Fig. 2
Cellular iron is associated with cell death triggered by BA1. Cells were treated with BA1 (LoVo: 2 nM, SW480: 1.5 nM) with or without iron (III)-citrate. (A) Cell viability was determined via the MTT assay. (B) Cell proliferation was analyzed by cell counting. Representative cell images were captured by a light microscope. Scale bar: 1 mm. (C) Cell death was analyzed by flow cytometry using annexin V/PI staining. (D) The expression of caspase-3 and caspase-9 was detected by western blotting. Etoposide (100 µM) treatment was used as a positive control for inducing cleaved caspase-3 and -9. (E,F) Intracellular iron was examined using by FerroOrange. Representative cell images were captured by a laser confocal scanning microscope, scale bar: 10 μm (E), and the relative fluorescence intensity was measured using a microplate reader (F) after cell staining with FerroOrange. (G) The expression of ferritin heavy chain was detected by western blotting. (H) Representative image of lipofuscin stained by Sudan Black B, captured by laser confocal microscope. Scale bar: 10 μm. #p > 0.05 (no significance), *p < 0.05, **p < 0.01, ***p < 0.001, compared with control.
