The Role of Thiols in Mitochondrial Susceptibility to Iron and tert-Butyl Hydroperoxide-Mediated Toxicity in Cultured Mouse Hepatocytes

• Published in: Chemical research in toxicology 1994-05, Vol.7 (3), p.358-366
• Main Authors: Shertzer, Howard G, Bannenberg, Gerard L, Zhu, Huan, Liu, Rui-Ming, Moldeus, Peter
• Format: Article
• Language: English
• Subjects: Animals; Biological and medical sciences; Calcium - metabolism; Cell Death - drug effects; Cell Line; Cell Membrane - metabolism; Cell Nucleus - drug effects; Cell Nucleus - metabolism; Chemical and Drug Induced Liver Injury - physiopathology; Chemical and industrial products toxicology. Toxic occupational diseases; DNA - chemistry; Glutathione - metabolism; Iron - toxicity; Lipid Peroxidation - drug effects; Medical sciences; Mice; Microscopy, Fluorescence; Mitochondria, Liver - drug effects; Oxidation-Reduction; Peroxides - toxicity; Reactive Oxygen Species - toxicity; Spectrometry, Fluorescence; Sulfhydryl Compounds - pharmacology; tert-Butylhydroperoxide; Toxicology; Various organic compounds
• ISSN: 0893-228X; 1520-5010
• DOI: 10.1021/tx00039a013

Cultured hepatocytes derived from the newborn mutant c14CoS/c14CoS mouse (14CoS/14CoS cells) have 3-fold higher levels of reduced glutathione (GSH) and greater resistance to menadione toxicity than hepatocytes derived from the wild-type cch/cch mouse (ch/ch cells). Therefore, we used these cell lines to examine mechanisms of oxidative stress produced by iron and tert-butyl hydroperoxide (TBHP). Both cell types were resistant to 25 microM Fe2+ toxicity in the absence of added TBHP. However, in the presence of Fe2+, striking differences in susceptibility to TBHP toxicity between the cell types were observed. With 25 microM Fe2+, ch/ch cells showed TBHP concentration-dependent toxicity, with total lethality at 500 microM; in contrast, 14CoS/14CoS cells were completely resistant to the lethal effects of this concentration of TBHP. Concentration-dependent TBHP-mediated increases in cytosolic Ca2+, pH, and GSSG/GSH ratios, and decreases in GSH levels, were evident in ch/ch cells. 14CoS/14CoS cells exhibited concentration-dependent TBHP-mediated changes in GSH and GSSG/GSH ratios, but cytosolic Ca2+ and pH remained at control levels. Mitochondrial GSH pools were also diminished by TBHP, although there was no selective depletion; mitochondrial GSH remained at about 14% of total cellular GSH. Both cell types exhibited the same time-dependent decrease in plasma membrane protein thiols and a time-dependent increase in plasma membrane protein carbonyls. However, only ch/ch cells displayed a time-dependent depletion of mitochondrial protein thiols, concomitant with an increase in mitochondrial protein carbonyls, while 14CoS/14CoS cells were resistant to such changes. All of the effects produced by TBHP were prevented by desferoxamine.