Figure 2. Cisplatin induces autophagy in H460wt cells and H460crp53 cells.

Cells were treated with cisplatin at the indicated doses for 24 h (day 0), after which cells were washed and incubated in fresh medium. A. Acridine orange staining was performed after exposure to 10 μM cisplatin. Cells were treated with cisplatin for 24 h (day 0) and stained 48 h post-drug removal (day 3, n=3). B. Quantification of acridine orange staining. Autophagy induction was quantified by flow cytometry in response to increasing concentration of cisplatin 2 days after drug exposure (n=3). C. Western blotting. Levels of p62 were determined by western blotting at the indicated times after 10 μM cisplatin exposure for 24 h (D0). Lysates were collected on indicated days. One of three representative experiments is shown (n=3). D. Western blot Densitometry. The bar graph in each panel indicates the relative band intensity generated from densitometric scans of two independent experiments in arbitrary densitometric units (n=3). E. Co-localization of LC3 and LAMP. Fluorescence microscopy showing LC3 and LAMP2 co-localization in response to 10 μM cisplatin exposure 2 days after cisplatin removal. (20X objective, n=2) Unless stated, otherwise data were from three independent experiments, *p < 0.05, cisplatin treated group vs untreated control group in each cell line.