Figure 2

ESI induces the formation of autophagic vacuoles in H1299 and A549 cells. (a) H1299 cells overexpressing mCherry-LC3 were treated with DMSO or ESI (1.6 μM) for 24 h, and the mCherry-LC3 punctate dots in cells were examined. Positive signals were defined if the cells have five or more mCherry-LC3 dots in the cytoplasm. (b and c) The number of mCherry-LC3 dots per cell (b) and the percentage of the cells with mCherry-LC3 dots (c) were counted under florescence microscope. (d) Ultrastructural features of the H1299 cells treated with ESI (1.6 μM) for 24 h were analyzed by electron microscopy. The typical images of nucleus (N), mitochondria (M) and autophagosomes (arrows) were shown at higher magnification. The number of autophagosomes in H1299 cells was presented. Twenty cross-sections were counted in each experiment. (e) H1299 and A549 cells were treated with DMSO or indicated concentrations of ESI for 24 h, and then compared for expression levels of Beclin1, ATG3 and LC3 by western blot. Actin was included as loading control. All data were representative of three independent experiments. Bars, S.E.M.; *P<0.05, **P<0.01, ***P<0.001