Analytical and Bioanalytical Chemistry

, Volume 403, Issue 2, pp 431–441 | Cite as

Glutathione peroxidase inhibitory assay for electrophilic pollutants in diesel exhaust and tobacco smoke

  • Norbert Staimer
  • Tran B. Nguyen
  • Sergey A. Nizkorodov
  • Ralph J. Delfino
Original Paper

Abstract

We developed a rapid kinetic bioassay demonstrating the inhibition of glutathione peroxidase 1 (GPx-1) by organic electrophilic pollutants, such as acrolein, crotonaldehyde, and p-benzoquinone, that are frequently found as components of tobacco smoke, diesel exhaust, and other combustion sources. In a complementary approach, we applied a high-resolution proton-transfer reaction time-of-flight mass spectrometer to monitor in real-time the generation of electrophilic volatile carbonyls in cigarette smoke. The new bioassay uses the important antioxidant selenoenzyme GPx-1, immobilized to 96-well microtiter plates, as a probe. The selenocysteine bearing subunits of the enzyme’s catalytic site are viewed as cysteine analogues and are vulnerable to electrophilic attack by compounds with conjugated carbonyl systems. The immobilization of GPx-1 to microtiter plate wells enabled facile removal of excess reactive inhibitory compounds after incubation with electrophilic chemicals or aqueous extracts of air samples derived from different sources. The inhibitory response of cigarette smoke and diesel exhaust particle extracts were compared with chemical standards of a group of electrophilic carbonyls and the arylating p-benzoquinone. GPx-1 activity was directly inactivated by millimolar concentrations of highly reactive electrophilic chemicals (including acrolein, glyoxal, methylglyoxal, and p-benzoquinone) and extracts of diesel and cigarette smoke. We conclude that the potential of air pollutant components to generate oxidative stress may be, in part, a result of electrophile-derived covalent modifications of enzymes involved in the cytosolic antioxidant defense.

Figure

Cu/Zn superoxide dismutase (SOD-1) and glutathione peroxidase (GPx) are linked together in the cytosolic defense against reactive oxygen and nitrogen species (RONS). Cu/Zn-SOD catalyzes the dismutation of superoxide to oxygen and hydrogen peroxide (H2O2). H2O2 and other hydroperoxides are subsequently reduced by the selenoenzyme GPx. The selenofunction is viewed as a cysteine analogue, and in comparison to other thiol enzymes, is even more vulnerable to electrophilic attack by chemicals such as acrolein at physiological conditions. Cu/Zn-SOD and GPx team up with a complex cellular antioxidant system that includes catalase, glutathione transferase and reduced glutathione (not shown). Environmental exposure to reactive electrophiles present in cigarette smoke and diesel exhaust emissions may add to the endogenous burden of oxidative stress by direct inactivation of GPx

Keywords

Air pollution Electrophiles Antioxidant enzymes Oxidative stress 

Notes

Acknowledgments

This work was supported by the US Environmental Protection Agency (USEPA) STAR Grant No. RD83241301, National Institute of Environmental Health Sciences (grant no. R01 ES12243), and UCI Multi-Investigator Faculty Research Grant MI 7 2008–2009. We are grateful to Professor Arthur Cho, University of California, Los Angeles, for useful comments after reading this manuscript, helpful discussions, and for the kind gift of DEP samples.

