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TLR4 enhances TGF-β signaling and hepatic fibrosis

Nature Medicine volume 13, pages 1324–1332 (2007)Cite this article

Abstract

Hepatic injury is associated with a defective intestinal barrier and increased hepatic exposure to bacterial products. Here we report that the intestinal bacterial microflora and a functional Toll-like receptor 4 (TLR4), but not TLR2, are required for hepatic fibrogenesis. Using Tlr4-chimeric mice and in vivo lipopolysaccharide (LPS) challenge, we demonstrate that quiescent hepatic stellate cells (HSCs), the main precursors for myofibroblasts in the liver, are the predominant target through which TLR4 ligands promote fibrogenesis. In quiescent HSCs, TLR4 activation not only upregulates chemokine secretion and induces chemotaxis of Kupffer cells, but also downregulates the transforming growth factor (TGF)-β pseudoreceptor Bambi to sensitize HSCs to TGF-β–induced signals and allow for unrestricted activation by Kupffer cells. LPS-induced Bambi downregulation and sensitization to TGF-β is mediated by a MyD88–NF-κB–dependent pathway. Accordingly, Myd88-deficient mice have decreased hepatic fibrosis. Thus, modulation of TGF-β signaling by a TLR4-MyD88–NF-κB axis provides a novel link between proinflammatory and profibrogenic signals.

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Figure 1: Reduction of hepatic fibrogenesis and macrophage infiltration in TLR4-mutant mice.
Figure 2: Gut sterilization prevents hepatic fibrosis after bile-duct ligation.
Figure 3: Bone marrow–derived cells do not mediate TLR4-dependent profibrogenic effects.
Figure 4: Quiescent HSCs are a target of LPS in vitro and in vivo.
Figure 5: LPS downregulates Bambi to enhance TGF-β signaling and HSC activation.
Figure 6: LPS downregulates Bambi through Myd88-dependent signals.

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References

  1. Bataller, R. & Brenner, D.A. Liver fibrosis. J. Clin. Invest. 115, 209–218 (2005).

    Article  CAS  Google Scholar 

  2. Friedman, S.L. Molecular regulation of hepatic fibrosis, an integrated cellular response to tissue injury. J. Biol. Chem. 275, 2247–2250 (2000).

    Article  CAS  Google Scholar 

  3. Akira, S. & Takeda, K. Toll-like receptor signalling. Nat. Rev. Immunol. 4, 499–511 (2004).

    Article  CAS  Google Scholar 

  4. Jiang, D. et al. Regulation of lung injury and repair by Toll-like receptors and hyaluronan. Nat. Med. 11, 1173–1179 (2005).

    Article  CAS  Google Scholar 

  5. Bjorkbacka, H. et al. Reduced atherosclerosis in MyD88-null mice links elevated serum cholesterol levels to activation of innate immunity signaling pathways. Nat. Med. 10, 416–421 (2004).

    Article  Google Scholar 

  6. Zhai, Y. et al. Cutting edge: TLR4 activation mediates liver ischemia/reperfusion inflammatory response via IFN regulatory factor 3–dependent MyD88-independent pathway. J. Immunol. 173, 7115–7119 (2004).

    Article  CAS  Google Scholar 

  7. Tsung, A. et al. Hepatic ischemia/reperfusion injury involves functional TLR4 signaling in nonparenchymal cells. J. Immunol. 175, 7661–7668 (2005).

    Article  CAS  Google Scholar 

  8. Uesugi, T., Froh, M., Arteel, G.E., Bradford, B.U. & Thurman, R.G. Toll-like receptor 4 is involved in the mechanism of early alcohol-induced liver injury in mice. Hepatology 34, 101–108 (2001).

    Article  CAS  Google Scholar 

  9. Seki, E. et al. Contribution of Toll-like receptor/myeloid differentiation factor 88 signaling to murine liver regeneration. Hepatology 41, 443–450 (2005).

