Effect of addition of etoposide and H2O2 on formation of DMPO-GS nitrone by purified myeloperoxidase and myeloperoxidase activity in HL60 cells
| Additions | DMPO-GS Nitrone |
|---|---|
| μM | |
| Purified myeloperoxidase | |
| DMPO alone | 0.00 |
| + H2O2 | 8.5 ± 1.9 |
| +Etoposide (100 μM) + H2O2 | 27.9 ± 1.02-a |
| + Etoposide (500 μM) + H2O2 | 39.4 ± 1.72-b |
| + Phenol (300 μM) + H2O2 | 391 ± 362-b |
| HL60 cells | |
| DMPO alone | 0.00 |
| + H2O2 | 18.5 ± 2.0 |
| + Etoposide (100 μM) | 18.5 ± 0.5 |
| + Etoposide (100 μM) + H2O2 | 52.5 ± 3.02-c |
Myeloperoxidase (0.1 U/in 100 μl) or HL60 cells (2 × 106 cells/ml) were preincubated with DMPO (100 mM) for 2 min and then incubated in the presence or absence of H2O2(25 μM was added every 10 min) or etoposide in L1210 buffer, pH 7.4, containing 3-amino-1,2,4-triazole (2.5 mM) and DTPA (100 μM) at 37°C for 1 h. Etoposide was added 15 min before the addition of other reagents. After incubation, HL60 cells were washed twice with L1210 buffer, pH 7.4, and DMPO-GS nitrone was determined by HPLC as described in Materials and Methods.