Inflamed pouch mucosa possesses altered tight junctions indicating recurrence of inflammatory bowel disease

  • Salah AmashehEmail author
  • Sonja Dullat
  • Michael Fromm
  • Jörg D. Schulzke
  • Heinz J. Buhr
  • Anton J. Kroesen
Original Article


Background and aims

The etiology of pouchitis after coloproctomucosectomy with ileal pouch-anal anastomosis in patients with ulcerative colitis is still unknown. Beside changes in luminal antigens, the immunological predisposition is assumed to be responsible. In previous electrophysiological studies, we showed that mucosal barrier and transport function in pouchitis is markedly reduced. Thus, the aim of the present study was to analyze barrier function on the molecular level.

Material and methods

Pouch biopsies of 36 ulcerative colitis patients were analyzed. Time points were (1) intraoperative immediately prior to ileal pouch-anal anastomosis (n = 13), (2) >1 year after ileostomy closure (pouch, n = 12), and (3) during pouchitis (n = 11). Control terminal ileum biopsies were obtained from eight patients undergoing hemicolectomy due to carcinoma. Expression of tight junction proteins was analyzed by Western blotting and confocal laser-scanning microscopy. To elucidate effects on epithelial barrier properties, impedance spectroscopy was performed in miniaturized Ussing chambers.


In pouchitis, epithelial resistance was markedly reduced compared to non-inflamed pouch and control ileum. Expression of tight junction proteins claudin-1, 3, 4, 5, and 7 and occludin revealed differential expression regulation with the tightening tight junction protein claudin-1 being decreased and an increase of the pore-forming claudin-2, whereas other claudins remained constant. Morphometry indicated the mucosal surface to be unchanged.


Pouchitis is characterized by a selective change of tight junction proteins in favor of opening the epithelial tight junction and, thus, the paracellular pathway, which contributes to the inflammatory process. This resembles changes in inflammatory bowel disease (IBD) and indicates IBD recurrence in pouchitis.


Pouchitis Ulcerative colitis Barrier Ileum IBD Tight junction 



inflammatory bowel disease


ileo pouchanal anastomosis


ulcerative colitis


pouchitis disease activity index


short circuit current





We thank A. Fromm, S. Schön and D. Sorgenfrei for their excellent technical assistance. The study has been supported by the DFG and the Sonnenfeld-Stiftung Berlin.


