Repair of chromate-induced DNA damage in chick embryo hepatocytes

Cupo, Doreen Y.; Wetterhahn, Karen E.

doi:10.1093/carcin/5.12.1705

Cited by 33 publications

(20 citation statements)

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“…Even though no change in GSH level was observed after chromium(VI) treatment, the steady-state amount of the chromium(VI) thioester in the cells is expected to be small (12), so the detection of any small change in GSH levels is unlikely. Also, previous studies have shown that chromium(VI) stimulates the synthesis of GSH in chicken embryo hepatocytes (6).…”

Section: Discussionmentioning

confidence: 92%

“…Chromium(VI) treatment is known to produce DNA damage in chicken embryo hepatocytes (5,6). The hepatocytes are well suited for these experiments because intracellular levels of the different isozymes of cytochrome P-450 can be increased by f naphthoflavone (16) and isopentanol (17).…”

Section: In Rodentsmentioning

confidence: 99%

“…Chromium(VI) causes DNA strand breaks and cross-links in vivo and in cultured cells (3)(4)(5)(6). It has been proposed that cellular metabolism of chromium(VI) compounds is necessary for DNA damage, since chromium(VI) does not react with isolated DNA under physiological conditions (7).…”

Section: Introductionmentioning

confidence: 99%

“…Alkaline Elution. After sodium chromate treatment, the hepatocytes were analyzed for DNA damage by using the alkaline elution technique based on the procedure of Kohn et al (19) as described previously (6). The alkaline elution technique measures the size distribution of long singlestranded DNA, assuming that the rate of elution of the DNA from a polyvinylchloride filter depends on the length of the DNA and is altered by filter absorption of proteins linked to the DNA (19).…”

mentioning

confidence: 99%

“…GSH was determined spectrofluorometrically according to Hissin and Hilf (22) as described previously (6). Up to 0.1 mM N-acetyl-L-cysteine did not interfere with this assay.…”

mentioning

confidence: 99%

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Modification of chromium(VI)-induced DNA damage by glutathione and cytochromes P-450 in chicken embryo hepatocytes.

Cupo¹,

Wetterhahn²

1985

Proc. Natl. Acad. Sci. U.S.A.

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The role ofglutathione and cytochrome P-450 in the production of DNA damage by chromium(VI) was examined In chicken embryo hepatocytes by the alkaline elution technique. Cellular levels of glutathione and cytochrome P-450 were altered by treating the hepatocytes with N-acetyl-Lcysteine, buthionine sulfoximine, isopentanol, or 8-naphthoflavone. A dramatic increase in chromium(VI)-induced DNA strand breaks was observed after increasing glutathione levels in the cells. Chromium(VI)-induced DNA strand breaks were even more numerous when the level of cytochrome P-450 was also increased. Upon depletion of glutathione levels and induction ofcytochrome P-450 or cytochrome P 448, little or no DNA strand breaks or DNA interstrand cross-links were observed after chromium(VI) treatment. Chromium(VI)-induced DNA-protein cross-links generally decreased after either increases or decreases in cellular levels of glutathione or cytochrome P-450 or P-448. These results suggest that glutathione enhances chromium(VI)-induced DNA damage through metabolic activation of chromium(VI). The possible production of reactive chromium species upon metabolism by glutathione and cytochrome P-450 or P448 and their involvement in DNA damage is discussed.Chromium(VI) compounds are recognized as carcinogens in humans and animals (1) and as mutagens in bacterial and mammalian cell systems (2). Chromium(VI) causes DNA strand breaks and cross-links in vivo and in cultured cells (3)(4)(5)(6). It has been proposed that cellular metabolism of chromium(VI) compounds is necessary for DNA damage, since chromium(VI) does not react with isolated DNA under physiological conditions (7). The DNA-damaging ability of other carcinogenic agents has been shown to be modulated by metabolic pathways such as the cytochrome P450 system, sulfotransferases, and glutathione (GSH) (8). Changes in cellular levels of cytochromes P-450 alter the levels of DNA damage produced by carbon tetrachloride (9), benzo-[a]pyrene (10), and dimethylnitrosamine (11) in rodents.In vitro studies have identified several cellular components that are capable of metabolizing chromium(VI) (12-14). Rat liver microsomes contain a NADPH-dependent chromium(VI) reductase activity (15). Induction of cytochrome(s) P-450 by phenobarbital enhances the chromium(VI) reductase activity ofliver microsomes; however, induction of cytochrome(s) P-448 by 3-methylcholanthrene has no effect (13). GSH also reduced chromium(VI) in vitro under physiological conditions (12,14). The reaction of GSH with chromium(VI) has been proposed to occur by a two-step mechanism that involves the rapid formation of a chromium(VI) thioester followed by the reduction of the chromium(VI) thioester to chromium(III) (12).The present report describes the effect of altering the intracellular concentrations of cytochrome P450 and P-448 and GSH on chromium(VI)-induced DNA damage in primary cultured chicken embryo hepatocytes. Chromium(VI) treatment is known to produce DNA damage in chicken embryo hepatocytes (5, 6). The hepatocytes ar...

