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Cell and Tissue Research

, Volume 336, Issue 1, pp 67–77 | Cite as

TNFα up-regulates claudin-2 expression in epithelial HT-29/B6 cells via phosphatidylinositol-3-kinase signaling

  • J. Mankertz
  • M. Amasheh
  • S. M. Krug
  • A. Fromm
  • S. Amasheh
  • B. Hillenbrand
  • S. Tavalali
  • M. Fromm
  • J. D. Schulzke
Regular Article

Abstract

Our aim has been to characterize the molecular mechanisms regulating the expression of the channel-forming tight-junctional protein claudin-2 in response to the pro-inflammatory cytokine tumor necrosis factor-α (TNFα), which is elevated, for example, in active Crohn’s disease. TNFα caused an 89% decrease of the paracellular resistance in colonic HT-29/B6 cells, whereas transcellular resistance was unaltered. The claudin-2 protein level was increased by TNFα without changes in subcellular tight-junctional protein localization as revealed by confocal laser scanning microscopy. Enhanced gene expression was identified as the source of this increase, since claudin-2-specific mRNA and promoter activity was elevated, whereas mRNA stability remained unaltered. Specific inhibitors and phospho-specific antibodies revealed that the increased gene expression of claudin-2 after TNFα treatment was mediated by the phosphatidylinositol-3-kinase pathway. Thus, the up-regulation of claudin-2 by TNFα is attributable to the regulation of the expression of the gene, as a result of which epithelial barrier function is disturbed, for example, during chronic intestinal inflammation.

Keywords

Barrier Claudin-2 Phosphatidylinositol-3-kinase Tight junction TNFα Human 

Notes

Acknowledgements

We thank Nicole Held and Susanna Schön for excellent technical assistance. The support of Detlef Sorgenfrei (electronic engineer) is gratefully acknowledged.