    Article  CAS  Google Scholar 

  10. Schwabe, R.F., Seki, E. & Brenner, D.A. Toll-like receptor signaling in the liver. Gastroenterology 130, 1886–1900 (2006).

    Article  CAS  Google Scholar 

  11. Seki, E. et al. Lipopolysaccharide-induced IL-18 secretion from murine Kupffer cells independently of myeloid differentiation factor 88 that is critically involved in induction of production of IL-12 and IL-1β. J. Immunol. 166, 2651–2657 (2001).

    Article  CAS  Google Scholar 

  12. Duffield, J.S. et al. Selective depletion of macrophages reveals distinct, opposing roles during liver injury and repair. J. Clin. Invest. 115, 56–65 (2005).

    Article  CAS  Google Scholar 

  13. Rivera, C.A. et al. Attenuation of CCl4-induced hepatic fibrosis by GdCl3 treatment or dietary glycine. Am. J. Physiol. Gastrointest. Liver Physiol. 281, G200–G207 (2001).

    Article  CAS  Google Scholar 

  14. Paik, Y.H. et al. Toll-like receptor 4 mediates inflammatory signaling by bacterial lipopolysaccharide in human hepatic stellate cells. Hepatology 37, 1043–1055 (2003).

    Article  CAS  Google Scholar 

  15. Brun, P., Castagliuolo, I., Pinzani, M., Palu, G. & Martines, D. Exposure to bacterial cell wall products triggers an inflammatory phenotype in hepatic stellate cells. Am. J. Physiol. Gastrointest. Liver Physiol. 289, G571–G578 (2005).

    Article  CAS  Google Scholar 

  16. Muhlbauer, M. et al. LPS-mediated NF-κB activation varies between activated human hepatic stellate cells from different donors. Biochem. Biophys. Res. Commun. 325, 191–197 (2004).

    Article  Google Scholar 

  17. Rakoff-Nahoum, S., Paglino, J., Eslami-Varzaneh, F., Edberg, S. & Medzhitov, R. Recognition of commensal microflora by Toll-like receptors is required for intestinal homeostasis. Cell 118, 229–241 (2004).

    Article  CAS  Google Scholar 

  18. Magilavy, D.B. & Rothstein, J.L. Spontaneous production of tumor necrosis factor alpha by Kupffer cells of MRL/lpr mice. J. Exp. Med. 168, 789–794 (1988).

    Article  CAS  Google Scholar 

  19. Yata, Y. et al. DNase I-hypersensitive sites enhance alpha1(I) collagen gene expression in hepatic stellate cells. Hepatology 37, 267–276 (2003).

    Article  CAS  Google Scholar 

  20. Onichtchouk, D. et al. Silencing of TGF-β signalling by the pseudoreceptor Bambi. Nature 401, 480–485 (1999).

    Article  CAS  Google Scholar 

  21. Iimuro, Y. et al. NF-κB prevents apoptosis and liver dysfunction during liver regeneration. J. Clin. Invest. 101, 802–811 (1998).

    Article  CAS  Google Scholar 

  22. Maeda, S., Kamata, H., Luo, J.L., Leffert, H. & Karin, M. IKKbeta couples hepatocyte death to cytokine-driven compensatory proliferation that promotes chemical hepatocarcinogenesis. Cell 121, 977–990 (2005).

    Article  CAS  Google Scholar 

  23. Pikarsky, E. et al. NF-κB functions as a tumour promoter in inflammation-associated cancer. Nature 431, 461–466 (2004).

    Article  CAS  Google Scholar 

  24. Isogawa, M., Robek, M.D., Furuichi, Y. & Chisari, F.V. Toll-like receptor signaling inhibits hepatitis B virus replication in vivo. J. Virol. 79, 7269–7272 (2005).

    Article  CAS  Google Scholar 

  25. Izuishi, K. et al. Cutting edge: high-mobility group box 1 preconditioning protects against liver ischemia-reperfusion injury. J. Immunol. 176, 7154–7158 (2006).

