The human embryonic genome is karyotypically complex, with chromosomally abnormal cells preferentially located away from the developing fetus
The human embryonic genome is karyotypically complex, with chromosomally abnormal cells preferentially located away from the developing fetus
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DOI：
10.1093/humrep/deac238
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发表时间：
2022-11-09
影响因子：
6.1
通讯作者：
Kearns, W. G.
中科院分区：
文献类型：
--
作者：
Griffin, D. K.;Brezina, P. R.;Kearns, W. G.
研究问题受精后第3天检测到的染色体异常是否主要保留在胚泡结构中，而不是内细胞团(ICM)，其中染色体正常的细胞优先保留？在人类胚胎中，非整倍体细胞在第3天到第5天的发育过程中，从ICM分离出来，部分地隔离到滋养外胚层(TE)，但更重要的是隔离到胚腔内的胚腔液(BC)和囊胚周围的外周细胞(PC)。已知的是，在所有二倍体真核生物中，一个普遍持有的教条是两个配子，每个配子都有n‘染色体(人类为23)，融合形成一个’2n‘合子(人类为46)；对于所有体细胞分裂来说，这种状态保持不变。然而，人类胚胎在早期阶段表现出高水平的染色体非整倍体，据报道，受精后第三天(卵裂期)到第五天(囊胚)染色体非整倍体水平下降。虽然这种观察可能部分是由于胚泡形成前的非整倍体胚胎停滞，但也可能是由于胚胎在胚泡形成过程中‘正常化’到整倍体状态。这种正常化是否会发生以及如何发生，需要进一步调查。研究设计、大小、持续时间共对964个卵裂期(第3天)胚胎进行了单细胞活组织检查和染色体构成诊断。所有细胞都保存在培养中，评估整倍体和非整倍体的胚胎率。对那些被确定为整倍体、胚胎化并随后转移的人的妊娠率进行了评估。对已确定的非整倍体和囊胚(174枚)，进行了ICM(全部174枚)、TE(全部174枚)、BC(47枚)和PC(38枚)的染色体组成分析。具体地说，评估了与最初的第三天诊断和确定是否在所有四个结构中有任何‘标准化’到整倍体核型的一致性。对象/材料、地点、方法所有患者(144对夫妇)均在三个IVF临床环境中进行常规植入前非整倍体遗传学检测。卵裂期活检先于下一代测序进行染色体分析。所有患者均提供知情同意。对囊胚进行了额外的分子测试，并对多达四种胚胎结构(ICM、TE、BC和PC)进行了分析。主要结果和463/964个胚胎(48%)在第3天被诊断为整倍体、70%囊胚化(导致59%的妊娠率)和30%退化的机会的作用。相反，在被诊断为非整倍体的501例(52%)中，65%是退化的，35%(174%)是胚胎化的，有非常显著的差异(P<；0.0001)。在174枚囊胚率中，ICM组为39%/57%/3%，TE组为49%/48%/3%，PC组为78%/21%/0%，BC组为83%/10%/5%。因此，TE核型对判断ICM的阳性预测值为86.7%，尽管异常率略高(P=.071)。BC/PC的异常水平显著高于ICM和TE的倍体(P<；0.0001)，几乎所有的染色体异常(至少部分)与第3天的诊断一致。局限性，谨慎的理由结果只与人类试管受精胚胎有关，因此对体内情况和其他物种的推断并不确定。我们承认(而不是我们这里建议的谱系特有的存活)在胚泡形成过程中存在其他机制的可能性，例如谱系特有的细胞运动。然而，伦理方面的考虑使得在人类胚胎上研究这一机制变得困难。这一发现的更广泛的含义是，马赛克人类卵裂阶段的胚胎可以分化为整倍体的ICM，其中整倍体细胞群占主导地位。将非整倍体细胞/核隔离到不再参与胎儿发育的结构中，对植入前和产前基因测试具有重要意义。这些结果也挑战了以前对人类早期发育中有丝分裂保真度的基本理解，并表明人类胚胎基因组的复杂和流动的性质。研究资金/竞争利益(S)这项研究由Organon制药公司和默克公司通过拨款给W.G.K.资助。W.G.K.也是AdvaGenix的员工，后者可能会从这篇手稿的出版中间接获得经济利益。RCM由美国国立卫生研究院国家普通医学科学研究所资助，获奖号为R35GM133747。内容完全由作者负责，不一定代表美国国立卫生研究院的官方观点。DK.G.为关爱生育提供有偿咨询服务。
STUDY QUESTION Are chromosome abnormalities detected at Day 3 post-fertilization predominantly retained in structures of the blastocyst other than the inner cell mass (ICM), where chromosomally normal cells are preferentially retained? SUMMARY ANSWER In human embryos, aneuploid cells are sequestered away from the ICM, partly to the trophectoderm (TE) but more significantly to the blastocoel fluid within the blastocoel cavity (Bc) and to peripheral cells (PCs) surrounding the blastocyst during Day 3 to Day 5 progression. WHAT IS KNOWN ALREADY A commonly held dogma in all diploid eukaryotes is that two gametes, each with 'n' chromosomes (23 in humans), fuse to form a '2n' zygote (46 in humans); a state that remains in perpetuity for all somatic cell divisions. Human embryos, however, display high levels of chromosomal aneuploidy in early stages that reportedly declines from Day 3 (cleavage stage) to Day 5 (blastocyst) post-fertilization. While this observation may be partly because of aneuploid embryonic arrest before blastulation, it could also be due to embryo 'normalization' to a euploid state during blastulation. If and how this normalization occurs requires further investigation. STUDY DESIGN, SIZE, DURATION A total of 964 cleavage-stage (Day 3) embryos underwent single-cell biopsy and diagnosis for chromosome