Fig. 6: DNMT3A regulates intestinal paracellular permeability.

a Representative images of ΔDNMT3A and WT spheroids cultured in 3D matrix for 14 days. Confocal microscopy was used to analyze spheroid organization. Phalloidin (red) and DRAQ5 (purple) was used to visualize actin filaments and nuclei, respectively. Spheroids area (µm²) was determined in WT and ΔDNMT3A (n = 40 over 3 independent experiments). b Representative images of Caco-2 cells, WT and ΔDNMT3A, at 0 h and 24 h after inducing the gap. The percentage of wound closure after 24 h was measured with a phase-contrast microscope using the absolute wound area normalized to the corresponding value at 0 h. (n = 20 over 3 independent experiments). c Transmission electron microscopy (TEM) was used to measure intercellular distance and apical-junctional complex length in WT and ΔDNMT3A Caco-2 cells cultivated on a transwell support (n = 8). d Trans-epithelial electrical resistance (TEER) measurements of differentiated ΔDNMT3A and WT Caco-2 cells (n = 12 over 3 independent experiments). e Representative TEM images and quantification of intercellular distance and apical-junctional complex length of small intestinal sections from Dnmt3a^ΔIEC and Dnmt3a^fl/fl mice (n = 3). f TEER measurements of colonic organoids from Dnmt3a^ΔIEC and Dnmt3a^fl/fl mice grown as monolayer on transwell support (n = 16 over 4 independent experiments). g Representative images and fluorescence intensity ratio between the luminal (L) and basolateral (BL) side of Dnmt3a^ΔIEC and Dnmt3a^fl/fl colonic organoids incubated with FITC-D4 for 24 h (n = 10). h FITC-dextran quantification in serum isolated from Dnmt3a^ΔIEC and Dnmt3a^fl/fl mice (n = 6) 1 h after oral administration. The values represent mean ± SEM. Statistical analysis was performed using two-tailed t-test with Mann–Whitney correction. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.