April 6, 2023

A study published in Cell Stem Cell looks at the generation of embryo-like structures from monkey embryonic stem cells.

Prof Magdalena Zernicka-Goetz, Bren Professor of Biology and Biological Engineering, California Institute of Technology; and Professor of Mammalian Development and Stem Cell Biology, University of Cambridge, said:

This is an exciting development building on work from our own and other labs showing the importance of establishing interactions between embryonic and extra-embryonic stem cells to establish models of the mammalian embryo at pre-and early post-implantation stages. The excitement of this study is not only that embryos generated from monkey stem cells provide a close model for human embryos, but monkeys are also experimentally tractable.

The authors follow approaches that have been previously used to direct embryonic stem cells into a naive state, and then use treatments that allow the nave monkey ES cells to form extra-embryonic cell types. Together these cells assemble into blastoids, structures resembling blastocysts, that are able to develop in vitro into structures with a striking resemblance to the embryonic disc at gastrulation, both in morphology and gene expression. The blastoids also appear to implant into foster monkey mothers but, in common with similar structures in the mouse, development appears restricted.

This study is a hugely encouraging development in the study of primate embryo models.

The paper is excellent and an important step forward but still the stem cell derived embryos have a limited developmental potential, as the authors state themselves. Nevertheless, it is an important step in the very exciting field of enormous potential for understanding how the embryo develops and why so many pregnancies fail.

Prof Roger Sturmey, Professor of Reproductive Medicine, Hull York Medical School, University of Hull, said:

The work by Li and colleagues is an impressive technical achievement that has demonstrated the possibility that embryonic stem cells from a primate can be persuaded to form structures that mirror many features of early embryos.

Similar achievements have already been reported in other species, however this work assesses the primate embryo-like structures in detail and gives new insights into how the cell lineages families of cells that constitute the early embryo can be generated from stem cells.

Remarkably, when cultured in a laboatory, the embryo-like structures are able to replicate a number of key developmental features, most notably the formation of cells that resemble the primordial germ cells the cells that can produce gametes as well as the formation of a structure similar to the so-called primitive streak. When transferred into a recipient macaque uterus, these embryo-like structures were able to generate components of a pregnancy response, but were unable to develop, indicating that while these structures do share many features with competent embryos, there are still aspects of early development that differ between competent embryos and stem-cell derived models, preventing full development.

The work by Li and colleagues will offer important new tools in our understanding of the earliest stages of embryo development, but also highlight the need for guidance in this area, something that scientists in the UK are actively working on.

Prof Alfonso Martinez Arias, ICREA Senior Research Professor, Department of Experimental and Health Sciences, Universitat Pompeu Fabra (UPF), said:

This is a timely study.

About half of human pregnancies fail during the proliferation of the zygote and the implantation of the blastocyst. Understanding the causes of this failure rate will impact human fertility and IVF success. In part to address this need, over the last few years, a number of Embryonic Stem (ES) cell models of early mammalian development have been created in the lab. Amidst these, mouse and human blastoids mimic mammalian blastocysts and as such can play an important role in understanding the process of implantation. Blastoids have been derived from mouse and human ES cells.

For these studies to go forward there is a need to develop a proper test for the function of the blastocyst: its implantation into the uterus. In the case of mouse blastoids this can be tested by implanting them into females. However, there is no such a test for human blastoids since, for obvious reasons, it is not possible to implant them into a human uterus. And yet there is a need to develop a system to study these structures in humans. Mouse reproductive biology and implantation are very different from human, which means that while an excellent system to find principles, the mouse is not useful for the specifics of this process; and this is what matters. It is this vacuum of a system to study human implantation and peri-implantation development that is addressed in the present study.

Following protocols established for human blastoids, macaque blastoids are made from nave stem cells and their potential is tested in two ways. One, by culturing them in vitro up to gastrulation stages and the other, by placing them in the uterus of a macaque foster mother. The idea behind this system is that it has reduced ethical barriers compared to human and therefore might provide an experimental system to test the potential of blastoids fully and, in the long term, to study infertility. The work is well conducted and the result is clear: although at the level of single cells macaque blastoids bear a strong resemblance to blastocysts, they do not behave as blastocysts. Although they implant and initiate gastrulation, they do not reach the end of this process. In vitro, blastoids cultured to form an epiblast and to undergo gastrulation, display progressive problems over time and, though they reach early stages of gastrulation, it is difficult to see in their data how faithful they are to an early gastrula. In one important experiment they implant some of these into female macaques and follow their progress with ultrasound. It appears as if they might perform well in the early stages of implantation, and the release of progesterone is a sign that something has gone well, but then, they disappear after about a week.

So, the important result of this work is that we are not close to generating blastoids that can be recognised as blastocysts by the mother. Definitely an important proof of principle but the lesson is that there is work to do.

