A faithful model of the human blastocyst has been developed which replicates key phases of very early development from 4 to 13 days post fertilization, including attaching to layers of hormone-stimulated endometrial cells in vitro, mimicking implantation in the uterus.

Implantation is extremely difficult to study, and the researchers who developed the blastoids say the ability to recapitulate aspects of this process has enabled them to discover molecules that could be developed as contraceptives or as fertility enhancers, to improve success rates in in vitro fertilization (IVF).

The new model, which was reported in the December 2, 2021, advance online edition of Nature, will provide a "powerful scientific and ethical alternative" to the use of fertilized eggs to study embryonic development, said Nicolas Rivron, group leader at the Institute of Molecular Biotechnology in Vienna, Austria, where the research was carried out.

Rivron lays claim to having coined the term blastoid, having derived a mouse version in 2018. That triggered interest in forming models using human stem cells, and other labs have generated blastoids using different types of adult and pluripotent stem cells as the starting point.

"Finding the right shade of human pluripotent stem cells and the right media turned out to be quite complicated. If these parameters are not correct, the cells start behaving and forming cellular states that do not reflect the blastocyst, the day 7 embryo that is composed of epiblast, hypoblast and trophectoderm cells," Rivron told BioWorld Science.

Rivron's formula involves inhibiting three signaling pathways, Hippo, TGF-beta and ERK, and culturing the cells in a chemically defined medium.

"What was really amazing is that under these conditions of triple inhibition, the cells remember where they came from," Rivron said. The blastoids "formed very efficiently" with more than 70% of the aggregates of human pluripotent stem cells forming the correct structure.

The aggregates also efficiently emulated normal blastocyst development timelines, forming "with the right pace, in the right time."

"Together, we came to the conclusion that we had formed blastocyst-like cells, but also formed these cells according to the sequence (trophoblast/epiblast first, hypoblast last) and to the pace (4 days) of human blastocyst. And they do this very efficiently," said Rivron.

Although blastoids derived in other labs were claimed as good models when the details were first published, subsequent independent transcriptomic analyses, in which the three cell lineages were examined separately, has called this into question.

In contrast, Rivron said single cell transcriptomics carried out in his lab and independently, show his blastoids match reference maps of human blastocysts and are clearly distinct from the post-implantation stages of embryonic development.

"This will ensure that we can trust that the molecular pathways and cellular behaviors we observe within those blastoids are the ones that are in play within the blastocyst," Rivron said.

On day 7 after fertilization, human blastocysts begin to implant into the endometrium lining the womb. In a further demonstration of their fidelity, when exposed to endometrial cells secreting estrogen and progesterone, the hormones that make the uterus receptive to implantation, the blastoids attach to the 2D layer.

Conversely, blastoids deposited on nonstimulated endometrial cell layers did not attach. "We concluded that human blastoids are capable of interacting specifically with endometrial cells that are made receptive," the researchers say in their paper, published in Nature.

As a demonstration of the potential utility of the blastoid model in drug discovery, it was shown that the contraceptive levonorgestrel impaired blastoid attachment.

A video of the interaction between a blastoid and a layer of endometrial cells showed the blastoid orientating and positioning itself to attach via one side only, mimicking the blastocyst, which attaches on its polar side.

Pregnancy is characterized by the detection of chorionic gonadotrophin. Upon attachment to the stimulated endometrial cell layer, the blastoids expressed this hormone at levels detectable using standard pregnancy tests.

The blastoids were cultured for up to 13 days, at which point they contained about 300 cells.

Using the blastoids in screening, the researchers found an orally available small molecule that inhibits attachment of blastoids to stimulated endometrial cells, pointing the way to a new, nonhormonal form of contraception.

They also have discovered a compound that improves the self-organizing ability of the stem cells into a blastoid, and suggest this might be used to boost the formation of natural embryos in IVF.

"We hope we can use such molecules to improve the number and quality of IVF embryos, and the chance of becoming pregnant," Rivron said.