References

  1. 1.
    Delfino RJ, Staimer N, Vaziri ND (2011) Air pollution and circulating biomarkers of oxidative stress. Air Qual Atmos Health 4:37–52CrossRefGoogle Scholar
  2. 2.
    Kregel KC, Zhang HJ (2007) An integrated view of oxidative stress in aging: basic mechanisms, functional effects, and pathological considerations. Am J Physiol Regul Integr Comp Physiol 292(1):R18–R36CrossRefGoogle Scholar
  3. 3.
    Mak JC, Chan-Yeung MM (2006) Reactive oxidant species in asthma. Curr Opin Pulm Med 12(1):7–11CrossRefGoogle Scholar
  4. 4.
    Dhalla NS, Temsah RM, Netticadan T (2000) Role of oxidative stress in cardiovascular diseases. J Hypertens 18(6):655–673CrossRefGoogle Scholar
  5. 5.
    Pai JK, Pischon T, Ma J, Manson JE, Hankinson SE, Joshipura K et al (2004) Inflammatory markers and the risk of coronary heart disease in men and women. N Engl J Med 351:2599–2610CrossRefGoogle Scholar
  6. 6.
    Vaziri ND, Rodríguez-Iturbe B (2006) Mechanisms of disease:oxidative stress and inflammation in the pathogenesis of hypertension. Nat Clin Pract Nephrol 2:582–593CrossRefGoogle Scholar
  7. 7.
    Ayres JG, Borm P, Cassee FR et al (2008) Evaluating the toxicity of airborne particulate matter and nanoparticles by measuring oxidative stress potential—a workshop report and consensus statement. Inhal Toxicol 20:75–99CrossRefGoogle Scholar
  8. 8.
    Li N, Hao M, Phalen RF, Hinds WC, Nel AE (2003) Particulate air pollutants and asthma. A paradigm for the role of oxidative stress in PM-induced adverse health effects. Clin Immunol 109:250–265CrossRefGoogle Scholar
  9. 9.
    Mills NL, Törnqvist H, Robinson SD, Gonzalez MC, Söderberg S, Sandström T, Blomberg A, Newby DE, Donaldson K (2007) Air pollution and atherothrombosis. Inhal Toxicol 19(Suppl 1):81–89, ReviewCrossRefGoogle Scholar
  10. 10.
    Pandya RJ, Solomon G, Kinner A, Balmes JR (2002) Diesel exhaust and asthma: hypotheses and molecular mechanisms of action. Environ Heal Perspect 110(Supp (1)):103–112CrossRefGoogle Scholar
  11. 11.
    Kode A, Yang S-R, Rahman I (2006) Differential effects of cigarette smoke on oxidative stress and proinflammatory cytokine release in primary human airway epithelial cells and in a variety of transformed alveolar epithelial cells. Respir Res 7:132CrossRefGoogle Scholar
  12. 12.
    Lannan S, Donaldson K, Brown D, MacNee W (1994) Effect of cigarette smoke and its condensates on alveolar epithelial cell injury in vitro. Am J Physiol 266:L92–L100Google Scholar
  13. 13.
    Oppermann U (2007) Carbonyl Reductases: the complex relationships of mammalian carbonyl- and quinone-reducing enzymes and their role in physiology. Annu Rev Pharmacol Toxicol 47:293–322CrossRefGoogle Scholar
  14. 14.
    Wooten JB, Chouchane S, McGrath TE (2006) Tobacco smoke constituents affecting oxidative stress. In: Halliwell BB, Poulsen HE (eds) Cigarette smoke and oxidative stress. Springer, BerlinGoogle Scholar
  15. 15.
    Wang H-T, Zhang S, Hu Y, M-s T (2009) Mutagenicity and sequence specificity of acrolein-DNA adducts. Chem Res Toxicol 22:511–517CrossRefGoogle Scholar
  16. 16.
    Zhang S, Villalta PW, Wang M, Hecht SS (2007) Detection and quantitation of acrolein-derived 1, N2-propanodeoxyguanosine adducts in human lung by liquid chromatography-electrospray ionization-tandem mass spectrometry. Chem Res Toxicol 20:565–571CrossRefGoogle Scholar
  17. 17.
    Beauchamp RO Jr, Andjelkovich DA, Kligerman AD, Morgan KT, Heck Hd’A (1985) A critical review of the literature on acrolein toxicity. Crit Rev Toxicol 14(4):309–380CrossRefGoogle Scholar
  18. 18.
    Fujioka K, Shibamoto T (2006) Determination of toxic carbonyl compounds in cigarette smoke. Environ Toxicol 21:47–54CrossRefGoogle Scholar
  19. 19.
    Bhatnagar A (2004) Cardiovascular pathophysiology of environmental pollutants. Am J Physiol Circ Physiol 286(2):H479–H485CrossRefGoogle Scholar
  20. 20.
