Abstract
Preimplantation genetic diagnosis (PGD) has become an established and successful IVF procedure screening for specific chromosomal abnormalities in human embryos. PGD involves the biopsy of polar bodies or embryonic cells for genetic analysis. Though successful, biopsy procedures are invasive to the embryo, possibly compromising development, and time consuming for the embryology laboratory. With recent advances in the field of proteomics it has become possible to analyze the proteins produced and secreted by human embryos into the surrounding medium (secretome). The objective of this study was to investigate whether embryos with chromosomal abnormalities could be distinguished from normal embryos by their respective secretomes Prospective Study Cryopreserved day 1 embryos and aneuploid human blastocysts from PGD cycles were donated with consent for research. Day 1 embryos were thawed and subsequently cultured in sequential media (G1/G2) supplemented with recombinant human albumin at 37°C, 5% O2 & 6% CO2 up to the hatching blastocyst stage. Trophectoderm biopsy was performed removing up to 12 cells for FISH (chromosomes 13, 14, 15, 16, 17, 18, 21, 22, X and Y). Aneuploid blastocysts (abnormalities involving any of the analyzed chromosomes) from PGD cycles were cultured under these same conditions for 24 hrs. Drops of media cultured in the same dishes without blastocysts were used as controls. All spent media (aneuploid n=18, normal n=6) and controls (n=12) were subsequently processed and analyzed by time-of-flight mass spectrometry. The secretome profiles from hatching blastocysts identified as normal for the 10 chromosomes tested exhibited notably different secretome profiles to hatching blastocysts identified as aneuploid. Significantly, five proteins/biomarkers were observed to be in higher abundance in the secretome of hatching aneuploid blastocysts (P < 0.05). In addition, aneuploid embryos arrested in culture prior to the blastocyst stage also showed higher abundance of several proteins/biomarkers in their secretome profiles compared with hatching aneuploid blastocysts (P < 0.05). This is the first study to describe the differences in the secretome between human blastocysts identified as either normal or aneuploid for 10 chromosomes tested. This study has also shown that degenerating aneuploid embryos exhibit a significantly different secretome profile to hatching aneuploid blastocysts. This approach of protein analysis will further increase our understanding of human blastocyst physiology and its association with chromosomal composition. This work could lead to the development of a non-invasive assay to identify human embryos with chromosomal abnormalities.
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