542 | Nature | Vol 577 | 23 January 2020
Article
To compare cynomolgus and human EPI development, we analysed
scRNA-seq from monkey 6–17-d.p.f. EPIs^39. Principal component analy-
sis (PCA) revealed that cynomolgus and human cells separated along the
PC1, representing a major species difference (Extended Data Fig. 10a).
We identified genes with significantly positive or negative PC1 scores for
cynomolgus and human genes (Extended Data Fig. 10b, Supplementary
Table 7). Along the PC2 and PC3 axes, cynomolgus and human EPIs
cells were plotted to reflect their similar developmental transitions
and conserved signalling pathways and Gene Ontology (GO) terms
(Extended Data Fig. 10a, c, Supplementary Table 7). To examine species
differences, we compared the dynamics of naive genes and signalling
pathways over EPST in human and reported monkey data^39. In con-
trast to human EPST, TBX3 and SOX15 gradually decreased, and UTF1
and NR0B1 were absent during monkey EPST^39 (Fig. 4c, Extended Data
Fig. 10d, e). Pathway analysis for upregulated genes during the EPST
revealed a similar enrichment trend for NOTCH, BMP and FGF signal-
ling pathways in humans and monkeys^39 (Extended Data Fig. 10f–h).
However, LEFTY1, LEFTY2 and NODAL in BMP signalling displayed dif-
ferent patterns between human and monkey EPST (Extended Data
Fig. 10f ). Together, human and monkey EPIs had unique phenotypes
and similarities during development.
Discussion
Here, we report a 3D-culture system that successfully cultured human
blastocyst growing to the PSA stage. These 3D embryos can recapitu-
late almost all key 3D architectures and developmental landmarks of
in vivo pre-gastrulation embryos. By contrast, many 3D structures
and developmental landmarks—such as AME–EPI separation, base-
ment membrane, SYS, anterior visceral endoderm, anterior–posterior
polarity initiation and PSA—were not found using in vitro implantation
platforms of 2D-cultured embryos^1 ,^2 and using human pluripotent
stem cells that model early developmental embryonic events^6 –^8. Our
3D-cultured embryos recapitulated the timing and outcome of lineage
segregation and development, which more authentically mimicked
early human embryonic development in vivo (Fig. 4d).
Because human embryogenesis is not well understood, we estab-
lished a platform to delineate EPIs, AMEs, PrEs and TrBs. We revealed
the unique characteristics of AME as the first differentiated cell group
emerging from an expanding EPI population^8. We uncovered specific
pathways and transcription factors for TrB subtype separation and
functional differences between subtypes and between pre-gastrulation
TrBs and fetal TrBs^32. Unlike mice, human EPI after implantation main-
tains its transcriptional properties for a steady and prolonged period
while acquiring properties for neuron differentiation and vasculature
development. Overall, we reveal the molecular and morphogenetic
developmental landscape of pre-gastrulation human embryos. These
data provide crucial insights into the pluripotency of human pluripo-
tent stem cells and uncover stem-cell self-renewal and differentiation
processes, and will inform future strategies to improve in vitro ferti-
lization success rates.
Online content
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availability are available at https://doi.org/10.1038/s41586-019-1875-y.
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