ERK1/2-HNF4α axis is involved in epigallocatechin-3-gallate inhibition of HBV replication

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Abstract

Epigallocatechin gallate (EGCG), a major polyphenol in green tea, exhibits diverse biological activities. Previous studies show that EGCG could effectively suppress HBV gene expression and replication, but the role of EGCG in HBV replication and its underlying mechanisms, especially the signaling pathways involved, remain unclear. In this study we investigated the mechanisms underlying EGCG inhibition on HBV replication with a focus on the signaling pathways. We showed that EGCG (12.5−50 μM) dose-dependently inhibited HBV gene expression and replication in HepG2.2.15 cells. Similar results were observed in HBV mice receiving EGCG (25 mg· kg−1· d−1, ip) for 5 days. In HepG2.2.15 cells, we showed that EGCG (12.5−50 μM) significantly activate ERK1/2 MAPK signaling, slightly activate p38 MAPK and JAK2/STAT3 signaling, while had no significant effect on the activation of JNK MAPK, PI3K/AKT/mTOR and NF-κB signaling. By using specific inhibitors of these signaling pathways, we demonstrated that ERK1/2 signaling pathway, but not other signaling pathways, was involved in EGCG-mediated inhibition of HBV transcription and replication. Furthermore, we showed that EGCG treatment dose-dependently decreased the expression of hepatocyte nuclear factor 4α (HNF4α) both at the mRNA and protein levels, which could be reversed by pretreatment with the ERK1/2 inhibitor PD98059 (20 μM). Moreover, we revealed that EGCG treatment dose-dependently inhibited the activity of HBV core promoter and the following HBV replication. In summary, our results demonstrate that EGCG inhibits HBV gene expression and replication, which involves ERK1/2-mediated downregulation of HNF4α.These data reveal a novel mechanism for EGCG to inhibit HBV gene expression and replication.

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References

  1. 1.

    Tsai KN, Kuo CF, Ou JJ. Mechanisms of hepatitis B virus persistence. Trends Microbiol. 2018;26:33–42.

  2. 2.

    Schweitzer A, Horn J, Mikolajczyk RT, Krause G, Ott JJ. Estimations of worldwide prevalence of chronic hepatitis B virus infection: a systematic review of data published between 1965 and 2013. Lancet. 2015;386:1546–55.

  3. 3.

    Wong GL, Wong VW, Chan HL. Virus and host testing to manage chronic hepatitis B. Clin Infect Dis. 2016;62(Suppl 4):298–305.

  4. 4.

    Ward H, Tang L, Poonia B, Kottilil S. Treatment of hepatitis B virus: an update. Future Microbiol. 2016;11:1581–97.

  5. 5.

    Narotzki B, Reznick AZ, Aizenbud D, Levy Y. Green tea: a promising natural product in oral health. Arch Oral Biol. 2012;57:429–35.

  6. 6.

    Xu J, Xu Z, Zheng W. A review of the antiviral role of green tea catechins. Molecules 2017;22: pii: E1337.

  7. 7.

    Xi J, Shen D, Zhao S, Lu B, Li Y, Zhang R. Characterization of polyphenols from green tea leaves using a high hydrostatic pressure extraction. Int J Pharm. 2009;382:139–43.

  8. 8.

    Guo Q, Zhao B, Li M, Shen S, Xin W. Studies on protective mechanisms of four components of green tea polyphenols against lipid peroxidation in synaptosomes. Biochim Biophys Acta. 1996;1304:210–22.

  9. 9.

    Reygaert WC. The antimicrobial possibilities of green tea. Front Microbiol. 2014;5:434.

  10. 10.

    Calland N, Albecka A, Belouzard S, Wychowski C, Duverlie G, Descamps V, et al. (-)-Epigallocatechin-3-gallate is a new inhibitor of hepatitis C virus entry. Hepatology. 2012;55:720–9.

  11. 11.

    Wang CY, Hour MJ, Lai HC, Chen CH, Chang PJ, Huang SH, et al. Epigallocatechin-3-gallate inhibits the early stages of Japanese encephalitis virus infection. Virus Res. 2018;253:140–6.

  12. 12.

    Yang ZF, Bai LP, Huang WB, Li XZ, Zhao SS, Zhong NS, et al. Comparison of in vitro antiviral activity of tea polyphenols against influenza A and B viruses and structure-activity relationship analysis. Fitoterapia. 2014;93:47–53.

  13. 13.

    Steinmann J, Buer J, Pietschmann T, Steinmann E. Anti-infective properties of epigallocatechin-3-gallate (EGCG), a component of green tea. Br J Pharmacol. 2013;168:1059–73.

