Abstract
The involvement of reactive oxygen species in chromate-induced genotoxicity has been postulated. Because intracellular antioxidants help in eliminating the reactive species of oxygen, we have investigated both the prooxidant and antioxidant status of human leukemic T-lymphocyte MOLT4 cells exposed to nontoxic levels of chromium(VI) in culture. The cells treated with 0 →200 μM potassium chromate in a salts/glucose medium for 2 h were found to contain significantly lower levels of both small molecular weight and macromolecular antioxidants. In particular, the levels of glutathione and ascorbate were found to decrease with increased doses of chromate exposure in a dose-dependent manner. As little as 10 μM chromate was found to decrease these small molecular weight antioxidants significantly (p<0.01). The macromolecular antioxidants, such as glutathione peroxidase, catalase, glutathione reductase, glucose-6-phosphate dehydrogenase and superoxide dismutase were also significantly (p<0.01) decreased by exposing the cells to as little as 10 μM chromate. Concomitantly there was a dose-dependent increase in intracellular H2O2 accumulation in cells exposed to chromium(VI). These results indicate that chromate-induced genotoxicity may be due, at least in part, to decreased levels of intracellular antioxidants in conjunction with an increased production of the reactive oxygen species.
Similar content being viewed by others
References
IARC Monograph on the Evaluation of Carcinogenic Risk to Humans. Chromium, Nickel and Welding. Vol 49, Lyon, France, 1990
Doll R: Problems of Epidemiological Evidence. Env Health Perspect 40: 11–20, 1981
ATSDR: Toxicological profiles of chromium. Report # ATSDR/TP-88/10, 1989
Cohen MD, Latta DM, Coogan TP, Costa M: The reaction of metals with nucleic acids. In: E. Foulkes (ed.) Biological Effect of Heavy Metals. Vol. 2: Mechanism of metal carcinogenesis. CRC Press, Boca Raton, Florida, 1990, pp 19–77
Jennette KW: Chromate metabolism in liver microsomes. Biol Trace Element Res 1: 55–62, 1962
Connett PH, Watterhahn KE: Metabolism of carcinogen chromate by cellular constituents. Struct Bonding 54: 93–124, 1983
Cupo DY, Watterhahn KE: Modification of chromium(VI) induced DNA damage by glutathione and cytochrome P-450 in chicken embryo hepatocytes. Proc Natl Acad Sci, USA 82: 6755–6759, 1985
O'Brien P, Barrett J, Swanson F: Chromium(V) can be generated in the reduction of chromium (VI) by glutathione. J Inorg Chim Acta 108: 19–20, 1985
Kawanishi S, Inoue S, Sano S: Mechanism of DNA cleavage induced by sodium chromate (VI) in the presence of hydrogen peroxide. J Biol Chem 261: 5952–5958, 1986
Kitagawa S, Seki H, Kametani F, Sakurai H: ESR study on the interaction of hexavalent chromium with glutathione or cysteine: production of pentavalent chromium and its stability. J Inorg Chim Acta 152: 251–255, 1988
Sugiyama M, Ando A, Ogura R: Vitamin B2-enhancement of sodium chromate (VI)-induced DNA single strand breaks: ESR study of action of vitamin B2. Biochem Biophys Res Commun 159: 1080–1085, 1989
Gruber JC, Jennette KW: Metabolism of the carcinogen chromate by rat liver microsomes. Biochem Biophys Res Commun 82: 700–706, 1978
Shi X, Dalal NS: On the hydroxyl radical formation in the reaction between hydrogen peroxide and biologically generated chromium (V) species. Arch Biochem Biophys 277: 342–350, 1990
Jennette KW: Microsomal reduction of the carcinogenic chromate produces chromium (V). J Am Chem Soc 104: 874–875, 1982
De Flora S, Morelli A, Basso C, Romano M, Serra D, De Flora A: Prominent role of DT-diaphorases as a cellular mechanism reducing chromium (VI) and reverting its mutagenicity. Cancer Res 45: 3188–3196, 1985
Banks RB, Cooke RT, Jr.: Chromate reduction by rabbit liver aldehyde oxidase. Biochem Biophys Res Commun 137: 8–14, 1986
Taspakos MJ, Wetterhahn KE: The interaction of chromium with nucleic acids. Chem Biol Interactions 46: 265–277, 1983
Sugiyama M, Ando A, Nakao K, Ueta H, Hidaka T, Ogura R: Influence of vitamin B2 on formation of chromium (V), alkali-labile sites, and lethality of sodium chromate (VI) in Chinese Hamster V-79 cells. Cancer Res 49: 6180–6184, 1989
