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

• Published in: Biological & pharmaceutical bulletin Vol. 23; no. 1; pp. 37 - 42
• Main Authors: KANNO, Shuuichi, ISHIKAWA, Masaaki, TAKAYANAGI, Motoaki, TAKAYANAGI, Yoshio, SASAKI, Ken-ichi
• Format: Journal Article
• Language: English
• Published: Tokyo The Pharmaceutical Society of Japan 01.01.2000; Maruzen; Japan Science and Technology Agency
• Subjects: Acetylcysteine - pharmacology; Ageing, cell death; Animals; Antioxidants - pharmacology; apoptosis; Apoptosis - drug effects; Ascorbic Acid - pharmacology; Biological and medical sciences; Cell physiology; Cell Survival - drug effects; Cells, Cultured; DNA Fragmentation - drug effects; Dose-Response Relationship, Drug; Fundamental and applied biological sciences. Psychology; hydrogen peroxide (H2O2); Hydrogen Peroxide - antagonists & inhibitors; Hydrogen Peroxide - toxicity; Liver - cytology; Liver - drug effects; Male; Mice; Molecular and cellular biology; oxidative stress; primary cultured hepatocyte; Cell culture; Oxidative stress; Digestive system; Toxicity; Liver; Rodentia; Fluorescence; Hydrogen Peroxides; In vitro; Vertebrata; Mammalia; Hepatocyte; Mouse; Cell death; Primary culture; Apoptosis
• ISSN: 0918-6158; 1347-5215
• DOI: 10.1248/bpb.23.37

The influence of oxidative stress by hydrogen peroxide (H2O2) was examined in mouse primary cultured hepatocytes. A change in morphology was observed in hepatocytes incubated for 30min in saline A containing H2O2. 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 H2O2. 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 H2O2-induced cell viability reduction suggested that oxidative stress by H2O2 itself or H2O2-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.