Science in China Series C: Life Sciences

, Volume 52, Issue 2, pp 147–154 | Cite as

Hyaluronic acid fragments evoke Kupffer cells via TLR4 signaling pathway

  • JinXiang Zhang
  • Hui Wang
  • Qing Xiao
  • HuiFang Liang
  • ZhuoYa Li
  • ChunFang Jiang
  • HeShui Wu
  • QiChang Zheng
In Memoriam: Professor Ray Wu


Kupffer cells, expressing toll-like receptor 4 (TLR4), play a central role in hepatic ischemia/reperfusion (I/R) injury. Hyaluronic acid (HA) fragments, degradative products of high-molecular-weight HA (HMW-HA), acquire the ability to activate immune cells under inflammatory conditions. Here we investigated whether HA fragments could activate Kupffer cells and analyzed the underlying mechanism. Kupffer cells were isolated from wild-type mice (WT, C3H/HeN) and TLR4 mutant mice (C3H/HeJ) and HA fragments were produced by the methods of enzyme digestion and chromatography. Then Kupffer cells were stimulated by HA fragments or other control stimuli. The activation of Kupffer cells was estimated as the release of pro-inflammatory cytokines. The activation of p38 MAPK pathway of Kupffer cells was checked and blocking experiments were done as well. The results indicated that HA fragments acquired the ability to activate Kupffer cells in vitro, which was TLR4 dependent and not due to contamination of lipopolysaccharide. Experiments of p38 MAPK kinase inhibition by SB-203580 verified p38 MAPK was required in HA fragments induced Kupffer cells activation. This suggests that HA fragments, degradative products of one of the major glycosaminoglycans of the extracellular matrix, play critical roles in Kupffer cell activation mediated by TLR4 signaling pathway, which is, at least partially, dependent on p38 MAPK activation.


Kupffer cells hyaluronic acid fragments toll-like receptor 4 p38 MAPK lipoplysaccharide reperfusion injury liver 


