Fig. 3.

DNA damage and DNA damage responses upon H₂O₂ challenge. a, c, e, g Confocal images showing the basal region of the organ of Corti cultures treated with either culture medium alone (a, e) or containing 0.5 mM H₂O₂ (c, g) for 5 h before being maintained in culture medium alone for 3 days. The samples were then immunolabeled for myosin 7A (red, a, c, e, g), γH2AX (green, a and c) and 53BP1 (green, e and g). Scale bars: a, c, e and g = 10 μm. b, d, f, h Higher magnification images of representative OHC and IHC nuclei from all conditions tested. Scale bar = 2.5 μm. i Representative Western blot analysis using antibodies against γH2AX, 53BP1, DDB2, p-Chk1, p-Chk2, p53, and β-actin in whole cochlear extracts. j, k Histograms representing the levels of γH2AX, 53BP1, DDB2, p-Chk1, p-Chk2, and p53 in control and in 0.4 and 0.5 mM H₂O₂-exposed groups (n = 6 cochleae per condition). β-Actin served as a loading control. Data are expressed as mean ± SEM. One-way ANOVA test followed by post hoc Tukey’s test (*P ≤ 0.04, **P ≤ 0.01, ***P ≤ 0.001 vs. H₂O₂ 0 mM). All experiments were performed in triplicate