Sugiyama M: Role of physiological antioxidants in chromium(VI)-induced cellular injury. Free Radical Biol Med 12: 397–407, 1992
Tietze F: Enzymatic method for quantitative determination of nanogram amounts of total and oxidized glutathione. Anal Biochem 27: 502–522, 1969
Sies H and Akerboom TPM: Glutathione disulfide (GSSG) efflux from cell and tissues. In: Packer L (ed.) Methods in Enzymology, Vol 105. Academic Press Inc., New York, 1984, pp 445–451
Griffith OW: Determination of glutathione and gutathione disulfide using glutathione reductase and 2-Vinylpyridine. Anal Biochem 106: 207–212, 1980
Jagota SK, Dani HM: A new colorimetric technique for the estimation of vitamin C using Folin Phenol reagent. Anal Biochem 127: 178–182, 1982
Bass DA, Parce JW, Dechatelet LR, Szejda P, Seeds MC, Thomas M: Flow cytometric studies of oxidative product formation by neutrophils. J Immunol 130: 1910–1917, 1983
Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72: 248–254, 1976
Beers RF, Sizer IW: A spectrophotometric method for measuring the breakdown fo hydrogen peroxide by catalase. J Biol Chem 195: 133–140, 1952
Kornburg A, Horecker, BL: Glucose-6-phosphate dehydrogenase. In: Colowick SP, Kaplan NO (eds.) Methods in Enzymoloy, Vol 1. Academic Press Inc., New York, 1955, pp 323–327
Gamble PE, Burke JJ: Effect of water stress on the chloroplast antioxidant system. 1. Alterations in glutathione reductase activity. Plant Physiol 76: 615–621, 1984
Tappel AL: Glutathione peroxidase and hydroperoxides. In: Fleischer S, Packer L (eds.) Methods in Enzymology, Vol. LII. Academic Press, New York, 1978, pp 506–513
McCord JM, Fridovich I: Superoxide dismutase: An enzymatic function for erythrocuprein (hemocuprein). J Biol Chem 244: 6049–6055, 1969
Kaplan J: Electrophoretic study of glutathione reductase in human Erythrocytes and Leukocytes. Nature 217: 256–258, 1968
Niedzwicki A, Reveillaud I, Fleming JE: Changes in superoxide dismutase and catalase in aging heat-shocked drosophilla. Free Rad Res Comms 17: 355–367, 1992
Gustafson TL: Epistatistics software. Gustafson, Round Rock, Texas, 1984
Suzuki Y, Fukuda K: Reducation of hexavalent chromium by ascorbic acid and glutathione with special reference to rat lung. Arch Toxicol 64: 169–176, 1990
Chorvatovicova D, Ginter E, Kosinova A, Zolch Z: Effect of vitamin C and E on toxicity and mutagenicity of hexavalent chromium in rat and guinea pig. Mutat Res 262: 41–46, 1991
Long WK: Glutathione reductase in red blood cells: Variant associated with gout. Science 155: 712–713, 1967
Standeven AM: Watterhanh KE: Possible role of glutathione in chromium (VI) metabolism and cytotoxicity in rats. Pharmacol Toxicol 68: 469–476, 1991
Sugiyama M, Ando A, Furuno H, Furlong NB, Hidaka T, Ogura, R: Effects of vitamin E, vitamin B2 and selenite on DNA single strand breaks induced by sodium chromate (VI). Cancer Lett 38: 1–7, 1987
Sugiyama M, Ando A, Ogura R: Effect of vitamin E on survival, glutathione reductase and formation of chromium(V) in Chinese Hamster V-79 cells treated with sodium chromate(VI). Carcinogenesis 10: 737–741, 1989
Sugiyama M: Effects of vitamin E and vitamin B2 on chromate-induced DNA lesions. Biol Trace Elem Res 21: 399–404, 1989
Samitz MH, Katz SA: Protection against inhalation of chromic acid mist. Use of filters impregnated with ascorbic acid. Arch Env Health 11: 770–773, 1965
Susa N, Ueno S, Furukawa Y, Michiba N, Minoura S: Induction of lipid peroxidation in mice by hexavalent chromium and its relation to the toxicity. Jpn J Vet Sci 51: 1103–1110, 1989
Ueno S, Susa N, Furukawa Y, Akikawa K, Itagaki I: Cellular injury and lipid peroxidation induced by hexavalent chromium in isolated rat hepatocytes. Jpn J Vet Sci 51: 137–145, 1989
Korralus U, Harzdorf C, Lewalter J: Experimental bases for ascorbic acid therapy of poisoning by hexavalent chromium compounds. Ind Arch Occup Environ Health 53: 247–256, 1984
Sugiyama M, Tsuzuki K, Ogura R: Effect of ascorbic acid on DNA damage, cytotoxicity, glutathione reductase, and formation of paramagnetic chromium in Chinese hamster V-79 cells treated with sodium chromate (VI). J Biol Chem 266: 3383–3386, 1991
