In their efforts to understand the very early stages of life and how they can go wrong, scientists face ethical issues surrounding the use of human embryos. The use of animal embryos is also subject to restrictions based on ethical considerations. To overcome these limitations, scientists have attempted to recreate early embryos using stem cells.
One of the challenges of creating these so-called synthetic embryos is to generate all of the cell types that are normally found in a young embryo before it implants itself in the lining of the uterus. Some of these cells eventually give rise to the placenta. Others become the amniotic sac in which the fetus grows. Both the placenta and the amniotic sac are essential for the survival of the fetus, and defects in these embryonic components are the main causes of early miscarriage.
A group of scientists from the Gladstone Institutes, the Center for iPS Cell Research and Application (CiRA) at Kyoto University and the RIKEN Center for Biosystems Dynamics Research in Kobe, Japan, have now demonstrated the presence of precursors of the placenta and amniotic sac in synthetic embryos they created from mouse stem cells.
“Our results provide strong evidence that our system is a good model for studying the early pre-implantation stages of embryonic development,” says Kiichiro Tomoda, PhD, researcher at the newly opened iPS Cell Research Center in Gladstone and first author of the study published in the journal Stem cell reports. “Using this model, we will be able to dissect the molecular events that take place during these early stages, and the signals that different embryonic cells send to each other.”
Ultimately, this knowledge could help scientists develop strategies to reduce infertility due to early embryonic development gone wrong.
The new findings could also shed light on a defining property of early embryonic cells that was difficult to capture in the lab: their ability to produce all of the cell types found in the embryo and, ultimately, throughout the body. Scientists call this property “totipotency”.
“Totipotency is a very unique and short-lived property of early embryonic cells,” says Cody Kime, PhD, researcher at the RIKEN Center for Biosystems Dynamics Research and lead author of the study.
“It has been much more difficult to exploit in the laboratory than pluripotency,” he adds, referring to the ability of some cells to give rise to several types of cells, but not all. “A very exciting prospect of our work is the ability to understand how we can reprogram cells in the lab to achieve totipotency.”
Cultivate the fundamental components of the first embryos in the laboratory
To generate synthetic embryos, scientists started with pluripotent mouse stem cells that normally give birth only to the fetus – not the placenta or amniotic sac. They can grow these cells, called epiblast stem cells, and multiply them indefinitely in the lab.
In previous work, the team had discovered a combination of nutrients and chemicals that could allow epiblast stem cells to assemble into small cellular structures that closely resemble pre-implantation embryos. In fact, the structures could even reach the implantation stage when transferred to female mice, although they degenerated soon after.
“This meant that we could successfully reprogram epiblast cells to return to an earlier stage, when embryonic cells are totipotent, and provided a clue as to how we might generate both the fetus and the tissues that support its implantation. Explains Tomoda, who is also a program-specific research center associate professor at CiRA.
To build on this work and better understand the reprogramming process, scientists needed molecular resolution. In their new study, they turned to single-cell RNA sequencing, a technique that allows scientists to study individual cells based on the genes they turn on or off.
After analyzing thousands of individual cells reprogrammed from epiblast stem cells and sifting the data by computerized analyzes, they confirmed that after 5 days of reprogramming, some cells closely resemble the three precursors of the fetus, placenta and of the amniotic sac. .
Additionally, as they grew in the lab for a few more days, all three cell types exhibited more distinct molecular profiles with striking similarity to real embryonic model cells. This is the same as one would expect during the growth of a normal embryo, when all three tissues acquire distinct physical properties and biological functions.
“Our unicellular RNA sequencing analysis confirms the emergence in our synthetic embryonic system of cell types that lead to the three fundamental components of an early mammalian embryo,” says Kime. “In addition, it reveals in surprising detail the genes and biological pathways involved in the development of these precursors and their maturation in specific tissues.”
This knowledge provides a comprehensive backdrop for understanding the mechanisms of early embryonic development and the possible causes of its failure.
For now, the scientists plan to work on ways to increase the efficiency of their reprogramming process, in order to reliably produce large quantities of pre-implantation-type synthetic embryos for further studies. This would allow them to conduct hitherto unthinkable experiments, such as large-scale screens for genetic mutations that disrupt early embryos. And it may shed light on the causes of pregnancy loss due to early embryonic failure.
They also want to better understand the molecular steps involved in reprogramming. In particular, they plan to look into the reprogramming process earlier than 5 days, in the hope of spotting the truly totipotent cells that are the source of their synthetic embryos.
“The discovery that we could reprogram cells to adopt earlier, more pluripotent states revolutionized developmental biology 15 years ago,” says Tomoda, referring to the discovery of pluripotent stem cells induced by his mentor and that of Kime, Nobel Laureate Shinya Yamanaka.
“In recent years, the field of synthetic embryology using stem cells has exploded,” he says. “Our method of generating synthetic embryos is simpler than others and quite efficient. We believe it will be a great resource for many laboratories.”