Systematic characterization of methods for inducing human naïve pluripotency [Registered Report Stage 1 Protocol]

ABSTRACT Since their derivation, human pluripotent stem cells (hPSCs) have been maintained in a “primed” pluripotent state in vitro, corresponding to embryonic cells of the post-implantation blastocyst. To mimic cells of the pre-implantation blastocyst, the field has devised protocols to revert primed hPSCs to an earlier embryonic, “naïve”, state that can be used to derive in vitro blastocyst-like structures, “blastoids”, providing an interesting model of human pre-implantation embryos. Despite their utility in understanding fundamental cell biology and early embryology, in vitro naïve hPSCs are prone to karyotypic abnormalities and the efficacy of their derivation varies across protocols and cell lines. To promote their reproducibility and reliability in the field, we here apply two widely adopted methods for naïve induction on several hPSC lines varying initial culture conditions while systematically assessing the quality of resulted cells by transcriptional profiling, karyotyping, and blastoid formation ability.</p