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  1. 1.
    Kupiec-Weglinski J W, Busuttil R W. Ischemia and reperfusion injury in liver transplantation. Transplant Proc, 2005, 37: 1653–1656 15919422, 10.1016/j.transproceed.2005.03.134, 1:STN:280:DC%2BD2M3nslWltQ%3D%3DCrossRefPubMedGoogle Scholar
  2. 2.
    Fondevila C, Busuttil R W, Kupiec-Weglinski J W. Hepatic ischemia/reperfusion injury—a fresh look. Exp Mol Pathol, 2003, 74: 86–93 12710939, 10.1016/S0014-4800(03)00008-X, 1:CAS:528:DC%2BD3sXjtFWms7g%3DCrossRefPubMedGoogle Scholar
  3. 3.
    Tsung A, Hoffman R A, Izuishi K, et al. Hepatic ischemia/reperfusion injury involves functional TLR4 signaling in nonparenchymal cells. J Immunol, 2005, 175: 7661–7668 16301676, 1:CAS:528:DC%2BD2MXht1ent7rICrossRefPubMedGoogle Scholar
  4. 4.
    Zhai Y, Shen X D, O’Connell R, et al. Cutting edge: TLR4 activation mediates liver ischemia/reperfusion inflammatory response via IFN regulatory factor 3-dependent MyD88-independent pathway. J Immunol, 2004, 173: 7115–7119 15585830, 1:CAS:528:DC%2BD2cXhtVCqtbfFCrossRefPubMedGoogle Scholar
  5. 5.
    Yu M, Wang H, Ding A, et al. HMGB1 signals through toll-like receptor (TLR) 4 and TLR2. Shock, 2006, 26: 174–179 16878026, 10.1097/01.shk.0000225404.51320.82, 1:CAS:528:DC%2BD28Xot1amtrg%3DCrossRefPubMedGoogle Scholar
  6. 6.
    Vabulas R M, Ahmad-Nejad P, Ghose S, et al. HSP70 as endogenous stimulus of the Toll/interleukin-1 receptor signal pathway. J Biol Chem, 2002, 277: 15107–15112 11842086, 10.1074/jbc.M111204200, 1:CAS:528:DC%2BD38XjslSgtLs%3DCrossRefPubMedGoogle Scholar
  7. 7.
    Kol A, Lichtman A H, Finberg R W, et al. Cutting edge: heat shock protein (HSP) 60 activates the innate immune response: CD14 is an essential receptor for HSP60 activation of mononuclear cells. J Immunol, 2000, 164: 13–17 10604986, 1:CAS:528:DC%2BD3cXisVKgCrossRefPubMedGoogle Scholar
  8. 8.
    Sun D, Chen D, Du B, et al. Heat shock response inhibits NF-kappa B activation and cytokine production in murine Kupffer cells. J Surg Res, 2005, 129: 114–121 16243048, 10.1016/j.jss.2005.05.028, 1:CAS:528:DC%2BD2MXhtFCjsb%2FMCrossRefPubMedGoogle Scholar
  9. 9.
    Kim J Y, Park J S, Strassheim D, et al. HMGB1 contributes to the development of acute lung injury after hemorrhage. Am J Physiol Lung Cell Mol Physiol, 2005, 288: L958–965 15640285, 10.1152/ajplung.00359.2004, 1:CAS:528:DC%2BD2MXktlSntLY%3DCrossRefPubMedGoogle Scholar
  10. 10.
    Powell J D, Horton M R. Threat matrix: low-molecular-weight hyaluronan (HA) as a danger signal. Immunol Res, 2005, 31: 207–218 15888912, 10.1385/IR:31:3:207, 1:CAS:528:DC%2BD2MXkslymtbo%3DCrossRefPubMedGoogle Scholar
  11. 11.
    McKee C M, Penno M B, Cowman M, et al. Hyaluronan (HA) fragments induce chemokine gene expression in alveolar macrophages: The role of HA size and CD44. J Clin Invest, 1996, 98: 2403–2413 8941660, 10.1172/JCI119054, 1:CAS:528:DyaK28XntVOhtr0%3DCrossRefPubMedCentralPubMedGoogle Scholar
  12. 12.
    Taylor K R, Yamasaki k, Radek K A, et al. Recognition of hyaluronan released in sterile injury involves a unique receptor complex dependent on Toll-like receptor 4, CD44, and MD-2. J Biol Chem, 2007, 282: 18265–18275 17400552, 10.1074/jbc.M606352200, 1:CAS:528:DC%2BD2sXmsVOlu7k%3DCrossRefPubMedGoogle Scholar
  13. 13.
    Voelcker V, Gebhardt C, Averbeck M, et al. Hyaluronan fragments induce cytokine and metalloprotease upregulation in human melanoma cells in part by signalling via TLR4. Exp Dermatol, 2008, 17: 100–107 18031543, 10.1111/j.1600-0625.2007.00638.x, 1:CAS:528:DC%2BD1cXjs1Cit7w%3DCrossRefPubMedGoogle Scholar
  14. 14.
    Cantor J O, Nadkarni P P. Hyaluronan: the jekyll and hyde molecule. Inflamm Allergy Drug Targets, 2006, 5: 257–260 17168797, 10.2174/187152806779010936, 1:CAS:528:DC%2BD28XhtlSjtb%2FECrossRefPubMedGoogle Scholar
  15. 15.
    Laurent T C, Fraser J R. Hyaluronan. FASEB J, 1992, 6: 2397–2404 1:CAS:528:DyaK38XltVKgu7o%3DGoogle Scholar
  16. 16.
    Reinders M E, van Wagensveld B A, van Gulik T M, et al. Hyaluronic acid uptake in the assessment of sinusoidal endothelial cell damage after cold storage and normothermic reperfusion of rat livers. Transpl Int, 1996, 9: 446–453 8875786, 10.1111/j.1432-2277.1996.tb00987.x, 1:CAS:528:DyaK2sXjtFGqsg%3D%3DCrossRefPubMedGoogle Scholar
  17. 17.
    Jeyabalan G, Tsung A, Billiar T R. Linking proximal and downstream signalling events in hepatic ischaemia/reperfusion injury. Biochem Soc Trans, 2006, 34: 957–959 17052236, 10.1042/BST0340957, 1:CAS:528:DC%2BD28XhtFWnsb3ICrossRefPubMedGoogle Scholar
  18. 18.
