Journal of Food Hygiene and Safety (한국식품위생안전성학회지)

The Korean Society of Food Hygiene and Safety (한국식품위생안전성학회)
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Protective Effect of Green Tea Extract and EGCG on Ethanol-induced Cytotoxicity and DNA Damage in NIH/3T3 and HepG2 Cells

  • Kim, Nam Yee (College of Pharmacy, Kangwon National University) ;
  • Kim, Hyun Pyo (College of Pharmacy, Kangwon National University) ;
  • Heo, Moon Young (College of Pharmacy, Kangwon National University)
  • Received : 2016.02.03
  • Accepted : 2016.02.19
  • Published : 2016.02.28
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Abstract

In the present study, our aim was to determine whether green tea extract (GTE) and its major constituent, epigallocatechin-3-gallate (EGCG) have a protective effect on ethanol-induced cytotoxicity and DNA damage in NIH/3T3 and HepG2 cells. The cell viability and DNA single strand breaks were examined by MTT assay and alkaline single cell gel electrophoresis (Comet assay), respectively. Ethanol decreased the cell viability and also increased DNA single strand breaks in a concentration-dependent manner. On the other hand, GTE showed the protective effect of cytotoxicity and DNA damage induced by ethanol in both cell lines. GTE and EGCG, were found to possess the anti-oxidative and anti-genotoxic activities by evaluation with DPPH test, LDL oxidation assay, oxidative DNA damage assay and 8OH-2'dG generation test. These results were also verified by the experimental results demonstrating the lower cytotoxicity and genotoxicity of commercial green tea liqueur compared to pure ethanol in same concentration. Thus it is concluded that the supplementation of GTE or EGCG may mitigate the ethanol-induced cytotoxicity and DNA damage.

본 연구의 목적은 NIH3T3와 HepG2 세포에서 에탄올 유도 세포독성 및 유전독성에 대하여 녹차엑기스(GTE)와 epigallocatechin-3-gallate (EGCG)의 보호작용을 평가하는데 있다. 세포생존율은 MTT assay를 실시하였으며 DNA 손상도는 Comet assay로 실시한 결과 에탄올은 농도의존적인 세포독성과 유전독성을 나타내었다. 한편 GTE와 EGCG는 에탄올 유도 세포독성 및 DNA 손상에 대하여 유의성 있는 억제효과를 나타내었으며 DPPH시험과 LDL oxidation 및 8OH-2'dG 생성시험에서 항산화효과를 나타내었다. 한편 녹차성분 함유 시판 리큐르주도 순수 에탄올에 비하여 세포독성억제 및 DNA 손상억제효과를 나타내었다. 이상의 시험결과 GTE와 함유 EGCG는 항산화성 유전독성억제기전을 통한 에탄올독성저감 물질로 판단된다.