  1. 1.
    Fazio VW, O'Riordain MG, Lavery IC, Church JM, Lau P, Strong SA, Hull T (1998) Long-term functional outcome and quality of life after stapled restorative proctocolectomy. Ann Surg 230:575–584CrossRefGoogle Scholar
  2. 2.
    Runkel N, Kroesen A, Buhr HJ (1998) Technique and results of ileoanal pouches in ulcerative colitis after colectomy and proctomucosectomy. Zentralbl Chir 123:375–380PubMedGoogle Scholar
  3. 3.
    Buhr HJ, Heuschen UA, Stern J, Herfarth C (1993) Continence preserving operation after proctocolectomy. Indications, technique and results. Chirurg 64:601–613PubMedGoogle Scholar
  4. 4.
    Heppell J, Kelly K (1998) Pouchitis. Curr Opin Gastroenterol 14:322–326CrossRefGoogle Scholar
  5. 5.
    Amasheh S, Barmeyer C, Koch CS, Tavalali S, Mankertz J, Epple HJ, Gehring MM, Florian P, Kroesen AJ, Zeitz M, Fromm M, Schulzke JD (2004) Cytokine-dependent transcriptional down-regulation of epithelial sodium channel in ulcerative colitis. Gastroenterology 126:1711–1720PubMedCrossRefGoogle Scholar
  6. 6.
    Kroesen AJ, Dullat S, Schulzke JD, Fromm M, Buhr H (2008) Permanently increased mucosal permeability in patients with backwash ileitis after ileoanal pouch for ulcerative colitis. Scand J. Gastroenterol. 43:704–711PubMedCrossRefGoogle Scholar
  7. 7.
    Kroesen AJ, Leistenschneider P, Lehmann K, Ransco C, Dullat S, Blaut M, Schulzke JD, Fromm M, Buhr HJ (2006) Increased bacterial permeation in long-lasting ileoanal pouches. Inflamm Bowel Dis 12:736–744PubMedCrossRefGoogle Scholar
  8. 8.
    Furuse M, Fujita K, Hiiragi T, Fujimoto K, Tsukita S (1998) Claudin-1 and -2: novel integral membrane proteins localizing at tight junctions with no sequence similarity to occludin. J Cell Biol 141:1539–1550PubMedCrossRefGoogle Scholar
  9. 9.
    Van Itallie C, Rahner C, Anderson JM (2001) Regulated expression of claudin-4 decreases paracellular conductance through a selective decrease in sodium permeability. J Clin Invest 107:1319–1327PubMedCrossRefGoogle Scholar
  10. 10.
    Sawada N, Murata M, Kikuchi K, Osanai M, Tobioka H, Kojima T, Chiba H (2003) Tight junctions and human diseases. Med Electron Microsc 36:147–156PubMedCrossRefGoogle Scholar
  11. 11.
    Furuse M, Hata M, Furuse K, Yoshida Y, Haratake A, Sugitani Y, Noda T, Kubo A, Tsukita S (2002) Claudin-based tight junctions are crucial for the mammalian epidermal barrier: a lesson from claudin-1-deficient mice. J Cell Biol 56:1099–1111CrossRefGoogle Scholar
  12. 12.
    Amasheh S, Schmidt T, Mahn M, Florian P, Mankertz J, Tavalali S, Gitter AH, Schulzke JD, Fromm M (2005) Expression of claudin-5 contributes to barrier properties in tight junctions of epithelial cells. Cell Tissue Res 321:89–96PubMedCrossRefGoogle Scholar
  13. 13.
    Amasheh S, Meiri N, Gitter AH, Schöneberg T, Mankertz J, Schulzke JD, Fromm M (2002) Claudin-2 expression induces cation-selective channels in tight junctions of epithelial cells. J Cell Sci 115:4969–4976PubMedCrossRefGoogle Scholar
  14. 14.
    Bürgel N, Bojarski C, Mankertz J, Zeitz M, Fromm M, Schulzke JD (2002) Mechanisms of diarrhea in collagenous colitis. Gastroenterology 123:433–443PubMedCrossRefGoogle Scholar
  15. 15.
    Heller F, Florian P, Bojarski C, Richter J, Christ M, Hillenbrand B, Mankertz J, Gitter AH, Bürgel N, Fromm M, Zeitz M, Fuss I, Strober W, Schulzke JD (2005) Interleukin-13 is the key effector Th2 cytokine in ulcerative colitis that affects epithelial tight junctions, apoptosis, and cell restitution. Gastroenterology 129:550–564PubMedGoogle Scholar
  16. 16.
    Zeissig S, Bürgel N, Günzel D, Richter J, Mankertz J, Wahnschaffe U, Kroesen AJ, Zeitz M, Fromm M, Schulzke JD (2007) Changes in expression and distribution of claudin 2, 5 and 8 lead to discontinuous tight junctions and barrier dysfunction in active Crohn's disease. Gut 56:61–72PubMedCrossRefGoogle Scholar
  17. 17.
    Sandborn WJ, Tremaine WJ, Batts KP, Pemberton JH, Phillips SF (1994) Pouchitis after ileal pouch-anal anastomosis: a pouchitis disease activity index. Mayo Clin Proc 69:409–415PubMedGoogle Scholar
  18. 18.
