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Influence of quercetin and rutin on growth and antioxidant defense system of a human hepatoma cell line (HepG2)

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Summary

Background

Dietary polyphenols like quercetin and rutin are considered beneficial because of their potential protective role in the pathogenesis of multiple diseases associated to oxidative stress such as cancer, coronary heart disease and atherosclerosis. However, many of these effects may depend on the concentration of the polyphenol utilized since high doses of some phenolic compounds may be prooxidant and negatively affect cell growth and viability.

Aim of the study

To test the potential chemoprotective effects of quercetin and rutin, two flavonols with high antioxidant capacity, on cell growth, viability and the response of the antioxidant defense system of a human hepatoma cell line (HepG2).

Methods

Cell growth was measured by diaminobenzoic acid and bromodeoxyuridine assays, cell toxicity by lactate dehydrogenase leakage assay, reduced glutathione was quantified by a fluorimetric assay, cellular malondialdehyde was analyzed by high–performance liquid chromatography, reactive oxygen species were quantified by the dichlorofluorescein assay, antioxidant enzyme activities were determined by spectrophotometric analysis and their gene expression by northern blot.

Results

Short-term exposure (4 h) to these flavonols had no antiproliferative nor cytotoxic effect. High doses of quercetin (50–100 µM) increased glutathione concentration and gene expression of Cu/Zn superoxide dismutase and catalase inhibiting the activity of the latter enzyme, whereas lower doses (0.1–1 µM) decreased gene expression of Cu/Zn superoxide dismutase and increased that of glutathione peroxidase. All doses of quercetin and rutin diminished reactive oxygen species and high doses (10–100 µM) decreased malondialdehyde concentration.

Conclusion

The results indicate that both natural antioxidants induce favorable changes in the antioxidant defense system of cultured HepG2 that prevent or delay conditions which favor cellular oxidative stress.

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References

  1. Bravo L (1998) Polyphenols: chemistry, dietary sources, metabolism and nutritional significance. Nutr Rev 56:317–333

    CAS  Google Scholar 

  2. Hertog MGL, Holland PCH (1996) Potential health effects of the dietary flavonol quercetin. Eur J Clin Nutr 50:63–71

    CAS  Google Scholar 

  3. Aherne SA, O’Brien NM (1999) The flavonoids, myricetin, quercetin and rutin, protect against cholestan-3β:5α:6β-triol – induced toxicity in Chinese hamster ovary cells in vitro. Nutr Res 19:749–760

    Article  CAS  Google Scholar 

  4. Middelton E (1996) Biological properties of plant flavonoids: an overview. Int J Pharmacol 34:344–348

    Google Scholar 

  5. Mora A, Paya M, Rios JL, Alcaraz MJ (1990) Structure activity relationship of polymethoxyflavones and other flavonoids as inhibitors of non-enzymatic lipid peroxidation. Biochem Pharmacol 40:793–797

    Article  CAS  Google Scholar 

  6. Hertog MGL, Kromhout D, Aravanis C, Blackburn H, Buzina R, Fidanza F, Giampaoli S, Jansen A, Menotti A, Nedeljkovic S, Pekkarinen M, Simic BS, Toshima H, Feskens EJM, Hollman PCH, Katan MB (1995) Flavonoid intake and long term risk of coronary heart disease and cancer in the seven countries study. Arch Intern Med 155:381–386

    Article  CAS  Google Scholar 

  7. Cook NC, Samman S (1996) Flavonoids – chemistry, metabolism, cardioprotective effects and dietary sources. J Nutr Biochem 7:66–76

    CAS  Google Scholar 

  8. Lapidot T, Walker MD, Kanner J (2002) Can apple antioxidants inhibit tumor cell proliferation? Generation of H2O2 during interaction of phenolic compounds with cell culture media. J Agric Food Chem 50:3156–3160

    CAS  Google Scholar 

  9. Narayanan VS, Fitch CA, Levenson CW (2001) Tumor suppressor protein p53 mRNA and subcellular localization are altered by changes in cellular copper in human Hep G2 cells. J Nutr 131: 1427–1432

    CAS  Google Scholar 

  10. Aden DP, Fogel A, Plotkin S, Damjanov I, Knowles BB (1979) Controlled synthesis of HBsAg in a differentiated human liver carcinoma-derived cell line. Nature 286:615–616

    Google Scholar 

  11. Hinegardner RT (1971) An improved fluorometric assay for DNA. Anal Biochem 39:197–201

    Article  CAS  Google Scholar 

  12. Vasault A (1987) Lactate dehydrogenase. UV-method with pyruvate and NADH. In: Bergmeyer HV (ed) Methods of enzymatic analysis. Weinheim: Verlag-Chemie, pp 118–133

  13. Welder AA, Acosta D (1994) Enzyme leakage as an indicator of cytotoxicity in culture cells. In: Tyson CA, Franzier JM (eds) In vitro toxicity indicators: methods in toxicology. New York: Academic press, pp 46–49

