O-205 Aneuploidy induces proteotoxic stress and autophagy-mediated apoptosis in human preimplantation embryos

Abstract Study question Are aneuploid cells in human preimplantation embryos eliminated by apoptosis due to proteotoxic stress and autophagy-mediated apoptosis? Summary answer Proteotoxic stress, autophagy and apoptosis are differentially activated in aneuploid embryos, showing that aneuploid cells are eliminated by these mechanisms during early human embryogenesis. What is known already Aneuploidies are a common feature of human preimplantation embryos which could explain low success rates after in vitro fertilization (IVF). While most aneuploidies of meiotic origin are detrimental, transfer of euploid-aneuploid mosaic embryos can lead to healthy live-births. Moreover, the proportion of aneuploid cells are lower in blastocysts when compared to cleavage stage embryos. In the mouse, aneuploid cells are eliminated from the epiblast by autophagy-mediated apoptosis in a p53-dependent manner. We propose that in human embryos, aneuploidy causes chronic protein misfolding which leads to autophagy-induced apoptosis. Study design, size, duration Eighty-one blastocysts that were diagnosed by PGT as euploid (n = 49) or uniformly combined abnormal (CA, n = 32), i.e. 2 or more chromosomes were abnormal in every cell, were warmed. Sixty-seven were suitable for trophectoderm (TE) biopsy, 54 biopsies were successfully tubed and sent for RNA-sequencing while the remainder of the embryos was fixed for immunostaining. Thirty-three day-3 embryos were overnight incubated in 0.5µM reversine allowed to develop into blastocysts and treated as the PGT embryos. Participants/materials, setting, methods After TE biopsy, we live-stained the embryos with either Caspase-3/7 or 8 and subsequently fixed them. The biopsies underwent RNA-sequencing using the SMART-seqv4 and the fixed embryos were immunostained for LC3B, p62 (autophagy) and HSP70 (proteotoxic stress). Confocal imaging was performed using a Zeiss LSM800 confocal microscope and the presence of signal was quantified using the Zen Blue 2.0 and Arivis software. Main results and the role of chance Forty-two percent of the embryos in which we induced aneuploidies using reversine developed into blastocysts, which is comparable to untreated embryos. After immunostaining, we observed that CA and reversine-treated (RT) embryos contained less cells than euploid embryos (median number of nuclei: 43.5, 47, 90, respectively). This correlates with a higher expression of apoptotic markers Caspase-3/7 in CA embryos (p = 0.0199) and Caspase-8