An important difference between a blastoid and a blatocyst is their origin. The blastocyst in the egg, the blastoid in the ES cells. There might be elements in the oocyte that are important for the viability of the blastocysts and that will not be provided by the ES cells. Furthermore, if about 50% of conceptions fail at implantation, it is difficult to gauge whether the failure of the high level goal of the experiment (long term development in the womb) is due to defects in the blastoid system or whether the failure mirrors the natural situation; eight experimental subjects, the numbers of the experiment, are not sufficient to make a judgement. Only more experiments will decide and the one reported here, within well established ethical footprints, is definitely one to watch.

Dr Darius Widera, Associate Professor in Stem Cell Biology and Regenerative Medicine, University of Reading, said:

This is an interesting study that demonstrates the successful generation of embryo-like structures from monkey embryonic stem cells. These structures resembled natural early embryonic structures and could generate cell types of all three germ layers. Although similar studies have been conducted using human stem cells, this is the first report showing that (in this case, monkey) embryo-like structures can induce signs of pregnancy if transplanted into females. Therefore, the method could be used as a model of primate and human development and potentially provide new insights into certain factors that contribute to miscarriages in humans.

However, the study has some limitations. Only 3 out of 8 embryo-like structures were successfully implanted into female monkeys, and none of these persisted for more than one week. Thus, the structures do not have full developmental potential.

In addition, the ethical implications of embryonic stem cell research in monkeys are complex. Primates are intelligent, social animals with complex cognitive and emotional lives. Therefore, it is important to carefully consider both the potential benefits and the ethical impact of primate embryonic stem cell research.

Prof Robin Lovell-Badge FRS FMedSci, Group Leader, Francis Crick Institute, said:

The paper by Jie Li et al is another demonstration of the remarkable ability of pluripotent stem cells, in this case embryonic stem cells derived from early Macaque (non-human primate) embryos, to self-organise and begin a process of embryo formation in culture that mirrors that of normal Macaque embryos. However, the paper also shows that these stem cell-based embryo models are not entirely normal they could be implanted in female macaques, appear to initiate a pregnancy, but then fail soon after.

The authors were able to culture these stem cell-based embryo models, which they refer to as blastoids, through to gastrulation stages, equivalent to post-implantation embryos developing in a uterus, with good signs of development of all the main extraembryonic and embryonic tissues, where the latter included ectoderm, mesoderm and endoderm organised in a similar fashion to normal embryos. They could also demonstrate the presence of primordial germ cell-like cells and cells that are early progenitors of the blood system. These stages would be equivalent to those of human embryos at about 16 -18 days of development, beyond the 14 day limit (or the beginning of gastrulation) which is the maximum period normal human embryos are allowed to be cultured by law in the UK and some other countries.

It has been shown by others that human pluripotent stem cells can also be used to form blastoids, but to date such cultures have been stopped prior to gastrulation, but the paper by Li et al suggests that they could indeed be taken beyond this and provide valuable information about these early stages of human development that are otherwise very difficult to obtain. The data from the Macaque embryos and blastoid cultures may also help to understand aspects of human development, but without direct comparisons this will always be tentative, given how much mammalian embryos can vary at these stages.

These embryo models are referred to as integrated stem cell-based embryo models because they include extraembryonic tissues that normally give rise to the placenta and yolk sac that in a normal conceptus would permit implantation into the uterus and support the development of the embryo proper. So how much like a real embryo are these Macaque blastoids and could they implant and develop much further in a uterus? Although all the detailed comparisons presented in the paper of gene expression in the various cell types between normal Macaque embryos and the embryo models suggests that they can be very similar, the proportion of the blastoids reaching advanced stages was very low, indicating that most are not normal, and those that did still showed some differences. Moreover, while some could implant, begin to develop some complexity, and induce a typical response in the host uterus and lead to production of the typical pregnancy hormones, chorionic gonadotrophin and progesterone, the embryos all failed before gastrulation. This suggests that they failed to form fully functional extraembryonic tissues that could adequately support the embryo and that these could not give rise to a placenta, which would be essential for more complex development. It is likely that the same would be true for human integrated stem cell-based embryo models, although it would be unethical and illegal (in the UK) to attempt to implant these into a woman.

It seems likely that the culture methods for these integrated stem cell-based embryo models will be improved, and who knows it may eventually be possible to have them implant and develop normally, but the failure of this to happen as reported in this paper will give regulators some breathing space to develop appropriate rules for the culture of such human models, notably whether they can be taken beyond the equivalent of gastrulation stages, which would be of immense importance in helping to understand not just normal development of the human embryo, but what so often goes wrong and leads to embryo failure and congenital disorders.

Cynomolgus monkey embryo model captures gastrulation and early pregnancy by Jie Li et al. was published in Cell Stem Cell at 16:00 UK time on Thursday 6 April 2023.

DOI: 10.1016/j.stem.2023.03.009

Declared interests

Prof Magdalena Zernicka-Goetz: I have no conflict of interest to declare.

Prof Roger Sturmey: None.

Prof Alfonso Martinez Arias: I have no conflict of interests.

Dr Darius Widera: I have no conflict of interest to declare.

Prof Robin Lovell-Badge: I have no conflicts of interest to declare, except I do serve on the HFEAs Scientific and Clinical Advances Advisory Committee and I am a member of their Legislative Reform Advisory Group.

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