    Kroll JH, Ng NL, Murphy SM, Varutbangkul V, Flagan RC, Seinfeld JH (2005) Chamber studies of secondary organic aerosol growth by reactive uptake of simple carbonyl compounds. J Geophys Res D 110:D23207/1CrossRefGoogle Scholar
  21. 21.
    Nguyen TB, Batemana AP, Bones DL, Nizkorodov SA, Laskin J, Laskin A (2010) High-resolution mass spectrometry analysis of secondary organic aerosol generated by ozonolysis of isoprene. Atmos Environ 44:1032–1042CrossRefGoogle Scholar
  22. 22.
    Shinyashiki M, Rodriguez CR, Di Stefano EM et al (2008) On the interaction between glyceraldehyde-3-phosphate dehydrogenase and airborne particles: evidence for electrophilic species. Atmos Environ 42:517–529CrossRefGoogle Scholar
  23. 23.
    Delfino RJ, Staimer N, Tjoa T, Polidori A, Arhami M, Gillen D, Kleinman MT, Vaziri N, Longhurst J, Zaldivar F, Sioutas C (2008) Circulating biomarkers of inflammation, antioxidant activity, and platelet activation are associated with primary combustion aerosols in subjects with coronary artery disease. Env Health Perspect 116:898–906CrossRefGoogle Scholar
  24. 24.
    Delfino RJ, Staimer N, Tjoa T, Gillen D, Polidori A, Arhami M, Kleinman MT, Vaziri N, Longhurst J, Sioutas C (2009) Air pollution exposures and circulating biomarkers of effect in a susceptible population: clues to potential causal component mixtures and mechanisms. Environ Health Perspect 117:1232–1238CrossRefGoogle Scholar
  25. 25.
    Carp H, Janoff A (1978) Possible mechanisms of emphysema in smokers. In vitro expression of serum elastase-inhibitory capacity by fresh cigarette smoke and its prevention by antioxidants. Am Rev Respir Dis 118:617–621Google Scholar
  26. 26.
    Church DF, Pryor WA (1985) Free radical chemistry of cigarette smoke and its toxicological implications. Environ Health Perspect 64:111–126CrossRefGoogle Scholar
  27. 27.
    Ritter D, Knebel J, Aufderheide M (2004) Comparative assessment of toxicities of mainstream smoke from commercial cigarettes. Inhal Toxicol 16:691–700CrossRefGoogle Scholar
  28. 28.
    Carmella SG, Chen M, Zhang Y, Zhang S et al (2007) Quantitation of acrolein-derived 3-hydroxypropylmercapturic acid in human urine by liquid chromatographyatmospheric pressure chemical ionization-tandem mass spectrometry: effects of cigarette smoking. Chem Res Toxicol 20:986–990CrossRefGoogle Scholar
  29. 29.
    Kehrer JP, Biswal SS (2000) The molecular effects of acrolein. Toxicol Sci 57(1):6–15CrossRefGoogle Scholar
  30. 30.
    Geiser M, Rothen-Rutishauser B, Kapp N, Schurch S, Kreyling W, Schulz H, Semmler M, Im HV, Heyder J, Gehr P (2005) Ultrafine particles cross cellular membranes by nonphagocytic mechanisms in lungs and in cultured cells. Environ Health Perspect 113:1555–1560CrossRefGoogle Scholar
  31. 31.
    Meacher DM, Menzel DB (1999) Glutathione depletion in lung cells by low-molecular-weight aldehydes. Cell Biol Toxicol 15:163–171CrossRefGoogle Scholar
  32. 32.
    Avissar N, Finkelstein JN, Horowitz S, Willey JC, Coy E, Frampton MW, Watkins RH, Khullar P, Xu Y-L, Cohen HJ (1996) Extracellular glutathione peroxidase in human lung epithelial lining fluid and in lung cells. Am J Physiol 270:L173–L182Google Scholar
  33. 33.
    Papp LV, Lu J, Holmgren A, Khanna KK (2007) From selenium to selenoproteins: synthesis, identity, and their role in human health. Antioxid Redox Signal 9(7):775–806CrossRefGoogle Scholar
  34. 34.
    Uchida K, Kanematsu M, Morimitsu Y, Osawa T, Noguchi N, Niki E (1998) Acrolein is a product of lipid peroxidation reaction. Formation of free acrolein and its conjugate with lysine residues in oxidized low density lipoproteins. J Biol Chem 273(26):16058–16066CrossRefGoogle Scholar
  35. 35.
    Uchida K (1999) Current status of acrolein as a lipid peroxidation product. Trends Cardiovasc Med 9(5):109–113CrossRefGoogle Scholar
  36. 36.
    Liebler DC (2008) Protein damage by reactive electrophiles: targets and consequences. Chem Res Toxicol 21:117–128CrossRefGoogle Scholar
  37. 37.
