Karyotype assessment of whole human blastocysts by aCGH

The use of in vitro fertilization (IVF) has revolutionized the treatment of infertility and is now responsible for 1-5% of all births in industrialized countries. During IVF, it is typical for patients to generate multiple embryos. However, only a small proportion of them possess the genetic and metabolic requirements needed in order to produce a healthy pregnancy. The identification of the embryo with the greatest developmental capacity represents a major challenge for fertility clinics. Current methods for the assessment of embryo competence are proven inefficient and the inadvertent transfer of non-viable embryos is the principal reason why most IVF treatments (approximately two-thirds) end in failure. In this study, we investigate how the application of proteomic measurements could improve success rates in clinical embryology. We describe a procedure that allows the identification and quantification of proteins of embryonic origin, present in attomole concentrations in the blastocoel, the enclosed fluid filled cavity that forms within five-day old human embryos. By using targeted proteomics, we demonstrate the feasibility of quantifying multiple proteins in samples derived from single blastocoels, and that such measurements correlate with aspects of embryo viability, such as chromosomal (ploidy) status. This study illustrates the potential of high-sensitivity proteomics to measure clinically relevant biomarkers in minute samples and, more specifically, suggests that key aspects of embryo competence could be measured using a proteomic-based strategy, with negligible risk of harm to the living embryo. Our work paves the way for the development of “next-generation” embryo competence assessment strategies, based on functional proteomics. This subset of experiments were used to determine embryonic karyotype. Differences in proteomic profiles of euploid and aneuploid embryos were then investigated. Karyotype was used as a marker for embryo viability to associate with blastocoel proteomic profiles generated using targeted mass spectrometry.

体外受精（in vitro fertilization, IVF）技术彻底革新了不孕症的治疗方案，目前在工业化国家中，经IVF诞生的婴儿占所有新生儿的1%至5%。在IVF流程中，患者通常会获得多枚胚胎，但仅有极少数胚胎具备健康妊娠所需的遗传与代谢条件。筛选出发育潜能最优的胚胎，是生殖诊所面临的核心挑战之一。当前用于评估胚胎发育潜能的方法被证实效率低下，而无意移植无存活能力胚胎，则是约三分之二的IVF治疗最终失败的主要原因。本研究探讨了蛋白质组学检测技术能否提升临床胚胎学的治疗成功率。我们建立了一套实验流程，可实现对囊胚腔（blastocoel，即5日龄人类胚胎内部形成的封闭液性空腔）中胚胎来源蛋白质的鉴定与定量，这类蛋白质的浓度仅为阿托摩尔级别。通过靶向蛋白质组学（targeted proteomics）技术，我们证实了从单个囊胚腔样本中定量多种蛋白质的可行性，且此类检测结果与胚胎存活能力的相关指标（如染色体倍性状态）存在关联。本研究证明了高灵敏度蛋白质组学（high-sensitivity proteomics）可用于在微量样本中检测临床相关生物标志物（clinically relevant biomarkers），更具体地说，该研究表明借助基于蛋白质组学的策略，可对胚胎发育潜能的关键特征进行评估，且不会对活胚胎造成可检测的损伤。本研究为开发基于功能蛋白质组学（functional proteomics）的“下一代”胚胎发育潜能评估方案铺平了道路。本研究中的部分实验用于确定胚胎核型（embryonic karyotype），随后我们对整倍体（euploid）与非整倍体（aneuploid）胚胎的蛋白质组学特征差异展开了探究。本研究以核型作为胚胎存活能力的标记物，将其与通过靶向质谱法（targeted mass spectrometry）获得的囊胚腔蛋白质组学特征进行关联分析。