TABLE 1

Kinetics of cell death induced by peroxynitrite in the absence or presence of AACOCF3 and cytoprotection by arachidonic acid, cyclosporin A, N-acetyl-l-cysteine, or catalase

Cells were exposed for 3 min to either 100 μM or 1.2 mM peroxynitrite, centrifuged, and then postincubated for a further 57 min in saline A in the absence or presence (only for cells exposed to 100 μM peroxynitrite) of AACOCF3 (50 μM). In some experiments, arachidonic acid (0.1 μM) or catalase (10 U/ml) was also added during the 57-min post-treatment incubation. In other experiments, cyclosporin A (0.5 μM) was given to the cultures 5 min before peroxynitrite and freshly readded during the 57-min post-treatment incubation. The last experimental condition involved a 60-min exposure to N-acetyl-l-cysteine (5 mM) before treatment with peroxynitrite, as detailed above. After treatments, cells were analyzed for viability using the trypan blue exclusion assay. Results represent the mean ± S.E.M. from at least four separate experiments.

Treatment Trypan Blue-Negative Cells
5 min 10 min 15 min 30 min 60 min
% of control
Arachidonate 97 ± 3.6 96 ± 3.9 98 ± 4.1 95 ± 4.8 97 ± 3.2
Cyclosporin A 95 ± 4.6 98 ± 3.3 96 ± 2.7 99 ± 2.7 95 ± 3.5
AACOCF3 95 ± 3.8 94 ± 2.9 94 ± 4.1 96 ± 4.4 94 ± 3.8
N-Acetyl-l-cysteine 99 ± 4.0 97 ± 2.8 95 ± 2.7 94 ± 3.3 93 ± 2.5
Catalase 96 ± 2.9 95 ± 3.4 93 ± 3.8 98 ± 4.1 96 ± 4.7
Peroxynitrite (100 μM) 94 ± 5.2 93 ± 6.3 95 ± 4.7 91 ± 3.8 91 ± 4.5
    + Arachidonate 96 ± 4.5 97 ± 4.2 91 ± 5.7 92 ± 2.7 93 ± 5.1
    + Cyclosporin A 97 ± 4.3 96 ± 3.6 93 ± 2.8 92 ± 5.8 93 ± 3.3
    + N-acetyl-l-cysteine 95 ± 5.0 94 ± 3.8 94 ± 4.5 90 ± 3.9 92 ± 4.9
    + Catalase 93 ± 5.8 95 ± 3.6 91 ± 3.3 94 ± 4.9 96 ± 4.1
    + AACOCF3 93 ± 5.5 89 ± 3.4 80 ± 4.5* 67 ± 5.2** 51 ± 2.9**
    + AACOCF3 + Arachidonate 93 ± 6.1 91 ± 2.4 91 ± 5.3 92 ± 4.6 91 ± 3.8
    + AACOCF3 + Cyclosporin A 93 ± 4.7 92 ± 3.5 90 ± 3.6 91 ± 2.4 92 ± 3.9
    + AACOCF3 + N-Acetyl-l-cysteine 90 ± 3.6 87 ± 4.8 81 ± 5.3* 62 ± 3.7** 54 ± 4.4**
    + AACOCF3 + Catalase 94 ± 2.9 83 ± 4.8 77 ± 3.1* 69 ± 4.0** 52 ± 5.1**
Peroxynitrite (1.2 mM) 94 ± 5.8 88 ± 3.6 81 ± 4.2* 64 ± 5.0** 55 ± 2.1**
    + Arachidonate 91 ± 4.0 91 ± 3.3 92 ± 4.5 89 ± 5.3 93 ± 6.7
    + Cyclosporin A 89 ± 4.9 91 ± 3.0 90 ± 5.8 93 ± 6.1 90 ± 4.6
    + N-acetyl-l-cysteine 93 ± 3.6 92 ± 2.5 94 ± 4.4 95 ± 5.8 91 ± 3.8
    + Catalase 94 ± 5.3 89 ± 4.7 92 ± 4.1 91 ± 5.3 93 ± 6.1

  • ↵* , P < 0.01; **, P < 0.001 compared with cells that did not receive peroxynitrite.