    Stockmann M, Gitter AH, Sorgenfrei D, Fromm M, Schulzke JD (1999) Low edge damage container insert that adjusts intestinal forceps biopsies into Ussing chamber systems. Pflügers Arch 438:107–112PubMedCrossRefGoogle Scholar
  19. 19.
    Gitter AH, Schulzke JD, Sorgenfrei D, Fromm M (1997) Ussing chamber for high-frequency transmural impedance analysis of epithelial tissues. J Biochem Biophys Methods 35:81–88PubMedCrossRefGoogle Scholar
  20. 20.
    Schulzke JD, Gitter AH, Mankertz J, Spiegel S, Seidler U, Amasheh S, Saitou M, Tsukita S, Fromm M (2005) Epithelial transport and barrier function in occludin-deficient mice. Biochim Biophys Acta 1669:34–42PubMedCrossRefGoogle Scholar
  21. 21.
    Kroesen AJ, Stockmann M, Ransco C, Schulzke JD, Fromm M, Buhr HJ (2002) Impairment of epithelial transport but not of barrier function in idiopathic pouchitis after ulcerative colitis. Gut 6:821–826CrossRefGoogle Scholar
  22. 22.
    Amasheh S, Milatz S, Krug SM, Bergs M, Amasheh M, Schulzke JD, Fromm M (2009) Na+ absorption defends from paracellular back-leakage by claudin-8 upregulation. Biochem Biophys Res Commun 378:45–50PubMedCrossRefGoogle Scholar
  23. 23.
    Furuse M, Hirase T, Itoh M, Nagafuchi A, Yonemura S, Tsukita S, Tsukita S (1993) Occludin: a novel integral membrane protein localizing at tight junctions. J Cell Biol 123:1777–1788PubMedCrossRefGoogle Scholar
  24. 24.
    Ikenouchi J, Furuse M, Furuse K, Sasaki H, Tsukita S, Tsukita S (2005) Tricellulin constitutes a novel barrier at tricellular contacts of epithelial cells. J Cell Biol 171:939–945PubMedCrossRefGoogle Scholar
  25. 25.
    Saitou M, Furuse M, Sasaki H, Schulzke JD, Fromm M, Takano H, Noda T, Tsukita S (2000) Complex phenotype of mice lacking occludin, a component of tight junction strands. Mol Biol Cell 11:4131–4142PubMedGoogle Scholar
  26. 26.
    Reyes JL, Lamas M, Martin D, del Carmen Namorado M, Islas S, Luna J, Tauc M, González-Mariscal L (2002) The renal segmental distribution of claudins changes with development. Kidney Int 62:476–487PubMedCrossRefGoogle Scholar
  27. 27.
    Dhawan P, Singh AB, Deane NG, No Y, Shiou SR, Schmidt C, Neff J, Washington MK, Beauchamp RD (2005) Claudin-1 regulates cellular transformation and metastatic behavior in colon cancer. J Clin Invest 115:1765–1776PubMedCrossRefGoogle Scholar
  28. 28.
    Troeger H, Epple HJ, Schneider T, Wahnschaffe U, Ullrich R, Burchard GD, Jelinek T, Zeitz M, Fromm M, Schulzke JD (2007) Effect of chronic Giardia lamblia infection on epithelial transport and barrier function in human duodenum. Gut 56:328–335PubMedCrossRefGoogle Scholar
  29. 29.
    Merrett MN, Soper N, Mortensen N, Jewell DP (1996) Intestinal permeability in the ileal pouch. Gut 39:226–230PubMedCrossRefGoogle Scholar
  30. 30.
    Mankertz J, Amasheh M, Krug SM, Fromm A, Amasheh S, Hillenbrand B, Tavalali S, Fromm M, Schulzke JD (2009) TNFalpha up-regulates claudin-2 expression in epithelial HT-29/B6 cells via phosphatidylinositol-3-kinase signaling. Cell Tissue Res 336:67–77PubMedCrossRefGoogle Scholar
  31. 31.
    Harten SK, Shukla D, Barod R, Hergovich A, Balda MS, Matter K, Esteban MA, Maxwell PH (2009) Regulation of renal epithelial tight junctions by the von Hippel-Lindau tumor suppressor gene involves occludin and claudin 1 and is independent of E-cadherin. Mol Biol Cell 20:1089–1101PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Salah Amasheh
    • 1
    Email author
  • Sonja Dullat
    • 2
  • Michael Fromm
    • 1
  • Jörg D. Schulzke
    • 3
    • 4
  • Heinz J. Buhr
    • 2
  • Anton J. Kroesen
    • 2
  1. 1.Institute of Clinical PhysiologyCBF, Charité, Freie Universität und Humboldt UniversitätBerlinGermany
  2. 2.Department of SurgeryCBF, Charité, Freie Universität und Humboldt UniversitätBerlinGermany
  3. 3.Department of General MedicineCBF, Charité, Freie Universität und Humboldt UniversitätBerlinGermany
  4. 4.Department of Gastroenterology, Infectious Diseases, and RheumatologyCBF, Charité, Freie Universität und Humboldt UniversitätBerlinGermany

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