Fig. 5

Analysis of fibrosis-related signaling pathway involving SMAD2 and SNAI2 transcription factors in human cardiac fibroblasts (hCFs) stimulated with TGF-β (1 ng/ml) and treated with EV-H5 (from hiPS-L3). A Confocal microscopy tile scans containing 5 × 5 images of cells, followed by high magnification Airyscan images of individual nuclei (B). C Quantification of pSMAD2 foci in the nucleus (upper panel) or cytoplasm (lower panel) in hCFs (n = 10). Fluorimetric analysis of pSMAD2 (n = 55) (D) and SNAI2 (n = 30) (E) protein level in cell nuclei. F Relative quantification of SNAI1 (n = 3) and SNAI2 (n = 4) genes expression in the tested samples by real-time qPCR. G Analysis of mRNA levels of pro-fibrotic genes in activated hCFs after EV-H5 treatment (n = 4). All data are presented as the mean ± SD. Statistical significance was tested using Welch ANOVA and Dunnet’s test (C) or with Kruskal–Wallis with Dunn’s post hoc test (D-G). *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001