Fig. 5

GT3 pre-treatment decreased droxinostat-induced apoptosis in colon cancer cells. a– HT-29 cells were treated with 10 μM GT3 followed by 21 μM droxinostat treatment for 24 h. Cellular apoptosis was measured with annexin V and PI staining. Apoptotic data are expressed as the means ± SD of three independent experiments. *p < 0.05 vs. vehicle, **p < 0.01 vs. vehicle, ^##p < 0.01 vs. GT3. b – HT-29 cells were treated with 10 μM GT3 followed by 21 μM droxinostat treatment for 24 h. The activity of caspase-3 was measured at 405 nm and is expressed as optical density (OD). c – HT-29 cells were treated with 10 μM GT3 followed by 21 μM droxinostat treatment for 24 h. Expression of c-FLIP was assessed via RT-PCR. GADPH was used as a housekeeping control. ***p < 0.001 vs. vehicle, ^##p < 0.01 vs. GT3. d – HT-29 cells were treated with 10 μM Z-VAD-FMK followed by 21 μM droxinostat treatment for 24 h. ROS production was measured using DCFDA and analyzed via flow cytometry. The levels of ROS are expressed as the means ± SD of three independent experiments. ***p < 0.001 vs. vehicle, ^#p < 0.05 vs. droxinostat. e – HT-29 cells were treated with 10 μM Z-VAD-FMK followed by 21 μM droxinostat treatment for 24 h. Expression of c-FLIP was assessed via RT-PCR. f – 500 HT-29 cells were seeded in 6-well plates and treated with or without Z-VAD-FMK and droxinostat. Ten days later, the colony numbers were counted and are expressed as the colony survival fraction. ***p < 0.001 vs. vehicle, ^##p < 0.01 vs. Z-VAD-FMK