Monitoring Human Lymphocytic DNA-Protein Cross-Links as Biomarkers of Biologically Active Doses of Chromate

Costa, Max; Zhitkovich, Anatoly; Toniolo, Paolo; Taioli, Emanuela; Popov, Todor A.; Lukanova, Annekatrin

doi:10.2307/3433010

Cited by 5 publications

(1 citation statement)

References 5 publications

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“…Contact with industrial sources has provided valuable information on the numerous toxic effects associated with exposure to hexavalent chromium, Cr(VI) [1][2][3][4][5][6][7][8]. Since greater than 10 5 tons of Cr are released annually into the environment, and since Cr is a significant contaminant at many sites (Superfund and other) across the U.S.A. [9][10][11], environmental exposure is of increasing concern.…”

Section: Introductionmentioning

confidence: 99%

Reductive activation of hexavalent chromium by human lung epithelial cells: Generation of Cr(V) and Cr(V)-thiol species

Borthiry

Antholine

Myers

et al. 2008

Journal of Inorganic Biochemistry

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Chromium(VI) compounds (e.g. chromates) are cytotoxic, mutagenic, and potentially carcinogenic. The reduction of Cr(VI) can yield reactive intermediates such as Cr(V) and reactive oxygen species. Bronchial epithelial cells are the primary site of pulmonary exposure to inhaled Cr(VI) and are the primary cells from which Cr(VI)-associated human cancers arise. BEAS-2B cells were used here as a model of normal human bronchial epithelium for studies on the reductive activation of Cr(VI). Cells incubated with Na 2 CrO 4 exhibited two Cr(V) ESR signals, g = 1.979 and 1.985, which persisted for at least one hour. The g = 1.979 signal is similar to that generated in vitro by human microsomes and by proteoliposomes containing P450 reductase and cytochrome b 5 . Unlike many cells in culture, these cells continued to express P450 reductase and cytochrome b 5 . Studies with the non-selective thiol oxidant diamide indicated that the g = 1.985 signal was thiol-dependent whereas the g = 1.979 signal was not. Pretreatment with phenazine methosulfate eliminated both Cr(V) signals suggesting that Cr(V) generation is largely NAD(P)H-dependent. ESR spectra indicated that a portion of the Cr (VI) was rapidly reduced to Cr(III). Cells incubated with an insoluble chromate, ZnCrO 4 , also generated both Cr(V) signals, whereas Cr(V) was not detected with insoluble PbCrO 4 . In clonogenic assays, the cells were very sensitive to Na 2 CrO 4 and ZnCrO 4 , but considerably less sensitive to PbCrO 4 .

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Section: Introductionmentioning

confidence: 99%

Reductive activation of hexavalent chromium by human lung epithelial cells: Generation of Cr(V) and Cr(V)-thiol species

Borthiry

Antholine

Myers

et al. 2008

Journal of Inorganic Biochemistry

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Carcinogenic chromium(VI)‐induced protein oxidation and lipid peroxidation: implications in DNA–protein crosslinking

Mattagajasingh

Misra

2008

J of Applied Toxicology

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Hexavalent chromium [Cr(VI)] compounds are Group-I human carcinogens. Cr(VI)-induced DNA-protein crosslinks (DPCs) have been implicated in the mutagenic and carcinogenic effects of Cr(VI). Although multiple mechanisms have been suggested for Cr(VI)-induced DNA-protein crosslinking, the mechanism of formation of DNA-protein crosslinks is not well understood. In this study, we explored the hypothesis that Cr(VI)-induced DPCs could be formed via generation of protein carbonyls and malonaldehyde (MDA) through protein oxidation and lipid peroxidation, respectively. Treatment of human leukemic T-lymphocyte MOLT4 cells with potassium chromate induced the formation of protein carbonyls and DPCs within 2 h, but increased the level of MDA only after 4 h, in a dose-dependent manner. Chromate treatment of MOLT4 cell homogenates also resulted in increased formation of MDA and protein carbonyls in a dose-dependent manner. EPR spectrometry in combination with spin trapping techniques revealed that reaction of Cr(VI) with biological reductants such as NADPH, glutathione reductase or H(2)O(2) generates Cr(V) and (*)OH radicals. Pretreatment of cells with antioxidants such as alpha-tocopherol or Tiron inhibited chromate-induced increase in formation of protein carbonyls, MDA and DPCs, but pretreatment of cells with riboflavin or 3-aminotriazole, a catalase inhibitor, had the opposite effect. Our results, for the first time, demonstrate that Cr(VI) exposure increases the cellular level of protein carbonyls and that Cr(VI)-induced DPCs may be formed, at least in part, via generation of protein carbonyls.

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Reduction of hexavalent chromium by human cytochrome b5: Generation of hydroxyl radical and superoxide

Borthiry

Antholine

Kalyanaraman

et al. 2007

Free Radical Biology and Medicine

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The reduction of hexavalent chromium, Cr(VI), can generate reactive Cr intermediates and various types of oxidative stress. The potential role of human microsomal enzymes in free radical generation was examined using reconstituted proteoliposomes (PLs) containing purified cytochrome b 5 and NADPH:P450 reductase. Under aerobic conditions, the PLs reduced Cr(VI) to Cr(V) which was confirmed by ESR using isotopically pure 53 Cr(VI). When 5-Diethoxyphos-phoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO) was included as a spin trap, a very prominent signal for the hydroxyl radical (HO • ) adduct was observed as well as a smaller signal for the superoxide (O 2 •− ) adduct. These adducts were observed even at very low Cr(VI) concentrations (10 μM). NADPH, Cr(VI), O 2 and the PLs were all required for significant HO • generation. Superoxide dismutase eliminated the O 2 • − adduct and resulted in a 30% increase in the HO • adduct. Catalase largely diminished the HO • adduct signal indicating its dependence on H 2 O 2 . Some sources of catalase were found to have Cr(VI)-reducing contaminants which could confound results, but a source of catalase free of these contaminants was used for these studies. Exogenous H 2 O 2 was not needed, indicating that it was generated by the PLs. Adding exogenous H 2 O 2 , however, did increase the amount of DEPMPO/ HO • adduct. The inclusion of formate yielded the carbon dioxide radical adduct of DEPMPO, and experiments with dimethylsulfoxide (DMSO) plus the spin trap α-phenyl-N-tert-butylnitrone (PBN) yielded the methoxy and methyl radical adducts of PBN, confirming the generation of HO • . Quantification of the various species over time was consistent with a stoichiometric excess of HO • relative to the net amount of Cr(VI) reduced. This also represents the first demonstration of a role for cytochrome b 5 in the generation of HO • . Overall, the simultaneous generation of Cr(V) and H 2 O 2 by the PLs and the resulting generation of HO • at low Cr(VI) concentrations could have important implications for Cr(VI) toxicity.

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Monitoring Human Lymphocytic DNA-Protein Cross-Links as Biomarkers of Biologically Active Doses of Chromate

Cited by 5 publications

References 5 publications

Reductive activation of hexavalent chromium by human lung epithelial cells: Generation of Cr(V) and Cr(V)-thiol species

Reductive activation of hexavalent chromium by human lung epithelial cells: Generation of Cr(V) and Cr(V)-thiol species

Carcinogenic chromium(VI)‐induced protein oxidation and lipid peroxidation: implications in DNA–protein crosslinking

Reduction of hexavalent chromium by human cytochrome b5: Generation of hydroxyl radical and superoxide

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