Fig. 5: Low levels of Notch signaling promote cochlear SC and KC proliferation, organoid formation, and organoid growth.

a Experimental scheme (b–d). Cochlear epithelial cells from stage P2 wild-type mice were infected with a lentivirus that expressed the intracellular domain of Notch1 (NICD) in a dox-dependent manner. Dox (0.25, 0.5, 5, 10 µg/mL) was added to the culture medium on day 2, and organoids were analyzed after 7 days of expansion. b Bright field (BF) and green fluorescent (GFP) images of control (0 dox) and NICD expressing organoids. GFP labels organoids infected with lentivirus. Scale bars = 400 µm. c, d Organoid diameter (c) and organoid forming efficiency (d) for (b) (n = 6 independent biological replicates, two independent experiments). e Experimental scheme (f, g). f Bright field (BF) images of DMSO (vehicle control) and LY411575-treated organoids on day 7 of expansion. LY411575 or DMSO (0.2%) was added to the culture medium on day 2 of expansion. Scale bars = 400 µm. g, h Organoid diameter (g) and organoid forming efficiency (h) for (f) (n = 6 independent biological replicates, two independent experiments). i Cell proliferation in control (DMSO) and LY411575-treated organoids. A single EdU pulse was given on day 7 of expansion, and EdU incorporation (red) was analyzed 1 hour later. Hoechst labels cell nuclei (blue). Scale bars = 25 µm. j Percentage of EdU⁺ cells in (i) (n = 8 animals, two independent experiments). Graphed are individual data points and mean ± standard deviation. One-way ANOVA with Tukey’s correction was used to calculate exact p-values. Source data are provided as a Source Data file.