    Article  CAS  Google Scholar 

  26. Lotze, M.T. & Tracey, K.J. High-mobility group box 1 protein (HMGB1): nuclear weapon in the immune arsenal. Nat. Rev. Immunol. 5, 331–342 (2005).

    Article  CAS  Google Scholar 

  27. Termeer, C. et al. Oligosaccharides of hyaluronan activate dendritic cells via Toll-like receptor 4. J. Exp. Med. 195, 99–111 (2002).

    Article  CAS  Google Scholar 

  28. Tsung, A. et al. The nuclear factor HMGB1 mediates hepatic injury after murine liver ischemia-reperfusion. J. Exp. Med. 201, 1135–1143 (2005).

    Article  CAS  Google Scholar 

  29. Isayama, F. et al. LPS signaling enhances hepatic fibrogenesis caused by experimental cholestasis in mice. Am. J. Physiol. Gastrointest. Liver Physiol. 290, G1318–G1328 (2006).

    Article  CAS  Google Scholar 

  30. Friedman, S.L. & Arthur, M.J. Activation of cultured rat hepatic lipocytes by Kupffer cell conditioned medium. Direct enhancement of matrix synthesis and stimulation of cell proliferation via induction of platelet-derived growth factor receptors. J. Clin. Invest. 84, 1780–1785 (1989).

    Article  CAS  Google Scholar 

  31. De Minicis, S. et al. Gene expression profiles during hepatic stellate cell activation in culture and in vivo. Gastroenterology 132, 1937–1946 (2007).

    Article  CAS  Google Scholar 

  32. Sasaki, T., Sasahira, T., Shimura, H., Ikeda, S. & Kuniyasu, H. Effect of Nma on growth inhibition by TGF-β in human gastric carcinoma cell lines. Oncol. Rep. 11, 1219–1223 (2004).

    CAS  PubMed  Google Scholar 

  33. Tsang, M. et al. Zebrafish nma is involved in TGF-β family signaling. Genesis 28, 47–57 (2000).

    Article  CAS  Google Scholar 

  34. Sekiya, T. et al. Identification of BMP and activin membrane-bound inhibitor (Bambi), an inhibitor of transforming growth factor-β signaling, as a target of the β-catenin pathway in colorectal tumor cells. J. Biol. Chem. 279, 6840–6846 (2004).

    Article  CAS  Google Scholar 

  35. Sekiya, T., Oda, T., Matsuura, K. & Akiyama, T. Transcriptional regulation of the TGF-β pseudoreceptor Bambi by TGF-β signaling. Biochem. Biophys. Res. Commun. 320, 680–684 (2004).

    Article  CAS  Google Scholar 

  36. Oakley, F. et al. Inhibition of inhibitor of κB kinases stimulates hepatic stellate cell apoptosis and accelerated recovery from rat liver fibrosis. Gastroenterology 128, 108–120 (2005).

    Article  CAS  Google Scholar 

  37. Thurman, R.G. et al. The role of gut-derived bacterial toxins and free radicals in alcohol-induced liver injury. J. Gastroenterol. Hepatol. 13 Suppl, S39–S50 (1998).

    Article  CAS  Google Scholar 

  38. Diehl, A.M., Li, Z.P., Lin, H.Z. & Yang, S.Q. Cytokines and the pathogenesis of non-alcoholic steatohepatitis. Gut 54, 303–306 (2005).

    Article  CAS  Google Scholar 

  39. Campbell, J.S. et al. Proinflammatory cytokine production in liver regeneration is Myd88-dependent, but independent of Cd14, Tlr2, and Tlr4. J. Immunol. 176, 2522–2528 (2006).

    Article  CAS  Google Scholar 

  40. Huang, H. et al. A 7 gene signature identifies the risk of developing cirrhosis in patients with chronic hepatitis C. Hepatology, 46, 297–306 (2007).

    Article  CAS  Google Scholar 

  41. Hoebe, K. et al. Identification of Lps2 as a key transducer of MyD88-independent TIR signalling. Nature 424, 743–748 (2003).