    Jordan A, Haidacher S, Hanel G, Hartungen E, Herbig J, Maerk L, Schottkowsky R, Seehauser H, Sulzer P, Maerk TD (2009) An online ultra-high sensitivity proton-transfer-reaction mass-spectrometer combined with switchable reagent ion capability (PTR+SRI-MS). Int J Mass Spectrom 286(1):32–38CrossRefGoogle Scholar
  38. 38.
    Shinyashiki M, Eiguren-Fernandez A, Schmitz DA et al (2009) Electrophilic and redox properties of diesel exhaust particles. Environ Res 109:239–244CrossRefGoogle Scholar
  39. 39.
    Paglia DE, Valentine WN (1967) Studies on the quantitative and qualitative characterization of erythrocyte glutathione peroxidase. J Lab Clin Med 70:158–169Google Scholar
  40. 40.
    Spahn C, Minteer SD (2008) Enzyme immobilization in biotechnology. Recent Patents Eng 2:195–200CrossRefGoogle Scholar
  41. 41.
    Chen PX, Moldoveanu SC (2003) Mainstream smoke chemical analyses for 2R4F Kentucky reference cigarette. Beiträge zur Tabakforschung Int 20(7):448–458Google Scholar
  42. 42.
    Hoffmann D, Hecht SS (1990) Chapter 3: advances in tobacco carcinogenesis. In: Cooper DS, Grover P (eds) Chemical carcinogenesis and mutagenesis. Springer, LondonGoogle Scholar
  43. 43.
    Eiserich JP, van der Vliet A, Handelman GJ, Halliwell B, Cross CE (1995) Dietary antioxidants and cigarette smoke-induced biomolecular damage: a complex interaction. Am J Clin Nutr 62(suppl 6):1490S–1500SGoogle Scholar
  44. 44.
    Wang X, Thomas B, Sachdeva R, Arterburn L, Frye L, Hatcher PG, Cornwell DG, Ma J (2006) Mechanism of arylating quinone toxicity involving Michael adduct formation and induction of endoplasmic reticulum stress. PNAS 103(10):3604–3609CrossRefGoogle Scholar
  45. 45.
    Rappaport SM, Waidyanatha S, Qu Q et al (2002) Albumin adducts of benzene oxide and 1,4-benzoquinone as measures of human benzene metabolism. Cancer Res 62:1330–1337Google Scholar
  46. 46.
    Dey N, Das A, Ghosh A, Chatterjee IB (2010) Activated charcoal filter effectively reduces p-benzosemiquinone from the mainstream cigarette smoke and prevents emphysema. J Biosci 35(2):217–230CrossRefGoogle Scholar
  47. 47.
    Temime B, Healy RM, Wenger JC (2007) A denuder-filter sampling technique for the detection of gas and particle phase carbonyl compounds. Environ Sci Technol 41:6514–6520CrossRefGoogle Scholar
  48. 48.
    Alt C, Eyer P (1998) Ring addition of the alpha-amino group of glutathione increases the reactivity of benzoquinone thioethers. Chem Res Toxicol 11:1223–1233CrossRefGoogle Scholar
  49. 49.
    Enoch SJ, Cronin MTD (2010) A review of the electrophilic reaction chemistry involved in covalent DNA binding. Crit Rev Toxicol 40(8):728–748CrossRefGoogle Scholar
  50. 50.
    Park YS, KohYH TM et al (2003) Identification of the binding site of methylglyoxal on glutathione peroxidase: methylglyoxal inhibits glutathione peroxidase activity via binding to glutathione binding sites Arg 184 and 185. Free Radic Res 37(2):205–211CrossRefGoogle Scholar
  51. 51.
    Lo TW, Westwood ME, McLellan AC et al (1994) Binding and modification of proteins by methylglyoxal under physiological conditions. A kinetic and mechanistic study with N alpha-acetylarginine, N alpha-acetylcysteine, and N alpha-acetyllysine, and bovine serum albumin. J Biol Chem 269(51):32299–32305Google Scholar
  52. 52.
    Shangari N, Bruce WR, Poon R, O’Brien PJ (2003) Toxicity of glyoxals—role of oxidative stress, metabolic detoxification and thiamine deficiency. Biochem Soc Trans 31(Pt 6):1390–1393CrossRefGoogle Scholar
  53. 53.
    Takahashi M, Okamiya H, Furukawa F et al (1989) Effects of glyoxal and methylglyoxal administration on gastric carcinogenesis in Wistar rats after initiation with N-methyl-N′-nitro-N-nitrosoguanidine. Carcinogenesis 10(10):1925–1927CrossRefGoogle Scholar
  54. 54.
    Knighton WB, Herndon SC, Shorter JH, Miake-Lye RC, Zahniser MS, Akiyama K, Shimono A, Kitasaka K, Shimajiri H, Sugihara K (2007) Laboratory evaluation of an aldehyde scrubber system specifically for the detection of acrolein. J Air Waste Manage Assoc 57:1370–1378CrossRefGoogle Scholar
