Available online at www.sciencedirect.com ScienceDirect Synthetic human embryology: towards a quantitative future 1 2,3,4 Yue Shao and Jianping Fu Study of early human embryo development is essential for interest, as it is not only linked to reproductive health, advancing reproductive and regenerative medicine. Traditional but is also key to tissue engineering and regenerative human embryological studies rely on embryonic tissue medicine. Despite its scientific and clinical significance, specimens, which are difficult to acquire due to technical human embryo development remains largely mysterious. challenges and ethical restrictions. The availability of human Most of our knowledge about human development stem cells with developmental potentials comparable to pre- comes from limited histological specimens and medical implantation and peri-implantation human embryonic and images, preventing detailed understanding of human extraembryonic cells, together with properly engineered in vitro development. culture environments, allow for the first time researchers to in vitro generate self-organized multicellular structures that Although recent success in prolonging in vitro culture of in vivo mimic the structural and molecular features of their human embryo has generated new insights on human counterparts. The development of these stem cell-based, development [1–3], it is limited to a narrow time window synthetic human embryo models offers a paradigm-shifting due to ethical restrictions, namely, the 14-day rule [4], experimental system for quantitative measurements and prohibiting studies of post-implantation developmental perturbations of multicellular development, critical for stages such as gastrulation, neurulation, and beyond. advancing human embryology and reproductive and While in vitro culture of mouse and non-human primate regenerative medicine without using intact human embryos. embryos are not subject to this ethical restriction and thereby have enabled studies of peri-gastrulation devel- Addresses opment [5–9], clear evidence of interspecies divergence 1 Institute of Biomechanics and Medical Engineering, Department of between humans and mice [10,11] or non-human Engineering Mechanics, School of Aerospace Engineering, Tsinghua primates [3,12] calls for caution when extrapolating University, Beijing 100084, China 2 Department of Mechanical Engineering, University of Michigan, Ann knowledge from other mammals to humans. Further- Arbor, MI 48109, USA more, the limited availability of mammalian embryos, 3 Department of Cell and Developmental Biology, University of Michigan especially from non-human primates, prevents large-scale Medical School, Ann Arbor, MI 48109, USA 4 genetic studies or screens. Department of Biomedical Engineering, University of Michigan, Ann Arbor, MI, 48109, USA Recently, several groups, ours included, have success- Corresponding authors: fully reconstructed human embryogenic events in vitro Shao, Yue ([email protected]), Fu, Jianping ([email protected]) using human pluripotent stem cells (hPSC), an unlim- ited cell source with developmental potential matching Current Opinion in Genetics & Development 2020, 63:30–35 that of the pluripotent epiblast in the peri-implantation human embryo. By creating properly engineered culture This review comes from a themed issue on Dev. mechanisms, pat- environments, hPSC have now been applied to create terning and evolution multicellular structures to model the human amniotic sac Edited by Richard W Carthew and Amy Shyer formation, gastrulation, and neurulation. These stem cell-based human embryo models form the basis of the emerging field of synthetic human embryology, https://doi.org/10.1016/j.gde.2020.02.013 wherein different aspects of human embryo develop- in vitro 0959-437X/ã 2020 Elsevier Ltd. All rights reserved. ment are recreated and studied in a controllable and quantitative manner. Besides as an integrative area combining stem cell and developmental biology and human embryology, the advances in synthetic human embryology now have a new edge. By merging with designer bioengineering technologies and computational Introduction modeling, it is now possible to quantitatively control in vitro For centuries, the development of the human embryo has dynamic developmental niche and recapitulate in silico attracted great interest, especially on how it could give intricate developmental patterning . In this rise to the shape, size, function, and intelligence that review, we summarize the progress of this emerging make humans unique in the Mammalia class. Human field and its rapid advancement towards a quantitative embryo development also elicits significant clinical future. Current Opinion in Genetics & Development 2020, 63:30–35 www.sciencedirect.com Synthetic