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Effect of the olive oil phenol hydroxytyrosol on human hepatoma HepG2 cells

Protection against oxidative stress induced by tert-butylhydroperoxide

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Abstract

Background

Scientific evidence suggests that olive oil’s beneficial effects are related to the high level of antioxidants, including phenolic compounds such as hydroxytyrosol. In vivo studies have shown that olive oil HTy is bioavailable and its biological activities, similar to those reported for other natural antioxidants such as quercetin, include prevention of LDL oxidation. Previous studies from our laboratory have shown that HTy and other phenolics in olive oil are absorbed and metabolized by cultured human hepatoma HepG2 cells where glucuronidated and methylated conjugates were the main derivatives formed, resembling the metabolic profile of olive oil phenols observed in human plasma and urine.

Aim of the study

The effect of olive oil phenol (HTy) on cell viability and redox status of cultured HepG2 cells, and the protective effect of HTy against an oxidative stress induced by tert-butylhydroperoxide (t-BOOH) were investigated.

Methods

Lactate dehydrogenase activity as marker for cell integrity, concentration of reduced glutathione (GSH), generation of reactive oxygen species (ROS) and activity of the antioxidant enzyme glutathione peroxidase (GPx) as markers of redox status and determination of malondialdehyde (MDA) as marker of lipid peroxidation were measured.

Results

No changes in cell integrity or intrinsic antioxidant status resulted from a direct treatment with 10–40 μM HTy. Pre-treatment of HepG2 with 10–40 μM HTy for 2 or 20 h completely prevented cell damage as well as the decrease of reduced glutathione and increase of malondialdehyde evoked by t-BOOH in HepG2 cells. Reactive oxygen species generation and the significant increase of glutathione peroxidase activity induced by t-BOOH were greatly reduced when cells were pretreated with HTy.

Conclusion

The results clearly show that treatment of HepG2 cells with the olive oil phenolic HTy may positively affect their antioxidant defense system, favoring cell integrity and resistance to cope with a stressful situation.

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References

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

    Article  CAS  Google Scholar 

  2. Visioli F, Galli C (1998) The effect of minor constituents of olive oil on cardiovascular disease: new findings. Nutr Rev 56:142–147

    Article  CAS  Google Scholar 

  3. Mateos R, Espartero JL, Trujillo M, Ríos JJ, León-Camacho M, Alcudia F, Cert A (2001) Determination of phenols, flavones, and lignans in virgin olive oils by solid-phase extraction and high-performance liquid chromatography with diode array ultraviolet detection. J Agric Food Chem 49:2185–2192

    Article  CAS  Google Scholar 

  4. Owen R, Mier W, Giacosa A, Hull WE, Spiegelhalder B, Bartsch H (2000) Identification of lignans as major components in the phenolic fraction of olive oil. Clin Chem 46:976–988

    CAS  Google Scholar 

  5. Visioli F, Galli C, Bornet F, Mattei A, Patelli R, Galli G, Caruso D (2000) Olive oil phenolics are dose-dependently absorbed in humans. FEBS Lett 468:159–160

    Article  CAS  Google Scholar 

  6. Caruso D, Visioli F, Patelli R, Galli C, Galli G (2001) Urinary excretion of olive oil phenols and their metabolites in humans. Metab Clin Exp 50:1426–1428

    CAS  Google Scholar 

  7. Vissers MH, Zock PL, Roodenburg AJ, Leenen R, Katan MB (2002) Olive oil phenols are absorbed in humans. J Nutr 132:409–417

    CAS  Google Scholar 

  8. Miró-Casas E, Covas M-I, Fito M, Farré-Albaladejo M, Marrugat J, de la Torre R (2003) Tyrosol and hydroxytyrosol are absorbed from moderate and sustained doses of virgin olive oil in humans. Eur J Clin Nutr 57:186–190

    Article  CAS  Google Scholar 

  9. D’Angelo S, Manna C, Migliardi V, Mazzoni O, Morrica P, Capasso G, Pontoni G, Galletti P, Zappia V (2001) Pharmacokinetics and metabolism of hydroxytyrosol, a natural antioxidant from olive oil. Drug Metab Dispos 29:1492–1498

    CAS  Google Scholar 

  10. Tuck KL, Freeman MP, Hayball PJ, Stretch GL, Stupans I (2001) The in vivo fate of hydroxytyrosol and tyrosol, antioxidant phenolic constituents of olive oil, following intravenous and oral dosing of labelled compounds to rats. J Nutr 131:1993–1996