    Google Scholar 

  14. Hissin PJ, Hilf R (1976) A fluorometric method for determination of oxidised and reduced glutathione in tissues. Anal Biochem 74:214–226

    Article  CAS  Google Scholar 

  15. Mateos R, Goya L, Bravo L (2004) Determination of malondialdehyde (MDA) by high-performance liquid chromatography as the 2,4-dinitrophenylhydrazine derivative. A marker for oxidative stress in cell cultures of human hepatoma HepG2. J Chrom B 805:33–39

    CAS  Google Scholar 

  16. Wang H, Joseph JA (1999) Quantifying cellular oxidative stress by dichlorofluorescein assay using microplate reader. Free Rad Biol Med 27:612–616

    CAS  Google Scholar 

  17. Aebi H (1987) Catalase. In: Bergmeyer HV (ed) Methods of enzymatic analysis. Weinheim: Verlag-Chemie, pp 273–282

  18. Goldberg DM, Spooner RJ (1987) Glutathione reductase. In: Bergmeyer HV (ed) Methods of enzymatic analysis. Weinheim: Verlag-Chemie, pp 258–265

  19. Gunzler WA, Kramers H, Flohe L (1974) An improved coupled test procedure for glutathione peroxidase. Klin Chem Klin Biochem 12:444

    CAS  Google Scholar 

  20. Bradford MM (1976) A rapid and sensitive method for the quantification of microgram quantities of protein, utilizing the principle of protein-dye binding. Anal Biochem 72:248

    Article  CAS  Google Scholar 

  21. Lipkin M, Reddy B, Newmark H, Lamprecht SA (1999) Dietary factors in human colorectal cancer. Annu Rev Nutr 19:545–586

    Article  CAS  Google Scholar 

  22. Duthie GG, Duthie SJ, Kyle JAM (2000) Plant polyphenols in cancer and heart disease: implications as nutritional antioxidants. Nutr Res Rev 13:79–106

    Article  CAS  Google Scholar 

  23. Agullo G, Gamet L, Besson C, Demigne C, Remesy C (1994) Quercetin exerts a preferential cytotoxic effect on active dividing colon carcinoma HT29 and Caco-2 cells. Cancer Lett 87:55–63

    Article  CAS  Google Scholar 

  24. Ranelletti FO, Ricci R, Larocca LM, Maggiano N, Capelli A, Scambia G, Benedetti-Panici P, Mancuso S, Rumi C, Piantelli M (1992) Growth-inhibitory effect of quercetin and presence of type-II estrogen-binding sites in human colon-cancer cell lines and primary colorectal tumors. Int J Cancer 50:486–492

    CAS  Google Scholar 

  25. Richter M, Ebermann R, Marian B (1999) Quercetin-induced apoptosis in colorectal tumor cells: possible role of EGF receptor signaling. Nutr Cancer 34:88–99

    CAS  Google Scholar 

  26. Singhal RL, Yelt YA, Praja N, Olah E, Sledge GW, Weber G (1995) Quercetin down-regulates signal transduction in human breast carcinoma cells. Biochem Biophys Res Commun 208:425–431

    Article  CAS  Google Scholar 

  27. Duthie SJ, Johnson W, Dobson VL (1997) The effect of dietary flavonoids on DNA damage (strand breaks and oxidized pyrimidines) and growth in human cells. Mutat Res 390:141–151

    CAS  Google Scholar 

  28. Noroozi M, Angerson WJ, Lean MEJ (1998) Effects of flavonoids and vitamin C on oxidative DNA damage to human lymphocytes. Am J Clin Nutr 67:1210–1218

    CAS  Google Scholar 

  29. Scambia G, Ranelletti FO, Panici PB, Devincenzo R, Bonanno G, Ferrandina G, Piantelli M, Bussa S, Rumi C, Cianfriglia M, Mancuso S (1994) Quercetin potentiates the effect of adriamycin in a multidrug-resistant mcf-7 human breast-cancer cell-line–p-glycoprotein as a possible target. Cancer Chem Pharmacol 34:459–464

    CAS  Google Scholar 

  30. Kang TB, Liang NC (1997) Studies on the inhibitory effects of quercetin on the growth of HL-60 leukemia cells. Biochem Pharmacol 54:1013–1018

    Article  CAS  Google Scholar 

  31. Uddin S, Choudhry MA (1995) Quercetin, a bioflavonoid, inhibits the DNA synthesis of human leukemia cells. Biochem Mol Bio Int 36:545–550

    CAS  Google Scholar 

  32. Kuo SM (1996) Antiproliferative potency of structurally distinct dietary flavonoids on human colon cancer cells. Cancer Lett 110:41–48

    Article  CAS  Google Scholar 

  33. Chi CW, Chang YF, Ou YR, Hsieh CC, Lui YW, Peng FK, Liu TY (1997) Effect of quercetin on the in vitro and in vivo growth of mouse hepatoma cells. Oncol Res 4:1021–1024

