Skip to main content
SpringerLink
Log in

Alarmin HMGB1 Is Released in the Small Intestine of Gnotobiotic Piglets Infected with Enteric Pathogens and Its Level in Plasma Reflects Severity of Sepsis

  • Published:
Journal of Clinical Immunology Aims and scope Submit manuscript

Abstract

Objectives

Alarmin high mobility group box 1 (HMGB1) is essential for correct DNA folding and transcription. It can be released from damaged cells or secreted by stimulated cells. HMGB1 has been detected in serum or plasma as a late marker of sepsis, but its suitability as a marker of sepsis has been disputed.

Methods

One-week-old germ-free piglets were orally infected/colonized with enteric bacterial pathogens (Salmonella Typhimurium or Escherichia coli O55) or with probiotic bacteria (E. coli Nissle 1917) for 24 h. The transcriptions of HMGB1, interleukin (IL)-8, tumor necrosis factor (TNF)-α, and IL-10 (quantitative reverse transcription and polymerase chain reaction), their protein levels (ELISA), and clinical state of the piglets (somnolence, anorexia, diarrhea, tachycardia, tachypnea, and tremor) were estimated.

Results

The piglets infected with enteric pathogens suffered from infections. HMGB1 was transcribed in the terminal ileum constitutively, regardless of any bacterial presence. In contrast, the transcription of cytokines was upregulated by virulent bacteria. HMGB1, IL-8, and TNF-α levels in the ileum were increased by both enteric pathogens, while IL-10 levels increased in E. coli O55-infected piglets only. HMGB1 significantly increased in the plasma of piglets infected with virulent E. coli only, but cytokine levels were in most cases increased by both virulent bacteria. HMGB1 and cytokine levels in ileum lavages and plasma of piglets colonized with probiotic E. coli remained comparable to those of the non-stimulated germ-free piglets.

Conclusion

The local and systemic expression of HMGB1, its relationship to the inflammatory cytokines, and clinical findings showed HMGB1 as a suitable marker of severity of sepsis in the gnotobiotic piglet infection model.

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
Fig. 6

Similar content being viewed by others

Abbreviations

EcN:

E. coli Nissle 1917

HMGB1:

High mobility group box 1

IL:

Interleukin

LT2:

LT2 strain of Salmonella enterica serovar Typhimurium

O55:

E. coli O55

MAMPs:

Microbe-associated molecular patterns

PAMPs:

Pathogen-associated molecular patterns

TNF:

Tumor necrosis factor

References

  1. Bianchi ME. DAMPs, PAMPs and alarmins: all we need to know about danger. J Leukoc Biol. 2007;81(1):1–5.

    Article  PubMed  CAS  Google Scholar 

  2. Kawai T, Akira S. The role of pattern-recognition receptors in innate immunity: update on Toll-like receptors. Nat Immunol. 2010;11(5):373–84.

    Article  PubMed  CAS  Google Scholar 

  3. Matzinger P. The danger model: a renewed sense of self. Science. 2002;296(5566):301–5.

    Article  PubMed  CAS  Google Scholar 

  4. Goodwin GH, Sanders C, Johns EW. A new group of chromatin-associated proteins with a high content of acidic and basic amino acids. Eur J Biochem. 1973;38(1):14–9.

    Article  PubMed  CAS  Google Scholar 

  5. Weir HM, Kraulis PJ, Hill CS, Raine AR, Laue ED, Thomas JO. Structure of the HMG box motif in the B-domain of HMG1. EMBO J. 1993;12(4):1311–9.

    PubMed  CAS  Google Scholar 

  6. Wang H, Bloom O, Zhang M, Vishnubhakat JM, Ombrellino M, Che J, et al. HMG-1 as a late mediator of endotoxin lethality in mice. Science. 1999;285(5425):248–51.

    Article  PubMed  CAS  Google Scholar 

  7. Li J, Kokkola R, Tabibzadeh S, Yang R, Ochani M, Qiang X, et al. Structural basis for the proinflammatory cytokine activity of high mobility group box 1. Mol Med. 2003;9(1–2):37–45.

