we evaluated standard morphological characteristics (intact survival, mitosis resumption, number of blastomeres, symmetry and fragmentation) of 986 warmed day 3 embryos and, from a subset of 654, we evaluated morphometric characteristics (fragmentation, symmetry and volume change). Secondly, we tested the hypothesis that IR of day 3 vitrified/warmed embryos is influenced by morphometric characteristics. IR per embryo transferred was calculated using embryos that were transferred in a single embryo transfer (SET) or a double embryo transfer (DET) with either 0 or 100 % implantation (830/986). We investigated the significant differences in IR between the different categories of a specific characteristic. These categories were based on our standard embryo evaluation system. The statistical tests Chi-square, Fisher's exact or Cochrane-Armitage were used according to the type and/or categories of the variable.The 986 embryos were transferred in 671 FET cycles with 16.9 % (167/986) IR. After exclusion of DET with 1 embryo implanted, IR per embryo transferred was 12.4 % (103/830). Embryo symmetry, fragmentation and volume change in vitrified/warmed day 3 embryos were not associated with IR. However, when mitosis resumption was present after overnight culture, intact embryos reached significantly higher IR than non-intact embryos and only when the embryo compacted after overnight culture the number of cells damaged after warming had no effect on IR. Concretely, embryos with 8 cells after warming or >9 cells after overnight culture-including compacted embryos-reached the highest IR (>15 %) while embryos with <6 cells after warming or with ?6 cells after overnight culture had extremely low IR (<1 %).IR of vitrified embryos is determined by the number of cells lost, by the occurrence of mitosis resumption, and by the specific number of blastomeres present but not by fragmentation, blastomere symmetry or volume change. Unselecting embryos for cryopreservation because of fragmentation >10 % and/or symmetry?

SUBMITTER: Fernandez Gallardo E

PROVIDER: S-EPMC4967519 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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Effect of embryo morphology and morphometrics on implantation of vitrified day 3 embryos after warming: a retrospective cohort study.

Fernandez Gallardo Elia E Spiessens Carl C D'Hooghe Thomas T Debrock Sophie S

Reproductive biology and endocrinology : RB&E 20160730 1

<h4>Background</h4>Characteristics routinely used to evaluate embryo quality after thawing include number of blastomeres survived and presence of mitosis resumption after overnight culture. It is unknown to which extent symmetry and fragmentation affect implantation after warming and whether application of stricter criteria either before vitrification or after warming would improve implantation rate (IR) of vitrified/warmed embryos. This study aimed to find new parameters to improve selection cr ...[more]

PMID: 27475526

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Project description:BackgroundMorphometric and morphokinetic evaluation of in vitro cultured human embryos allows evaluation without time restriction and reduces intra- and inter-observer variability. Even though these technologies have been reported to improve the quality of cleavage stage embryo evaluation during fresh culture, possible advantages in the evaluation of cryopreserved embryos have been scarcely explored. This study aims to compare morphometric and morphokinetic parameters between slow frozen and vitrified embryos and to determine their relationship to embryo survival and implantation rate (IR) after thawing/warming.MethodsDuring fresh culture, morphometric characteristics (Total Cell Volume (TCV), symmetry, fragmentation and number of blastomeres) were measured in 286 thawed/warmed embryos. Likewise, after thawing/warming, similar morphometric characteristics were measured in 135 survived embryos. Moreover, morphokinetic parameters (time to mitosis resumption and time to compaction) were measured in 90 embryos after thawing/warming. Then, using linear regression, we investigated the differences between vitrified and slow frozen embryos and the relation of the measured characteristics to embryo survival and IR. Statistical corrections were applied to account for data clustering and for multiple testing.ResultsVitrified embryos resume mitosis and start compaction significantly earlier than slow frozen embryos. Mitosis resumption rate was 82% for vitrified and 63% for slow frozen embryos and median time to mitosis resumption was 7.6 h and 13.1 h (p = 0.02), respectively. Compaction rate was 62% in vitrified and only 23% in slow frozen embryos. Median time to compaction was 18.1 h for vitrified embryos but, for slow frozen could not be computed since less than half of the slow frozen embryos reached compaction (p = 0.0001). Moreover, intact embryos resume mitosis significantly earlier than not intact ones regardless of the freezing method (rate: 79% vs. 66%, median time: 7.6 h vs 14.6 h, respectively, p = 0.03). Regarding morphometrics, slow frozen embryos showed lower TCV and higher blastomere symmetry after thawing than vitrified embryos despite having similar blastomere number. IR was related to blastomere number at cryopreservation in slow frozen embryos, but not in vitrified ones.ConclusionsInterestingly, vitrified/warmed embryos undergo mitosis resumption and compaction significantly earlier than slow frozen/thawed embryos. However, the clinical use of this morphokinetic parameters still remains to be investigated in larger studies.Trial registrationRetrospectively registered on December 15, 2015 NCT02639715 .