    Article  CAS  Google Scholar 

  42. Adachi, O. et al. Targeted disruption of the MyD88 gene results in loss of IL-1- and IL-18-mediated function. Immunity 9, 143–150 (1998).

    Article  CAS  Google Scholar 

  43. Kisseleva, T. et al. Bone marrow-derived fibrocytes participate in pathogenesis of liver fibrosis. J. Hepatol. 45, 429–438 (2006).

    Article  CAS  Google Scholar 

  44. Kennedy, D.W. & Abkowitz, J.L. Kinetics of central nervous system microglial and macrophage engraftment: analysis using a transgenic bone marrow transplantation model. Blood 90, 986–993 (1997).

    CAS  PubMed  Google Scholar 

  45. Van Rooijen, N. & Sanders, A. Liposome mediated depletion of macrophages: mechanism of action, preparation of liposomes and applications. J. Immunol. Methods 174, 83–93 (1994).

    Article  CAS  Google Scholar 

  46. Bataller, R. et al. NADPH oxidase signal transduces angiotensin II in hepatic stellate cells and is critical in hepatic fibrosis. J. Clin. Invest. 112, 1383–1394 (2003).

    Article  CAS  Google Scholar 

  47. Siegmund, S.V., Uchinami, H., Osawa, Y., Brenner, D.A. & Schwabe, R.F. Anandamide induces necrosis in primary hepatic stellate cells. Hepatology 41, 1085–1095 (2005).

    Article  CAS  Google Scholar 

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Acknowledgements

The authors thank S. Akira (Osaka University, Osaka, Japan) for providing MyD88-deficient mice, J. Massague (Memorial Sloan-Kettering Cancer Center, New York, NY, USA) for providing dominant-negative Bambi plasmid, T. Akiyama (University of Tokyo, Tokyo, Japan) for providing Bambi-reporter plasmid, Y. Zhang for assistance with microarray analysis and A. Mencin (Columbia University, New York, New York, USA) for assistance with mouse studies. This study was supported by a Research Scholar Award from the American Gastroenterological Association (to R.F.S.), grants from the Uehara Memorial Foundation and the American Liver Foundation (to E.S.), an Alimenti e Salute grant from the University of Ancona (to S.D.M.), US National Institutes of Health grant R01GM041804 (to D.A.B.) and the Austrian Science Foundation (to C.H.Ö.).

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Authors and Affiliations

  1. Department of Medicine, Columbia University, College of Physicians and Surgeons, New York, 10032, New York, USA

    Ekihiro Seki, Samuele De Minicis, Christoph H Österreicher, Johannes Kluwe, Yosuke Osawa & Robert F Schwabe

  2. Department of Medicine, University of California, San Diego, School of Medicine, La Jolla, 92093, California, USA

    Ekihiro Seki, Samuele De Minicis, Christoph H Österreicher & David A Brenner

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Contributions

E.S. performed animal surgery and fibrosis evaluation, cell isolations, cell-culture studies, reporter assays and real-time PCR, generated recombinant adenoviruses, created figures and contributed to the design of the study and the writing of the manuscript. S.D.M. performed cell isolations, real-time PCR and microarray studies. Y.O. generated recombinant adenoviruses. C.H.Ö. generated recombinant adenoviruses. J.K. performed animal surgeries and cell isolations, performed western blots and assisted in fibrosis evaluation and in manuscript preparation. D.A.B. contributed to the design of the study, experimental design and the writing of the manuscript. R.F.S. (the principal investigator) designed and coordinated the study, contributed to experimental setup, data analysis and interpretation, and drafted and edited the manuscript.

Corresponding author

Correspondence to Robert F Schwabe.

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Seki, E., De Minicis, S., Österreicher, C. et al. TLR4 enhances TGF-β signaling and hepatic fibrosis. Nat Med 13, 1324–1332 (2007). https://doi.org/10.1038/nm1663

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