Effects of vitamin E on the killing of cultured hepatocytes by tert-butyl hydroperoxide.
The disposition of vitamin E was examined in cultured rat hepatocytes intoxicated with tert-butyl hydroperoxide (TBHP). Culturing of the cells overnight (18-20 hr) with approximately 60 nM alpha-tocopherol (alpha-T) equivalents [Williams' E medium, 18 nM tocopherol phosphate (alpha-TP), 9% fetal calf serum, 43 nM alpha-T] resulted in a content of alpha-T that was 16% of the concentration of vitamin E measured in freshly isolated hepatocytes. Supplementation of the medium with 1 microM alpha-TP maintained the alpha-T concentration of the cultured cells at the level of freshly isolated hepatocytes. Supplemented hepatocytes exposed to TBHP showed decreased lipid peroxidation and delayed cell killing, compared with hepatocytes not cultured overnight with alpha-TP. Killing of the supplemented cells by TBHP was accompanied by a loss of alpha-T. Pretreatment of supplemented hepatocytes with the iron chelator deferoxamine prevented much of the loss of alpha-T. At the same time, deferoxamine inhibited both the lipid peroxidation and cell killing. The antioxidant N,N'-diphenyl-1,4-phenylenediamine reduced the loss of alpha-T and significantly decreased lipid peroxidation. In the presence of N,N'-diphenyl-1,4-phenylenediamine, cell killing was delayed by 15 min and reduced in extent. Overnight supplementation of hepatocytes with nonesterified alpha-T, or vitamin E esters other than alpha-TP, similarly rendered the cells less sensitive to TBHP. The nonesterified alpha-T produced a higher cell-associated vitamin E concentration than did the esters; however, nonesterified alpha-T did not result in greater protection against TBHP. These data indicate that the mechanisms of the cell killing by TBHP are the same in cultured hepatocytes that contain low or physiological concentrations of vitamin E.
Glascott
P A
PA
Department of Pathology, Thomas Jefferson University, Philadelphia, Pennsylvania 19107.
Gilfor
E
E
Farber
J L
JL
D000818:Animals; D016923:Cell Death / Q000187:drug effects; D002478:Cells, Cultured; D004952:Esters / Q000494:pharmacology; D007501:Iron / Q000378:metabolism; D008099:Liver / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D010084:Oxidation-Reduction; D010545:Peroxides / Q000633:toxicity*; D051381:Rats; D011919:Rats, Inbred Strains; D014810:Vitamin E / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D020122:tert-Butylhydroperoxide
D004952:Esters; D010545:Peroxides; D014810:Vitamin E; D020122:tert-Butylhydroperoxide; D007501:Iron