Skip to main content

Advertisement

SpringerLink
Log in

Human β-defensin-2 increases cholinergic response in colon epithelium

  • Transport Physiology
  • Published:
Pflügers Archiv - European Journal of Physiology Aims and scope Submit manuscript

Abstract

The human β-defensin-2 (hBD-2) is expressed in epithelial cells of skin and respiratory and gastrointestinal tracts. Defensins are arginine-rich small cationic peptides with six intramolecular disulfide bonds and are antimicrobially active against a broad spectrum of pathogens. In addition, they have cytokine-like immunomodulatory properties. We hypothesized that hBD-2 also might influence epithelial cells themselves, thereby altering fluid composition in the gastrointestinal tract. We therefore tested its impact on electrogenic ion transport properties of distal colon in Ussing chamber experiments. Application of hBD-2 did not affect transepithelial voltage or resistance in cAMP-stimulated distal colon. However, it increased cholinergic Ca2+-dependent Cl− secretion. After 20 min of incubation with hBD-2, the effect of carbachol (CCh) on the equivalent short circuit current (I′sc) was enhanced twofold compared to vehicle-treated colon. Modulation of Ca2+ signaling by hBD-2 was validated by Fura-2 measurements in human colon carcinoma HT29 cells. Twenty-minute incubation with hBD-2 increased the CCh-induced Ca2+ transient by 20–30% compared to either vehicle-treated cells or cells treated with the defensins hBD-1, hBD-3, or HD-5. This effect was concentration-dependent, with an EC50 of 0.043 µg/ml, and still present in the absence of extracellular Ca2+. Also, the ionomycin-induced Ca2+ transient was increased by hBD-2 treatment. We conclude that hBD-2 facilitates cholinergic Ca2+-regulated epithelial Cl− secretion. These findings contribute to the concept of a specific interaction of antimicrobial peptides with epithelial function.

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

References

  1. Ballard ST, Spadafora D (2007) Fluid secretion by submucosal glands of the tracheobronchial airways. Respir Physiol Neurobiol 159:271–277

    Article  CAS  PubMed  Google Scholar 

  2. Bals R, Wang X, Meegalla RL, Wattler S, Weiner DJ, Nehls MC, Wilson JM (1999) Mouse beta-defensin 3 is an inducible antimicrobial peptide expressed in the epithelia of multiple organs. Infect Immun 67:3542–3547

    CAS  PubMed  Google Scholar 

  3. Bals R, Wang X, Wu Z, Freeman T, Bafna V, Zasloff M, Wilson JM (1998) Human beta-defensin 2 is a salt-sensitive peptide antibiotic expressed in human lung. J Clin Invest 102:874–880

    Article  CAS  PubMed  Google Scholar 

  4. Biragyn A, Ruffini PA, Leifer CA, Klyushnenkova E, Shakhov A, Chertov O, Shirakawa AK, Farber JM, Segal DM, Oppenheim JJ, Kwak LW (2002) Toll-like receptor 4-dependent activation of dendritic cells by beta-defensin 2 1. Science 298:1025–1029

    Article  CAS  PubMed  Google Scholar 

  5. Bleich M, Ecke D, Schwartz B, Fraser G, Greger R (1997) Effects of the carcinogen dimethylhydrazine (DMH) on the function of rat colonic crypts. Pflugers Arch 433:254–259

    Article  CAS  PubMed  Google Scholar 

  6. Drin G, Scarlata S (2007) Stimulation of phospholipase Cbeta by membrane interactions interdomain movement, and G protein binding—how many ways can you activate an enzyme? Cell Signal 19:1383–1392

    Article  CAS  PubMed  Google Scholar 

  7. Ecke D, Bleich M, Schwartz B, Fraser G, Greger R (1996) The ion conductances of colonic crypts from dexamethasone-treated rats. Pflugers Arch 431:419–426

    Article  CAS  PubMed  Google Scholar 

  8. Field M (2003) Intestinal ion transport and the pathophysiology of diarrhea. J Clin Invest 111:931–943

    CAS  PubMed  Google Scholar 

  9. Ganz T (2003) Defensins: antimicrobial peptides of innate immunity 2. Nat Rev Immunol 3:710–720

    Article  CAS  PubMed  Google Scholar 

  10. Goldman MJ, Anderson GM, Stolzenberg ED, Kari UP, Zasloff M, Wilson JM (1997) Human beta-defensin-1 is a salt-sensitive antibiotic in lung that is inactivated in cystic fibrosis. Cell 88:553–560

    Article  CAS  PubMed  Google Scholar 

  11. Greger R, Allert N, Frobe U, Normann C (1993) Increase in cytosolic Ca2+ regulates exocytosis and Cl− conductance in HT29 cells. Pflugers Arch 424:329–334