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Section: Discussionmentioning

confidence: 92%

Section: In Rodentsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

mentioning

confidence: 99%

“…GSH was determined spectrofluorometrically according to Hissin and Hilf (22) as described previously (6). Up to 0.1 mM N-acetyl-L-cysteine did not interfere with this assay.…”

mentioning

confidence: 99%

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Modification of chromium(VI)-induced DNA damage by glutathione and cytochromes P-450 in chicken embryo hepatocytes.

Cupo¹,

Wetterhahn²

1985

Proc. Natl. Acad. Sci. U.S.A.

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Evaluation of the genotoxic potential of glutaraldehyde

Clair

Bermudez

Gross

et al. 1991

Environ and Mol Mutagen

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The cytotoxic and genotoxic effects of glutaraldehyde were studied in vitro in the human TK6 lymphoblast cell line and in primary cultures of rat hepatocytes. TK6 lymphoblasts were exposed to glutaraldehyde for 2 hr in serum-free GSH-free media. Cytotoxic effects were observed at concentrations as low as 10 microM with only 10% cell survival at 20 microM. Alkaline elution studies indicated that glutaraldehyde-induced DNA-protein crosslinking increased linearly over the concentration range from 0 to 25 microM. Glutaraldehyde-induced mutations were assessed at the thymidine kinase locus over the same concentration range and reached a plateau at 10 microM of about six times the background mutant frequency. At equivalent levels of DNA-protein crosslinks and cytolethality, glutaraldehyde was mutagenic at approximately a one-seventh lower concentration than the rodent nasal carcinogen formaldehyde (Craft et al.; Mutation Research 176:147-155, 1987). Glutaraldehyde induced a marginal increase in unscheduled DNA synthesis in the in vitro hepatocyte DNA repair assay, but only at the two highest concentrations of 50 and 100 microM, indicating the induction of some DNA excision-repair activity. These data demonstrate that glutaraldehyde exhibits DNA-reactive genotoxic activity that may involve, at least in part, DNA-protein crosslinking in these cell culture models. These findings suggest the need to examine the potential carcinogenic activity of glutaraldehyde in appropriate inhalation studies.

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Characterization of DNA‐protein complexes induced in intact cells by the carcinogen chromate

Miller

Costa

1988

Molecular Carcinogenesis

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Potassium chromate induced the formation of DNA-protein complexes in cultured Chinese hamster ovary cells. The DNA-protein complexes were isolated by ultracentrifugal sedimentation in the presence of 2% sodium dodecyl sulfate (SDS) and 5 M urea. Two-dimensional SDS-polyacrylamide gel electrophoresis analysis of the chromate-induced DNA-protein complexes revealed that two acidic proteins of 53 and 45 kDa and a basic protein of 54 kDa were selectively complexed to the DNA. Numerous other proteins also became associated with the DNA to a lesser degree as the chromate concentration was increased. Nuclease digestion was not a prerequisite for the resolution of the protein component of the DNA-protein complexes using two-dimensional gel electrophoresis. Ultracentrifugal analysis of the DNA-protein complexes in the presence of proteinase K, nucleases, or a chelating agent demonstrated that protein aggregation was not responsible for the increased protein recovery in chromate-treated samples and that the complexes were disrupted by EDTA. These data suggest that the selectively complexed proteins were associated with the DNA through strong interactions that may be mediated by the trivalent form of chromium.

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Repair of chromate-induced DNA damage in chick embryo hepatocytes

Cited by 33 publications

References 0 publications

Modification of chromium(VI)-induced DNA damage by glutathione and cytochromes P-450 in chicken embryo hepatocytes.

Modification of chromium(VI)-induced DNA damage by glutathione and cytochromes P-450 in chicken embryo hepatocytes.

Evaluation of the genotoxic potential of glutaraldehyde

Characterization of DNA‐protein complexes induced in intact cells by the carcinogen chromate

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