    PubMed  CAS  Google Scholar 

  8. Yang H, Ochani M, Li J, Qiang X, Tanovic M, Harris HE, et al. Reversing established sepsis with antagonists of endogenous high-mobility group box 1. Proc Natl Acad Sci U S A. 2004;101(1):296–301.

    Article  PubMed  CAS  Google Scholar 

  9. Scaffidi P, Misteli T, Bianchi ME. Release of chromatin protein HMGB1 by necrotic cells triggers inflammation. Nature. 2002;418(6894):191–5.

    Article  PubMed  CAS  Google Scholar 

  10. Lantos J, Foldi V, Roth E, Weber G, Bogar L, Csontos C. Burn trauma induces early HMGB1 release in patients: its correlation with cytokines. Shock. 2010;33(6):562–7.

    PubMed  CAS  Google Scholar 

  11. Peltz ED, Moore EE, Eckels PC, Damle SS, Tsuruta Y, Johnson JL, et al. HMGB1 is markedly elevated within 6 h of mechanical trauma in humans. Shock. 2009;32(1):17–22.

    Article  PubMed  CAS  Google Scholar 

  12. Cohen MJ, Brohi K, Calfee CS, Rahn P, Chesebro BB, Christiaans SC, et al. Early release of high mobility group box nuclear protein 1 after severe trauma in humans: role of injury severity and tissue hypoperfusion. Crit Care. 2009;13(6):R174.

    Article  PubMed  Google Scholar 

  13. Angus DC, Linde-Zwirble WT, Lidicker J, Clermont G, Carcillo J, Pinsky MR. Epidemiology of severe sepsis in the United States: analysis of incidence, outcome, and associated costs of care. Crit Care Med. 2001;29(7):1303–10.

    Article  PubMed  CAS  Google Scholar 

  14. Palumbo R, Galvez BG, Pusterla T, De MF, Cossu G, Marcu KB, et al. Cells migrating to sites of tissue damage in response to the danger signal HMGB1 require NF-kappaB activation. J Cell Biol. 2007;179(1):33–40.

    Article  PubMed  CAS  Google Scholar 

  15. Bone RC, Balk RA, Cerra FB, Dellinger RP, Fein AM, Knaus WA, et al. Definitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis. The ACCP/SCCM Consensus Conference Committee. American College of Chest Physicians/Society of Critical Care Medicine. Chest. 1992;101(6):1644–55.

    Article  PubMed  CAS  Google Scholar 

  16. Cai B, Deitch EA, Ulloa L. Novel insights for systemic inflammation in sepsis and hemorrhage. Mediat Inflamm. 2010;2010:642462.

    Google Scholar 

  17. Castellheim A, Brekke OL, Espevik T, Harboe M, Mollnes TE. Innate immune responses to danger signals in systemic inflammatory response syndrome and sepsis. Scand J Immunol. 2009;69(6):479–91.

    Article  PubMed  CAS  Google Scholar 

  18. Rittirsch D, Flierl MA, Ward PA. Harmful molecular mechanisms in sepsis. Nat Rev Immunol. 2008;8(10):776–87.

    Article  PubMed  CAS  Google Scholar 

  19. Cinel I, Opal SM. Molecular biology of inflammation and sepsis: a primer. Crit Care Med. 2009;37(1):291–304.

    Article  PubMed  CAS  Google Scholar 

  20. Yang H, Tracey KJ. Targeting HMGB1 in inflammation. Biochim Biophys Acta. 2010;1799(1–2):149–56.

    PubMed  CAS  Google Scholar 

  21. Kato S, Hussein MH, Kakita H, Goto T, Daoud GA, Kato T, et al. Edaravone, a novel free radical scavenger, reduces high-mobility group box 1 and prolongs survival in a neonatal sepsis model. Shock. 2009;32(6):586–92.

    Article  PubMed  CAS  Google Scholar 

  22. Rothkotter HJ, Sowa E, Pabst R. The pig as a model of developmental immunology. Hum Exp Toxicol. 2002;21(9–10):533–6.

