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A fascinating new study by Jose Silva (Thor’s former PhD advisor in Cambridge) and his team at Guangzhou Laboratory reports on the remarkable ability of naive human pluripotent stem cells to spontaneously form blastocyst-like structures (“blastoids”) in 3D suspension culture. These spontaneous blastoids mimic early-stage human blastocysts in terms of structure, size, and transcriptome characteristics and are capable of progressing to post-implantation stages on appropriate matrices, taking advantage of the recent methodology developed by Rowan Karvas and her colleagues in the Theunissen lab. This spontaneous blastoid potential of naive stem cells property is conferred by the glycogen synthase kinase-3 signalling inhibitor IM-12 present in 5iLAF self-renewing naive medium. IM-12 upregulates oxidative phosphorylation-associated genes that underly the capacity of naive stem cells to generate blastoids spontaneously. Starting from day one of self-organization, naive stem cells at the boundary of all 3D aggregates dedifferentiate into E5 embryo-like intermediates. Intermediates co-express SOX2/OCT4 and GATA6 and by day 3 specify trophoblast fate, which coincides with cavitation and blastoid formation. In summary, spontaneous blastoid formation results from 3D culture triggering dedifferentiation of naive stem cells into earlier embryo-like intermediates which are then competent to segregate blastocyst fates. Check out the paper in Nature Communications!

The featured image shows an example of a TBXT-positive primitive streak-like structure in spontaneous blastoids maintained on an optimized 3D matrix until D14 (courtesy of the Silva lab). SOX2 is indicated in red, TBXT in green, and GATA3 in grey.

Our paper on extended human blastoid culture was published today in Cell Stem Cell. In a transformative advance, several groups have reported that naive human pluripotent stem cells (hPSCs) can form blastocyst-like structures (also known as “blastoids”) that model the human pre-implantation embryo. However, the extent to which blastoids can recapitulate defining features of post-implantation development remained unexplored. In this study, we optimized the conditions for blastoid generation from naïve hPSCs and investigated their capacity for extended culture on thick 3D extracellular matrices, which better mimic the physical environment of the human endometrium compared to flat surfaces. We developed an experimental methodology that supports human blastoid culture for up to day 21 (D21), including the formation of complex embryonic and placental structures. By performing a detailed single cell transcriptome analysis at three distinct time points (D7, D14, and D21), we benchmarked our model system to human embryos at pre-implantation, early post-implantation, and early gastrulation stages.

3D-cultured human blastoids display several molecular and morphogenetic hallmarks of early post-implantation development, including lumenogenesis of the epiblast compartment, rapid expansion and diversification of trophoblast lineages, and robust invasion of extravillous trophoblast cells by D14. Extended blastoid culture resulted in the formation of a primitive streak-like structure, as evidenced by the localized activation of TBXT (Brachyury) by D18. Blastoids maintained until D21 acquired a single cell transcriptome profile that closely resembled that of a gastrulating human embryo analyzed at Carnegie Stage 7. This included the emergence of blastoid primordial germ cells, definitive endoderm, and various mesodermal lineages, and a diverse array of extraembryonic cell types, including blastoid amnion, cytotrophoblast, extravillous trophoblast, extraembryonic mesoderm, syncytiotrophoblast, and yolk sac endoderm. Thus, the 3D-cultured human blastoids described herein model embryonic and extraembryonic development from pre-implantation to early gastrulation stages, offering a continuous and integrated in vitro model system of early embryogenesis.

Congratulations to Rowan, our collaborators in the Dietmann and Zhou labs, and the entire team!

Note: The generation of integrated models of human development in our laboratory is entirely supported by private foundation grants and does not involve federal funding from the National Institutes of Health.

Our review on stem-cell-based models of X-chromosome inactivation and reactivation during human development was published in the Special Issue on Early Embryonic Development Models in Current Opinion in Genetics & Development. Congrats Shafqat!

Our review article on the many recently uncovered paths for generating human trophoblast stem cells is now online at Cellular and Molecular Life Sciences. Congratulations to Rowan Karvas and our collaborator in France, Laurent David!

Our study describing a genome-wide CRISPR-Cas9 knockout screen for essential and growth-restricting genes in human trophoblast stem cells (hTSCs) was published in Nature Communications today. By cross-referencing our results to those from similar genetic screens performed in other cell types, as well as gene expression data from early human embryos, we define hTSC-specific and -enriched regulators. These include both well-established and previously uncharacterized trophoblast regulators. Integrated analysis of chromatin accessibility, gene expression, and genome-wide location data reveals that the hTSC-specific essential transcription factor TEAD1 regulates the expression of many trophoblast regulators in hTSCs. In the absence of TEAD1, hTSCs fail to complete faithful differentiation into extravillous trophoblast (EVT) cells and instead show a bias towards syncytiotrophoblast (STB) differentiation. Overall, our study provides a valuable resource for dissecting the molecular regulation of human placental development and pregnancy-related diseases. Congratulations to Chen Dong, who spearheaded this project in collaboration with Shuhua Fu in Bo Zhang’s lab at WashU, and all other contributors!

Our paper on stem-cell-derived trophoblast organoids (SC-TOs) was published in Cell Stem Cell today. In this work we describe a methodology for deriving self-renewing 3D trophoblast organoids from naive human pluripotent stem cells (hPSCs). Using single cell transcriptome analysis, we demonstrate that these organoids contain diverse trophoblast progenitor and specialized cell types that closely correspond to trophoblast identities in the post-implantation embryo. These organoid cultures also model placental X inactivation dynamics and selective vulnerability to emerging pathogens (SARS-CoV-2 and Zika virus). Big congrats to all members of the team, especially Rowan Karvas, and our collaborators Indira Mysorekar, Sabine Dietmann, Jacco Boon, and Liang Ma!

Our paper mapping the interactomes of the master transcription factor OCT4 in naive and primed human embryonic stem cells (hESCs) was published today in Nature Communications. We report that OCT4 cooperates with cell state-specific chromatin modifiers in naive and primed hESCs to promote an open chromatin architecture at blastocyst-associated and pan-ectodermal genes, respectively. This work was a collaboration between our team at WashU, Xin Huang and Jianlong Wang at Columbia University, Cigall Kadoch at the Dana-Farber Cancer Institute, and Rudolf Jaenisch at the Whitehead Institute. The WashU team was spearheaded by Kyoung-mi Park in the Theunissen lab in collaboration with the labs of Bo Zhang, Sabine Dietmann, and Ting Wang.

Our paper investigating the signaling requirements for naive human pluripotency by high-throughput chemical screening was published today in Cell Reports. We report that naive hESCs can be maintained by blocking distinct nodes in the FGF signaling pathway and that dual MEK/ERK inhibition promotes efficient primed-to-naive resetting in combination with Activin A. This study was a collaboration between Washington University School of Medicine, the Novartis Institutes for Biomedical Research, and the Whitehead Institute for Biomedical Research

Our paper examining the trophoblast potential of distinct human stem cell states was published today in eLife. We report that naive, but not primed, human pluripotent stem cells (hPSC) can directly differentiate into trophoblast stem cells (TSC). The derivation of TSCs from naïve hPSCs presents a new model system to elucidate early mechanisms  governing placental development and associated pathologies. This study was a collaboration with the Solnica-Krezel, Wang and Kommagani labs at Washington University.

Image of naive hPSC-derived human TSCs stained with a KRT7 antibody by Chen Dong (Theunissen Lab).