Fig 6. Inhibition of Foxo3 signal reversed the antitumor effects of CAPE in MDA-MB-231 cells.

(A) Cells were treated singly or in combination with Compc (10 μM) and CAPE (50 μM) for 12 h, and western blot was used for cytoplasm and nuclear Foxo3 detection. (B-D) Cells were transfected with siRNA, and CCK8 was used for the cell viability detection at 72 h, the colony was investigated by colony formation assay at 5 days and the expression of Cle-caspase3, PARP, and Bcl-xl were detected by western blot at 24 h. (E-H) Cells were treated singly or in combination with TIC10 (1, 5, 10 μM) and CAPE (50 μM) for 72 h. The cell viability was determined by CCK8. The colony was investigated by colony formation assay at 5 days. Apoptosis was determined by Annexin V-FITC/PI dual staining at 48 h. The expression of Cle-caspase3, PARP, and Bcl-xl were detected by western blot at 24 h. (I-N) Cells were transfected with siRNA or treated in combination with TIC10 (10 μM) for 24 h, DCFH-DA was used for ROS detection. The lipid peroxidation was determined by C11-BODIPY staining. The expression of GPX4 and Ferritin were detected using western blot. Values represent the mean ± SD from three independent experiments; ***p<0.001: CAPE compared with control, ^#p <0.05: combinatorial TIC10+CAPE groups compared with the single CAPE group.