Fig. 2.

Hemigramicidin S–conjugated 4-amino-2,2,6,6-tetra-methyl-piperidine-N-oxyl (hemi-GS-TEMPO) 5-125 inhibits γ–irradiation induced (A) superoxide generation. Mouse embryonic cells were exposed to 10 Gy of γ-irradiation. The hemi-GS-TEMPO (20 μM) was added to cells either 10 minutes before or 1 hour after irradiation and removed after 5 hours of incubation. For comparison, cells were incubated with 100 U/ml of polyethylene glycol-superox-ide dismutase (PEG-SOD) 10 minutes before irradiation. Cells were incubated with 5 μM of dihydroethidium (DHE) for 30 minutes at the indicated times. Ethidium fluorescence was analyzed by using a FACScan flow cytometer supplied with CellQuest software. Mean fluorescence intensity from 10,000 cells was acquired by using a 585-nm band-pass filter. (B) Cardiolipin oxidation. Cardiolipin hydroperoxides were determined by using a fluorescent high-performance liquid chromatography (HPLC)-based Amplex Red assay. Data presented as mean ± SE (n = 3). *p < 0.01 vs. nonirradiated cells; # (&)p < 0.01 (0.05) vs. irradiated cells without 5-125 treatment under the same condition. Insert is a typical 2D-high performance thin layer chromatography profile of phospholipids from cells.