Fig. 4

Reactive oxygen species (ROS) production were involved in CTB-induced hepatoma cells apoptosis. a, b Intracellular reactive oxygen species (ROS) was detected by Flow cytometry analysis and microscopic observation, using fluorescent probe DCFH-DA Scale bar: 50 μm. c MitoSOX Red immunofluorescence staining evaluated mitochondrial superoxide. Scale bar: 50 μm. d SOD2 enzyme activity was detected with Total Superoxide Dismutase Assay Kit with WST-8. e Cells were treated with CTB at the indicated concentrations (0, 1, 2, 4 μΜ) or NAC (10 mΜ) for 24 h. Cell viability was determined by the MTT assay. f TUNEL staining evaluated cells apoptosis. Data are represented as mean ± SD. Significance: *P < 0.05, **P < 0.01 and ***P < 0.001 vs Control; ^#P < 0.05, ^##P < 0.01 and ^###P < 0.01 vs CTB (2 μΜ) treatment