Keywords

References

  1. Bondy, S.C.: Ethanol toxicity and oxidative stress. Toxicology Letter 63, 231-41 (1992). https://doi.org/10.1016/0378-4274(92)90086-Y
  2. Cederbaum, A.I., Wu, D., Mari, M., Bai, J.: CYP2E1-dependent toxicity and oxidative stress in HepG2 cells. Free Radic. Biol. Med. 31, 1539-43 (2001). https://doi.org/10.1016/S0891-5849(01)00743-2
  3. French, S.W.: Intragastric ethanol infusion model for cellular and molecular studies of alcoholic liver disease. J. Biomed. Sci. 8, 20-7 (2001). https://doi.org/10.1007/BF02255967
  4. Adachi, M. and Ishii H.: Role of mitochondria in alcoholic liver injury. Free Radi. Biol. Med. 32, 487-91 (2002). https://doi.org/10.1016/S0891-5849(02)00740-2
  5. Bailey, S.M. and Cunningham, C.C.: Contribution of mitochondria to oxidative stress associated with alcoholic liver disease. Free Radi. Biol. Med. 32, 11-6 (2002). https://doi.org/10.1016/S0891-5849(01)00769-9
  6. Arteel, G.E.: Oxidants and antioxidants in alcohol-induced liver disease. Gastroenterology 124, 778-90 (2003). https://doi.org/10.1053/gast.2003.50087
  7. De Groot, H.: Reactive oxygen species in tissue injury. Hepato-Gastroenterology 41, 328-32 (1994).
  8. Fernandez-Checa, J.C., Kaplowiz, N., Colell, A., Garcia-Ruiz, C.: Oxidative stress and alcoholic liver disease. Alcohol Health & Research World 21, 321-4 (1997).
  9. Kim, N.Y., Lee, J.H., Heo, M.Y.: Protective effect of green tea extracts on oxidative stress. Korean J. Medicinal Crop Sci. 14(6), 322-328 (2006).
  10. Zheng, W. and Wang, S.Y.: Antioxidant activity and phenolic compounds in selected herbs. J. Agric. Food. Chem. 49, 5165-5170 (2001). https://doi.org/10.1021/jf010697n
  11. Cole, S.P.C.: Rapid chemosensitivity testing of human lung tumor cells using the MTT assay. Cancer Chemother. Pharmacol. 17, 259-63 (1986).
  12. Collins, A.R.: The comet assay for DNA damage and repair: principles, applications, and limitations. Mol. Biotechnol. 26(3), 249-61 (2004).
  13. Singh, N.P., McCoy, M.T., Tice, R.R., Schneider E.L.: A simple technique for quantitation of low levels of DNA damage in individual cells. Experimental Cell Research 175, 184-91 (1988). https://doi.org/10.1016/0014-4827(88)90265-0
  14. Olive, P.L., Banath, R.E., Durand, R.E.: Heterogenecity in radiation-induced DNA damage and repair in tumor and normal cells measured using the comet assay. Radiat. Res. 122, 86-94 (1990). https://doi.org/10.2307/3577587
  15. Fugita, Y, Uera, I., Morimoto, Y., Nakajima, M., Hatano, C., Okuda, T.: Studies on inhibition mechanism of auto-oxidation by tannins and flavonoids. Yakugaku Zasshi 108, 129-35 (1988). https://doi.org/10.1248/yakushi1947.108.2_129
  16. Ohkawa, H., Ohishi, N., Yagi, K.: Assay for lipid peroxides in animal tissues by thiobarbituric acid reaction. Anal. Biochem. 95, 351-88 (1979). https://doi.org/10.1016/0003-2697(79)90738-3
  17. Park, J.W., Cundy, K.C., Ames, B.N.: Detection of DNA adducts high-performance liquid chromatography with electrochemical detection. Carcinogenesis, 10, 827-32 (1989). https://doi.org/10.1093/carcin/10.5.827
  18. Song, E.J., Kim, N.Y., Heo, M.Y.: Protective effect of Korean medicinal plants on ethanol-induced cytotoxicity in HepG2 cells. Natural Product Sciences, 19, 329-36 (2013).
  19. Nordmann, R., Ribiere, C., Rouach, H.: Implication of free radical mechanisms in ethanol-induced cellular injury. Free Radic. Biol. Med. 12, 219-40 (1992). https://doi.org/10.1016/0891-5849(92)90030-K
  20. Tsukamoto, H., Horne, W., Kamimura, S.: Experimental liver cirrhosis induced by alcohol and iron. Journal of Clinical Investigation 96, 620-30 (1995). https://doi.org/10.1172/JCI118077
  21. Nanji, A.A., Zhao, S., Sadrazadeh, S.M.H.: Markedly enhanced cytochrome P450 2E1 induction and lipid peroxidation is associated with severe liver injury in fish oil.treated ethanolfed rats. Alcoholism : Clinical and Experimental Research 18, 1280-5 (1994). https://doi.org/10.1111/j.1530-0277.1994.tb00119.x
  22. Corrao, G., Torchio, P., Zambon, A., D'Amicis, A., Lepore, A.R., di Orio, F.: Alcohol consumption and micronutrient intake as risk factors for liver cirrhosis: a case-control study. The Provincial Group for the Study of Chronic Liver Disease. Ann. Epidemiol. 8, 154-9 (1998). https://doi.org/10.1016/S1047-2797(97)00193-2
  23. Feng, Q., Kumagai, T., Torii, Y., Nakamura, Y., Osawa, T., Uchida, K.: Anticarcinogenic antioxidants as inhibitors against intracellular oxidative stress. Free Radic. Res. 35(6), 779-88 (2001). https://doi.org/10.1080/10715760100301281
  24. Arteel, G.E., Uesugi, T., Bevan, L.N., Gabele, E., Wheele,r M.D., McKim, S.E., Thurman, R.G.: Green tea extract protects against early alcohol-induced liver injury in rats. Biol. Chem. 383, 663-70 (2002).
  25. Jimenez-Lopez, J.M. and Cederbaum, A.I.: Green tea polyphenol epigallocatechin-3-gallate protects HepG2 cells against CYP2E1-dependent toxicity. Free Radic. Biol. Med. 36(3), 359-70 (2004). https://doi.org/10.1016/j.freeradbiomed.2003.11.016
  26. Thangapazham, R.L., Singh, A.K., Sharma, A., Warren, J., Gaddipati, J.P., Maheshwari, R.K.: Green tea polyphenols and its constituent epigallocatechin gallate inhibit proliferation of human breast cancer cells in vitro and in vivo. Cancer Lett. 245, 232-41 (2007). https://doi.org/10.1016/j.canlet.2006.01.027
  27. Lee, S.I., Kim, H.J., Boo, Y.C.;Effect of green tea and (-)-epigallocatechin gallate on ethanol-induced toxicity in HepG2 cells. Phytother Res. 22(5), 669-74 (2008). https://doi.org/10.1002/ptr.2390
  28. Garcia-Rodriguez Mdel, C., Nicolas-Mendez, T., Montano- Rodriguez, A.R., Altamirano-Lozano, M,A.: Antigenotoxic effects of (-)-epigallocatechin-3-gallate (EGCG), quercetin, and rutin on chromium trioxide-induced micronuclei in the polychromatic erythrocytes of mouse peripheral blood. J. Toxicol. Environ. Health A, 77(6), 324-36 (2014). https://doi.org/10.1080/15287394.2013.865006