    CAS  Google Scholar 

  11. Tuck KL, Hayball PJ, Stupans I (2002) Structural characterization of the metabolites of hydroxytyrosol, the principal phenolic component in olive oil, in rats. J Agric Food Chem 50:2404–2409

    Article  CAS  Google Scholar 

  12. Saija A, Trombetta D, Tomaino A, Lo Cascio R, Princi P, Uccella N, Bonina F, Castelli F (1998) In vitro evaluation of the antioxidant activity and biomembrane interaction of the plant phenols oleuropein and hydroxytyrosol. Int J Pharm 166:123–133

    Article  CAS  Google Scholar 

  13. Gordon MH, Paiva-Martins F, Almeida M (2001) Antioxidant activity of hydroxytyrosol acetate compared with that of other olive oil polyphenols. J Agric Food Chem 49:2480–2485

    Article  CAS  Google Scholar 

  14. De la Puerta R, Martínez-Domínguez ME, Ruiz-Gutierrez V, Flavill JA, Hoult JR (2001) Effects of virgin olive oil phenolics on scavenging of reactive nitrogen species and upon nitrergic neurotransmission. Life Sci 69:1213–1222

    Article  Google Scholar 

  15. Manna C, Galletti P, Cucciolla V, Moltedo O, Leone A, Zappia V (1997) The protective effect of the olive oil polyphenols (3,4-dihydroxyphenyl)-ethanol counteracts reactive oxygen metabolite-induced cytotoxicity in Caco-2 cells. J Nutr 127:286–292

    CAS  Google Scholar 

  16. Mateos R, Domínguez MM, Espartero JL, Cert A (2003) Antioxidant effect of phenolic compounds, α-tocopherol, and other minor components in virgin olive oil. J Agric Food Chem 51:7170–7175

    Article  CAS  Google Scholar 

  17. Salami M, Galli C, De Angelis L, Visioli F (1995) Formation of F2-isoprostanes in oxidized low-density lipoprotein: inhibitory effect of hydroxytyrosol. Pharmacol Res 31:275–279

    Article  CAS  Google Scholar 

  18. Caruso D, Berra B, Giavarini F, Cortesi N, Fedeli E, Galli G (1999) Effect of virgin olive oil phenolic compounds on in vitro oxidation of human low density lipoproteins. Nutr Metab Cardiovasc Dis 9:102–107

    CAS  Google Scholar 

  19. Scaccini C, Nardini M, D’Aquino M, Gentili V, Di Felice M, Tomassi G (1992) Effect of dietary oils on lipid peroxidation and on antioxidant parameters of rat plasma and lipoprotein fractions. J Lipid Res 33:627–633

    CAS  Google Scholar 

  20. Coni E, Di Benedetto R, Di Pasquale M, Masella R, Modesti D, Mattei R, Carlini EA (2000) Protective effect of oleuropein, an olive oil biophenol, on low density lipoprotein oxidizability in rabbits. Lipids 35:45–54

    Article  CAS  Google Scholar 

  21. Mateos R, Goya L, Bravo L (2005) Metabolism of the olive oil phenols hydroxytyrosol, tyrosol and hydroxytyrosyl acetate by human hepatoma HepG2 cells. J Agric Food Chem 53:9897–9905

    Article  CAS  Google Scholar 

  22. Alía M, Mateos R, Ramos S, Lecumberri E, Bravo L, Goya L (2006) Influence of quercetin and rutin on growth and the antioxidant defense system in a human hepatoma cell line (HepG2). Eur J Nutr 45:19–28

    Article  CAS  Google Scholar 

  23. Baraldi PG, Simoni D, Manfredini S, Menziani E (1983) Preparation of 3,4-dihydroxy-1-benzeneethanol: a reinvestigation. Liebigs Ann Chem 684–686

  24. Alía M, Ramos S, Mateos R, Bravo L, Goya L (2005) Response of the antioxidant defense system to t-butyl hydroperoxide and hydrogen peroxide in a human hepatoma cell line (HepG2). J Biochem Mol Toxicol 19:119–128

