Skip to main content
SpringerLink
Log in

NF-κB mediates the induction of Fas receptor and Fas ligand by microcystin-LR in HepG2 cells

  • Published:
Molecular and Cellular Biochemistry Aims and scope Submit manuscript

Abstract

Microcystin-LR (MC-LR) is the most frequent and most toxic microcystin identified. This natural toxin has multiple features, including inhibitor of protein phosphatases 1 and 2A, inducer of oxidative stress, as well as, tumor initiator and promoter. One unique character of MC-LR is this chemical can accumulate into liver after contacting and lead to severe damage to hepatocytes, such as apoptosis. Fas receptor (Fas) and Fas ligand (FasL) system is a critical signaling system initiating apoptosis. In current study, we explored whether MC-LR could induce Fas and FasL expression in HepG2 cells, a well used in vitro model for the study of human hepatocytes. The data showed MC-LR induced Fas and FasL expression, at both mRNA and protein levels. We also found MC-LR induced apoptosis at the same incubation condition at which it induced Fas and FasL expression. The data also revealed MC-LR promoted nuclear translocation and activation of p65 subunit of NF-κB. By applying siRNA to knock down p65 in HepG2 cells, we successfully impaired the activation of NF-κB by MC-LR. In these p65 knockdown cells, we also observed significant reduction of MC-LR-induced Fas expression, FasL expression, and apoptosis. These findings demonstrate that the NF-κB mediates the induction of Fas and FasL as well as cellular apoptosis by MC-LR in HepG2 cells. The results bring important information for understanding how MC-LR induces apoptosis in hepatocytes.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  1. Dittmann E, Wiegand C (2006) Cyanobacterial toxins—occurrence, biosynthesis and impact on human affairs. Mol Nutr Food Res 50(1):7–17

    Article  PubMed  CAS  Google Scholar 

  2. van Apeldoorn ME, van Egmond HP, Speijers GJ, Bakker GJ (2007) Toxins of cyanobacteria. Mol Nutr Food Res 51(1):7–60

    Article  PubMed  Google Scholar 

  3. Honkanen RE, Zwiller J, Moore RE, Daily SL, Khatra BS, Dukelow M, Boynton AL (1990) Characterization of microcystin-LR, a potent inhibitor of type 1 and type 2A protein phosphatases. J Biol Chem 265(32):19401–19404

    PubMed  CAS  Google Scholar 

  4. Goldberg J, Huang HB, Kwon YG, Greengard P, Nairn AC, Kuriyan J (1995) Three-dimensional structure of the catalytic subunit of protein serine/threonine phosphatase-1. Nature 376(6543):745–753

    Article  PubMed  CAS  Google Scholar 

  5. Craig M, Luu HA, McCready TL, Williams D, Andersen RJ, Holmes CF (1996) Molecular mechanisms underlying he interaction of motuporin and microcystins with type-1 and type-2A protein phosphatases. Biochem Cell Biol 74(4):569–578

    Article  PubMed  CAS  Google Scholar 

  6. Fischer WJ, Hitzfeld BC, Tencalla F, Eriksson JE, Mikhailov A, Dietrich DR (2000) Microcystin-LR toxicodynamics, induced pathology, and immunohistochemical localization in livers of blue-green algae exposed rainbow trout (Oncorhynchus mykiss). Toxicol Sci 54(2):365–373

    Article  PubMed  CAS  Google Scholar 

  7. Ding WX, Shen HM, Ong CN (2000) Critical role of reactive oxygen species and mitochondrial permeability transition in microcystin-induced rapid apoptosis in rat hepatocytes. Hepatology 32(3):547–555

    Article  PubMed  CAS  Google Scholar 

  8. Bouaicha N, Maatouk I (2004) Microcystin-LR and nodularin induce intracellular glutathione alteration, reactive oxygen species production and lipid peroxidation in primary cultured rat hepatocytes. Toxicol Lett 148(1–2):53–63

    Article  PubMed  CAS  Google Scholar 

  9. Gupta N, Pant SC, Vijayaraghavan R, Rao PV (2003) Comparative toxicity evaluation of cyanobacterial cyclic peptide toxin microcystin variants (LR, RR, YR) in mice. Toxicology 188(2–3):285–296

    Article  PubMed  CAS  Google Scholar 

  10. Eriksson JE, Gronberg L, Nygard S, Slotte JP, Meriluoto JA (1990) Hepatocellular uptake of 3H-dihydromicrocystin-LR, a cyclic peptide toxin. Biochim Biophys Acta 1025(1):60–66

    Article  PubMed  CAS  Google Scholar 

  11. Fischer WJ, Altheimer S, Cattori V, Meier PJ, Dietrich DR, Hagenbuch B (2005) Organic anion transporting polypeptides expressed in liver and brain mediate uptake of microcystin. Toxicol Appl Pharmacol 203(3):257–263

    Article  PubMed  CAS  Google Scholar 

  12. Earnshaw WC, Martins LM, Kaufmann SH (1999) Mammalian caspases: structure, activation, substrates, and functions during apoptosis. Annu Rev Biochem 68:383–424