    CAS  Google Scholar 

  34. Brusick D (1993) Genotoxicity of phenolic antioxidants. Toxicol Ind Health 9:223–230

    CAS  Google Scholar 

  35. Ahmed MS, Ramesh V, Nagaraja V, Parish JH, Hadi SM (1994) Mode of binding of quercetin to DNA. Mutagenesis 9:193–197

    CAS  Google Scholar 

  36. Viña J (1990) Glutathione: metabolism and physiological functions.CRC Press, Boston

  37. Rodgers EH, Grant MH (1998) The effect of the flavonoids, quercetin, myricetin and epicatechin on the growth and enzyme activities of MCF7 human breast cancer cells. Chem Biol Interact 116:213–228

    CAS  Google Scholar 

  38. Myhrstad MC, Carlsen H, Nordstrom O, Blomhoff R, Moskaug JO (2002) Flavonoids increase the intracellular glutathione level by transactivation of the gamma-glutamylcysteine synthetase catalytical subunit promoter. Free Rad Biol Med 32:386–393

    Article  CAS  Google Scholar 

  39. Scharf G, Prustomersky S, Knasmuller S, Schulte-Hermann R, Huber WW (2003) Enhancement of glutathione and g-glutamylcysteine synthetase, the rate limiting enzyme of glutathione synthesis, by chemoprotective plant-derived food and beverage components in the human hepatoma cell line HepG2. Nutr Cancer 45:74–83

    CAS  Google Scholar 

  40. Pilz J,Meineke I, Gleiter CH (2000) Measurement of free and bound malondialdehyde in plasma by high-performance liquid chromatography as the 2,4-dinitrophenylhydrazine derivative. J Chrom B 742:315–325

    CAS  Google Scholar 

  41. Suttnar J, Cermak J, Dyr E (1997) Solidphase extraction in malondialdehyde analysis. Anal Biochem 249:20–23

    Article  CAS  Google Scholar 

  42. Suttnar J, Masova L, Dyr E (2001) Influence of citrate and EDTA anticoagulants on plasma malondialdehyde concentrations estimated by high-performance liquid chromatography. J Chrom B 751:193–197

    CAS  Google Scholar 

  43. Holley AE, Cheeseman KH (1993) Measuring free radical reactions in vivo. Br Med Bull 49:494–505

    CAS  Google Scholar 

  44. LeBel CP, Ishiropoulos H, Bondy SC (1992) Evaluation of the probe 2’,7’- dichlorofluorescein as an indicator of reactive oxygen species formation and oxidative stress. Chem Res Toxicol 5:227–231

    Article  CAS  Google Scholar 

  45. Lores-Arnaiz S, Llesuy S, Cutrin JC, Boveris A (1995) Oxidative stress by acute acetaminophen administration in mouse liver. Free Rad Biol Med 19:303–310

    CAS  Google Scholar 

  46. Röhrdanz E, Ohler S, Tran-Thi Q-H, Kahl R (2002) The phytoestrogen Daidzein affects the antioxidant enzyme system of rat hepatoma H4IIE cells. J Nutr 132:370–375

    Google Scholar 

  47. Ursini F, Maiorino M, Brigelius-Flohé R, Aumann KD, Roveri A, Schomburg D, Flohé L (1995) Diversity of Glutathione peroxidases. Methods Enzymol 252:38–114

    CAS  Google Scholar 

  48. Nagata H, Takekoshi S, Takagi T, Honma T, Watanabe K (1999) Antioxidative action of flavonoids, quercetin and catechin, mediated by the activation of glutathione peroxidase. Tokai J Exp Clin Med 24:1–11

    CAS  Google Scholar 

  49. Breinholt V (1999) Desirable versus harmful levels of intake of flavonoids and phenolic acids. In: Kumpulainen JT, Salonen JT (eds) Natural antioxidants in nutrition, health and disease. Cambridge: Royal Society of Chemistry, pp 93–105

  50. Park EY, Rho HM (2002) The transcriptional activation of the human copper/ zinc superoxide dismutase gene by 2,3,7,8-tetrachlorodibenzo-p-dioxin through two different regulator sites, the antioxidant responsive element and xenobiotic responsive element. Mol Cell Biochem 240:47–55

    Article  CAS  Google Scholar 

  51. Zhou LZ, Johnson AP, Rando TA (2001) NF kappa B and AP-1 mediate transcriptional responses to oxidative stress in skeletal muscle cells. Free Rad Biol Med 31:1405–1416

    Article  CAS  Google Scholar 

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Correspondence to L. Goya.

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Alía, M., Mateos, R., Ramos, S. et al. Influence of quercetin and rutin on growth and antioxidant defense system of a human hepatoma cell line (HepG2). Eur J Nutr 45, 19–28 (2006). https://doi.org/10.1007/s00394-005-0558-7

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  • DOI: https://doi.org/10.1007/s00394-005-0558-7

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