    Article  CAS  PubMed  Google Scholar 

  12. Harder J, Bartels J, Christophers E, Schroder JM (1997) A peptide antibiotic from human skin. Nature 387:861

    Article  CAS  PubMed  Google Scholar 

  13. Harder J, Bartels J, Christophers E, Schroder JM (2001) Isolation and characterization of human beta-defensin-3, a novel human inducible peptide antibiotic. J Biol Chem 276:5707–5713

    Article  CAS  PubMed  Google Scholar 

  14. Kagan BL, Selsted ME, Ganz T, Lehrer RI (1990) Antimicrobial defensin peptides form voltage-dependent ion-permeable channels in planar lipid bilayer membranes. Proc Natl Acad Sci USA 87:210–214

    Article  CAS  PubMed  Google Scholar 

  15. Kerst G, Fischer KG, Normann C, Kramer A, Leipziger J, Greger R (1995) Ca2+ influx induced by store release and cytosolic Ca2+ chelation in Ht29 colonic carcinoma cells. Pflugers Arch 430:653–665

    Article  CAS  PubMed  Google Scholar 

  16. Kourie JI, Shorthouse AA (2000) Properties of cytotoxic peptide-formed ion channels. Am J Physiol Cell Physiol 278:C1063–C1087

    CAS  PubMed  Google Scholar 

  17. Krause KH, Fivaz M, Monod A, van der Goot FG (1998) Aerolysin induces G-protein activation and Ca2+ release from intracellular stores in human granulocytes. J Biol Chem 273:18122–18129

    Article  CAS  PubMed  Google Scholar 

  18. Kunzelmann K, Mall M (2002) Electrolyte transport in the mammalian colon: mechanisms and implications for disease. Physiol Rev 82:245–289

    CAS  PubMed  Google Scholar 

  19. Kunzelmann K, Schreiber R, Boucherot A (2001) Mechanisms of the inhibition of epithelial Na(+) channels by CFTR and purinergic stimulation. Kidney Int 60:455–461

    Article  CAS  PubMed  Google Scholar 

  20. Leipziger J, Fischer KG, Greger R (1994) Voltage-dependent Ca2+ influx in the epithelial cell line HT29: simultaneous use of intracellular Ca2+ measurements and nystatin perforated patch-clamp technique. Pflugers Arch 426:427–432

    Article  CAS  PubMed  Google Scholar 

  21. Lencer WI, Cheung G, Strohmeier GR, Currie MG, Ouellette AJ, Selsted ME, Madara JL (1997) Induction of epithelial chloride secretion by channel-forming cryptdins 2 and 3. Proc Natl Acad Sci USA 94:8585–8589

    Article  CAS  PubMed  Google Scholar 

  22. Mall M, Bleich M, Schurlein M, Kuhr J, Seydewitz HH, Brandis M, Greger R, Kunzelmann K (1998) Cholinergic ion secretion in human colon requires coactivation by cAMP. Am J Physiol 275:G1274–G1281

    CAS  PubMed  Google Scholar 

  23. Matthews JB (2002) Molecular regulation of Na+-K+-2Cl− cotransporter (NKCC1) and epithelial chloride secretion. World J Surg 26:826–830

    Article  PubMed  Google Scholar 

  24. Merlin D, Yue G, Lencer WI, Selsted ME, Madara JL (2001) Cryptdin-3 induces novel apical conductance(s) in Cl− secretory, including cystic fibrosis, epithelia. Am J Physiol Cell Physiol 280:C296–C302

    CAS  PubMed  Google Scholar 

  25. Morgera F, Antcheva N, Pacor S, Quaroni L, Berti F, Vaccari L, Tossi A (2008) Structuring and interactions of human beta-defensins 2 and 3 with model membranes. J Pept Sci 14:518–523

    Article  CAS  PubMed  Google Scholar 

  26. Morrison GM, Semple CA, Kilanowski FM, Hill RE, Dorin JR (2003) Signal sequence conservation and mature peptide divergence within subgroups of the murine beta-defensin gene family. Mol Biol Evol 20:460–470

    Article  CAS  PubMed  Google Scholar 

  27. Nitschke R, Benning N, Ricken S, Leipziger J, Fischer KG, Greger R (1997) Effect of intracellular pH on agonist-induced [Ca2+]i transients in HT29 cells. Pflugers Arch 434:466–474

    Article  CAS  PubMed  Google Scholar 

  28. Nitschke R, Leipziger J, Greger R (1993) Agonist-induced intracellular Ca2+ transients in HT29 cells. Pflugers Arch 423:519–526

    Article  CAS  PubMed  Google Scholar 

  29. O’Neil DA (2003) Regulation of expression of beta-defensins: endogenous enteric peptide antibiotics. Mol Immunol 40:445–450

    Article  PubMed  Google Scholar 

  30. O’Neil DA, Porter EM, Elewaut D, Anderson GM, Eckmann L, Ganz T, Kagnoff MF (1999) Expression and regulation of the human beta-defensins hBD-1 and hBD-2 in intestinal epithelium. J Immunol 163:6718–6724

