CAFFEIC ACID PHENETHYL ESTER STIMULATES HUMAN ANTIOXIDANT RESPONSE ELEMENT-MEDIATED EXPRESSION OF THE NAD(P)H-QUINONE OXIDOREDUCTASE (NQO1)GENE

Caffeic acid phenethyl ester (CAFE) is a phenolic antioxidant derived from the propolis of honeybee hives, CAFE was shown to inhibit the for mation of intracellular hydrogen peroxide and oxidized bases in DNA of 12-O-tetradecanoylphorbol-13-acetate (TPA)-treated HeLa cells and was also found to induce a redox change that correlated with differential growth effects in transformed cells but not the nontumorigenic parent al ones, Mediated via the electrophile or human antioxidant response e lement (hARE), induction of the expression of NAD(P)H quinone oxidored uctase (NQO1) and glutathione S-transferase Ya subunit genes by certai n phenolic antioxidants has been correlated with the chemopreventive p roperties of these agents, Here, we determined by Northern analysis th at CAFE treatment of hepatoma cells stimulates NQO1 gene expression in cultured human hepatoma cells (HepG2), and we characterized the effec ts of CAFE treatment on the expression of a reporter gene either conta ining or lacking the hARE or carrying a mutant version of this element in rodent hepatoma (Hepa-1) transfectants. A dose-dependent transacti vation of human hARE-mediated chloramphenicol acetyltransferase (cat) gene expression was observed upon treatments of the Hepa-1 transfectan ts with TPA, a known inducer, as well as with CAFE, The combined treat ments resulted in an apparent additive stimulation of the reporter exp ression, To learn whether this activation of cat gene expression was e ffected by protein kinase C in CAFE-treated cells, a comparison was ma de of cat gene activity after addition of calphostin, a protein kinase C inhibitor, Calphostin reduced the cat gene induction by TPA but not by CAFE, suggesting that stimulation of gene expression in this syste m by these agents proceeds via distinct mechanisms, Band-shift experim ents to examine binding of transactivator proteins from nuclear extrac ts of treated and untreated cells to a hARE DNA probe showed that TPA exposure increased the binding level, In contrast, binding of factors to this probe was inhibited after either in vivo treatment of cells wi th CAFE or in vitro addition of this compound to the nuclear extract, In view of the clear stimulation by CAFE of gene expression mediated b y hARE, possible explanations of this result are discussed.