  14. 14.

    Li S, Hattori T, Kodama EN. Epigallocatechin gallate inhibits the HIV reverse transcription step. Antivir Chem Chemother. 2011;21:239–43.

  15. 15.

    Zhong L, Hu J, Shu W, Gao B, Xiong S. Epigallocatechin-3-gallate opposes HBV-induced incomplete autophagy by enhancing lysosomal acidification, which is unfavorable for HBV replication. Cell Death Dis. 2015;6:e1770.

  16. 16.

    Lai YH, Sun CP, Huang HC, Chen JC, Liu HK, Huang C. Epigallocatechin gallate inhibits hepatitis B virus infection in human liver chimeric mice. BMC Complement Alter Med. 2018;18:248.

  17. 17.

    Huang HC, Tao MH, Hung TM, Chen JC, Lin ZJ, Huang C. (-)-Epigallocatechin-3-gallate inhibits entry of hepatitis B virus into hepatocytes. Antivir Res. 2014;111:100–11.

  18. 18.

    Xu J, Gu W, Li C, Li X, Xing G, Li Y, et al. Epigallocatechin gallate inhibits hepatitis B virus via farnesoid X receptor alpha. J Nat Med. 2016;70:584–91.

  19. 19.

    Trepo C, Chan HL, Lok A. Hepatitis B virus infection. Lancet. 2014;384:2053–63.

  20. 20.

    Quasdorff M, Protzer U. Control of hepatitis B virus at the level of transcription. J Viral Hepat. 2010;17:527–36.

  21. 21.

    Schieck A, Schulze A, Gahler C, Muller T, Haberkorn U, Alexandrov A, et al. Hepatitis B virus hepatotropism is mediated by specific receptor recognition in the liver and not restricted to susceptible hosts. Hepatology. 2013;58:43–53.

  22. 22.

    Yan H, Zhong G, Xu G, He W, Jing Z, Gao Z, et al. Sodium taurocholate cotransporting polypeptide is a functional receptor for human hepatitis B and D virus. Elife. 2012;1:e49.

  23. 23.

    Tang H, McLachlan A. Transcriptional regulation of hepatitis B virus by nuclear hormone receptors is a critical determinant of viral tropism. Proc Natl Acad Sci USA. 2001;98:1841–6.

  24. 24.

    Chen EQ, Sun H, Feng P, Gong DY, Liu C, Bai L, et al. Study of the expression levels of hepatocyte nuclear factor 4 alpha and 3 beta in patients with different outcome of HBV infection. Virol J. 2012;9:23.

  25. 25.

    Raney AK, Johnson JL, Palmer CN, McLachlan A. Members of the nuclear receptor superfamily regulate transcription from the hepatitis B virus nucleocapsid promoter. J Virol. 1997;71:1058–71.

  26. 26.

    Zhao Z, Hong W, Zeng Z, Wu Y, Hu K, Tian X, et al. Mucroporin-M1 inhibits hepatitis B virus replication by activating the mitogen-activated protein kinase (MAPK) pathway and down-regulating HNF4alpha in vitro and in vivo. J Biol Chem. 2012;287:30181–90.

  27. 27.

    Huang LR, Wu HL, Chen PJ, Chen DS. An immunocompetent mouse model for the tolerance of human chronic hepatitis B virus infection. Proc Natl Acad Sci USA. 2006;103:17862–7.

  28. 28.

    Zhong L, Shu W, Dai W, Gao B, Xiong S. Reactive oxygen species-mediated c-Jun NH2-terminal kinase activation contributes to hepatitis B virus X protein-induced autophagy via regulation of the beclin-1/Bcl-2 interaction. J Virol. 2017;91:e00001–17 pii.

  29. 29.

    Gao B, Duan Z, Xu W, Xiong S. Tripartite motif-containing 22 inhibits the activity of hepatitis B virus core promoter, which is dependent on nuclear-located RING domain. Hepatology. 2009;50:424–33.

  30. 30.

    Sato S, Li K, Kameyama T, Hayashi T, Ishida Y, Murakami S, et al. The RNA sensor RIG-I dually functions as an innate sensor and direct antiviral factor for hepatitis B virus. Immunity. 2015;42:123–32.

  31. 31.

    Li Y, Ying C, Zuo X, Yi H, Yi W, Meng Y, et al. Green tea polyphenols down-regulate caveolin-1 expression via ERK1/2 and p38MAPK in endothelial cells. J Nutr Biochem. 2009;20:1021–7.

  32. 32.

    Zhao X, Liu F, Jin H, Li R, Wang Y, Zhang W, et al. Involvement of PKCalpha and ERK1/2 signaling pathways in EGCG’s protection against stress-induced neural injuries in Wistar rats. Neuroscience. 2017;346:226–37.

