Fig. 6

Contribution of ROS-dependent ATF3 activation to TC-HW-mediated apoptosis. a and b HCT116 and SW480 cells were treated with TC-HW at the indicated concentrations for 24 h. For Western blot analysis, cell lysates were subjected to SDS-PAGE and the Western blot was performed using antibodies against ATF3. Actin was used as internal control for Western blot analysis. For RT-PCR analysis of the gene expression of ATF3, total RNA was prepared. GAPDH was used as internal control for RP-PCR. b HCT116 cells were co-transfected with ATF3 promoter and pRL-null for 24 h, and then treated with TC-HW at the indicated concentrations for 24 h. Luciferase activity for ATF3 promoter activity was measured as a ratio of firefly luciferase signal/renilla luciferase signal using a dual luciferase assay kit. *P < 0.05 compared to cell without TC-HW. c HCT116 cells were treated with TC-HW (100 μg/ml) in presence/absence of NAC. Cell lysates were subjected to SDS-PAGE and the Western blot was performed using antibody against ATF3. Actin was used as internal control for Western blot analysis. d HCT116 cells were co-transfected with ATF3 promoter and pRL-null for 24 h, and then treated with TC-HW (100 μg/ml) in presence/absence of NAC. Luciferase activity for ATF3 promoter activity was measured as a ratio of firefly luciferase signal/renilla luciferase signal using a dual luciferase assay kit. *P < 0.05 compared to cell without TC-HW. e HCT116 cells were transfected with control and ATF3 siRNA and then treated with TC-HW (100 μg/ml). Cell lysates were subjected to SDS-PAGE and the Western blot was performed using antibody against ATF3 or cleaved PARP. Actin was used as internal control for Western blot analysis