Journal: Cell Death Discovery

Article Title: Transcriptome-based prediction of drugs, inhibiting cardiomyogenesis in human induced pluripotent stem cells

doi: 10.1038/s41420-023-01616-6

Figure Lengend Snippet: The hiPSCs were cultured as a monolayer on matrigel-coated plates for 2 days under pluripotent conditions and on day 0 exposed to GSK3 inhibitor, CHIR (10 µM) for 24 h. After 48 h exposed to Wnt inhibitor, IWP2 (5 µM). Spontaneously beating cardiac clusters were observed from day 9 onwards. Simultaneously, cells were exposed to test substances for a single exposure of 24 h (day1). The cells were harvested for gene array analysis on day1, day4 and day14 (Fig. ). Medium changes were done as indicated every alternate date. A Representative phase-contrast images of control and ISO treated hiPSC at day1-, 4 -and 14day. Scale bar, 100 µm. B PCA blot of 54,675 probe sets for three timepoints during the differentiation. C PCA blot of the 500 SPS with the highest variance across the mean of the condition‐wise samples. The respective day is indicated by the shape and the respective measured compound is indicated by the color of the dot, as labels are shown next to the plots. The distribution of the data points on the x-axis is given by the PC 1 and on the y-axis by PC2. The percentages in parentheses denote the proportion of explained variance for the respective PC.

Article Snippet: The medium was then changed to basal RPMI/B-27-ins medium and cells were kept for further 24 h. At day 2, RPMI/B-27-ins medium with small molecule WNT inhibitor IWP2 (Tocris, United Kingdom) 5 μM was added and cells were kept for 48 h (day 2 to day4).

Techniques: Cell Culture, Control

Journal: National Science Review

Article Title: Inhibition of Wnt activity improves peri-implantation development of somatic cell nuclear transfer embryos

doi: 10.1093/nsr/nwad173

Figure Lengend Snippet: Wnt inhibition greatly improves the development of SCNT embryos. (a) Comparison of Wnt-related genes in differential gene expression profiles of the ICM of IVF and SCNT blastocysts. The genes marked are Wnt antagonists or activators. (b) TCF/Lef: H2B-GFP IVF (left) and SCNT (right) embryos at late E4.5 stained for GFP, Oct4, Gata6 and DAPI. Scale bar, 20 μm. (c) Percentage of GFP-positive cells in the EPI of IVF and SCNT embryos. (d) Percentage of rosette-like SCNT embryos generated with no treatment, Wnt activator CHIR treatment and Wnt inhibitor IWP2 treatment. (e) Comparison of pluripotent marker gene expression among the IVF-, SCNT- and Wnti-treated SCNT embryos. (f) Absolute value comparison of the relative expression of markers in (d) to late-E4.5 IVF EPI between groups of IVF-, SCNT- and Wnti-SCNT embryos. P -values were determined using the Wilcoxon signed-rank test. (g) GSEA of specific genes expressed in naïve and primed ESCs between the late-E4.5 EPI of untreated and Wnti-SCNT embryos. NES, normalized enrichment score. (h) PCA comparison of gene expression profiles among the late-E4.5 EPI of IVF-, SCNT- and Wnti-SCNT embryos, naïve ESCs, primed ESCs and RSCs. Published transcriptome data of stem cell lines were obtained from GSE145727. (i) Hierarchical clustering analysis of the late-E4.5 EPI of IVF, untreated and Wnti-SCNT embryos, naïve ESCs, primed ESCs and RSCs. (j) PCA plot of E3.5 ICM, E5.5 EPI, E6.5 EPI, late-E4.5 EPI of IVF, SCNT and Wnti-SCNT embryos based on the H3K27me3 levels. (k) Heat map showing the H3K27me3 levels for the naïve and primed markers between late-E4.5 EPI of IVF, SCNT and Wnti-SCNT embryos. For (c) and (d), P -values were determined using the unpaired two-tailed t -test; error bars and means ± SD are shown for n ≥ 3 experiments.

Article Snippet: Small-molecule inhibitors IWP2 (Selleck, 686770-61-6), IWR1-endo (Selleck, 1127442-82-3) and Wnt activator CHIR99021 (Sigma, SML1046) were applied to inhibit or activate the canonical Wnt/β-catenin signaling pathway.

Techniques: Inhibition, Comparison, Gene Expression, Staining, Generated, Marker, Expressing, Two Tailed Test