ABSTRACT
Flavonoids are ubiquitous substances in fruits and vegetables. Among them, the flavonol kaempferol contributes up to 30% of total dietary flavonoid intake. Flavonoids are assumed to exert beneficial effects on human health, e.g., anticancer properties. For this reason, they are used in food supplements at high doses. The aim of this project was to determine the effects of kaempferol on oxidative stress and apoptosis in H4IIE rat hepatoma cells over a broad concentration range. Kaempferol is rapidly taken up and glucuronidated by H4IIE cells. The results demonstrate that kaempferol protects against H2O2-induced cellular damage at concentrations which lead to cell death and DNA strand breaks in the absence of H2O2-mediated oxidative stress. Preincubation with 50 microM kaempferol exerts protection against the loss of cell viability induced by 500 microM H2O2 (2 h) while the same concentration of kaempferol reduces cell viability by 50% in the absence of H2O2 (24 h). Preincubation with 50 microM kaempferol ameliorates the strong DNA damage induced by 500 microM H2O2 while 50 microM kaempferol leads to a significant increase of DNA breakage in the absence of H2O2. Preincubation with 50 microM kaempferol reduces H2O2-mediated caspase-3 activity by 40% (4 h) while the same concentration of kaempferol leads to the formation of a DNA ladder in the absence of H2O2 (24 h). It is concluded that the intake of high dose kaempferol in food supplements may not be advisable because in our cellular model protective kaempferol concentrations can also induce DNA damage and apoptosis by themselves.
Subject(s)
Antioxidants/pharmacology , Apoptosis/drug effects , Kaempferols/pharmacology , Oxidants/pharmacology , Oxidative Stress/drug effects , Animals , Chromans/pharmacology , DNA/drug effects , DNA/metabolism , DNA Damage , Dose-Response Relationship, Drug , Fluoresceins/chemistry , Formazans/chemistry , Hydrogen Peroxide/pharmacology , Kaempferols/pharmacokinetics , Kaempferols/toxicity , Lipid Peroxides/metabolism , Liver Neoplasms, Experimental/metabolism , Liver Neoplasms, Experimental/pathology , Malondialdehyde/analysis , Neutral Red/chemistry , Rats , Tetrazolium Salts/chemistryABSTRACT
Phytoestrogens such as the soy isoflavonoid daidzein have potential health benefits. The antioxidant properties of phytoestrogens are considered to be responsible in part for their protective effects. The antioxidant enzyme (AOE) system plays an important role in the defense of cells against oxidative insults. To determine whether flavonoids can exert antioxidative effects not only directly but also indirectly by modulating the AOE system, we investigated the influence of the flavonoid daidzein on the expression of different AOE. Daidzein treatment of hepatoma H4IIE cells increased catalase mRNA expression two- to threefold. Expression levels of copper zinc superoxide dismutase (CuZnSOD) were not affected by exposure to daidzein. Manganese superoxide dismutase (MnSOD) mRNA expression levels decreased slightly and glutathione peroxidase (GPx) levels increased slightly after daidzein exposure. Changes in AOE mRNA expression levels were significant at 300 micromol/L daidzein. To elucidate the mechanisms underlying the strong increase in catalase mRNA, transfection experiments were performed. Transient transfection of hepatoma cells with reporter plasmids containing different parts of the upstream region of the catalase gene showed a significant one- to threefold increase in reporter gene activity after daidzein exposure. This indicates that daidzein can directly activate the rat catalase promoter region. Despite the increase in catalase mRNA, daidzein pretreatment of cells did not protect against oxidative stress resulting from H(2)O(2) exposure. On the contrary, daidzein itself exerted a mild oxidative stress. In conclusion, the changes in the AOE system provoked by daidzein affected the oxidant rather than the antioxidant properties of daidzein.
