The significance of this research
Two research groups, Yu et al. and Liu et al., have succeeded in generating a three-dimensional structure (blastoid) that resembles a human blastocyst (the stage after the fertilized egg has grown) from human iPS cells (iPSCs) in vitro, and their papers have been published in Nature. This means that a human embryo could be reconstructed from iPSCs in vitro, though imperfectly. A blastocyst is a spherical cell mass formed in the early development of living beings and from which the entire body of them is formed. Models of human blastocysts are needed to study human embryology. A proper understanding of early human development leads to improve assisted reproductive technologies and prevent pregnancy loss and congenital diseases.

Early human development
A few days after fertilization, the fertilized egg forms a blastocyst, which contains an outer cell layer called the trophectoderm and a cell cluster called the inner cell mass (ICM). The trophectoderm surrounds a cavity containing ICM. As the blastocyst develops, the ICM becomes segregated into two cell populations, the epiblast and the hypoblast. Then, the blastocyst implants into the uterine wall, setting the stage for an event called gastrulation. During gastrulation, epiblast cells give rise to the three basic cell layers, the ectoderm, mesoderm, and endoderm, that will form the entire fetus. The trophectoderm forms most of the placenta, and the hypoblast forms some layers of a structure called the yolk sac. The pre-implantation ICM and the post-implantation epiblast are pluripotent stem cells, which are capable to differentiate into all the cells of the body.

Studies about early human development
There are challenges of research on early development of the human body — few human embryos are available, and research is subject to ethical and legal constraints. Therefore, the development of techniques that use cells cultured in vitro to construct models of mammalian embryos has been promoted. In previous studies, human stem cells which are developmentally similar to post-implantation, pre-gastrulation epiblast cells were used for models of early human development. That is why cells associated with the trophectoderm, hypoblast or both were not formed, although some stages of post-implantation human development could be recapitulate.

Study 1: Generation of blastoids using embryonic or induced pluripotent stem cells
Yu et al. used either human embryonic stem cells, which are derived from blastocysts, or induced pluripotent stem cells, which are generated from adult cells, to generate blastoids. Both of these types of cell are developmentally similar to epiblast cells. By placing these cells in 3D culture dishes and treating the cells with two different types of culture medium, they have succeeded in generating blastoids that have the characteristics of epiblast cells and can differentiate into the trophectoderm and hypoblast.

Study2: Generation of blastoids using reprogrammed fibroblasts
Liu et al. reprogrammed adult skin cells called fibroblasts to generate a mixed cell population containing cells with gene-expression profiles similar to those of the epiblast, trophectoderm and hypoblast. These cells were then cultured in 3D culture dishes to generate blastoids.

Characteristics of blastoids generated in Study1 and Study2
In both studies, blastoids emerged after 6–8 days of culture, with a formation efficiency of up to almost 20%. The blastoids had a similar size and shape to human blastocysts, with a similar total number of cells. In the blastoids, a cavity and an ICM-like cluster were observed. Detailed analyses of the blastoids revealed that their cell lineages are molecularly similar to those of the pre-implantation human blastocyst. (A cell lineage denotes the developmental history of cells from a fertilized egg to an adult.) The groups also founded that the blastoid cells have key properties of blastocyst lineages. In other words, cells isolated from the blastoids could be used to generate various stem-cell types.

Next, an experiment to mimic implantation into the uterus was conducted in culture dishes. When blastoids were grown in a culture dishes for 4-5 days, some attached to the dish and continued to develop. In a part of these attached blastoids, cells representative of the epiblast were reorganized into a structure enclosing a cavity. This cavity resembles the pro-amniotic cavity, which forms in the post-implantation epiblast. In some blastoids, the trophectoderm-related cells showed signs of differentiation into specialized placental cell types. Yu et al. also reported a second cavity in the hypoblast-related cells in some blastoids, similar to the yolk-sac cavity.

These two studies demonstrate that human blastoids are valuable in vitro models of pre-implantation and early post-implantation blastocyst development.

Additional information for researchers
Differentiation into cells related to the epiblast, hypoblast, and trophectoderm was observed in the early post-implantation blastoids, but development of the blastoids is limited in post-implantation stages. Further optimization of culture protocols will be needed to improve post-implantation-stage culturing of human blastoids, up to the equivalent of 14 days in vivo. Today, culturing human embryos for longer than 14 days is internationally prohibited on ethical grounds (the 14-day rule). Whether this rule will be applied to blastoids is an important concern for future research. In response to the rapid progress in stem cell research, the International Society for Stem Cell Research (ISSCR) will release the society’s updated Guidelines for Stem Cell Research and Clinical Translation in May 2021. The guidelines will include guidance for stem cell-based embryo models.

Title: First complete model of the human embryo
Authors: First complete model of the human embryo