Fig 10.

mΔψ hyperpolarization-dependent mitochondrial ROS generation is involved in death induced by LeTx. (A) RAW264.7 cells were pretreated with DPI (2 μM), rotenone (1 μM), or antimycin A (10 μg/ml) for 1 h and further incubated with or without LeTx (500 ng/ml LF and 1 μg/ml PA) or LeTx plus inhibitors for 3 h. Cells were then washed with prewarmed culture medium and stained with TMRM (top) or Mitosox red (bottom) as described in Materials and Methods. Data are representative of four independent experiments. (B) RAW264.7 cells were pretreated with apocynin (0.5 mM) or Mito-Tempo (10 μM) for 1 h and treated with LeTx as described above (A). Cells were stained with TMRM (top) or Mitosox red (bottom) as described in Materials and Methods. Data are representative of three independent experiments. (C) Cells were preincubated as for panels A and B and incubated with LeTx (250 ng/ml LF and 500 μg/ml PA) for 5 h. Cell death was measured by MTT assay, and data are expressed as means ± SD (n = 3).