Inhibition of the opening of the PTP antagonizes POP effects on mitochondrial function and cell viability. Jurkat cells (10⁶/mL) were treated with POP (50 μM) in the absence or presence of the PTP inhibitor cyclosporin A (CsA; 1 μL) or with nonirradiated psoralen (PS; 50 μM). CT indicates untreated cells. (A) After 4 hours of incubation, cells were stained with 25 nM TMRM to detect changes in mitochondrial membrane potential (Δψ_m). The images were acquired before and after the addition of the mitochondrial uncoupler FCCP (2 μM). The differences between the fluorescence values of each cell reflect the actual extent of Δψ_m. The values obtained in treated cells were normalized to those obtained in untreated cells at time 0. (B) After 20 hours of incubation, apoptosis and necrosis were evaluated by means of Hoechst 33 258 and PI fluorescence, respectively. Values are means ± SD of at least 4 experiments. (C) Mitochondrial swelling, which reflects PTP opening, was monitored as the decrease in light absorbance at 540 nm. RLM (0.5 mg protein/mL) were incubated with 80 μM Ca²⁺, which triggered PTP opening only upon the further addition of POP (10 μM). Since Ca²⁺ did not cause mitochondrial swelling in untreated mitochondria (CT), POPs caused an increased sensitivity of the PTP to Ca²⁺. Mitochondrial swelling was prevented by CsA (1 μM) demonstrating the involvement of the PTP.