Sugiyama M, Tsuzuki K, Haramaki N: DNA single-strand breaks and cytotoxicity induced by sodium chromate(VI) in hydrogen peroxideresistant cell lines. Mutat Res 299: 95–102, 1993
Shi X, Dalal NS: NADPH-dependent flavoenzymes catalyze one electron reduction of metal ions and molecular oxygen and generato hydroxyl radicals. FEBS 276: 189–191, 1990
Aiyar J, Berkovits JH, Floyd RA, Wetterhahn KE: Reaction of cromium(VI) with hydrogen peroxide in the presence of glutathione: Reactive intermediates and resulting DNA damage. Chem Res Toxicol 3: 595–603, 1990
Shi X, Dalal NS: Evidence for a Fenton-Type mechanism for the generation of OH radicals in the reduction of Cr(VI) in cellular media. Arch Biochem Biophys 281: 90–95, 1990
Ketterer B: Protective role of glutathione and glutathione transferases in mutagenesis and carcinogenesis. Mutat Res 202: 323–361, 1988
Shi X, Dalal NS: The mechanisms of the chromate reduction by glutathione: ESR evidence for the glutathionyl radical and an isolable Cr(V) intermediate. Biochem Biophys Res Commun 156: 137–142, 1988
Misra HP: Generation of superoxide free radical during the autoxidation of thiols. J Biol Chem 249: 2151–2155, 1974
Shi X, Dalal NS: Chromium(V) and hydroxyl radical formation during the glutathione-reductase catalyzed reduction of chromium(VI). Biochem Biophys Res Commun 163: 627–634, 1989
Horning D: Distribution of ascorbic acid, metabolism and analogus in man and animals. Ann N Y Acad Sci 258: 103–118, 1975
Sengupta T, Chattopadhyay D, Ghosh N, Maulik G, Chatterjee GC: Impact of chromium on lipoperoxidative processes and subsequent operation of the glutathione cycle in rat renal system. Indian J Biochem Biophys 29: 287–290, 1992
Koutras GA, Schneider AS, Hattori M, Valentine WN: Studies on chromated erythrocytes. Mechanism of chromate inhibition of glutathione reductase. Brit J Haematol 2: 360–369, 1965
Sengupta T, Chattopadhyay D, Ghosh N, Das M, Chatterjee, GC: Effect of chromium administration on glutathione cycle of rat intestinal epithelial cells. Indian J Exp Biol 28: 1132–1135, 1990
Scott JA, Fischman AJ, Homcy CJ, Fallon JT, Khaw BA, Peto CA, Rabito CA: Morphologic and functional correlates of plasma membrane injury during oxidant exposure. Free Radic Biol Med 6: 361–367, 1989
Tirmenstein MA, Reed DJ: Characterization of glutathione-dependent inhibition of lipid peroxidation of isolated rat liver nuclei. Arch Biochem Biophys 261: 1–11, 1988
Ursini F, Maiorino M, Hochstein P, Ernster L: Microsomal lipid peroxidation: mechanism of initiation: The role of iron and iron chelators. Free Radic Biol Med 6: 31–36, 1989
Binodoli A: Lipid peroxidation in mitochondria. Free Radic Biol Med 5: 247–261, 1988
Dizdaroglu M, Bergtold DS: Characterization of free radical induced base damage in DNA at biologically relevant levels. Anal Biochem 156: 182–188, 1986
Cross CE, Halliwell B, Borish ET, Pryor WA, Ames BA, Saul RA, McChord JM, Harman D: Oxygen radicals and human disease. Ann Int Med 107: 526–545, 1987
Ames BN: Endogenous oxidative DNA damage, aging, and cancer. Free Radic Res Comms 7: 121–128, 1989
Cerruti PA: Prooxidant states and tumor promotion. Science 227: 375–381, 1985
Patierno SR, Bahn D, Landolph JR: Transformation of C3H/10T1/2 mouse embryo cells to focus formation and anchorage independence by insoluble lead chromate but not soluble calcium chromate: relationship to mutagenesis and internalization of lead chromate particles. Cancer Res 48: 5280–5288, 1988
Tursh MA, Kensler TW: An overview of the relationship between oxidative stress and chemical carcinogenesis. Free Radical Biol Med 10: 201–209, 1991
Faux SP, Gao M, Chipman JK, Levy LS: Production of 8-hydroxydeoxyguanosine in isolated DNA by chromium(VI) and chromium(V). Carcinogenesis 13: 1667–1669, 1992
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Mattagajasingh, S.N., Misra, H.P. Alterations in the prooxidant and antioxidant status of human leukemic T-lymphocyte MOLT4 cells treated with potassium chromate. Mol Cell Biochem 142, 61–70 (1995). https://doi.org/10.1007/BF00928914
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF00928914