References

  1. 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
  2. Bojarski C, Weiske J, Schöneberg T, Schröder W, Mankertz J, Schulzke JD, Florian P, Fromm M, Tauber R, Huber O (2004) The specific fates of tight junction proteins in apoptotic epithelial cells. J Cell Sci 117:2097–2107PubMedCrossRefGoogle Scholar
  3. Bruewer M, Utech M, Ivanov AI, Hopkins AM, Parkos CA, Nusrat A (2005) Interferon-γ induces internalization of epithelial tight junction proteins via a macropinocytosis-like process. FASEB J 19:923–933PubMedCrossRefGoogle Scholar
  4. Cario E, Gerken G, Podolsky DK (2007) Toll-like receptor 2 controls mucosal inflammation by regulating epithelial barrier function. Gastroenterology 132:1359–1374PubMedCrossRefGoogle Scholar
  5. Clayburgh DR, Musch MW, Leitges M, Fu YX, Turner JR (2006) Coordinated epithelial NHE3 inhibition and barrier dysfunction are required for TNF-mediated diarrhea in vivo. J Clin Invest 116:2682–2694PubMedCrossRefGoogle Scholar
  6. Escaffit F, Boudreau F, Beaulieu JF (2005) Differential expression of claudin-2 along the human intestine: implication of GATA-4 in the maintenance of claudin-2 in differentiating cells. J Cell Physiol 203:15–26PubMedCrossRefGoogle Scholar
  7. Fasano A, Baudry B, Pumplin DW, Wasserman SS, Tall BD, Ketley JM, Kaper JB (1991) Vibrio cholerae produces a second enterotoxin, which affects intestinal tight junctions. Proc Natl Acad Sci USA 88:5242–5246PubMedCrossRefGoogle Scholar
  8. 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
  9. 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
  10. Furuse M, Furuse K, Sasaki H, Tsukita S (2001) Conversion of zonulae occludentes from tight to leaky strand type by introducing claudin-2 into Madin-Darby canine kidney I cells. J Cell Biol 153:263–272PubMedCrossRefGoogle Scholar
  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 156:1099–1111PubMedCrossRefGoogle Scholar
  12. Fuss IJ, Neurath M, Boirivant M, Klein JS, de la Motte C, Strong SA, Fiocchi C, Strober W (1996) Disparate CD4+ lamina propria (LP) lymphokine secretion profiles in inflammatory bowel disease. Crohn’s disease LP cells manifest increased secretion of IFN-gamma, whereas ulcerative colitis LP cells manifest increased secretion of IL-5. J Immunol 157:1261–1270PubMedGoogle Scholar
  13. Gitter AH, Bendfeldt K, Schulzke JD, Fromm M (2000) Leaks in the epithelial barrier caused by spontaneous and TNF-α-induced single-cell apoptosis. FASEB J 14:1749–1753PubMedCrossRefGoogle Scholar
  14. Gonzalez-Mariscal L, Tapia R, Chamorro D (2008) Crosstalk of tight junction components with signaling pathways. Biochim Biophys Acta 1778:729–756PubMedCrossRefGoogle Scholar
  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. Ikenouchi J, Furuse M, Furuse K, Sasaki H, Tsukita S (2005) Tricellulin constitutes a novel barrier at tricellular contacts of epithelial cells. J Cell Biol 171:939–945PubMedCrossRefGoogle Scholar
  17. Inai T, Kobayashi J, Shibata Y (1999) Claudin-1 contributes to the epithelial barrier function in MDCK cells. Eur J Cell Biol 78:849–855PubMedGoogle Scholar
  18. Kinugasa T, Sakaguchi T, Gu X, Reinecker HC (2000) Claudins regulate the intestinal barrier in response to immune mediators. Gastroenterology 118:1001–1011PubMedCrossRefGoogle Scholar
  19. Kreusel KM, Fromm M, Schulzke JD, Hegel U (1991) Cl- secretion in epithelial monolayers of mucus-forming human colon cells (HT-29/B6). Am J Physiol 261:C574–C582PubMedGoogle Scholar
  20. Leverrier Y, Thomas J, Mathieu AL, Low W, Blanquier B, Marvel J (1999) Role of PI3-kinase in Bcl-X induction and apoptosis inhibition mediated by IL-3 or IGF-1 in Baf-3 cells. Cell Death Differ 6:290–296PubMedCrossRefGoogle Scholar
  21. Lipschutz JH, Li S, Arisco A, Balkovetz DF (2005) Extracellular signal-regulated kinases 1/2 control claudin-2 expression in Madin-Darby canine kidney strain I and II cells. J Biol Chem 280:3780–3788PubMedCrossRefGoogle Scholar
  22. Mankertz J, Schulzke JD (2007) Altered permeability in inflammatory bowel disease: pathophysiology and clinical implications. Curr Opin Gastroenterol 23:379–383PubMedCrossRefGoogle Scholar
  23. Mankertz J, Tavalali S, Schmitz H, Mankertz A, Riecken EO, Fromm M, Schulzke JD (2000) Expression from the human occludin promoter is affected by tumor necrosis factor alpha and interferon gamma. J Cell Sci 113:2085–2090PubMedGoogle Scholar
