Relationship of the metabolism of vitamins C and E in cultured hepatocytes treated with tert-butyl hydroperoxide.
The relationship between the metabolism of alpha-tocopherol (alpha-T) (vitamin E) and that of ascorbic acid (vitamin C) was examined in cultured hepatocytes intoxicated with tert-butyl hydroperoxide (TBHP). Unlike vitamin E, the cellular content of vitamin C did not decline after overnight culturing of freshly prepared hepatocytes. In addition, this basal vitamin C content was not affected by the presence of alpha-T phosphate in the overnight culture medium. Supplementation of the overnight culture medium with vitamin C (10 microM to 10 mM) increased the cellular content of vitamin C by > 1 order of magnitude. Increasing the cellular content of ascorbate increased the protection against TBHP toxicity, with or without the presence of a physiological content of vitamin E. In vitamin E-supplemented cells, a loss of alpha-T occurred within 15 min of exposure to TBHP and before the decrease in cellular ascorbate content. The vitamin C content declined in parallel with the loss of cell viability. Supplementation of the overnight culture medium with increasing concentrations of ascorbate progressively spared the depletion of alpha-T while preventing the cell killing. Pretreatment with the ferric iron chelator deferoxamine or the antioxidant N,N'-diphenyl-1,4-phenylenediamine prevented the loss of ascorbate and the cell killing by TBHP in hepatocytes either sufficient or deficient in alpha-T. However, neither alpha-T nor ascorbate prevented the accumulation of DNA single-strand breaks caused by TBHP, indicating that these vitamins do not effectively scavenge the TBHP-derived radicals responsible for DNA damage. The data in the present study indicate that vitamins E and C act as independent antioxidants and that ascorbate does not regenerate alpha-T in cultured rat hepatocytes.
Glascott
P A
PA
Department of Pathology and Cell Biology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
Gilfor
E
E
Farber
J L
JL
D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D001205:Ascorbic Acid / Q000378:metabolism*; D002478:Cells, Cultured; D004249:DNA Damage; D004277:DNA, Single-Stranded / Q000187:drug effects; D003676:Deferoxamine / Q000494:pharmacology; D015227:Lipid Peroxidation; D008099:Liver / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D010545:Peroxides / Q000494:pharmacology*; D010655:Phenylenediamines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D014810:Vitamin E / Q000378:metabolism*; D020122:tert-Butylhydroperoxide
D000975:Antioxidants; D004277:DNA, Single-Stranded; D010545:Peroxides; D010655:Phenylenediamines; D014810:Vitamin E; D020122:tert-Butylhydroperoxide; C004845:N,N'-diphenyl-4-phenylenediamine; D003676:Deferoxamine; D001205:Ascorbic Acid