P-128 Non-invasive morphometric assessment of embryo quality: Effectiveness of the number of trophectoderm cells at the equatorial region of expanded blastocysts
Authors: Morishita N , Miura M , Kobayashi Y , Matsunaga R , Ochi M , Horiuchi T 
Published: July 2024

Abstract

Study question
Does the number of trophectoderm cells at the equatorial region of expanded blastocysts (eTEs) affects pregnancy rate in single vitrified blastocyst transfer (SVBT)?

Summary answer
The number of eTEs highly correlated with the total number of TEs. The greater number of eTEs was higher pregnancy rate became.

What is known already
The increased use of blastocyst transfer has emphasized the importance of selecting high-quality embryos. Morphological grading: Gardner’s criteria, or time-lapse imaging of embryo morpho kinetics is established a tool for selection of high-quality blastocysts. The Gardner’s criteria is one of the most used evaluation according to grading scores of developmental stage, trophectoderm (TE) and inner cell mass (ICM). Although, it cannot completely exclude subjectivity, and it varies from observer to observer. A quantitative evaluation method for blastocyst selection is necessary to solve these problems and to improve the pregnancy rate of SVBT.

Study design, size, duration
First, we checked the correlation between the number of eTEs and the total number of TEs using 23 discarded blastocysts by immunostaining of CDX2. Then, we analyzed 1103 vitrified-warmed blastocysts from July 2017 to June 2021 for pregnancy rates. Expanded blastocysts which reached the blastocoel cavity of 160 µm or more with obvious ICM were vitrified and cryopreserved. Relationship between the number of eTEs and pregnancy rate after SVBT was analyzed by logistic regression.

Participants/materials, setting, methods
CDX2-positive cells were defined as TE, and the difference between total cells by Hoechst staining and CDX2-positive cells as ICM. The correlation between the number of eTEs and the total number of TEs was examined. In SVBT, all embryos were monitored by a time-lapse system. eTEs were counted in the time-lapse image just before cryopreservation. Pregnancy and abortion rates were examined for each eTEs. Also, logistic regression analysis was performed on eTEs and pregnancy.

Main results and the role of chance
There was a strong positive correlation between the number of eTEs and the total number of TEs (CDX2-positive cells) in the whole blastocysts (r = 0.945). In addition, there was a significant correlation between the number of eTEs and the total cell number of blastocysts (r = 0.73). On the other hand, there was no correlation between the number of eTEs and the cell number of ICM (r = 0.008). The number of eTEs was presented as quartiles: Pregnancy rates for &lt;5, 5-6, 6-7 and ≥7 eTEs were 25.9, 29.4, 38.4, and 64.2% (P &lt; 0.05), respectively, and the miscarriage rates were 50.0, 28.3, 14.1 and 19.2% (P &lt; 0.05), respectively. Pregnancy rate for ≥7 eTEs was significantly higher than that for the other groups (P &lt; 0.05), and miscarriage rate was lower than that for &lt;6 eTEs. That is, increased eTEs significantly increased the pregnancy rate and decreased the miscarriage rate. Significant relationship between the number of eTEs and pregnancy was shown by logistic regression analysis (P &lt; 0.0001), with an AUC of 0.68 and a cutoff value of 6.5.

Limitations, reasons for caution
In this study, expanded blastocysts at 160 µm or more in diameter were analyzed and cryopreserved. The different size of blastocysts may be different number of eTEs, and the relationship between the number of eTEs and pregnancy rate will be needed to confirm in individual clinic.

Wider implications of the findings
Quantitative analysis of eTEs makes it possible to select highly fertile blastocysts. This assessment is simply and easy. It can be the additional possible evaluation before cryopreservation to allows for quantitative treatment of continuous variables by classifying them in more detail than Gardner’s criteria.

Trial registration number
N/A