Derivation of Pluripotent Stem Cells with In Vivo …

Figure 7. Analyses of Molecular Features of EPS Cells and the Roles of DiM and

(A and B) PCA analysis of RNA-seq and microarray data from EPS cells and known PSC types. Log2 expression values were normalized to mES cells (A) or primed hPSCs (B) in each study. For (A), data from mEPS cells (this study), mES cells (each study), 2C-like cells (Macfarlan et al. (2012)), and epiblast stem cells (Najm et al. (2011)) were analyzed, and a total of 17,243 genes were selected. For (B), data from hEPS cells (this study), naive hPSCs (Takashima et al. (2014), Chan et al. (2013), Gafni et al. (2013), and Theunissen et al. (2014)), and primed hPSCs (each study) were analyzed, and a total of 15,958 genes were selected. Circles, RNA-seq data; triangles, microarray data. (C and D) Heatmaps showing the presence of EPS-specific gene modules in mEPS (C) and hEPS (D) cells when compared to mES cells (C) and primed hPSCs (D) respectively. Correlations between genes and samples were calculated using Euclidean distance (complete linkage). Log2 expression values were normalized to mES cells (C) or primed hPSCs (D) in each study. hESC, primed hPSCs. To compare EPS cells with embryonic cells from preimplantation stages, data from Tang et al. (2011) (C) and Yan et al. (2013) (D) were analyzed. (E) Analyses of the influence of DiM or MiH substitution and Parp1 knockout on the chimeric ability of mEPS cells. Multiple cells were injected. The bar chart shows the percentage of chimeras among the recovered blastocysts. n indicates numbers of recovered blastocysts. ICM & TE, embryos with the integration of mouse cells into both ICM and TE. (F) Representative images of hEPS colonies after the omission of DiM, MiH or both from the LCDM condition. Scale bar, 100 m. (G) Representative images of hEPS colonies at passage 7 after DiM or MiH substitution. Scale bars, 100 m. (H) Analyses of the influence of DiM or MiH substitution on the chimeric ability of hEPS cells. Multiple cells were injected. The bar chart shows the percentage of chimeras among the recovered blastocysts. n indicates numbers of recovered blastocysts. ICM & TE, embryos with the integration of human cells into both ICM and TE. TH, tripelennamine HCL; DES, desloratadine; NAM, nicotinamide. PD, PD 0325901; SB, SB 203580; SP: SP 600125. See also Figure S7.

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Derivation of Pluripotent Stem Cells with In Vivo ...