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. 2017 Mar 28;8(25):40741–40755. doi: 10.18632/oncotarget.16580

Figure 9. Effect of antioxidant NAC and cathepsin B inhibitor CA-074 Me on the mitochondrial apoptosis pathway in SW480 cells.

Figure 9

SW480 cells were pretreated with or without 2mM N-acetyl-L-cysteine (NAC) or 50μM CA-074 Me followed by exposure to43°C for 2h and further incubation at 37°C for 6h. A. Flow cytometry analysis heat stress-induced mitochondrial depolarization (low ΔΨm). B. The histogram represents the quantification of the percentage of low ΔΨm analyzed by flow cytometry. C. Intracellular location of cytochrome C was determined by Western blots. β-actin was run as an internal control. COX IV was used as a mitochondrial loading control. D. Quantification of Western blots for cytochrome C. E. Bcl-2 and Bax were determined by Western blots. β-actin was run as an internal control. F. Quantification of Western blots for Bax/Bcl-2 ratio. G. Enzymatic activity of caspase-9 was measured in cell lysates using a fluorogenic substrate, Ac-LEHD-AFC. H. Enzymatic activity of caspase-3 was measured in cell lysates using a fluorogenic substrate, Ac-DEVD-AMC. The data shown represent the mean ±SD of at least three independent experiments, performed in triplicate. *P < 0.05, statistically significant relative to control (37°C),#P < 0.05, statistically significant relative to heat stress group (HS).