Colitis facilitates neurogenesis in the colonic myenteric plexus. (A) Confocal images of colonic myenteric plexus isolated from tamoxifen- and 1×DSS-treated Nes^ER-CreR26-STOP^fl/fltdTomato (Cre⁺) mice on day 45 (6.5 wk) and stained with S100β (enteric glia, top) or CD31 (blood endothelial cells, bottom), tdTomato (newborn cells), and βIII-tubulin (nerve fibers) antibodies. Scale bars, 100 μm. (B) Confocal image of full-length colon Swiss roll cross-section from tamoxifen- and 3×DSS-treated Cre⁺ mice at week 10, stained with antibodies to Hu (neuronal somas), tdTomato (newborn cells), and DAPI (nuclei). Overview image—scale bar, 500 μm. Magnified images—scale bars, 50 μm. (C) Percentage of newly generated tdTomato⁺Hu ⁺ neurons among total Hu⁺ myenteric neurons in proximal to middle regions of the colon from tamoxifen- and 3×DSS- or water-treated Cre⁺ mice at week 10 after starting DSS, quantified by confocal microscopy of muscularis whole mounts (water, n =8–9; and DSS, n = 21–30, n—microscopy fields of view in the same mouse). (D) Complementary to Fig. 2 E: percentage of newly generated tdTomato⁺Hu⁺ neurons among total colonic myenteric neurons per field per mouse in tamoxifen- and 3×DSS- or water-treated Cre⁺ mice at week 10, quantified by confocal microscopy of muscularis whole mounts. Data represent the mean ± SEM of 5–10 fields of view analyzed per mouse per group. Each violin plot represents a mouse (n = 6 per group). (E) Complementary to Fig. 2 F: FC plots showing percentages of newly generated tdTomato⁺ cells among total CD45^–CD90⁺CD24⁺ myenteric neurons in colonic muscularis isolated from tamoxifen- and 3×DSS- or water-treated Nes^ER-CreR26-STOP^fl/fltdTomato (Cre⁺) mice as compared to control R26-STOP^fl/fltdTomato (WT) mice. (F) Complementary to Fig. 2 F: percentage of newly generated tdTomato⁺ cells among CD45^– nonhematopoietic CD90⁺, CD24⁺, and CD90^–CD24^– stromal cells in colonic myenteric plexus from mice described in E, quantified by FC (control, n = 3–6; DSS, n = 4–6). Week 10 data are from the same mice as in D. All graphs show the mean ± SEM, n—mouse. Data are representative of at least two independent experiments with similar results. Statistical analyses: unpaired Student’s t test (C and F) and two-way ANOVA for multiple comparisons (D), **P ≤ 0.01, ***P ≤ 0.001, and ****P ≤ 0.0001.