Generation of reactive oxygen species in the enzymatic reduction of PbCrO4 and related DNA damage.
Leonard-SS; Vallyathan-V; Castranova-V; Shi-X
Mol Cell Biochem 2002 May-Jun, 234-235(1-2):309-315
Free radical reactions are believed to play an important role in the mechanism of Cr(VI)-induced carcinogenesis. Most studies concerning the role of free radical reactions have been limited to soluble Cr(VI). Various studies have shown that solubility is an important factor contributing to the carcinogenic potential of Cr(VI) compounds. Here, we report that reduction of insoluble PbCrO4 by glutathione reductase in the presence of NADPH as a cofactor generated hydroxyl radicals (.OH) and caused DNA damage. The OH radicals were detected by electron spin resonance (ESR) using 5,5-dimethyl-N-oxide as a spin trap. Addition of catalase, a specific H2O2 scavenger, inhibited the OH radical generation, indicating the involvement of H2O2 in the mechanism of Cr(VI)-induced OH generation. Catalase reduced OH radicals measured by electron spin resonance and reduced DNA strand breaks, indicating OH radicals are involved in the damage measured. The H2O2 formation was measured by change in fluorescence of scopoletin in the presence of horseradish peroxidase. Molecular oxygen was used in the system as measured by oxygen consumption assay. Chelation of PbCrO4 impaired the generation of OH radical. The results obtained from this study show that reduction of insoluble PbCrO4 by glutathione reductase/NADPH generates OH radicals. The mechanism of OH generation involves reduction of molecular oxygen to H2O2, which generates OH radicals through a Fenton-like reaction. The OH radicals generated by PbCrO4 caused DNA strand breakage.
Free-radical-generation; Free-radicals; Carcinogenesis; DNA-damage; Occupational-exposure; Lung-irritants; Toxins; Respiratory-system-disorders; Lung-cancer; Cancer; Tumors
X. Shi, Pathology and Physiology Research Branch, Health Effects Laboratory Division, National Institute for Occupational Safety and Health, 1095 Willowdale Road, Morgantown, WV 26505, USA
Research Tools and Approaches: Cancer Research Methods
Molecular and Cellular Biochemistry