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. 2013 Sep 3;104(11):1476–1482. doi: 10.1111/cas.12253

Figure 6.

Figure 6

Mitochondrial‐derived reactive oxygen species (ROS) production plays an essential role in peneciraistin C (Pe‐C)‐induced autophagic cell death. (a) A549 cells were treated with 3 μM Pe‐C for the indicated time, then ROS generation was detected using chloromethyl‐2′,7′‐dichlorofluorescein diacetate (CM‐H2‐DCFDA, 10 μM) by flow cytometry. Data were processed with CellQuest software and analyzed by densitometry. (b) A549 cells were treated with Pe‐C (3 μM) in the absence (–) or presence of antioxidants N‐acetyl‐l‐cysteine (NAC, 10 mM) or Mito‐TEMPO (100 μM) for 12 h; CM‐H2‐DCFDA was added 30 min before end of treatment. The ROS levels were detected with a flow cytometer and data were analyzed as described in (a). (c, d) A549 cells were treated with Pe‐C (3 μM) in the absence (–) or presence of antioxidants NAC (10 mM) or Mito‐TEMPO (100 μM) for 48 h. (c) Cell viabilities were determined by MTT assay. (d) Cells were fixed for immunofluorescence staining with anti‐LC3 antibody and observed under a fluorescence microscope.