Project description:BackgroundCryopreservation of embryos plays a major role in the in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) treatment. However, the storage condition of the cryopreserved embryo can change temporarily due to repeated retrieval of the embryo from the liquid nitrogen (LN2) tank during the practical application during cryopreservation. Whether the implantation potential of a cryopreserved embryo will be damaged when the cane containing it is temporarily exposed to air due to the transfer between the LN2 tank and LN2 container is yet to be elucidated. Also, whether the exposed-to-air frequency (EAF) of cryopreserved embryos influences the clinical outcomes is unclear.ObjectiveTo investigate whether the EAF of cryopreserved embryo affects the clinical outcomes of vitrified-warmed embryo transfer.MethodsA total of 9200 vitrified-warmed embryo transfer cycles were included in this study. All cycles were divided into five groups according to different EAFs (2, 4, 6, 8, or ≥ 10). Post-warming survival rates and clinical outcomes, including implantation, clinical pregnancy and live birth rates were investigated. Kruskal-Wallis test and Pearson's chi-squared tests were used to compare the patient characteristics and clinical outcomes among the five groups. Furthermore, multivariate logistic regression analyses were conducted to investigate the association between EAF and clinical outcomes.ResultsNo significant differences were observed in the positive HCG rate, implantation rate and live birth rate (P > 0.05) among five EAF groups with respect to D3 embryo, D5 blastocyst and D6 blastocyst. Post-warmed survival rate of D3 embryos (P = 0.015) differed significantly among the five EAF groups, but it was not EAF-dependent. Although clinical pregnancy was different among the five groups with respect to D5 blastocyst (P = 0.042), multivariate logistic regression analysis adjusted for confounding variables suggested that EAF did not adversely affect clinical pregnancy or live birth.ConclusionThese findings indicated that human vitrified embryos in the open system could be repeatedly retrieved from the LN2 tank without affecting the implantation potential of the embryo.

Project description:The ability to study human post-implantation development remains limited owing to ethical and technical challenges associated with intrauterine development after implantation1. Embryo-like models with spatially organized morphogenesis and structure of all defining embryonic and extra-embryonic tissues of the post-implantation human conceptus (that is, the embryonic disc, the bilaminar disc, the yolk sac, the chorionic sac and the surrounding trophoblast layer) remain lacking1,2. Mouse naive embryonic stem cells have recently been shown to give rise to embryonic and extra-embryonic stem cells capable of self-assembling into post-gastrulation structured stem-cell-based embryo models with spatially organized morphogenesis (called SEMs)3. Here we extend those findings to humans using only genetically unmodified human naive embryonic stem cells (cultured in human enhanced naive stem cell medium conditions)4. Such human fully integrated and complete SEMs recapitulate the organization of nearly all known lineages and compartments of post-implantation human embryos, including the epiblast, the hypoblast, the extra-embryonic mesoderm and the trophoblast layer surrounding the latter compartments. These human complete SEMs demonstrated developmental growth dynamics that resemble key hallmarks of post-implantation stage embryogenesis up to 13-14 days after fertilization (Carnegie stage 6a). These include embryonic disc and bilaminar disc formation, epiblast lumenogenesis, polarized amniogenesis, anterior-posterior symmetry breaking, primordial germ-cell specification, polarized yolk sac with visceral and parietal endoderm formation, extra-embryonic mesoderm expansion that defines a chorionic cavity and a connecting stalk, and a trophoblast-surrounding compartment demonstrating syncytium and lacunae formation. This SEM platform will probably enable the experimental investigation of previously inaccessible windows of human early post implantation up to peri-gastrulation development.

Project description:Background:Extended embryo culture has been reported to affect perinatal outcome regarding higher risks of large for gestational age (LGA) and preterm birth (PTB) yet decreased risk of small for gestational age (SGA). However, existing data about the obstetric outcome and the safety for offspring resulting from the transfer of day 7 blastocysts is rare. Objectives:To compare obstetric and perinatal outcome using embryos vitrified on day 7 with those vitrified on day 3, day 5, and day 6. Methods:Data were collected from 4489 infertile women who gave birth to live-born singletons after vitrified-warmed embryo transfer cycles from January 1, 2006 to December 31, 2017. Singletons were compared depending on the age of embryos. Main perinatal outcome parameters included PTB (gestational age < 37 weeks), very PTB (VPTB, gestational age < 32 weeks), LGA (birthweights > 90th percentiles), and SGA (birthweights < 10th percentiles). Obstetric outcomes included gestational diabetes (GDM), pregnancy-induced hypertension (PIH), preterm premature rupture of membranes (PPROM), pre-eclampsia, placenta previa, placental abruption, and postpartum hemorrhage. Propensity score matching (PSM) was used to adjust the confounding factors across groups and then analyze the association between in vitro culture period and the outcome measures. Results:After PSM, the transfer of day 7 blastocysts was associated with higher birth weight Z-scores and increased incidence of very large for gestational age (VLGA) compared with the transfer of day 3 cleavage-stage embryos while the incidence of PTB, low birth weight (LBW), SGA did not reach statistical significance. Moreover, comparable perinatal outcome was found in the comparison of day 7 vs. day 5 and day 7 vs. day 6. Day 7 blastocysts did not result in adverse obstetric outcome compared with day 3, day 5, and day 6 embryos, respectively. Conclusion:In vitrified-warmed transfer cycles, day 7 blastocysts were associated with adverse perinatal outcome regarding higher risk of VLGA compared with day 3 cleavage-stage embryo, while blastocysts with diverse growth rates embrace similar developmental viability regardless of blastocysts vitrified on day 5, day 6, or day 7.

Dataset Information

Effect of embryo morphology and morphometrics on implantation of vitrified day 3 embryos after warming: a retrospective cohort study.

Publications

Effect of embryo morphology and morphometrics on implantation of vitrified day 3 embryos after warming: a retrospective cohort study.

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