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Lipoate/ascorbate-dependent recycling prevents myeloperoxidase-catalyzed oxidation of vitamin e homologues phenoxyl radical in live HL-60 cells.
Yalowich JC; Kuzmenko AI; Tyurina YY; Kisin E; Shvedova AA; Kagan VE
Toxicologist 2001 Mar; 60(1):40
Effectiveness of the antioxidant system in cells is believed to be due to antioxidant recycling mechanisms. Several antioxidant cascades including ascorbate/thiols and enzymatic electron carriers have been suggested to participate in vitamin E recycling based on experiments in cell-free systems. We report that thiol-driven redox-cycling of vitamin E operates in live HL-60 cells. Myeloperoxidase-rich HL-60 cells were able to generate, in the presence of H2O2, phenoxyl radicals of 2,2,5,7,8-pentamethyl-6-hydroxy chromane (PMC), a vitamin E homologue devoid of the hydrocarbon side-chain. The steady-state concentrations of PMC phenoxyl radicals were high enough to be directly detectable by EPR spectroscopy. Pretreatment of HL-60 cells with succinyl acetone, an inhibitor of heme synthesis, resulted in a decreased activity of myeloperoxidase and an alleviated levels of PMC phenoxyl radicals. Preloading with ascorbate (achieved by the addition of dehydroascorbate to the growth medium for 1 h) resulted in disappearance of PMC phenoxyl radical signal and appearance of ascorbate radical signal in the cells exposed to PMC. HPLC measurements confirmed that myeloperoxidase-catalyzed PMC consumption was delayed and oxidation of ascorbate was detected during this period. Pretreatment of the cells with lipoate and dehydroascorbate further postponed PMC oxidation and increased the effectiveness of ascorbate in recycling PMC phenoxyl radical as evidenced by both EPR measurements and HPLC assays. We conclude that electron transfer reactions in the recycling cascade (dihydro)lipoate->ascorbate>PMC phenoxyl radical was operational in live HL-60 cells.
Oxidation; Vitamins; Antioxidants; Thiols; Exposure-levels; Exposure-assessment; Cell-cultures
Issue of Publication
The Toxicologist. Society of Toxicology 40th Annual Meeting, March 25-29, 2001, San Francisco, California
Page last reviewed: September 2, 2020
Content source: National Institute for Occupational Safety and Health Education and Information Division