Fig. 4. DAPT treatment of cochleae in culture induces hair cell differentiation. (A) Real-time PCR analysis of myosin VIIa transcripts is presented in terms of average Ct (see Materials and methods). A greater Ct value represents a lower transcript expression level. Histogram and bars represent average Ct±s.d. (^*P<0.05). High DAPT doses increase myosin VIIa expression, with significant induction at 5 µM DAPT. (B) Western blot confirmation of myosin VIIa protein expression changes with DAPT dose. The myosin band of Kaleidoscope prestained standard is used as control. (C) Expression of COUP-TFI and COUP-TFII transcripts in cochleae treated with DAPT. COUP-TFI and COUP-TFII expression was not changed with DAPT, even at the highest concentration of 5 µM. Because DAPT can induce hair cell differentiation (in terms of myosin VIIa expression as shown in Fig. 4A), the value is presented here after further normalization with the myosin VIIa values. (D) Expression of Hes5 in cochleae after culture with DAPT. Hes5 expression is reduced with increasing concentrations of DAPT, with statistically significant differences at 2.5 and 5 µM (^*P<0.05). A statistically significant reduction was also found between COUP-TFI^-/- and wild type at 5 µM (^**P<0.05).