Cr(IV) causes activation of nuclear transcription factor-kappa B, DNA strand breaks and dG hydroxylation via free radical reactions

Electrophoretic mobility shift, DNA strand breakage assays and electron spi n resonance (ESR) spin trapping were used to investigate the activation of nuclear transcription factor (NF)-kappa B, DNA strand breakage and 2'-deoxy guanosine hydroxylation induced by Cr(IV), as well the role of free radical reactions in these processes. Incubation of synthesized Cr(IV)-glutathione complex with cultured Jurkat cells resulted in activation of DNA binding a ctivity of NF-kappa B. Cr(VI) is also able to induce NF-kappa B activation through Cr(V) and Cr(IV) intermediates generated during the reduction of Cr (VI) by the cells. Cr(III) did not cause observable NF-kappa B activation d ue to its inability to cross cell membranes. Cr(IV)-induced NF-kappa B acti vation is dose-dependent. Catalase inhibited the activation while superoxid e dismutase enhanced it. The metal chelator, deferoxamine, and hydroxyl ((O H)-O-.) radical scavengers, sodium formate and aspirin, also inhibited the NF-KB activation. Electrophoretic assays using lambda Hind III linear DNA s howed that, in the presence of H2O2, Cr(IV) is capable of causing DNA stran d breaks. Deferoxamine, sodium formate and aspirin inhibited the DNA strand breaks. HPLC measurements also show that (OH)-O-. radical generated by the Cr(IV)-mediated reaction with H2O2 was capable of causing 2'-deoxyguanosin e (dG) hydroxylation to generate 8-hydroxyguanosine (8-OHdG). The relative magnitude of 8-OHdG formation correlated with the generation of (OH)-O-. ra dicals. ESR spin trapping measurements showed that reaction of Cr(IV) with H2O2 generated (OH)-O-. radicals, which were inhibited by deferoxamine, sod ium formate and aspirin. The results show that Cr(IV) can cause NF-kappa B activation, DNA strand breaks and dG hydroxylation through (OH)-O-. radical -initiated reactions. This reactive chromium intermediate may play an impor tant role in the mechanism of Cr(VI)-induced carcinogenesis. The results al so suggest that the Cr(IV)-glutathione complex may be used as a model compo und to study the role of Cr(IV) in Cr(VI) carcinogenicity. (C) 1999 Elsevie r Science Inc. All rights reserved.