    Munthe-Kaas A C, Berg T, Seglen P O, et al. Mass isolation and culture of rat Kupffer cells. J Exp Med, 1975, 141: 1–10 1090696, 10.1084/jem.141.1.1, 1:STN:280:DyaE2M7hsFykug%3D%3DCrossRefPubMedGoogle Scholar
  19. 19.
    Termeer C C, Hennies J, Voith U, et al. Oligosaccharides of hyaluronan are potent activators of dendritic cells. J Immunol, 2000, 165: 1863–1870 10925265, 1:CAS:528:DC%2BD3cXls1Kmsrg%3DCrossRefPubMedGoogle Scholar
  20. 20.
    Jaeschke H. Molecular mechanisms of hepatic ischemia-reperfusion injury and preconditioning. Am J Physiol Gastrointest Liver Physiol, 2003, 284: G15–26 12488232, 1:CAS:528:DC%2BD3sXntVSgug%3D%3DCrossRefPubMedGoogle Scholar
  21. 21.
    Wang H, Li Z Y, Wu H S, et al. Endogenous danger signals trigger hepatic ischemia/reperfusion injury through toll-like receptor 4/nuclear factor-kappa B pathway. Chin Med J (Engl), 2007, 120: 509–514 1:CAS:528:DC%2BD2sXkvVKqtrs%3DGoogle Scholar
  22. 22.
    Teoh N C, Farrell G C. Hepatic ischemia reperfusion injury: Pathogenic mechanisms and basis for hepatoprotection. J Gastroenterol Hepatol, 2003, 18: 891–902 12859717, 10.1046/j.1440-1746.2003.03056.x, 1:CAS:528:DC%2BD3sXnvFSlurk%3DCrossRefPubMedGoogle Scholar
  23. 23.
    Nanashima A, Yamaguchi H, Tanaka K, et al. Preoperative serum hyaluronic acid level as a good predictor of posthepatectomy complications. Surg Today, 2004, 34: 913–919 15526125, 10.1007/s00595-004-2845-yCrossRefPubMedGoogle Scholar
  24. 24.
    Rockey D C, Chung J J, McKee C M, et al. Stimulation of inducible nitric oxide synthase in rat liver by hyaluronan fragments. Hepatology, 1998, 27: 86–92 9425922, 10.1002/hep.510270115, 1:CAS:528:DyaK1cXlvVeqsg%3D%3DCrossRefPubMedGoogle Scholar
  25. 25.
    Mummert M E. Immunologic roles of hyaluronan. Immunol Res, 2005, 31: 189–206 15888911, 10.1385/IR:31:3:189, 1:CAS:528:DC%2BD2MXkslymtb0%3DCrossRefPubMedGoogle Scholar
  26. 26.
    Soltés L, Mendichi R, Kogan G, et al. Degradative Action of Reactive Oxygen Species on Hyaluronan. Biomacromolecules, 2006, 7: 659–668 16529395, 10.1021/bm050867vCrossRefPubMedGoogle Scholar
  27. 27.
    Termeer C, Benedix F, Sleeman J, et al. Oligosaccharides of Hyaluronan Activate Dendritic Cells via Toll-like Receptor 4. J Exp Med, 2002, 195: 99–111 11781369, 10.1084/jem.20001858, 1:CAS:528:DC%2BD38XjslOjtw%3D%3DCrossRefPubMedCentralPubMedGoogle Scholar
  28. 28.
    Beg A A. Endogenous ligands of Toll-like receptors: implications for regulating inflammatory and immune responses. Trends Immunol, 2002, 23: 509–512 12401394, 10.1016/S1471-4906(02)02317-7, 1:CAS:528:DC%2BD38XotVKksbs%3DCrossRefPubMedGoogle Scholar
  29. 29.
    Tsan M F, Gao B. Endogenous ligands of Toll-like receptors. J Leukoc Biol, 2004, 76: 514–519 15178705, 10.1189/jlb.0304127, 1:CAS:528:DC%2BD2cXnsVaksrk%3DCrossRefPubMedGoogle Scholar
  30. 30.
    Maung A A, Fujimi S, Miller M L, et al. Enhanced TLR4 reactivity following injury is mediated by increased p38 activation. J Leukoc Biol, 2005, 78: 565–573 15857937, 10.1189/jlb.1204698, 1:CAS:528:DC%2BD2MXntFCjsbs%3DCrossRefPubMedGoogle Scholar
  31. 31.
    Takeuchi O, Akira S. Toll-like receptors: their physiological role and signal transduction system. Int Immunopharmacol, 2001, 1: 625–635 11357875, 10.1016/S1567-5769(01)00010-8, 1:CAS:528:DC%2BD3MXit1OhsLw%3DCrossRefPubMedGoogle Scholar
  32. 32.
    Wolf D, Schumann J, Koerber K, et al. Low-molecular-weight hyaluronic acid induces nuclear factor-kappa B-dependent resistance against tumor necrosis factor alpha-mediated liver injury in mice. Hepatology, 2001, 34: 535–547 11526540, 10.1053/jhep.2001.27218, 1:CAS:528:DC%2BD3MXmvV2kt7w%3DCrossRefPubMedGoogle Scholar

Copyright information

© Science in China Press and Springer-Verlag GmbH 2009

Authors and Affiliations

  • JinXiang Zhang
    • 1
  • Hui Wang
    • 2
  • Qing Xiao
    • 3
  • HuiFang Liang
    • 4
  • ZhuoYa Li
    • 4
  • ChunFang Jiang
    • 1
  • HeShui Wu
    • 5
  • QiChang Zheng
    • 5
  1. 1.Department of Emergency SurgeryUnion Hospital Affiliated to Huazhong University of Science and TechnologyWuhanChina
  2. 2.Department of Medical GeneticsTongji Medical College Affiliated to Huazhong University of Science and TechnologyWuhanChina
  3. 3.Department of OphthalmologyUnion Hospital Affiliated to Huazhong University of Science and TechnologyWuhanChina
  4. 4.Department of Medical ImmunologyTongji Medical College Affiliated to Huazhong University of Science and TechnologyWuhanChina
  5. 5.Department of General SurgeryUnion Hospital Affiliated to Huazhong University of Science and TechnologyWuhanChina

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