  55. 55.
    Graus M, Müller M, Hansel A (2010) High resolution PTR-TOF quantification and formula confirmation of VOC in real time. J Am Soc Mass Spectrom 21:1037–1044CrossRefGoogle Scholar
  56. 56.
    See SW, Wang YH, Balasubramanian R (2007) Contrasting reactive oxygen species and transition metal concentrations in combustion aerosols. Environ Res 103:317–324CrossRefGoogle Scholar
  57. 57.
    Rafter GW (1982) Copper inhibition of glutathione reductase and its reversal with gold thiolates, thiol, and disulfide compounds. Biochem Med 27:381–391CrossRefGoogle Scholar
  58. 58.
    Fenner ML, Braven J (1968) The mechanisms of carcinogenesis by tobacco smoke. Further experimental evidence and a prediction from the thiol-defence hypothesis. Br J Cancer 22:474–479CrossRefGoogle Scholar
  59. 59.
    Leuchtenberger C, Leuchtenberger R, Zbinden I (1974) Gas vapour phase constituents and SH reactivity of cigarette smoke influence lung cultures. Nature 247:565–567CrossRefGoogle Scholar
  60. 60.
    Leuchtenberger C, Leuchtenberger R, Zbinden I, Schleh E (1976) SH reactivity of cigarette smoke and its correlation with carcinogenic effects on hamster lung cells. Soz Praev Med 21:47–50CrossRefGoogle Scholar
  61. 61.
    Stauffer HP (1974) The interaction of cigarette smoke with thiol groups, a model study. Soz Praev Med 19:55–58CrossRefGoogle Scholar
  62. 62.
    Mueller T, Haussmann H-J, Schepers G (1997) Evidence for peroxynitrite as an oxidative stress-inducing compound of aqueous cigarette smoke fractions. Carcinogenesis 18(2):295–301CrossRefGoogle Scholar
  63. 63.
    Asahi M, Fujii J, Takao T et al (1997) The oxidation of selenocysteine is involved in the inactivation of glutathione peroxidase by nitric oxide donor. J Biol Chem 272(31):19152–19157CrossRefGoogle Scholar
  64. 64.
    Fujii J, Taniguchi N (1999) Down regulation of superoxide dismutases and glutathione peroxidase by reactive oxygen and nitrogen species. Free Radic Res 31:301–308CrossRefGoogle Scholar
  65. 65.
    Klein I, Nagler RM, Toffler R, van DER Vliet A, Reznick A (2003) Effect of cigarette smoke on oral peroxidase activity in human saliva: role of hydrogen cyanide. Free Radic Biol Med 35(11):1448–1452CrossRefGoogle Scholar
  66. 66.
    Kraus RJ, Prohaska JR, Ganther HE (1980) Oxidized forms of ovine erythrocyte glutathione peroxidase. Cyanide inhibition of a 4-glutathione:4-selenoenzyme. Biochim Biophys Acta 615(1):19–26Google Scholar
  67. 67.
    Kraus RJ, Ganther HE (1980) Reaction of cyanide with glutathione peroxidase. Biochem Biophys Res Commun 96(3):1116–1122CrossRefGoogle Scholar
  68. 68.
    Rickert WS, Stockwell PB (1979) Automated determination of hydrogen cyanide acrolein and total aldehydes in the gas phase of tobacco smoke. J Autom Chem 1(3):152–154CrossRefGoogle Scholar
  69. 69.
    Ganther HE (1999) Selenium metabolism, selenoproteins and mechanisms of cancer prevention: complexities with thioredoxin reductase. Carcinogenesis 20(9):1657–1666CrossRefGoogle Scholar
  70. 70.
    Rahman I, MacNee W (1996) Role of oxidants/antioxidants in smoking-induced lung diseases. Free Radic Biol Med 21(5):669–681CrossRefGoogle Scholar
  71. 71.
    Miyamoto Y, Koh YH, Park YS et al (2003) Oxidative stress caused by inactivation of glutathione peroxidase and adaptive responses. Biol Chem 384(4):567–574Google Scholar
  72. 72.
    Boss G, Sharama V, Broderick KE (2008) Methods and compositions for treatment of excess nitric oxide or cyanide toxicity. US Patent 2008 (Pub. NO US 2008/0227746 A1)Google Scholar

Copyright information

© Springer-Verlag 2012

Authors and Affiliations

  • Norbert Staimer
    • 1
  • Tran B. Nguyen
    • 2
  • Sergey A. Nizkorodov
    • 2
  • Ralph J. Delfino
    • 1
  1. 1.Department of Epidemiology, School of MedicineUniversity of CaliforniaIrvineUSA
  2. 2.Department of ChemistryUniversity of CaliforniaIrvineUSA

Personalised recommendations