Abstract
Oltipraz protects cells from chemical-induced carcinogenesis partly because of phase 2 enzyme induction. Certain oltipraz metabolites also induce phase 2 enzymes. This study investigated the cytoprotective effects of the oxidized metabolites of oltipraz against arachidonic acid (AA), a proinflammatory fatty acid that causes cellular reactive oxygen species (ROS) production and mitochondrial impairment, and the mechanistic basis of their action in HepG2 cells. Treatment with 4-methyl-5-(pyrazin-2-yl)-3H-1,2-dithiol-3-one (M1) or 7-methyl-6,8-bis(methylthio)H-pyrrolo[1,2-a]-pyrazine (M2), but not 7-methyl-8-(methylsulfinyl)-6-(methylthio)H-pyrrolo[1,2-a]pyrazine (M3) or 7-methyl-6,8-bis(methylsulfinyl)H-pyrrolo[1,2-a]pyrazine (M4), enabled cells to protect against AA-induced apoptosis. M1 and M2 treatment protected cells from ROS produced by AA and inhibited AA-induced glutathione depletion. Moreover, both M1 and M2 effectively inhibited mitochondrial dysfunction induced by AA, although M2 alone slightly elicited it at a relatively high concentration. M1 and M2 activated AMP-activated protein kinase (AMPK), but M3 and M4 failed to do so. AMPK activation by M1 and M2 contributed to cell survival against AA through a decrease in cellular ROS production and prevention of mitochondrial dysfunction, as shown by the reversal of the metabolites' restoration of mitochondrial membrane potential by compound C treatment or overexpression of a dominant-negative mutant AMPK. Consistently, 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside, an AMPK activator, also had a cytoprotective and antioxidant effect against AA. Our results demonstrate that, of the major metabolites of oltipraz, M1 and M2 are capable of protecting cells from AA-induced ROS production and mitochondrial dysfunction, which may be associated with AMPK activation.
Footnotes
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This work was supported by the Korea Science and Engineering Foundation [Grant R11-2007-107-01001-0] funded by the Korea government (Ministry of Education, Science and Technology Development).
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Y.N.K. and S.M.S. contributed equally to this work.
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.108.025908.
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ABBREVIATIONS: Nrf2, NF-E2-related factor-2; GST, glutathione S-transferase; C/EBP, CCAAT/enhancer binding protein; AMPK, AMP-activated protein kinase; AA, arachidonic acid; M1, 4-methyl-5-(pyrazin-2-yl)-3H-1,2-dithiol-3-one; M2, 7-methyl-6,8-bis(methylthio)H-pyrrolo[1,2-a]pyrazine; M3, 7-methyl-8-(methylsulfinyl)-6-(methylthio)H-pyrrolo[1,2-a]pyrazine; M4, 7-methyl-6,8-bis(methylsulfinyl)H-pyrrolo[1,2-a]pyrazine; GSH, glutathione; ROS, reactive oxygen species; ACC, acetyl-CoA carboxylase; PARP, poly(ADP-ribose)polymerase; TUNEL, terminal deoxynucleotidyl transferase dUTP nick-end labeling; AICAR, 5-aminoimidazole-4-carboxamide-1-β-d-ribofuranoside; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide; Rh123, rhodamine 123; DCFH-DA, 2′,7′-dichlorofluorescein diacetate; NAC, N-acetyl-l-cysteine; DMEM, Dulbecco's modified Eagle's medium; FBS, fetal bovine serum; PBS, phosphate-buffered saline; MMP, mitochondrial membrane potential; Ad-LacZ, adenoviral LacZ; Ad-DN, adenoviral dominant negative; PPD, pyrrolopyrazine thione; cyt c, cytochrome c; CsA, cyclosporin A.
- Received December 1, 2008.
- Accepted March 18, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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