Fig. 3

Involvement of EGFR in cell viability of CYLD-knockdown OSCC cells. A EGFR phosphorylation was assessed by immunoblotting in OSCC cells. Cells were transfected with siRNA and then incubated for 72 h before harvesting. Cell lysate was immunoblotted with antibodies against the indicated proteins. B Cells were treated with gefitinib (10 µM) and cisplatin (8 µg/mL), and cells survival rate was assessed after 72 h. Relative cell survival rate of siCYLD cells compared to that of siN cells after treatment with each anticancer drug was shown. Values are means ± S.D. of triplicate samples. **p < 0.01 in Tukey–Kramer method. C (left panels) Representative images of cells treated with 10 µM gefitinib for 72 h. Scale bars show 100 µm. (right panel). Cell survival rates of OSCC cells were evaluated 72 h after treatment with various concentrations of gefitinib. Values are means ± S.D. of triplicate samples. **p < 0.01 vs siN group in Tukey–Kramer method. D Cells were treated with gefitinib (10 µM) and the cell survival rates were evaluated 24–72 h after treatment. Values are means ± S.D. of triplicate samples. * p < 0.05, ** p < 0.01 vs siN group in Tukey–Kramer method. (E) The gefitinib (10 µM)-induced apoptosis was assessed by Annexin-V/7-AAD staining using flow cytometry. Values are means ± S.D. of triplicate samples. n.s.: not significant, **p < 0.01 vs siN group in Tukey–Kramer method. F Phosphorylation of Akt and ERK were assessed by immunoblotting in OSCC cells treated with 10 µM gefitinib