human embryology Shao and Fu 31 Models of human amniotic sac development including the specification of primordial germ cells. Upon implantation, the first developmental milestone of The microfluidic amniotic sac model is compatible with the human embryo is the formation of the asymmetric, live imaging, thus allowing tracking developmental lumenal amniotic sac from the epiblast of the human signaling dynamics. Given its controllability and scal- blastocyst. The amniotic sac is enveloped by the squa- ability, the microfluidic amniotic sac model could assist mous amniotic ectoderm at the dorsal pole and the in high-throughput drug and toxicity screens. Leverag- columnar epiblast at the ventral pole, respectively. The ing bioengineering tools for scalable tissue patterning, development of the amniotic sac is initiated by an apical another amniotic microtissue array system was recently lumen formation in the initial ball of pluripotent epiblast, reported [21], providing another promising platform for followed by patterning of the amniotic ectoderm at its high-throughput toxicity screens. dorsal pole [13] (Figure 1). Being an embryogenic event too early to detect, the development of the amniotic sac in Models of human gastrulation humans has been inaccessible for study until very After the amniotic sac formation, gastrulation is the next recently. Two recent studies report that hPSC have an developmental milestone. Gastrulation is initiated by the intrinsic lumenogenic property consistent with that of the formation of the primitive streak, culminating in the peri-implantation epiblast in vivo [2,14]. A hierarchical delineation of the three germ layers. The first human lumenogenic program in hPSC was further revealed, gastrulation model, termed ‘gastruloid’, was developed by mediated by actomyosin tension and driven by fusion culturing hPSC on 2D adhesive micropatterns of various of multiple pre-formed extracellular lumens or intracel- sizes. Treatments of 2D hPSC colonies with BMP4 lular apicosomes [15]. Importantly, such a ‘vesicle-fusion’ resulted in the emergence of concentric regions expres- mechanism of lumenogenesis was recently shown in both sing SOX2, T, SOX17, and CDX2, from colony center to mouse and human embryos [16,17]. Lumenal, pluripotent edge, mimicking the development of the three germ epithelial sacs of different shapes have also been gener- layers as well as the trophectoderm [22 ,23]. However, ated by culturing hPSC onto 2D culture surfaces contain- the gastruloid bypasses a few essential steps of gastrula- ing microscale grooves or trenches [14]. tion, namely, the formation of the primitive streak and its organizer. By replacing exogenous BMP4 with WNT3A, To recapitulate amniotic ectoderm development, two a morphologically distinguishable primitive streak-like culture systems were developed recently with tunable structure develops next to SOX2+ cells in the gastruloid mechanical stiffness of the culture environment [18 ,19 ]. [24 ]. Modulation of ACTIVIN-NODAL signaling in this Using both systems, a threshold of mechanical stiffness of streak model showed biased induction of posterior versus the culture environment was shown to control symmetry anterior primitive streak-like structures, as reflected by breaking of spherical lumenal hPSC tissues, resulting in differential SNAIL expression. Importantly, anterior prim- spontaneous differentiation of hPSC into squamous amni- itive streak-like structure induced in the model is GSC+, an otic cells and the formation of an asymmetric amniotic organizer marker. When such anterior primitive streak-like sac-like structure. Such a mechanical cell fate switch for structure was grafted into host chick embryos, it induced a amniotic differentiation of hPSC provides a potential secondary axis and neural tissue, confirming its role as an explanation for the absence of extraembryonic BMP organizer [24 ]. sources adjacent to the prospective amniotic ectoderm in vivo [13]. Amniogenic differentiation of hPSC requires Although 2D human gastruloid models provide trackable endogenous BMP-SMAD signaling [18 ]. By adjusting systems to study human gastrulation, they still deviate the initial cell number within each hPSC colony, syn- significantly from the actual human embryo in term of 3D thetic human amniotic sac model was created that not topology. To address this limitation, lumenal, pluripotent only resemble post-implantation amniotic sac develop- epithelial tissues formed by hPSC have recently been ment, but sequentially exhibit gastrulation-like features utilized for modeling human gastrulation [25 ]. With [19 ]. To date, this synthetic amniotic sac model remains precisely controlled, uniform BMP4 treatment, the 3D the only system that can recapitulate spontaneous