Trophectoderm biopsies of blastocysts for preimplantation genetic testing following in vitro fertilization and embryo culture increases epigenetic dysregulation in a mouse model [RNA-seq]

Trophectoderm biopsy for preimplantation genetic testing (TEBx) impacts placental and embryonic health during early development, with some alterations resolving while others worsening later in development. We observed that at E12.5, IVF + TEBx had a worse outcome in terms of changes in DNA methylation and differential gene expression in placentas and whole embryos compared with IVF alone. These changes were reflected in alterations in placental morphology and blood vessel density. At E18.5, early changes in embryos were maintained or worsened, while molecular and morphological changes in the placenta were not different from the IVF group, except for changes in blood vessel density, which persisted. Of note is that most differences were sex specific. We conclude that TEBx has more detrimental effects in both mid-gestation placental and embryonic tissues, with changes in embryonic tissues persisting or worsening in later developmental stages compared to IVF alone. Additionally, consistent with previous work, we report sex differences in the observed effects. Finally, we identified that the addition of vitrification after TEBx has a more pronounced detrimental effect on placental and fetal health. Our findings provide additional support that more studies should be done to assess the impact of new procedures during ART to ensure healthy pregnancies and offspring outcomes. Gene expression analysis using Total RNA extracted from E12.5 placenta n=30 (F=15 and M=15), E12.5 whole fetus n=30 (F=15 and M=15), E18.5 placentas (F=15 and M=15) and E18.5 fetal livers (F=15 and M=15) from 3 different groups Natural, IVF and IVF+TEBx were sequenced using Illumina NovaSeq6000用于植入前遗传学检测的滋养层活检（Trophectoderm biopsy for preimplantation genetic testing，TEBx）会影响早期发育阶段的胎盘与胚胎健康，部分异常可在后续发育中得到修复，另有部分则随发育进程加重。我们观察到，在胚胎发育第12.5天（E12.5），相较于单纯体外受精（In Vitro Fertilization，IVF）组，体外受精联合TEBx组的胎盘及全胚胎的DNA甲基化改变与差异基因表达情况均表现出更差的结局。上述变化也体现在胎盘形态学与血管密度的改变上。至胚胎发育第18.5天（E18.5），胚胎早期出现的变化仍持续存在甚至进一步加重；而胎盘的分子与形态学变化则与单纯IVF组无显著差异，仅血管密度的改变仍持续存在。值得注意的是，绝大多数差异具有性别特异性。我们得出结论：相较于单纯IVF组，TEBx对妊娠中期的胎盘与胚胎组织均存在更显著的有害影响，且胚胎组织的相关变化在后续发育阶段仍持续存在甚至加重。此外，与既往研究结果一致，本研究也证实了观测到的效应存在性别差异。最后，我们发现TEBx术后联合玻璃化冷冻（vitrification）会对胎盘与胎儿健康产生更为显著的有害影响。本研究结果进一步支持需开展更多研究以评估辅助生殖技术（Assisted Reproductive Technology，ART）中新操作流程的影响，从而保障妊娠结局与子代健康。本研究针对3个实验组（自然妊娠组、单纯IVF组、IVF+TEBx组）的样本进行了基因表达分析：提取E12.5胎盘（n=30，雌鼠15例、雄鼠15例）、E12.5全胚胎（n=30，雌鼠15例、雄鼠15例）、E18.5胎盘（n=30，雌鼠15例、雄鼠15例）以及E18.5胎肝（n=30，雌鼠15例、雄鼠15例）的总RNA，随后采用Illumina NovaSeq6000平台进行测序。