The protective effects of epigallocatechin gallate on lipopolysa ccharide-induced hepatotoxicity: an in vitro study on Hep3B cells

Document Type: Original Article


1 Department of Medical Microbiology, Medical Faculty, Kafkas University, Kars, Turkey

2 Department of General Surgery, Medical Faculty, Kafkas University, Kars, Turkey

3 Department of Histology and Embryology, Medical Faculty, Kafkas University, Kars, Turkey

4 Department of Pharmacology, Medical Faculty, Ataturk University, Erzurum, Turkey

5 Department of Physiology, Medical Faculty, Kafkas University, Kars, Turkey

6 Department of Biochemistry, Veterinary Faculty, Ataturk University, Erzurum, Turkey

7 Department of Virology, Veterinary Faculty, Ataturk University, Erzurum, Turkey

8 Department of Physiology, Veterinary Faculty, Kafkas University, Kars, Turkey

9 Department of Pharmacology and Toxicology, Veterinary Faculty, Ataturk University, Erzurum, Turkey


Objective(s): In the present study, our aim was to investigate the possible protective effects of epigallocatechin gallate (EGCG) on lipopolysaccharide (LPS)-induced hepatotoxicity by using Hep3B human hepatoma cells. Specifically, the study examines the role of some proinflammatory markers and oxidative damage as possible mechanisms of LPS-associated cytotoxicity. Consequently, the hepatocellular carcinoma cell line Hep3B was chosen as a model for investigation of LPS toxicity and the effect of EGCG on LPS-exposed cells.
Materials and Methods:The Hep3B human hepatoma cells were used for this study. The cytotoxic effects of chemicals (EGCG and LPS), AST and ALT levels, SOD and CAT activities, GSH-Px level and TNF-alpha and IL-6 levels were detected by using different biochemical and molecular methods. LPS and EGCG were applied to cells at various times and doses.
Results:The highest treatment dose of EGCG (400 µM) led to a dramatic decrease in SOD level and increase in CAT and GSH levels. Additionally, the highest dose of EGCG also led to a dramatic increase in TNF-alpha and IL-6 levels. On the other hand, effective doses of EGCG (200 and 100 µM) normalized all related parameters levels.
Conclusion:LPS caused hepatotoxicity, but interestingly, a high dose of EGCG was found to be a cytotoxic agent in this study. However, other two doses of EGCG led to a decrease in both inflammatory cytokine levels and antioxidant enzyme levels. Further studies should examine the effect of EGCG on secondary cellular signaling pathways.


1.Riordan SM, Williams R. Mechanisms of hepatocyte injury, multiorgan failure, and prognostic criteria in acute liver failure. Semin Liver Dis 2003; 23:203-215.

2.Betteridge DJ. What is oxidative stress? Metabolism. 2000; 49(2 Suppl 1):3-8.

3.Dinarello CA. Proinflammatory cytokines. Chest 2000; 118:503-508.

4.Elmarakby AA, Sullivan JC. Relationship between oxidative stress and inflammatory cytokines in diabetic nephropathy. Cardiovasc Ther 2012; 30:49-59.

5.Liu Q, Qian Y, Chen F, Chen X, Chen Z, Zheng M. EGCG attenuates pro-inflammatory cytokines and chemokines production in LPS-stimulated L02 hepatocyte. Acta Biochim Biophys Sin (Shanghai) 2014; 46:31-39.

6.Ghosh S, Latimer RD, Gray BM, Harwood RJ, Oduro A. Endotoxin-induced organ injury. Crit Care Med 1993; 21:19-24.

7.Bansal AK, Bansal M, Soni G, Bhatnagar D. Protective role of Vitamin E pre-treatment on N-nitrosodiethylamine induced oxidative stress in rat liver. Chem Biol Interact 2005; 156:101-111.

8.Kim HS, Quon MJ, Kim JA. New insights into the mechanisms of polyphenols beyond antioxidant properties; lessons from the green tea polyphenol, epigallocatechin 3-gallate. Redox Biol 2014; 2:187-195.

9.Gloro R, Hourmand-Ollivier I, Mosquet B, Mosquet L, Rousselot P, Salame E, et al. Fulminant hepatitis during self-medication with hydroalcoholic extract of green tea. Eur J Gastroenterol Hepatol 2005; 17:1135-1137.

10. Molinari M, Watt KD, Kruszyna T, Nelson R, Walsh M, Huang WY, et al. Acute liver failure induced by green tea extracts: case report and review of the literature. Liver Transpl 2006; 12:1892-1895.

11. Peairs A, Dai R, Gan L, Shimp S, Rylander MN, Li L, et al. Epigallocatechin-3-gallate (EGCG) attenuates inflammation in MRL/lpr mouse mesangial cells. Cell Mol Immunol 2010; 7:123-132.

12. Lambert JD, Sang S, Hong J, Yang CS. Anticancer and anti-inflammatory effects of cysteine metabolites of the green tea polyphenol, (-)-epigallocatechin-3-gallate. J Agric Food Chem 2010; 58:10016-10019.

13. Fridovich I. Superoxide dismutases. Adv Enzymol Relat Areas Mol Biol 1986; 58:61-97.

14. Mates JM, Perez-Gomez C, Nunez de Castro I. Antioxidant enzymes and human diseases. Clin Biochem 1999; 32:595-603.

15. Zhao Y, Zhou P, Liu B, Bambakidis T, Mazitschek R, Alam HB, et al. Protective effect of suberoylanilide hydroxamic acid against lipopolysaccharide-induced liver damage in rodents. J Surg Res 2015; 194:544-550.

