Glinsky Gennadi, Durruthy-Durruthy Jens, Wossidlo Mark, Grow Edward J, Weirather Jason L, Au Kin Fai, Wysocka Joanna, Sebastiano Vittorio
Institute of Engineering in Medicine, University of California, San Diego, 9500 Gilman Dr. MC 0435, La Jolla, CA 92093-0435, USA.
Department of Obstetrics and Gynecology, Institute for Stem Cell Biology & Regenerative Medicine, Stanford University, Stanford, CA 94305, USA.
Heliyon. 2018 Jun 28;4(6):e00667. doi: 10.1016/j.heliyon.2018.e00667. eCollection 2018 Jun.
Chromosome instability and aneuploidies occur very frequently in human embryos, impairing proper embryogenesis and leading to cell cycle arrest, loss of cell viability, and developmental failures in 50-80% of cleavage-stage embryos. This high frequency of cellular extinction events represents a significant experimental obstacle challenging analyses of individual cells isolated from human preimplantation embryos. We carried out single cell expression profiling of 241 individual cells recovered from 32 human embryos during the early and late stages of viable human blastocyst (VHB) differentiation. Classification of embryonic cells was performed solely based on expression patterns of human pluripotency-associated transcripts (), which represent a family of primate-specific transposable element-derived lincRNAs highly expressed in human embryonic stem cells and regulating nuclear reprogramming and pluripotency induction. We then validated our findings by analyzing transcriptomes of 1,708 individual cells recovered from more than 100 human embryos and 259 mouse cells from more than 40 mouse embryos at different stages of preimplantation embryogenesis. 's expression-guided spatiotemporal reconstruction of human embryonic development inferred from single-cell expression analysis of VHB differentiation enabled identification of telomerase-positive embryonic cells co-expressing key pluripotency regulatory genes and genetic markers of three major lineages. Follow-up validation analyses confirmed the emergence in human embryos prior to lineage segregation of telomerase-positive cells co-expressing genetic markers of multiple lineages. Observations reported in this contribution support the hypothesis of a developmental pathway of creation embryonic lineages and extraembryonic tissues from telomerase-positive pre-lineage cells manifesting multi-lineage precursor phenotype.
染色体不稳定性和非整倍体在人类胚胎中非常频繁地发生，损害正常的胚胎发育，并导致50-80%的卵裂期胚胎出现细胞周期停滞、细胞活力丧失和发育失败。这种高频率的细胞消亡事件是一个重大的实验障碍，挑战了对从人类植入前胚胎中分离的单个细胞的分析。我们对从32个人类胚胎中回收的241个单个细胞进行了单细胞表达谱分析，这些胚胎处于可行人类囊胚（VHB）分化的早期和晚期。胚胎细胞的分类完全基于人类多能性相关转录本的表达模式，这些转录本代表了一类灵长类特异性转座元件衍生的长链非编码RNA，在人类胚胎干细胞中高度表达，并调节核重编程和多能性诱导。然后，我们通过分析从100多个人类胚胎中回收的1708个单个细胞以及从40多个处于植入前胚胎发育不同阶段的小鼠胚胎中回收的259个小鼠细胞的转录组，验证了我们的发现。通过对VHB分化的单细胞表达分析推断出的人类胚胎发育的时空重建，能够识别共表达关键多能性调节基因和三个主要谱系遗传标记的端粒酶阳性胚胎细胞。后续的验证分析证实，在谱系分离之前，人类胚胎中出现了共表达多个谱系遗传标记的端粒酶阳性细胞。本研究报告的观察结果支持了从表现出多谱系前体表型的端粒酶阳性前谱系细胞产生胚胎谱系和胚外组织的发育途径假说。
