Fig. 3. Induction of cellular senescence in intestinal epithelial cells by gut bacteria.

a, b Early passage normal human colonic epithelial cells (CCD 841 CoN) were cultured with tissue culture media containing the indicated bacterial conditioned media or the plain bacterial culture media (Mock) at a ratio of 1/30 for 9 days, changing the medium every 3 days, and then subsequently cultured with plain tissue culture medium for another 3 days. Cell numbers were counted throughout the experiments, and representative photographs of the cells in the indicated culture conditions on day 12 are shown at the top of the panels. These assays were performed in triplicate (both biological and technical replicates) and representative data were shown (a). Cells on day 9 were subjected to RT-qPCR analysis for indicated genes (b). c Early passage normal human colonic epithelial cells (CCD 841 CoN) were cultured with tissue culture media containing the indicated bacterial conditioned media or the plain bacterial culture media (Mock) at a ratio of 1/30 for 9 days, changing the medium every 3 days, and then subsequently cultured with plain tissue culture medium for another 7 days. These cells were then subjected to EdU incorporation analysis. EdU (red) and DNA staining with 4′, 6-diamidino-2-phenylindole (DAPI) (blue) were shown. Representative photographs of the cells in the indicated culture conditions are shown. The histograms indicate the percentages of cells that were positive for EdU. These assays were performed in triplicate (both biological and technical replicates) and representative data were shown (b, c). For all graphs, error bars indicate mean ± s.d. with three biologically independent replicates. Statistical significance was determined with one-way ANOVA followed by Tukey’s test. P values < 0.05 were considered significant (b, c). Source data are provided as a Source Data file.