    PubMed  Google Scholar 

  31. Patil AA, Cai Y, Sang Y, Blecha F, Zhang G (2005) Cross-species analysis of the mammalian beta-defensin gene family: presence of syntenic gene clusters and preferential expression in the male reproductive tract. Physiol Genomics 23:5–17

    Article  CAS  PubMed  Google Scholar 

  32. Sahly H, Schubert S, Harder J, Rautenberg P, Ullmann U, Schroder J, Podschun R (2003) Burkholderia is highly resistant to human beta-defensin 3. Antimicrob Agents Chemother 47:1739–1741

    Article  CAS  PubMed  Google Scholar 

  33. Salzman NH, Underwood MA, Bevins CL (2007) Paneth cells, defensins, and the commensal microbiota: a hypothesis on intimate interplay at the intestinal mucosa. Semin Immunol 19:70–83

    Article  CAS  PubMed  Google Scholar 

  34. Selsted ME, Ouellette AJ (2005) Mammalian defensins in the antimicrobial immune response. Nat Immunol 6:551–557

    Article  CAS  PubMed  Google Scholar 

  35. Semple CA, Rolfe M, Dorin JR (2003) Duplication and selection in the evolution of primate beta-defensin genes. Genome Biol 4:R31

    Article  PubMed  Google Scholar 

  36. Sorensen OE, Borregaard N, Cole AM (2008) Antimicrobial peptides in innate immune responses. Contrib Microbiol 15:61–77

    Article  CAS  PubMed  Google Scholar 

  37. Suresh A, Verma C (2006) Modelling study of dimerization in mammalian defensins. BMC Bioinformatics 7(Suppl 5):S17

    Article  PubMed  Google Scholar 

  38. Tieleman DP, Shrivastava IH, Ulmschneider MR, Sansom MS (2001) Proline-induced hinges in transmembrane helices: possible roles in ion channel gating. Proteins 44:63–72

    Article  CAS  PubMed  Google Scholar 

  39. Turner JR (2006) Molecular basis of epithelial barrier regulation: from basic mechanisms to clinical application. Am J Pathol 169:1901–1909

    Article  CAS  PubMed  Google Scholar 

  40. Warth R, Bleich M (2000) K+ channels and colonic function. Rev Physiol Biochem Pharmacol 140:1–62

    Article  CAS  PubMed  Google Scholar 

  41. Yang D, Biragyn A, Kwak LW, Oppenheim JJ (2002) Mammalian defensins in immunity: more than just microbicidal. Trends Immunol 23:291–296

    Article  CAS  PubMed  Google Scholar 

  42. Yang D, Chertov O, Bykovskaia SN, Chen Q, Buffo MJ, Shogan J, Anderson M, Schroder JM, Wang JM, Howard OM, Oppenheim JJ (1999) Beta-defensins: linking innate and adaptive immunity through dendritic and T cell CCR6. Science 286:525–528

    Article  CAS  PubMed  Google Scholar 

  43. Yue G, Merlin D, Selsted ME, Lencer WI, Madara JL, Eaton DC (2002) Cryptdin 3 forms anion selective channels in cytoplasmic membranes of human embryonic kidney cells. Am J Physiol Gastrointest Liver Physiol 282:G757–G765

    CAS  PubMed  Google Scholar 

  44. Zasloff M (2002) Antimicrobial peptides of multicellular organisms. Nature 415:389–395

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We thank the department of animal care of the Christian-Albrechts-Universität Kiel (CAU) for their excellent support in breeding and animal handling. We thank T. Stegmann and J. Brdon for technical assistance. This work was supported by CAU grants to M.B. and by SFB617.

Author information

Authors and Affiliations

  1. Physiologisches Institut der Christian-Albrechts-Universität, Kiel, Germany

    Nina Himmerkus, Veit Vassen, Birte Sievers, Boeren Goerke, Qixian Shan & Markus Bleich

  2. Hautklinik, Universitätsklinikum Schleswig-Holstein, Campus Kiel, Kiel, Germany

    Jürgen Harder & Jens-Michael Schröder

  3. Physiologisches Institut, Olshausenstraβe 40, 24098, Kiel, Germany

    Nina Himmerkus

Authors
  1. Nina Himmerkus
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Veit Vassen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Birte Sievers
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Boeren Goerke
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Qixian Shan
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Jürgen Harder
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Jens-Michael Schröder
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Markus Bleich
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Nina Himmerkus.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Himmerkus, N., Vassen, V., Sievers, B. et al. Human β-defensin-2 increases cholinergic response in colon epithelium. Pflugers Arch - Eur J Physiol 460, 177–186 (2010). https://doi.org/10.1007/s00424-009-0780-x

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00424-009-0780-x

Keywords

Search

Navigation