International team of researchers define criteria for elusive goal of stem cell biology

A team of researchers from The Hospital for Sick Children (SickKids), the Karolinska Institutet and the KU Leuven has provided a set of criteria to assess whether a mouse stem cell line shows true totipotency. Totipotency is the ability of a cell to generate every cell type in the early embryo as well as the embryo’s supporting structures, including the placenta. In normal fetal development, this ability is typically only found during the first few cell divisions of an embryo.

Replicating mouse stem cells in a totipotent state within a laboratory has been a long-sought goal of stem cell biologists around the world. If successfully created, such a stem cell line could have profound impact on the field, enabling the study of normal development and regenerative medicine. To date, mouse stem cell lines in a pluripotent state have been made but these stem cells can only make the cell types that build the embryo itself. Other stem cells have also been made that can give rise to a placenta. However, a stem cell with totipotency, that could make cells leading to both an embryo and placenta, has remained elusive.

“Creating stem cells with totipotency is truly the ‘holy grail’ of stem cell biology. There is a lot of interest in pushing the boundaries of stem cell potential in order to capture the totipotent state so we needed criteria to assess whether a stem cell line has actually achieved this goal,” says Dr. Janet Rossant, Chief of Research Emeritus and Senior Scientist in the Developmental & Stem Cell Biology program at SickKids.

Rossant, who is also a Professor in the Departments of Molecular Genetics, Obstetrics/Gynaecology and Paediatrics at the University of Toronto, along with Drs. Eszter Posfai, former postdoctoral fellow in the Rossant lab and now an Assistant Professor at Princeton University, collaborated with colleagues at the Karolinska Institutet and the KU Leuven to develop these criteria, which were published in Nature Cell Biology on January 8, 2021.

The scientists from the Karolinska Institutet included Drs. Fredrik Lanner, Assistant Professor in the Department of Obstetics and Gynecology and John Schell, PhD. The scientists from the KU Leuven included Drs. Vincent Pasque, Associate Professor in the Department of Development and Regeneration at the KU Leuven and Adrian Janiszewski, PhD fellow. This international team came together as all have expertise in embryo development and were skeptical of previously published research on these cell types. They joined forces to reexamine the published data and determined three criteria for a mouse stem cell line to be totipotent:

1. The cells’ genetic activity, or gene expression profiles, need to be closer to that of an earlier embryo rather than pluripotent stem cells.
2. The cells should be able to readily transform into placental stem cells or into an early embryo-like structure in an artificial environment.
3. Most importantly, the cells should be able to contribute to the placenta as well as the fetus when returned into the environment of an early embryo.

The team tested two different mouse stem cell lines that had been reported as potentially totipotent and assessed them using their criteria. While both lines showed some gene expression differences from pluripotent stem cells, they didn’t look like totipotent early embryo cells and didn’t make certain functional cells in the placenta.

“Ultimately, we found the search for a totipotent stem cell is not over so it’s back to the drawing boards in our respective labs to find better ways of capturing these cell types,” says Posfai. “Now, we have a clear set of criteria to validate any new cell lines, which will need to be considered in all future publications.”

In addition to establishing the criteria, the team also developed a comprehensive analysis of gene expression and regulation at the single cell level for embryos and different pluripotent stem cell lines. The researchers say this will be an invaluable resource for the scientific community.

This work was supported by the Canadian Institutes of Health Research, Genome Canada, Ontario Genomics, Programme de bourses de chercheur-boursier FRQS Junior 1, the Swedish Research Council, Ragnar Söderberg Foundation, Ming Wai Lau Center for Reparative Medicine, Center for Innovative Medicine, Wallenberg Academy Fellow, Natural Sciences and Engineering Research Council, The Research Foundation-Flanders, the KU Leuven Research Fund and SickKids Foundation.