Derivation of trophoblast stem cells from naïve human pluripotent stem cells [ATAC-seq]

Naïve human pluripotent stem cells (hPSCs) provide a unique experimental platform of cell fate decisions during pre-implantation development, but their lineage potential remains incompletely characterized. As naïve hPSCs share transcriptional and epigenomic signatures with trophoblast cells, it has been proposed that the naïve state may have enhanced predisposition for differentiation along this extraembryonic lineage. Here we examined the trophoblast potential of isogenic naïve and primed hPSCs. We found that naïve hPSCs can directly give rise to human trophoblast stem cells (hTSCs) and undergo further differentiation into both extravillous and syncytiotrophoblast. In contrast, primed hPSCs do not support hTSC derivation, but give rise to non-self-renewing cytotrophoblasts in response to BMP4. Global transcriptome and chromatin accessibility analyses indicate that hTSCs derived from naïve hPSCs acquire features of pre-implantation trophectoderm. The derivation of hTSCs from naïve hPSCs will enable elucidation of early mechanisms that govern normal human trophoblast development and associated pathologies. Overall design: ATAC-seq of one cell type (human trophoblast stem cells). The samples are human trophoblast stem cells derived from the blastocyst (BT5) or naive hPSCs (H9 and AN). hTSCs were cultured as previously described (Okae et al., 2018). Briefly, a 6-well plate was coated with 5 µg/mL Collagen IV (Corning, 354233) at 37°C overnight. Cells were cultured in 2mL TS medium [DMEM/F12 supplemented with 0.1 mM 2-mercaptoethanol, 0.2% FBS, 0.5% Penicillin-Streptomycin, 0.3% BSA, 1% ITS-X (Gibco, 51500), 1.5 mg/ml L-ascorbic acid (Wako, 013-12061), 50 ng/ml EGF (Rockland, 009-001-C26), 2 mM CHIR99021 (Stemgent, 04-0004), 0.5 mM A83-01 (BioVision, 1725), 1 mM SB431542 (BioVision, 1674), 0.8 mM VPA (Tocris, 2815), and 5 mM Y-27632] and in 5% CO2 and 20% O2. Media was changed every 2 days,and cells were passaged using TrypLE Express every 3 days at a ratio of 1:4.