O-006 Oocyte factors as predictors of abnormal, mononucleated and trinucleated, zygotes and assessment of their morphokinetic differences
Authors: De Pablo Franco J L , Pérez-Cárceles M D , Almunia-Santiago N , Valcarcel-Andreu M , Salmerón-Martínez D , Montero-Venegas L , Diaz R , Vicente Llosá-Esteban V , Villaquirán A M , Gómez-Sánchez E 
Published: July 2024

Abstract

Study question
Are there oocyte factors associated with the occurrence of mononucleated and trinucleated zygotes after ICSI? Does morphokinetics differ in embryos from normal and abnormal zygotes?

Summary answer
Formation of 1PN zygotes relates to vitrificated and donor oocytes. 3PN links to age and own oocytes. 1PN embryo morphokinetics are slower than 2PN, 3PN.

What is known already
In assisted reproduction, after monitoring fertilization, it is common to observe zygotes with one or more than 2 pronuclei. These abnormal zygotes are often discarded due to potential chromosomal abnormalities. The formation of these anomalous zygotes can be attributed to various causes, such as non-activation of the male pronucleus, non-extrusion of the second polar body, etc. Simultaneously, limited data exist on the morphokinetics of abnormal zygotes, explored in a few studies. The embryonic kinetics of 1PN zygotes seem slower than those of 2PN and 3PN zygotes, but further research is needed to confirm this aspect.

Study design, size, duration
Retrospective multicentre study involved time-lapse monitoring of 28,218 ICSI zygotes (26,252 2PN, 1,190 1PN, and 776 &gt;3PN) generated between January 2019 and December 2023. The study includes donor and patient oocytes with various infertility diagnoses, whether fresh or vitrified. The comparison of embryonic morphokinetics from abnormal/normal zygotes was conducted on 254 embryos from a single centre, with 45 originating from 1PN zygotes, 198 from 2PN zygotes, and 11 from zygotes with 3 or more pronuclei.

Participants/materials, setting, methods
The study involved 1,053 couples treated in four different centres. Uniform culture media were used, and oocytes were cultivated in time-lapse technology-equipped incubators across all centres. A logistic regression analysis aimed to identify oocyte factors linked to an abnormal pronuclei count. Embryonic morphokinetics were analysed using the CHLOE artificial intelligence system (Fairtility, Israel). Evaluated parameters included: tPNa, tPNf, t2, t3, t4, t5, t6, t7, t8, t9+, tM, tSB, tB, and tEB.

Main results and the role of chance
Donor oocytes exhibited a 1.32 times higher risk of producing 1PN zygotes than 2PN or 3PN zygotes (p &lt; 0.001). Similarly, vitrified oocytes had a 1.63-fold higher risk of generating 1PN zygotes than 2PN or 3PN zygotes (p &lt; 0.001). Additionally, we found that both the use of own oocytes and advanced age were significantly associated with 3PN zygotes (p &lt; 0.001). Regarding age, the risk increases by 10% for each additional year. The average durations for all morphokinetic parameters of 2PN and 3PN embryos were similar. In contrast, all events, except t3 (p = 0.2), occurred more slowly in embryos derived from 1PN zygotes compared to embryos from 2PN zygotes. Thus, the disappearance of pronuclei had a p-value &lt;0.005, and from t4 to tEB, the p-value was &lt;0.001. When comparing embryos from 1PN and 3PN zygotes, only tPNa, t4, t8, t9+, and tM were significantly different. If only zygotes that reached the blastocyst stage were used for analysis, the results were less conclusive due to a significantly reduced number of cases.

Limitations, reasons for caution
Embryos were cultured in an EmbryoScope incubator at three centres, while Geri was used in the fourth.

Wider implications of the findings
Using donor or vitrified oocytes may lead to an increased occurrence of zygotes with 1PN, as will occur with 3PN when patient or aged oocytes are used. The kinetics of embryos from 3PN oocytes resemble those from 2PN, while those from 1PN zygotes exhibit slower kinetics.

Trial registration number
not applicable