Characterization of Hydrogen Peroxide-Induced Apoptosis in Mouse Primary Cultured Hepatocytes

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The influence of oxidative stress by hydrogen peroxide (H_2O_2) was examined in mouse primary cultured hepatocytes. A change in morphology was observed in hepatocytes incubated for 30min in saline A containing H_2O_2. The percentage of dead cells, as measured by the fluorescence method, was increased in a dose-dependent manner. In addition, a ladder-like DNA fragmentation pattern was detected by agarose gel electrophoresis 1 h after exposure to 3 mM H_2O_2. This phenomenon was prolonged for 24 h. Hydrogen peroxide-induced cell viability reduction and DNA fragmentation were dose-dependently protected by the addtion of antioxidants (N-acetylcysteine, L-ascorbic aicd), a metal-chelator (1,10-phenanthroline), iron-chelator (deferoxamine) and intracellular calcium ion chelator (quin 2-AM). No influence, however, was detected by endonuclease inhibitors (zinc, aurintricarboxylic acid) and poly (ADP-ribose) polymerase inhibitors (3-aminobenzamide, theophylline). These results following H_2O_2-induced cell viability reduction suggested that oxidative stress by H_2O_2 itself or H_2O_2-derived changes involved in ferrous or intracellular calcium ions resulted in apoptosis in mouse primary cultured hepatocytes. These phenomena are not likely to be associated with endonuclease or poly (ADP-ribose) polymerase.
社団法人日本薬学会の論文
2000-01-01