    Article  PubMed  CAS  Google Scholar 

  23. Rothkotter HJ. Anatomical particularities of the porcine immune system—a physician’s view. Dev Comp Immunol. 2009;33(3):267–72.

    Article  PubMed  Google Scholar 

  24. Castellheim A, Thorgersen EB, Hellerud BC, Pharo A, Johansen HT, Brosstad F, et al. New biomarkers in an acute model of live Escherichia coli-induced sepsis in pigs. Scand J Immunol. 2008;68(1):75–84.

    Article  PubMed  CAS  Google Scholar 

  25. Nielsen EW, Hellerud BC, Thorgersen EB, Castellheim A, Pharo A, Lindstad J, et al. A new dynamic porcine model of meningococcal shock. Shock. 2009;32(3):302–9.

    Article  PubMed  Google Scholar 

  26. Thorgersen EB, Hellerud BC, Nielsen EW, Barratt-Due A, Fure H, Lindstad JK, et al. CD14 inhibition efficiently attenuates early inflammatory and hemostatic responses in Escherichia coli sepsis in pigs. FASEB J. 2010;24(3):712–22.

    Article  PubMed  CAS  Google Scholar 

  27. Becker KL, Nylen ES, Snider RH, Muller B, White JC. Immunoneutralization of procalcitonin as therapy of sepsis. J Endotoxin Res. 2003;9(6):367–74.

    PubMed  CAS  Google Scholar 

  28. Mandel L, Travnicek J. The minipig as a model in gnotobiology. Nahrung. 1987;31(5–6):613–8.

    Article  PubMed  CAS  Google Scholar 

  29. Trebichavsky I, Schulze J, Dlabac V, Cukrowska B, Tlaskalova-Hogenova H, Rehakova Z. Salmonellosis: lessons drawn from a germ-free pig model. Folia Microbiol (Praha). 1998;43(6):697–701.

    Article  CAS  Google Scholar 

  30. Schultz M. Escherichia coli. In: Versalovic J, Wilson M, editors. Therapeutic microbiology: probiotics and related strategies. Washington, DC: ASM; 2008. p. 83–96.

    Google Scholar 

  31. Zambon M, Ceola M, Almeida-de-Castro R, Gullo A, Vincent JL. Implementation of the surviving sepsis campaign guidelines for severe sepsis and septic shock: we could go faster. J Crit Care. 2008;23(4):455–60.

    Article  PubMed  Google Scholar 

  32. Lever A, Mackenzie I. Sepsis: definition, epidemiology, and diagnosis. BMJ. 2007;335(7625):879–83.

    Article  PubMed  CAS  Google Scholar 

  33. Marshall JC, Reinhart K. Biomarkers of sepsis. Crit Care Med. 2009;37(7):2290–8.

    Article  PubMed  CAS  Google Scholar 

  34. Pierrakos C, Vincent JL. Sepsis biomarkers: a review. Crit Care. 2010;14(1):R15.

    Article  PubMed  Google Scholar 

  35. Horn DL, Morrison DC, Opal SM, Silverstein R, Visvanathan K, Zabriskie JB. What are the microbial components implicated in the pathogenesis of sepsis? Report on a symposium. Clin Infect Dis. 2000;31(4):851–8.

    Article  PubMed  CAS  Google Scholar 

  36. Poli-de-Figueiredo LF, Garrido AG, Nakagawa N, Sannomiya P. Experimental models of sepsis and their clinical relevance. Shock. 2008;30 Suppl:153–9.

    Google Scholar 

  37. Kato T, Hussein MH, Sugiura T, Suzuki S, Fukuda S, Tanaka T, et al. Development and characterization of a novel porcine model of neonatal sepsis. Shock. 2004;21(4):329–35.

    Article  PubMed  Google Scholar 

  38. Hussein MH, Kato T, Sugiura T, Daoud GA, Suzuki S, Fukuda S, et al. Effect of hemoperfusion using polymyxin B-immobilized fiber on IL-6, HMGB-1, and IFN gamma in a neonatal sepsis model. Pediatr Res. 2005;58(2):309–14.