    Article  PubMed  CAS  Google Scholar 

  13. Maher S, Toomey D, Condron C, Bouchier-Hayes D (2002) Activation-induced cell death: the controversial role of Fas and Fas ligand in immune privilege and tumour counterattack. Immunol Cell Biol 80(2):131–137

    Article  PubMed  CAS  Google Scholar 

  14. Chinnaiyan AM, O’Rourke K, Tewari M, Dixit VM (1995) FADD, a novel death domain-containing protein, interacts with the death domain of Fas and initiates apoptosis. Cell 81(4):505–512

    Article  PubMed  CAS  Google Scholar 

  15. Kischkel FC, Hellbardt S, Behrmann I, Germer M, Pawlita M, Krammer PH, Peter ME (1995) Cytotoxicity-dependent APO-1 (Fas/CD95)-associated proteins form a death-inducing signaling complex (DISC) with the receptor. EMBO J 14(22):5579–5588

    PubMed  CAS  Google Scholar 

  16. Guicciardi ME, Gores GJ (2009) Life and death by death receptors. FASEB J 23(6):1625–1637

    Article  PubMed  CAS  Google Scholar 

  17. Ogasawara J, Watanabe-Fukunaga R, Adachi M, Matsuzawa A, Kasugai T, Kitamura Y, Itoh N, Suda T, Nagata S (1993) Lethal effect of the anti-Fas antibody in mice. Nature 364(6440):806–809

    Article  PubMed  CAS  Google Scholar 

  18. Rouquet N, Carlier K, Briand P, Wiels J, Joulin V (1996) Multiple pathways of Fas-induced apoptosis in primary culture of hepatocytes. Biochem Biophys Res Commun 229(1):27–35

    Article  PubMed  CAS  Google Scholar 

  19. Muller M, Strand S, Hug H, Heinemann EM, Walczak H, Hofmann WJ, Stremmel W, Krammer PH, Galle PR (1997) Drug-induced apoptosis in hepatoma cells is mediated by the CD95 (APO-1/Fas) receptor/ligand system and involves activation of wild-type p53. J Clin Invest 99(3):403–413

    Article  PubMed  CAS  Google Scholar 

  20. French LE, Hahne M, Viard I, Radlgruber G, Zanone R, Becker K, Muller C, Tschopp J (1996) Fas and Fas ligand in embryos and adult mice: ligand expression in several immune-privileged tissues and coexpression in adult tissues characterized by apoptotic cell turnover. J Cell Biol 133(2):335–343

    Article  PubMed  CAS  Google Scholar 

  21. Aoudjehane L, Podevin P, Scatton O, Jaffray P, Dusanter-Fourt I, Feldmann G, Massault PP, Grira L, Bringuier A, Dousset B, Chouzenoux S, Soubrane O, Calmus Y, Conti F (2007) Interleukin-4 induces human hepatocyte apoptosis through a Fas-independent pathway. FASEB J 21(7):1433–1444

    Article  PubMed  CAS  Google Scholar 

  22. Javitt NB (1990) Hep G2 cells as a resource for metabolic studies: lipoprotein, cholesterol, and bile acids. FASEB J 4(2):161–168

    PubMed  CAS  Google Scholar 

  23. van ISC, Zegers MM, Kok JW, Hoekstra D (1997) Segregation of glucosylceramide and sphingomyelin occurs in the apical to basolateral transcytotic route in HepG2 cells. J Cell Biol 137(2):347–357

    Article  Google Scholar 

  24. van der Wouden JM, van ISC, Hoekstra D (2002) Oncostatin M regulates membrane traffic and stimulates bile canalicular membrane biogenesis in HepG2 cells. EMBO J 21(23):6409–6418

    Article  PubMed  Google Scholar 

  25. Mersch-Sundermann V, Knasmuller S, Wu XJ, Darroudi F, Kassie F (2004) Use of a human-derived liver cell line for the detection of cytoprotective, antigenotoxic and cogenotoxic agents. Toxicology 198(1–3):329–340

    Article  PubMed  CAS  Google Scholar 

  26. Ohgaki R, Matsushita M, Kanazawa H, Ogihara S, Hoekstra D, van Ijzendoorn SC (2010) The Na+/H+ exchanger NHE6 in the endosomal recycling system is involved in the development of apical bile canalicular surface domains in HepG2 cells. Mol Biol Cell 21(7):1293–1304

    Article  PubMed  CAS  Google Scholar 

  27. Baichwal VR, Baeuerle PA (1997) Activate NF-κB or die? Curr Biol 7(2):R94–R96

    Article  PubMed  CAS  Google Scholar 

  28. Guttridge DC, Albanese C, Reuther JY, Pestell RG, Baldwin AS Jr (1999) NF-κB controls cell growth and differentiation through transcriptional regulation of cyclin D1. Mol Cell Biol 19(8):5785–5799

