P-743 Semen samples quality and blastocysts diversity after PGT-A
Authors: Albanese C , Perruzza D , Azzena S , Crippa A , Graziosi S , Bilotta G , Magli M C , Gianaroli L 
Published: June 2023

Abstract

Study question
Does semen samples quality contribute to blastocyst diversity by correlating with the presence of DNA in the blastocoelic fluid (BF)?

Summary answer
Semen samples quality is associated with the detection of DNA in BFs from both euploid and aneuploid blastocysts

What is known already
DNA was detected by whole genome amplification (WGA) in BFs from expanded blastocysts dependently on the blastocyst chromosome condition (assessed by PGT-A on trophectoderm (TE) cells), being more frequent in aneuploid blastocysts. TE-euploid blastocysts with positive BF-WGA also seem to have less chances to implant compared to those with failed BF-WGA. The extrusion of abnormal cells from the embryo proper was proposed as a mechanism triggered by the mosaic embryo to eliminate abnormal cells. Severe male factor samples are at risk of causing embryo mosaicism. Possible association between semen quality and DNA in BFs has not yet been investigated.

Study design, size, duration
This retrospective cohort study included 115 patients (maternal age 37.9±4.1), which underwent PGT-A in the last three years. The aim of the study was to evaluate whether the detection of DNA in BFs was correlated with semen samples indices. Both TE-euploid and TE-aneuploid blastocysts were considered for the analysis and stratified according to the semen samples quality as normozoospermic (N), moderate oligoasthenoteratozoospermic (m-OAT) and severe OAT (s-OAT) following the 2021 WHO criteria.

Participants/materials, setting, methods
BF and trophectoderm (TE) biopsies were collected from high grade expanded blastocysts before vitrification, and submitted to WGA. Amplification after WGA was evaluated by loading an aliquot of the amplified product onto a 1.5% agarose gel. 24-chromosome analysis was performed on TE biopsies. To assess the impact of semen samples quality on the detection of DNA in BFs, a multiple logistic regression analysis was performed correcting for maternal age as a confounding factor.

Main results and the role of chance
The BF from 590 blastocysts (265 TE-euploid and 325 TE-aneuploid) were submitted to WGA. In TE-euploid blastocysts, 125 derived from N semen samples, 52 from m-OAT, and 88 from s-OAT. The incidence of positive BF-WGA was proportional to the severity of the male factor condition and occurred in 48.0% of blastocysts originated from N samples, in 55.8% of blastocysts derived from m-OAT, and in 60.2% of blastocysts from s-OAT. In TE-aneuploid blastocysts, 178 originated from N semen samples, 80 from m-OAT, and 67 from s-OAT. The incidence of positive BF-WGA showed an opposite trend compared with the TE-euploid group being detected in 64.0% of blastocysts originated from N samples, in 66.3% of blastocysts derived from m-OAT, and in 52.2% of blastocysts obtained from s-OAT. The relationship between semen quality and positive BF-DNA amplification was confirmed by a multiple logistic regression model setting the BF-WGA as the outcome variable. In the TE-euploid blastocyst cohort, s-OAT showed an odd ratio (OR) of 1.78 (95% CI: 1.02-3.14; p=0.045), whereas in the TE-aneuploid blastocyst group, s-OAT displayed an OR of 0.55 (95% CI: 0.31-0.98; p=0.043).

Limitations, reasons for caution
This is a retrospective cohort study including a limited number of cases where the different categories of sperm samples were not equally represented. Although the results of BF-WGA were evaluated per single blastocyst, they cannot be considered independent variables.

Wider implications of the findings
In s-OAT samples, abnormal cell elimination is especially active in TE-euploid blastocysts, whereas in TE-aneuploid blastocysts the efficiency of this process is reduced suggesting the difficulty of eliminating abnormal cells in an aneuploid background. BF-DNA amplification is a minimally invasive tool contributing additional knowledge to elucidate blastocyst diversity.

Trial registration number
None