Generation of transient totipotent blastomere-like stem cells by short-term high-dose Pladienolide B treatment
As an alternative model for study of the dynamic process of early mammalian embryonic development, multiple progresses have been made in using mouse embryonic stem cells (mESCs) to generate embryo-like structure, especially by modifying the starting cells. A previous study has shown that as long as continuously treating mESCs with a low-dose splicing inhibitor, pladienolide B (PlaB), a totipotent state comparable to 2- and 4-cell embryos at molecular levels can be reached. However, these totipotent mESCs mare less proliferative, with the culture time being limited to about 10-15 passages. Here we reported that transiently PlaB-treated mESCs were sufficient for generating a novel type of totipotent-like stem cells from classical mESCs, and these cells were endowed with the ability to differentiate into both embryonic and extra-embryonic cell lineages, we termed them as transient totipotent-like stem cells (TTLCs). Similarly, TTLCs could self-organize to blastoids and recapitulated key preimplantation developmental processes. Transcriptome analysis also showed that TTLC-blastoids bear similarities with mouse E3.5, E4.5 blastocyst and TBLC-blastoids. Moreover, we found that TTLC-blastoids could develop beyond implantation stage, forming egg-cylinder structures both in vivo and in vitro. In summary, our findings provided a rapidly alternative cell type to generate blastoids for the study of early mouse embryogenesis. Overall design: We chose TT2-mESCs treated with 0 nM or 20nM PlaB for 6 h in 2D culture, and TTLC-blastoids from day 1 to day 5 in 3D microwell culture. We compared the gene expression profiles between these samples as well as transcriptome data sets from the literature.