    Article  CAS  Google Scholar 

  25. Alía M, Ramos S, Mateos R, Bravo L, Goya L (2006) Quercetin protects human hepatoma cell line (HepG2) against oxidative stress induced by tertbutyl hydroperoxide. Toxicol Appl Pharm 212:110–118

    Article  CAS  Google Scholar 

  26. 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

    Google Scholar 

  27. 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. Academic press, New York, pp. 46–49

    Google Scholar 

  28. 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 

  29. 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 Chromatogr B 805:33–39

    Article  CAS  Google Scholar 

  30. 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 72:248–254

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  32. Gunzler WA, Kremers H, Flohe L (1974) An improved coupled test procedure for glutathione peroxidase. Klin Chem Klin Biochem 12:444–448

    CAS  Google Scholar 

  33. Petroni A, Blasevich M, Salami M, Papini N, Montedoro GF, Galli C (1995) Inhibition of platelet aggregation and eicosanoid production by phenolic components of olive oil. Throm Res 78:151–160

    Article  CAS  Google Scholar 

  34. De la Puerta R, Ruiz-Gutierrez V, Hoult RS (1999) Inhibition of leukocyte 5-lipoxygenase by phenolics from virgin olive oil. Biochem Pharmacol 57:445–449

    Article  Google Scholar 

  35. Manna C, Galletti P, Cucciolla V, Montedoro G, Zappia V (1999) Olive oil hydroxytyrosol protects human erythrocytes against oxidative damages. J Nutr Biochem 10:159–165

    Article  CAS  Google Scholar 

  36. Chen L, Yang X, Jiao H, Zhao B (2002) Tea catechins protect against lead-induced cytotoxicity, lipid peroxidation, and membrane fluidity in HepG2 cells. Toxicol Sci 69:149–156

    Article  CAS  Google Scholar 

  37. Murakami C, Hirakawa Y, Inui H, Nakano Y, Yoshida H (2002) Effects of epigallocatechin 3-O-gallate on cellular antioxidative system in HepG2 cells. J Nutr Sci Vitaminol 48:89–94

    CAS  Google Scholar 

  38. Murakami C, Hirakawa Y, Inui H, Nakano Y, Yoshida H (2002) Effect of tea catechins on cellular lipid peroxidation and cytotoxicity in HepG2 cells. Biosci Biotechnol Biochem 66:1559–1562

    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

    Article  CAS  Google Scholar 

  40. 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

    Article  CAS  Google Scholar 

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

    Google Scholar 

  42. 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 

  43. 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 Chromatogr B 742:315–325

    CAS  Google Scholar 

  44. Suttnar J, Cermak J, Dyr E (1997) Solid-phase extraction in malondialdehyde analysis. Anal Biochem 249:20–23

    Article  CAS  Google Scholar 

  45. Suttnar J, Masova L, Dyr E (2001) Influence of citrate and EDTA anticoagulants on plasma malondialdehyde concentrations estimated by high-performance liquid chromatography. J Chromatogr B 751:193–119

    CAS  Google Scholar 

  46. Courtois F, Delvin E, Ledoux M, Seidman E, Lavoie JC, Levy E (2002) The antioxidant BHT normalizes some oxidative effects of iron + ascorbate on lipid metabolism in Caco-2 cells. J Nutr 132:1289–1292

    CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  48. 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

    Article  CAS  Google Scholar 

  49. 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 

  50. 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 

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Acknowledgments

R. Mateos was a postdoctoral fellow from the Ministerio de Educación y Ciencia. We thank Dr. J.L. Espartero (University of Seville, Spain) for kindly providing hydroxytyrosol.

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Authors and Affiliations

  1. Depto. de Metabolismo y Nutrición, Instituto del Frío (CSIC), C/José Antonio Novais, 10 Ciudad Universitaria, 28040, Madrid, Spain

    Luis Goya PhD & Laura Bravo PhD

  2. CIFA Venta del Llano, IFAPA, Menjíbar, Jaén, Spain

    Raquel Mateos PhD

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  1. Luis Goya PhD
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Correspondence to Luis Goya PhD.

Additional information

This work was supported by grants AGL2000-1314 and AGL2004-00302 from the Spanish Ministry of Science and Technology (CICYT).

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Goya, L., Mateos, R. & Bravo, L. Effect of the olive oil phenol hydroxytyrosol on human hepatoma HepG2 cells. Eur J Nutr 46, 70–78 (2007). https://doi.org/10.1007/s00394-006-0633-8

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