Figure 5

Helenalin induces autophagy cell death. A2750 cells were treated with increasing concentrations of helenalin for 24 h, following which cells were harvested and cell lysates prepared and subjected to immunoblot analysis for (A) BCL2, BAX and Bid or (B) Atg12 and LC3-B. Actin was used as the loading control. A2750 cells were treated with DMSO, 0.5uM, 1uM, 2uM helenalin or pretreated with 200nM bafilomycin A1 for 45 min before treatment with 2uM helenalin for 24 h. Following which, cells were stained with Acridine Orange Solution and photographed using a florescence microscope (positive AVO staining in red) (C) or (D) trypsinized and harvested for FACS analysis to quantitate levels of cell staining representing autophagy. Figures represent data from three independent experiments.