Journal: Cell Stem Cell

Article Title: Amniogenesis occurs in two independent waves in primates

doi: 10.1016/j.stem.2022.03.014

Figure Lengend Snippet: hPSCs transiently gain the ability to form epithelial cavitating structures during the naive-to-primed transition (A) A scheme of human peri-implantation development and correspondent hPSC states (PreEPI, preimplantation epiblast; PostEPI-E and PostEPI-L, early and late postimplantation epiblast, respectively; PS, primitive streak). (B) Experimental setup. Partially primed hPSCs were treated with either an inhibitor of ALK4/5/7, MAPK, or their combination. (C) qRT-PCR for markers after differentiation in indicated conditions; results of two independent experiments. (D) Stitched images of the cells in 24-well plates after differentiation in indicated conditions and staining with Phalloidin. (E) Bright-field image of 3D epithelial cavitating spheres obtained in AP condition. Note that the spheres remain attached to the surface of culture plates. (F) Immunofluorescence for GATA3 in combination with E-cadherin and CDX2 in combination with POU5F1, of partially primed hPSCs differentiated in AP condition (“AP”) and undifferentiated control (“undiff”). (G) Experimental setup. hPSCs on different days of the formative transition were differentiated in AP. (H) Stitched images of scanned 24-well plates showing hPSCs after different periods of the formative transition differentiated in AP (4′,6-diamidino-2-phenylindole (DAPI) staining). (I) Bright-field images of hPSCs after different periods of the formative transition and conventional H9 hPSCs differentiated in AP. (J) qRT-PCR for markers during the time course of AP treatment of naive and partially primed hPSCs. (K) Immunofluorescence of naive and partially primed hPSCs during the time course of differentiation in AP. See also Figures S3 and .

Article Snippet: Human: H9 (WA09) , WiCell , hPSC line H9.

Techniques: Quantitative RT-PCR, Staining, Immunofluorescence

Journal: Cell Stem Cell

Article Title: Amniogenesis occurs in two independent waves in primates

doi: 10.1016/j.stem.2022.03.014

Figure Lengend Snippet:

Article Snippet: Human: H9 (WA09) , WiCell , hPSC line H9.

Techniques: Recombinant, In Vitro, Cell Culture, Derivative Assay, Software

Journal: Science Advances

Article Title: Epigenetic dynamics during capacitation of naïve human pluripotent stem cells

doi: 10.1126/sciadv.adg1936

Figure Lengend Snippet: ( A ) Percentage of all ChIP-seq and ATAC-seq peaks mapped to the X chromosome. ( B ) Distribution of scaled log-transformed normalized counts of H3K27me3 and H3K9me3 in 10-kb windows across the X chromosome and autosomes. The red line indicates the threshold used to categorize genomic regions as high or low H3K27me3 and H3K9me3. ( C ) H3K27me3 and H3K9me3 read count profiles across the X chromosome for naïve, capacitated day 10, and conventional H9 cells. ( D ) Gene expression levels of the long ncRNAs XIST and XACT . * P < 0.05, ** P < 0.01, and *** P < 0.001, Student’s paired t test. Error bars indicate 1 SD of the mean values from three independent experiments. ( E ) Flow chart illustrating the number of 10-kb windows in the X chromosome classified based on H3K27me3 and H3K9me3 scaled log 2 -normalized counts threshold of 0.5. The classification includes windows containing high levels of H3K27me3 marks (H3K27me3), high levels of H3K9me3 marks (“H3K9me3”), high levels of both H3K27me3 and H3K9me3 marks (“H3K27me3 + H3K9me3”), and low levels of both marks (Low). ( F ) Flow chart illustrating the number of 10-kb windows in the X chromosome in the cR-H9 day 10 (capacitated) and H9 primed cells (conventional). On the right side is the enrichment analysis for genomic features and repeats of the regions represented in red. Only results with an FDR below 0.05, absolute odds ratio value above 2, and a −log P value above 10 are shown. ( G ) Immuno-FISH for H3K27me3, XIST and XACT in conventional primed hPSCs, cells after a round of resetting and capacitation, and cells after additional expansion in either XAF or E8. ( H ) Quantification of the immuno-FISH. Scale bar, 10 μm. ( I ) qRT-PCR for XIST . GAPDH , glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: The experiments were conducted using the embryo-derived HNES1 and the chemically reset cR-H9-EOS naïve hPSC lines ( , ), as well as conventional primed H9 hPSC (WA09, WiCell).

Techniques: ChIP-sequencing, Transformation Assay, Expressing, Quantitative RT-PCR

Journal: Science Advances

Article Title: Epigenetic dynamics during capacitation of naïve human pluripotent stem cells

doi: 10.1126/sciadv.adg1936

Figure Lengend Snippet: Naïve hPSC can be derived from preimplantation hICM cells or through (chemical) resetting of conventional hPSCs. Naïve hPSC can be capacitated, a process for which PRC2 activity appears to be dispensable. While naïve hPSCs remain transcriptionally close to preimplantation epiblast cells, capacitated hPSCs are transcriptionally closer to postimplantation epiblast than conventional hPSCs. During capacitation, the hPSC epigenetic landscape is globally remodeled, including genome-wide DNA methylation increase or H3K27me3 decrease. While genomic imprinting is not maintained in naïve hPSC nor re-established during capacitation, X chromosome erosion, frequently observed in conventional female hPSCs, is reversed by resetting and subsequent capacitation. The roles of H3K27ac and H3K4me3 in gene expression control appear to be distinct between the naïve and the primed hPSCs. XCI, X chromosome inactivation; DNAme, DNA methylation.

Article Snippet: The experiments were conducted using the embryo-derived HNES1 and the chemically reset cR-H9-EOS naïve hPSC lines ( , ), as well as conventional primed H9 hPSC (WA09, WiCell).

Techniques: Derivative Assay, Activity Assay, Genome Wide, DNA Methylation Assay, Expressing