    PubMed  CAS  Google Scholar 

  29. Barkett M, Gilmore TD (1999) Control of apoptosis by Rel/NF-κB transcription factors. Oncogene 18(49):6910–6924

    Article  PubMed  CAS  Google Scholar 

  30. Dutta J, Fan Y, Gupta N, Fan G, Gelinas C (2006) Current insights into the regulation of programmed cell death by NF-κB. Oncogene 25(51):6800–6816

    Article  PubMed  CAS  Google Scholar 

  31. Chan H, Bartos DP, Owen-Schaub LB (1999) Activation-dependent transcriptional regulation of the human Fas promoter requires NF-κB p50–p65 recruitment. Mol Cell Biol 19(3):2098–2108

    PubMed  CAS  Google Scholar 

  32. Fu WY, Chen JP, Wang XM, Xu LH (2005) Altered expression of p53, Bcl-2 and Bax induced by microcystin-LR in vivo and in vitro. Toxicon 46(2):171–177

    Article  PubMed  CAS  Google Scholar 

  33. Fladmark KE, Brustugun OT, Hovland R, Boe R, Gjertsen BT, Zhivotovsky B, Doskeland SO (1999) Ultrarapid caspase-3 dependent apoptosis induction by serine/threonine phosphatase inhibitors. Cell Death Differ 6(11):1099–1108

    Article  PubMed  CAS  Google Scholar 

  34. Holtz-Heppelmann CJ, Algeciras A, Badley AD, Paya CV (1998) Transcriptional regulation of the human FasL promoter-enhancer region. J Biol Chem 273(8):4416–4423

    Article  PubMed  CAS  Google Scholar 

  35. Mondor I, Schmitt-Verhulst AM, Guerder S (2005) RelA regulates the survival of activated effector CD8 T cells. Cell Death Differ 12(11):1398–1406

    Article  PubMed  CAS  Google Scholar 

  36. Novac N, Baus D, Dostert A, Heinzel T (2006) Competition between glucocorticoid receptor and NFκB for control of the human FasL promoter. FASEB J 20(8):1074–1081

    Article  PubMed  CAS  Google Scholar 

  37. Campbell KJ, Rocha S, Perkins ND (2004) Active repression of antiapoptotic gene expression by RelA(p65) NF-κB. Mol Cell 13(6):853–865

    Article  PubMed  CAS  Google Scholar 

  38. Ashikawa K, Shishodia S, Fokt I, Priebe W, Aggarwal BB (2004) Evidence that activation of nuclear factor-κB is essential for the cytotoxic effects of doxorubicin and its analogues. Biochem Pharmacol 67(2):353–364

    Article  PubMed  CAS  Google Scholar 

  39. Campbell KJ, Witty JM, Rocha S, Perkins ND (2006) Cisplatin mimics ARF tumor suppressor regulation of RelA (p65) nuclear factor-κB transactivation. Cancer Res 66(2):929–935

    Article  PubMed  CAS  Google Scholar 

  40. Jones BE, Lo CR, Liu H, Pradhan Z, Garcia L, Srinivasan A, Valentino KL, Czaja MJ (2000) Role of caspases and NF-κB signaling in hydrogen peroxide- and superoxide-induced hepatocyte apoptosis. Am J Physiol Gastrointest Liver Physiol 278(5):G693–G699

    PubMed  CAS  Google Scholar 

  41. Roman J, Colell A, Blasco C, Caballeria J, Pares A, Rodes J, Fernandez-Checa JC (1999) Differential role of ethanol and acetaldehyde in the induction of oxidative stress in HEP G2 cells: effect on transcription factors AP-1 and NF-κB. Hepatology 30(6):1473–1480

    Article  PubMed  CAS  Google Scholar 

  42. Ghosh S, May MJ, Kopp EB (1998) NF-κB and Rel proteins: evolutionarily conserved mediators of immune responses. Annu Rev Immunol 16:225–260

    Article  PubMed  CAS  Google Scholar 

  43. DiDonato JA, Hayakawa M, Rothwarf DM, Zandi E, Karin M (1997) A cytokine-responsive IκB kinase that activates the transcription factor NF-κB. Nature 388(6642):548–554

    Article  PubMed  CAS  Google Scholar 

Download references

Conflict of interest

None declared.

Author information

Authors and Affiliations

  1. Department of Pathology, Northwestern University, 303 E Chicago Ave, Ward 3-070, Chicago, IL, 60613, USA

    Gong Feng

  2. Anong Biotech Institute, Tianjin, People’s Republic of China

    Gong Feng, Ying Li & Yansheng Bai

  3. Ross University School of Medicine, Roseau, Dominica

    Musa Abdalla

Authors
  1. Gong Feng
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Musa Abdalla
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ying Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Yansheng Bai
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Gong Feng.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Feng, G., Abdalla, M., Li, Y. et al. NF-κB mediates the induction of Fas receptor and Fas ligand by microcystin-LR in HepG2 cells. Mol Cell Biochem 352, 209–219 (2011). https://doi.org/10.1007/s11010-011-0756-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11010-011-0756-y

Keywords

Search

Navigation