    Article  PubMed  CAS  Google Scholar 

  39. Goto T, Hussein MH, Kato S, Daoud GA, Kato T, Kakita H, et al. Endothelin receptor antagonist attenuates inflammatory response and prolongs the survival time in a neonatal sepsis model. Intensive Care Med. 2010;36(12):2132–9.

    Article  PubMed  CAS  Google Scholar 

  40. Smith K, McCoy KD, Macpherson AJ. Use of axenic animals in studying the adaptation of mammals to their commensal intestinal microbiota. Semin Immunol. 2007;19(2):59–69.

    Article  PubMed  CAS  Google Scholar 

  41. van der Waaij D. Colonization resistance of the digestive tract—mechanism and clinical consequences. Nahrung. 1987;31(5–6):507–17.

    Article  PubMed  Google Scholar 

  42. Grozdanov L, Zahringer U, Blum-Oehler G, Brade L, Henne A, Knirel YA, et al. A single nucleotide exchange in the wzy gene is responsible for the semirough O6 lipopolysaccharide phenotype and serum sensitivity of Escherichia coli strain Nissle 1917. J Bacteriol. 2002;184(21):5912–25.

    Article  PubMed  CAS  Google Scholar 

  43. Pachot A, Monneret G, Voirin N, Leissner P, Venet F, Bohe J, et al. Longitudinal study of cytokine and immune transcription factor mRNA expression in septic shock. Clin Immunol. 2005;114(1):61–9.

    Article  PubMed  CAS  Google Scholar 

  44. Calogero S, Grassi F, Aguzzi A, Voigtlander T, Ferrier P, Ferrari S, et al. The lack of chromosomal protein Hmg1 does not disrupt cell growth but causes lethal hypoglycaemia in newborn mice. Nat Genet. 1999;22(3):276–80.

    Article  PubMed  CAS  Google Scholar 

  45. Splichal I, Trebichavsky I, Muneta Y, Mori Y. Early cytokine response of gnotobiotic piglets to Salmonella enterica serotype Typhimurium. Vet Res. 2002;33(3):291–7.

    Article  PubMed  CAS  Google Scholar 

  46. Splichalova A, Trebichavsky I, Rada V, Vlkova E, Sonnenborn U, Splichal I. Interference of Bifidobacterium choerinum or E. coli Nissle 1917 with Salmonella Typhimurium in gnotobiotic piglets correlates with cytokine patterns in blood and intestine. Clin Exp Immunol. 2011. doi:10.1111/j.1365-2249.2010.04283.x.

    PubMed  Google Scholar 

  47. Skjolaas KA, Burkey TE, Dritz SS, Minton JE. Effects of Salmonella enterica serovars Typhimurium (ST) and Choleraesuis (SC) on chemokine and cytokine expression in swine ileum and jejunal epithelial cells. Vet Immunol Immunopathol. 2006;111(3–4):199–209.

    Article  PubMed  CAS  Google Scholar 

  48. Girard F, Oswald IP, Taranu I, Helie P, Appleyard GD, Harel J, et al. Host immune status influences the development of attaching and effacing lesions in weaned pigs. Infect Immun. 2005;73(9):5514–23.

    Article  PubMed  CAS  Google Scholar 

  49. Jesmok G, Lindsey C, Duerr M, Fournel M, Emerson Jr T. Efficacy of monoclonal antibody against human recombinant tumor necrosis factor in E. coli-challenged swine. Am J Pathol. 1992;141(5):1197–207.

    PubMed  CAS  Google Scholar 

  50. Grozdanov L, Raasch C, Schulze J, Sonnenborn U, Gottschalk G, Hacker J, et al. Analysis of the genome structure of the nonpathogenic probiotic Escherichia coli strain Nissle 1917. J Bacteriol. 2004;186(16):5432–41.

    Article  PubMed  CAS  Google Scholar 

  51. Miller I, Cerna J, Travnicek J, Rejnek J, Kruml J. The role of immune pig colostrum, serum and immunoglobulins IgG, IgM, and IgA, in local intestinal immunity against enterotoxic strain in Escherichia coli O55 in germfree piglets. Folia Microbiol (Praha). 1975;20(5):433–8.

