Alteration of the mitochondrial transmembrane potential and ROS production induced by the C-terminal domain of TSP-1. (A, B) NB4-LR1 cells untreated (gray shaded curves) or treated with ATRA (1 μM; filled curves) were cultured for 3 days without or with the TSP-1 T3C1 recombinant fragment (3 μM; open curve). At each day of the treatment, cells were stained with the lipophilic fluorochrome DiOC₆(3) (A) or the oxidant-sensitive dye DCFH-DA (B), as described in “Materials and methods,” and then analyzed by flow cytometry. Percentage of positive cells are indicated. (C, left) Cells were preincubated for 2 hours with noncytotoxic doses of ebselen (Eb; 10 μM and 50 μM) and then treated or not with H₂O₂ (50 μM; 6 hours) for detection of ROS production upon cell staining with DCFH-DA. (Right) Untreated or ATRA-treated (1 μM) cells were cultured for 3 days with T3C1 without or with selenium (Se; 0.06 μM) or ebselen (Eb; 50 μM), and cell death, as quantified by annexin V and DiOC₆(3) staining, was analyzed at day 3. Results are expressed as mean ± SEM of 2 experiments.