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. 2025 Jun 6;13(2):101708. doi: 10.1016/j.gendis.2025.101708

Figure 1.

Figure 1

TMEM16F knockout prevented hearing loss and hair cell death in mice. (A) Western blotting confirmed the depletion of TMEM16F in mouse cochleae with Cre recombinase. (B) Auditory brainstem response (ABR) thresholds evoked by click stimuli in control (FF) and knockout (KO) mice during 3 months of non-diabetic or diabetic conditions. n = 5 (non-diabetic) or 6 (diabetic) mice in each group. (C) Click-evoked ABR waveforms at the end of the 3 months. (D) Distortion product otoacoustic emission (DPOAE) thresholds of the mice at month 3. n = 5 or 6 mice in each group. (E) Immunofluorescence (myosin-7a) of hair cells in mouse cochleae. (F) The ratio of the numbers of outer and inner hair cells. n = 5 or 6 mice in each group. (G, H) Click-evoked ABR thresholds in hair cell-specific cKO and FF mice during 3 months of diabetic condition, and DPOAE thresholds at month 3. n = 7 (FF) and 6 (cKO) mice. (I, J) Immunofluorescence and the ratio of the outer and inner hair cells. n = 7 (FF) and 6 (cKO) mice. (K, L) Click-evoked ABR thresholds in FF and cKO mice treated with 120-dB noise, and DPOAE thresholds after the treatment. n = 9 (FF) and 11 (cKO) mice. (M, N) Immunofluorescence and the ratio of the outer and inner hair cells. n = 9 (FF) and 11 (cKO) mice. (O) Western blotting confirmed TMEM16F KO in two lines of HEI-OC1 cells (immortalized mouse hair cells). (P) Viability of wild-type (WT) and TMEM16F–KO HEI-OC1 cells treated with ionomycin for 12 h (n = 14 wells) or H2O2 for 6 h (n = 16 wells) assessed by WST-1 assay. (Q) Percentage of cells with phosphatidylserine exposure detected by flow cytometry after treatment with 5 μM ionomycin or 500 μM H2O2 for 3.5 h. n = 3 dishes of cells in each group. #P < 0.05, ##P < 0.01, ###P < 0.001, and ####P < 0.0001 versus month 0 (B and G) or before (K), by 2-way ANOVA and Sidak's test; ∗P < 0.05, ∗∗P < 0.01, ∗∗∗P < 0.001, and ∗∗∗∗P < 0.0001 between FF and KO (B, D and F) or cKO (G, H, J, K, L and N), or between WT and KO (P and Q), by 2-way ANOVA and Sidak's test (B, D, F, G, H, J, K, L, N, and P) or 1-way ANOVA and Tukey's test (Q).