constitution. All were maintained in culture, assessing blastulation rate, both for those assessed euploid and aneuploid. Pregnancy rate was assessed for those determined euploid, blastulated and subsequently transferred. For those determined aneuploid and blastulated (174 embryos), ICM (all 174 embryos), TE (all 174), Bc (47 embryos) and PC (38 embryos) were analyzed for chromosome constitution. Specifically, concordance with the original Day 3 diagnosis and determination if any 'normalized' to euploid karyotypes within all four structures was assessed. PARTICIPANTS/MATERIALS, SETTING, METHODS All patients (144 couples) were undergoing routine preimplantation genetic testing for aneuploidy in three IVF clinical settings. Cleavage-stage biopsy preceded chromosome analysis by next-generation sequencing. All patients provided informed consent. Additional molecular testing was carried out on blastocyst embryos and was analyzed for up to four embryonic structures (ICM, TE, Bc and PC). MAIN RESULTS AND THE ROLE OF CHANCE Of 463/964 embryos (48%) diagnosed as euploid at Day 3, 70% blastulated (leading to a 59% pregnancy rate) and 30% degenerated. Conversely, of the 501 (52%) diagnosed as aneuploid, 65% degenerated and 35% (174) blastulated, a highly significant difference (P < 0.0001). Of the 174 that blastulated, the ratio of '(semi)concordant-aneuploid' versus 'normalized-euploid' versus 'other-aneuploid' embryos was, respectively, 39%/57%/3% in the ICM; 49%/48%/3% in the TE; 78%/21%/0% in the PC; and 83%/10%/5% in the Bc. The TE karyotype therefore has a positive predictive value of 86.7% in determining that of the ICM, albeit with marginally higher aneuploid rates of abnormalities (P = .071). Levels of abnormality in Bc/PC were significantly higher (P < 0.0001) versus the ploidy of the ICM and TE and nearly all chromosome abnormalities were (at least partially) concordant with Day 3 diagnoses. LIMITATIONS, REASONS FOR CAUTION The results only pertain to human IVF embryos so extrapolation to the in vivo situation and to other species is not certain.We acknowledge (rather than lineage-specific survival, as we suggest here) the possibility of other mechanisms, such as lineage-specific movement of cells, during blastulation. Ethical considerations, however, make investigating this mechanism difficult on human embryos. WIDER IMPLICATIONS OF THE FINDINGS Mosaic human cleavage-stage embryos can differentiate into a euploid ICM where euploid cell populations predominate. Sequestering of aneuploid cells/nuclei to structures no longer involved in fetal development has important implications for preimplantation and prenatal genetic testing. These results also challenge previous fundamental understandings of mitotic fidelity in early human development and indicate a complex and fluid nature of the human embryonic genome. STUDY FUNDING/COMPETING INTEREST(S) This research was funded by Organon Pharmaceuticals and Merck Serono by grants to W.G.K. W.G.K. is also an employee of AdvaGenix, who could, potentially, indirectly benefit financially from publication of this manuscript. R.C.M. is supported by the National Institute of General Medical Sciences of the National Institutes of Health under award number R35GM133747. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. D.K.G. provides paid consultancy services for Care Fertility.