  33. 33.

    Mi Y, Qi G, Fan R, Qiao Q, Sun Y, Gao Y, et al. EGCG ameliorates high-fat- and high-fructose-induced cognitive defects by regulating the IRS/AKT and ERK/CREB/BDNF signaling pathways in the CNS. FASEB J. 2017;31:4998–5011.

  34. 34.

    Liu S, Sun Z, Chu P, Li H, Ahsan A, Zhou Z, et al. EGCG protects against homocysteine-induced human umbilical vein endothelial cells apoptosis by modulating mitochondrial-dependent apoptotic signaling and PI3K/Akt/eNOS signaling pathways. Apoptosis. 2017;22:672–80.

  35. 35.

    Zhou X, Wang Y, Metselaar HJ, Janssen HL, Peppelenbosch MP, Pan Q. Rapamycin and everolimus facilitate hepatitis E virus replication: revealing a basal defense mechanism of PI3K-PKB-mTOR pathway. J Hepatol. 2014;61:746–54.

  36. 36.

    Zheng Y, Li J, Johnson DL, Ou JH. Regulation of hepatitis B virus replication by the ras-mitogen-activated protein kinase signaling pathway. J Virol. 2003;77:7707–12.

  37. 37.

    Guo H, Zhou T, Jiang D, Cuconati A, Xiao GH, Block TM, et al. Regulation of hepatitis B virus replication by the phosphatidylinositol 3-kinase-AKT signal transduction pathway. J Virol. 2007;81:10072–80.

  38. 38.

    Veto B, Bojcsuk D, Bacquet C, Kiss J, Sipeki S, Martin L, et al. The transcriptional activity of hepatocyte nuclear factor 4 alpha is inhibited via phosphorylation by ERK1/2. PLoS One. 2017;12:e172020.

  39. 39.

    Tipoe GL, Leung TM, Hung MW, Fung ML. Green tea polyphenols as an anti-oxidant and anti-inflammatory agent for cardiovascular protection. Cardiovasc Hematol Disord Drug Targets. 2007;7:135–44.

  40. 40.

    Li MJ, Yin YC, Wang J, Jiang YF. Green tea compounds in breast cancer prevention and treatment. World J Clin Oncol. 2014;5:520–8.

  41. 41.

    de Boussac H, Ratajewski M, Sachrajda I, Koblos G, Tordai A, Pulaski L, et al. The ERK1/2-hepatocyte nuclear factor 4alpha axis regulates human ABCC6 gene expression in hepatocytes. J Biol Chem. 2010;285:22800–8.

  42. 42.

    Hatzis P, Kyrmizi I, Talianidis I. Mitogen-activated protein kinase-mediated disruption of enhancer-promoter communication inhibits hepatocyte nuclear factor 4alpha expression. Mol Cell Biol. 2006;26:7017–29.

  43. 43.

    Chou YC, Chen ML, Hu CP, Chen YL, Chong CL, Tsai YL, et al. Transforming growth factor-beta1 suppresses hepatitis B virus replication primarily through transcriptional inhibition of pregenomic RNA. Hepatology. 2007;46:672–81.

  44. 44.

    Hosel M, Quasdorff M, Wiegmann K, Webb D, Zedler U, Broxtermann M, et al. Not interferon, but interleukin-6 controls early gene expression in hepatitis B virus infection. Hepatology. 2009;50:1773–82.

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Acknowledgements

This work was supported by grants from the National Natural Science Foundation of China (31470839, 31872731, and 21334001), the Jiangsu Natural Science Youth Fund (No. BK20170209), the China Postdoctoral Research Foundation (No. 180942), the Jiangsu Province Postdoctoral Research Foundation (1701012A) and the Key Talents of Young Medical Science Project in Jiangsu Province (No. QNRC2016169).

Author information

ZYW, YQL, ZWG, and XHZ conducted the experiments; ZYW, YQL, MDL, and BG analyzed the data; BG and TCX supervised the project; and BG, ZYW, and TCX wrote the paper.

Correspondence to Tong-chun Xue or Bo Gao.

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The authors declare no competing interests.

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Cite this article

Wang, Z., Li, Y., Guo, Z. et al. ERK1/2-HNF4α axis is involved in epigallocatechin-3-gallate inhibition of HBV replication. Acta Pharmacol Sin (2019) doi:10.1038/s41401-019-0302-0

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Keywords

  • HBV
  • EGCG
  • ERK1/2
  • HNF4α
  • core promoter
  • PD98059
  • HepG2.2.15 cells
  • HBV mice