    Article  CAS  Google Scholar 

  52. Trebichavsky I. Early immunological events in germ-free piglets monoassociated with nonpathogenic or virulent strain of Salmonella typhimurium. Vet Med-Czech. 2000;45:125–8.

    Google Scholar 

  53. Tzipori S, Gibson R, Montanaro J. Nature and distribution of mucosal lesions associated with enteropathogenic and enterohemorrhagic Escherichia coli in piglets and the role of plasmid-mediated factors. Infect Immun. 1989;57(4):1142–50.

    PubMed  CAS  Google Scholar 

  54. Gao H, Leaver SK, Burke-Gaffney A, Finney SJ. Severe sepsis and Toll-like receptors. Semin Immunopathol. 2008;30(1):29–40.

    Article  PubMed  CAS  Google Scholar 

  55. Bambou JC, Giraud A, Menard S, Begue B, Rakotobe S, Heyman M, et al. In vitro and ex vivo activation of the TLR5 signaling pathway in intestinal epithelial cells by a commensal Escherichia coli strain. J Biol Chem. 2004;279(41):42984–92.

    Article  PubMed  CAS  Google Scholar 

  56. Foster N, Lovell MA, Marston KL, Hulme SD, Frost AJ, Bland P, et al. Rapid protection of gnotobiotic pigs against experimental salmonellosis following induction of polymorphonuclear leukocytes by avirulent Salmonella enterica. Infect Immun. 2003;71(4):2182–91.

    Article  PubMed  CAS  Google Scholar 

  57. Splichal I, Trebichavsky I, Splichalova A, Barrow PA. Protection of gnotobiotic pigs against Salmonella enterica serotype Typhimurium by rough mutant of the same serotype is accompanied by the change of local and systemic cytokine response. Vet Immunol Immunopathol. 2005;103(3–4):155–61.

    Article  PubMed  CAS  Google Scholar 

  58. Angus DC, Yang L, Kong L, Kellum JA, Delude RL, Tracey KJ, et al. Circulating high-mobility group box 1 (HMGB1) concentrations are elevated in both uncomplicated pneumonia and pneumonia with severe sepsis. Crit Care Med. 2007;35(4):1061–7.

    Article  PubMed  Google Scholar 

  59. Azevedo MS, Yuan L, Pouly S, Gonzales AM, Jeong KI, Nguyen TV, et al. Cytokine responses in gnotobiotic pigs after infection with virulent or attenuated human rotavirus. J Virol. 2006;80(1):372–82.

    Article  PubMed  CAS  Google Scholar 

  60. Splichal I, Muneta Y, Mori Y, Takahashi E. Development and application of a pig IL-8 ELISA detection system. J Immunoassay Immunochem. 2003;24(2):219–32.

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

This work was supported by the grant ME915 of the Ministry of Education, Youth and Sport of the Czech Republic and by the Institutional Research Concept AV0Z50200510 of the Institute of Microbiology of the ASCR. Technical assistance of Marie Zahradnickova, Hana Sychrovska, Jarmila Jarkovska, and Jana Machova is greatly appreciated. We are grateful to Dr. R. Alexander for her language correction of the manuscript.

Conflicts of Interest

The authors have no conflicting financial interests.

Author information

Authors and Affiliations

  1. Department of Immunology and Gnotobiology, Institute of Microbiology of the Academy of Sciences of the Czech Republic, Doly 183, 549 22, Novy Hradek, Czech Republic

    Alla Splichalova, Igor Splichal, Petra Chmelarova & Ilja Trebichavsky

Authors
  1. Alla Splichalova
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Igor Splichal
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Petra Chmelarova
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ilja Trebichavsky
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Igor Splichal.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Splichalova, A., Splichal, I., Chmelarova, P. et al. Alarmin HMGB1 Is Released in the Small Intestine of Gnotobiotic Piglets Infected with Enteric Pathogens and Its Level in Plasma Reflects Severity of Sepsis. J Clin Immunol 31, 488–497 (2011). https://doi.org/10.1007/s10875-010-9505-3

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10875-010-9505-3

Keywords

Search

Navigation