  24. Mankertz J, Hillenbrand B, Tavalali S, Huber O, Fromm M, Schulzke JD (2004) Functional crosstalk between Wnt signaling and Cdx-related transcriptional activation in the regulation of the claudin-2 promoter activity. Biochem Biophys Res Commun 314:1001–1007PubMedCrossRefGoogle Scholar
  25. Martin-Padura I, Lostaglio S, Schneemann M, Williams L, Romano M, Fruscella P, Panzeri C, Stoppacciaro A, Ruco L, Villa A, Simmons D, Dejana E (1998) Junctional adhesion molecule, a novel member of the immunoglobulin superfamily that distributes at intercellular junctions and modulates monocyte transmigration. J Cell Biol 142:117–127PubMedCrossRefGoogle Scholar
  26. Ozes ON, Mayo LD, Gustin JA, Pfeffer SR, Pfeffer LM, Donner DB (1999) NF-kappaB activation by tumour necrosis factor requires the Akt serine-threonine kinase. Nature 401:82–85PubMedCrossRefGoogle Scholar
  27. Patrick DM, Leone AK, Shellenberger JJ, Dudowicz KA, King JM (2006) Proinflammatory cytokines tumor necrosis factor-alpha and interferon-gamma modulate epithelial barrier function in Madin-Darby canine kidney cells through mitogen activated protein kinase signaling. BMC Physiol 6:2PubMedCrossRefGoogle Scholar
  28. Prasad S, Minigrino R, Kaukinen K, Hayes KL, Powell RM, MacDonald TT, Collins JE (2005) Inflammatory processes have differential effects on claudins 2, 3 and 4 in colonic epithelial cells. Lab Invest 85:1139–1162PubMedCrossRefGoogle Scholar
  29. Rahner C, Mitic LL, Anderson JM (2001) Heterogeneity in expression and subcellular localization of claudins 2, 3, 4, and 5 in the rat liver, pancreas, and gut. Gastroenterology 120:411–422PubMedCrossRefGoogle Scholar
  30. Reiter B, Kraft R, Günzel D, Zeissig S, Schulzke JD, Fromm M, Harteneck C (2006) TRPV4-mediated regulation of epithelial permeability. FASEB J 20:1802–1812PubMedCrossRefGoogle Scholar
  31. Sakaguchi T, Gu X, Golden HM, Suh E, Rhoads DB, Reinecker HC (2002) Cloning of the human claudin-2 5′-flanking region revealed a TATA-less promoter with conserved binding sites in mouse and human for caudal-related homeodomain proteins and hepatocyte nuclear factor-1alpha. J Biol Chem 277:21361–21370PubMedCrossRefGoogle Scholar
  32. Schmitz H, Fromm M, Bentzel CJ, Scholz P, Detjen K, Mankertz J, Bode H, Epple HJ, Riecken EO, Schulzke JD (1999) Tumor necrosis factor-alpha (TNF α) regulates the epithelial barrier in the human intestinal cell line HT-29/B6. J Cell Sci 112:137–146PubMedGoogle Scholar
  33. Tsukita S, Furuse M, Itoh M (2001) Multifunctional strands in tight junctions. Nat Rev Mol Cell Biol 2:285–293PubMedCrossRefGoogle Scholar
  34. Utech M, Ivanov AI, Samarin SN, Bruewer M, Turner JR, Mrsny RJ, Parkos CA, Nusrat A (2005) Mechanism of IFN-γ-induced endocytosis of tight junction proteins: myosin II-dependent vacuolarization of the apical plasma membrane. Mol Biol Cell 16:5040–5052PubMedCrossRefGoogle Scholar
  35. Van Itallie CM, Anderson JM (2006) Claudins and epithelial paracellular transport. Annu Rev Physiol 68:403–429PubMedCrossRefGoogle Scholar
  36. Wang F, Schwarz BT, Graham WV, Wang Y, Su L, Clayburgh DR, Abraham C, Turner JR (2006) IFN-gamma-induced TNF receptor 2 expression is required for TNF-dependent intestinal barrier dysfunction. Gastroenterology 131:1153–1163PubMedCrossRefGoogle Scholar
  37. Willemsen LE, Hoetjes JP, Deventer SJ van, Tol EA van (2005) Abrogation of IFN-gamma mediated epithelial barrier disruption by serine protease inhibition. Clin Exp Immunol 142:275–284PubMedCrossRefGoogle Scholar
  38. Ye D, Ma I, Ma TY (2006) Molecular mechanism of tumor necrosis factor-alpha modulation of intestinal epithelial tight junction barrier. Am J Physiol Gastrointest Liver Physiol 290:G496–G504PubMedCrossRefGoogle Scholar
  39. Youakim A, Ahdieh M (1999) Interferon-gamma decreases barrier function in T84 cells by reducing ZO-1 levels and disrupting apical actin. Am J Physiol 276:G1279–G1288PubMedGoogle Scholar
  40. Zeissig S, Bürgel N, Günzel D, Richter JF, 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

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • J. Mankertz
    • 1
  • M. Amasheh
    • 1
  • S. M. Krug
    • 2
  • A. Fromm
    • 2
  • S. Amasheh
    • 2
  • B. Hillenbrand
    • 1
  • S. Tavalali
    • 1
  • M. Fromm
    • 2
  • J. D. Schulzke
    • 1
    • 3
  1. 1.Department of Gastroenterology, CharitéCampus Benjamin FranklinBerlinGermany
  2. 2.Institute of Clinical Physiology, CharitéCampus Benjamin FranklinBerlinGermany
  3. 3.Department of General Medicine, CharitéCampus Benjamin FranklinBerlinGermany

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