Correlation among blastocoel fluid DNA level, apoptotic genes expression and preimplantation aneuploidy.

Abstract

It is believed that aneuploid embryos release cell-free DNA (cfDNA) into the blastocyst cavity during the self-correction process through the apoptotic mechanism. This study aimed to develop less invasive methods for predicting ploidy status by investigating how ploidy status affects blastocoel fluid DNA (BF-DNA) levels and apoptotic gene expression as indicators of embryo viability. Human blastocysts were classified into three groups; Survivable Embryo (SE), Fatal Single and double Aneuploidy (FSDA), and Multiple Aneuploidy (MA) using array comparative genomic hybridization (array-CGH) by trophectoderm (TE) biopsy. Following BFs aspiration and whole genome amplification (WGA), BF-DNA level was quantified. Apoptotic activity was assessed by measuring the genes TNFRSF10B, CASP2, BAX, and CASP3 using Real-Time quantitative PCR. Day-5 intra-cytoplasmic sperm injection blastocysts were scored according to the Gardner and Schoolcraft system. BF-DNA levels were significantly higher in the MA vs. SE group (P=.01), while these were not statistically significant differences between MA and FSDA groups, and also, FSDA and SE groups, P=.17 and P=.38, respectively. TNFRSF10B, CASP2, and CASP3 were overexpressed in the MA and FSDA groups vs. SE, while BAX was downregulated. We found a significant correlation between the amount of BF-DNA and apoptosis marker genes. No significant correlation was found between embryo morphology score and BF-DNA level, BF volume, or apoptosis marker gene expression levels. We observed that the correlation between apoptotic activity and BF-DNA levels is influenced by the embryo's ploidy status. These findings suggest that BF-DNA level evaluation can be applicable in selecting viable embryos for transfer.