Differences in Morphokinetic Parameters and Incidence of Multinucleations in Human Embryos of Genetically Normal, Abnormal and Euploid Embryos Leading to Clinical Pregnancy

Abstract

The selection of the best embryo for embryo transfer (ET) is one of the most important steps in IVF (in vitro fertilisation) treatment. Preimplantation genetic testing (PGT) is an invasive method that can greatly facilitate the decision about the best embryo. An alternative way to select the embryo with the greatest implantation potential is by cultivation in a time-lapse system, which can offer several predictive factors. Non-invasive time-lapse monitoring can be used to select quality embryos with high implantation potential under stable culture conditions. The embryo for ET can then be selected based on the determined morphokinetic parameters and morphological features, which according to our results predict a higher implantation potential. This study included a total of 1027 morphologically high-quality embryos (552 normal and 475 abnormal PGT-tested embryos) from 296 patients (01/2016–06/2021). All embryos were cultivated in a time-lapse incubator and PGT biopsy of trophectoderm cells on D5 or D6 was performed. Significant differences were found in the morphological parameters cc2, t5 and tSB and the occurrence of multinucleations in the stage of two-cell and four-cell embryos between the group of genetically normal embryos and abnormal embryos. At the same time, significant differences in the morphological parameters cc2, t5 and tSB and the occurrence of multinucleations in the two-cell and four-cell embryo stage were found between the group of genetically normal embryos that led to clinical pregnancy after ET and the group of abnormal embryos. From the morphokinetic data found in the PGT-A group of normal embryos leading to clinical pregnancy, time intervals were determined based on statistical analysis, which should predict embryos with high implantation potential. Out of a total of 218 euploid embryos, which were transferred into the uterus after thawing (single frozen embryo transfer), clinical pregnancy was confirmed in 119 embryos (54.6%). Our results show that according to the morphokinetic parameters (cc2, t5, tSB) and the occurrence of multinucleations during the first two cell divisions, the best euploid embryo for ET can be selected with high probability.

Prediction of live birth - selection of embryos using morphokinetic parameters

“…A statistically significant difference was detected in the values of tSB and t5 parameters in the group of PGT-A normal embryos, which led to the birth of a healthy child after FET, and in the group of PGT-A normal embryos without achiewing a fetal heartbeat. This is consistent with the results of our previous study 28 , where it was found that the values of morphokinetic parameters cc2, t5 and tSB are significantly shorter in genetically normal embryos than in genetically abnormal embryos; furthermore, tSB and t5 parameters were significantly shorter in the group of genetically normal embryos with a proven clinical pregnancy compared to genetically normal embryos with a negative pregnancy test after FET.…”

“…The aim of this study was to follow up and confirm the results of our previous study 28 , where it was found that morphokinetic predictive parameters for achieving clinical pregnancy with a high probability are t5 and tSB; and also that the presence of multinucleations in 2-cell and 4-cell embryos is associated with a higher probability of embryonic aneuploidy. This study, unlike the one mentioned above, used patients who gave birth to a healthy child after FET of one time-lapse cultured euploid embryo.…”

“…All laboratory procedures and materials were used the same as in the previous study 28 , to which this current work follows.…”

Nucleation status of Day 2 pre-implantation embryos, acquired by time-lapse imaging during IVF, is associated with live birth

“…The nucleation errors present in the 2-cell stage, even if corrected in the 4-cell embryo, maybe evident in the quality of blastocysts. The higher incidence of 2-cell multinucleation and a slightly reduced incidence of the nucleation error during the transition to 4- cell stage and the possible self-correction mechanism provides a more significant prediction of aneuploidy in the embryo at 4- cell stage [ 62 ]. This is further strengthened by the conclusion drawn by Tvrdonova and colleagues [ 62 ] that the incidence of nucleation error in the group of euploid embryos that resulted in a clinical pregnancy in the four-cell stage is 7 times lower than in the group of aneuploid embryos.…”

Binucleated embryos at the two-cell stage show higher blastocyst formation rates and higher pregnancy and live birth rates compared to non-multinucleated embryos

“…The origin of multinucleated blastomeres has been suggested to involve mechanisms including karyokineses without cytokinesis, erroneous migration of chromosomes at anaphase, erroneous packaging of chromosomes and/or fragmentation of nuclei ( Alpha Scientists in Reproductive Medicine and ESHRE Special Interest Group of Embryology, 2011 , Hardy et al , 1993 ; Munné and Cohen, 1993 ; Pickering et al , 1995 ; Staessen and Van Steirteghem, 1998 ). Furthermore, researchers have found a correlation between multinucleation displayed in the early embryo and aneuploidy ( Kligman et al , 1996 ; Ambroggio et al , 2011 ; Yilmaz et al , 2014 ; Tvrdonova et al , 2021 ) and Hardarson et al (2001) concluded that unevenly sized blastomeres were an indicator for multinucleation and aneuploidy. In spite of this, the association between multinucleation and aneuploidy is not yet clear and studies using preimplantation genetic screening have reported comparable aneuploidy rates for multinucleated and non-multinucleated embryos ( Balakier et al , 2016 ; Desai et al , 2018 ), and consequently suggested that multinucleation at the two-cell stage should not be used as an indicator for aneuploidy and embryo selection.…”