16. Fang JL, Beland FA. Long-term exposure to zidovudine delays cell cycle progression, induces apoptosis, and decreases telomerase activity in human hepatocytes. Toxicol Sci 2009; 111:120-130.

17. Westerink WM, Schoonen WG. Phase II enzyme levels in HepG2 cells and cryopreserved primary human hepatocytes and their induction in HepG2 cells. Toxicol In Vitro 2007; 21:1592-1602.

18. Sahu SC, O'Donnell MW, Jr., Sprando RL. Interactive toxicity of usnic acid and lipopolysaccharides in human liver HepG2 cells. J Appl Toxicol 2012; 32:739-749.

19. Jennen DG, Magkoufopoulou C, Ketelslegers HB, van Herwijnen MH, Kleinjans JC, van Delft JH. Comparison of HepG2 and HepaRG by whole-genome gene expression analysis for the purpose of chemical hazard identification. Toxicol Sci 2010; 115:66-79.

20. O'Brien PJ, Irwin W, Diaz D, Howard-Cofield E, Krejsa CM, Slaughter MR, et al. High concordance of drug-induced human hepatotoxicity with in vitro cytotoxicity measured in a novel cell-based model using high content screening. Arch Toxicol 2006; 80:580-604.

21. Mayr C, Wagner A, Neureiter D, Pichler M, Jakab M, Illig R, et al. The green tea catechin epigallocatechin gallate induces cell cycle arrest and shows potential synergism with cisplatin in biliary tract cancer cells. BMC Complement Altern Med 2015; 15:194.

22. Kim MJ, Kim HI, Chung J, Jeong TS, Park HR. (-)-Epigallocatechin-3-gallate (EGCG) increases the viability of serum-starved A549 cells through its effect on Akt. Am J Chin Med 2009; 37:723-734.

23. Atli G, Ariyurek SY, Kanak EG, Canli M. Alterations in the serum biomarkers belonging to different metabolic systems of fish (Oreochromis niloticus) after Cd and Pb exposures. Environ Toxicol Pharmacol 2015; 40:508-515.

24. Kemelo MK, Wojnarova L, Kutinova Canova N, Farghali H. D-galactosamine/lipopolysaccharide-induced hepatotoxicity downregulates sirtuin 1 in rat liver: role of sirtuin 1 modulation in hepatoprotection. Physiol Res 2014; 63:615-623.

25. Hammad MA, Abdel-Bakky MS, Walker LA, Ashfaq MK. Oxidized low-density lipoprotein and tissue factor are involved in monocrotaline/lipopolysaccharide-induced hepatotoxicity. Arch Toxicol 2011; 85:1079-1089.

26. Lyakhovich VV, Vavilin VA, Zenkov NK, Menshchikova EB. Active defense under oxidative stress. The antioxidant responsive element. Biochemistry (Mosc) 2006; 71:962-974.

27. Wang D, Wang Y, Wan X, Yang CS, Zhang J. Green tea polyphenol (-)-epigallocatechin-3-gallate triggered hepatotoxicity in mice: responses of major antioxidant enzymes and the Nrf2 rescue pathway. Toxicol Appl Pharmacol 2015; 283:65-74.

28. Gardner CR, Laskin JD, Dambach DM, Sacco M, Durham SK, Bruno MK, et al. Reduced hepatotoxicity of acetaminophen in mice lacking inducible nitric oxide synthase: potential role of tumor necrosis factor-alpha and interleukin-10. Toxicol Appl Pharmacol 2002; 184:27-36.

29. Chen J, Xu J, Li J, Du L, Chen T, Liu P, et al. Epigallocatechin-3-gallate attenuates lipopolysaccharide-induced mastitis in rats via suppressing MAPK mediated inflammatory responses and oxidative stress. Int Immunopharmacol 2015; 26:147-152.

30. Spets H, Stromberg T, Georgii-Hemming P, Siljason J, Nilsson K, Jernberg-Wiklund H. Expression of the bcl-2 family of pro- and anti-apoptotic genes in multiple myeloma and normal plasma cells: regulation during interleukin-6(IL-6)-induced growth and survival. Eur J Haematol 2002; 69:76-89.

31. Wen WC, Kuo PJ, Chiang CY, Chin YT, Fu MM, Fu E. Epigallocatechin-3-gallate attenuates Porphyromonas gingivalis lipopolysaccharide-enhanced matrix metalloproteinase-1 production through inhibition of interleukin-6 in gingival fibroblasts. J Periodontol 2014; 85:868-875.

32. Zhong Y, Chiou YS, Pan MH, Shahidi F. Anti-inflammatory activity of lipophilic epigallocatechin gallate (EGCG) derivatives in LPS-stimulated murine macrophages. Food Chem 2012; 134:742-748.

33. Lombardo Bedran TB, Palomari Spolidorio D, Grenier D. Green tea polyphenol epigallocatechin-3-gallate and cranberry proanthocyanidins act in synergy with cathelicidin (LL-37) to reduce the LPS-induced inflammatory response in a three-dimensional co-culture model of gingival epithelial cells and fibroblasts. Arch Oral Biol 2015; 60:845-853.