Abstract
Background
Diminished intestinal epithelial barrier function contributes to the pathogenesis of Crohn’s disease. Clinical and experimental studies propose that increased tumor necrosis factor (TNF)-α promotes barrier dysfunction. The aim of this study was to investigate the effects of nutritional and other therapies upon intestinal barrier function in the presence of TNF-α in an in vitro model.
Methods
Caco-2 monolayers were grown to confluence on membrane supports and then exposed to TNF-α in the presence of polymeric formula, hydrocortisone or infliximab. Monolayer permeability was evaluated by measuring epithelial resistance, short-circuit current and horseradish peroxidase flux in an Ussing chamber. Tight junction and myosin II regulatory light-chain kinase gene expression was analysed by real-time PCR, with protein expression and localization analysed by Western blot and immunofluorescence.
Results
TNF-α increased monolayer permeability and diminished tight junction integrity. However both polymeric formula and infliximab completely abrogated the effects of TNF-α. These monolayers displayed unchanged permeability and tight junction integrity compared to untreated cells (media-no-TNF-α controls). In contrast, hydrocortisone only partially abrogated the effects of TNF-α, with these monolayers having increased permeability and altered tight junction integrity compared to media-no-TNF-α controls.
Conclusions
Both polymeric formula and infliximab completely prevent epithelial barrier dysfunction in the presence of TNF-α, whereas hydrocortisone partially prevents barrier dysfunction. These results provide evidence that superior mucosal healing can be achieved with both polymeric formula and infliximab compared to hydrocortisone.
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References
Forster C. Tight junctions and the modulation of barrier function in disease. Histochem Cell Biol. 2008;130:55–70.
Zeissig S, Burgel N, Gunzel D, Richter J, Mankertz J, Wahnschaffe U, et al. 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. 2007;56:61–72.
Edelblum KL, Turner JR. The tight junction in inflammatory disease: communication breakdown. Curr Opin Pharmacol. 2009;9:715–20.
Day AS, Whitten KE, Sidler M, Lemberg DA. Systematic review: nutritional therapy in paediatric Crohn’s disease. Aliment Pharmacol Ther. 2008;27:293–307.
de Ridder L, Rings EHHM, Damen GM, Kneepkens CMF, Schweizer JJ, Kokke FTM, et al. Infliximab dependency in pediatric Crohn’s disease: long-term follow-up of an unselected cohort. Inflamm Bowel Dis. 2008;14:353–8.
Yee AM, Pochapin MB. Treatment of complicated sarcoidosis with infliximab anti-tumor necrosis factor-alpha therapy. Ann Intern Med. 2001;135:27–31.
Escher JC, Taminiau JAJM, Nieuwenhuis EES, Buller HA, Grand RJ. Treatment of inflammatory bowel disease in childhood: best available evidence. Inflamm Bowel Dis. 2003;9:34–58.
Rosh JR, Gross T, Mamula P, Griffiths A, Hyams J. Hepatosplenic T-cell lymphoma in adolescents and young adults with Crohn’s disease: a cautionary tale? Inflamm Bowel Dis. 2007;13:1024–30.
Borrelli O, Cordischi L, Cirulli M, Paganelli M, Labalestra V, Uccini S, et al. Polymeric diet alone versus corticosteroids in the treatment of active pediatric Crohn’s disease: a randomized controlled open-label trial. Clin Gastroenterol Hepatol. 2006;4:744–53.
Day AS, Whitten KE, de Jong NSH. Nutrition and nutritional management of Crohn’s disease in children and adolescents. Curr Nutr Food Sci. 2006;2:3–14.
Berni Canani R, Terrin G, Borrelli O, Romano MT, Manguso F, Coruzzo A, et al. Short- and long-term therapeutic efficacy of nutritional therapy and corticosteroids in paediatric Crohn’s disease. Dig Liver Dis. 2006;38:381–7.
Fell JM, Paintin M, Arnaud-Battandier F, Beattie RM, Hollis A, Kitching P, et al. Mucosal healing and a fall in mucosal pro-inflammatory cytokine mRNA induced by a specific oral polymeric diet in paediatric Crohn’s disease. Aliment Pharmacol Ther. 2000;14:281–9.
Forster C, Burek M, Romero IA, Weksler B, Couraud PO, Drenckhahn D. Differential effects of hydrocortisone and TNF-alpha on tight junction proteins in an in vitro model of the human blood–brain barrier. J Physiol. 2008;586:1937–49.
Baldassano R, Braegger CP, Escher JC, Dewoody K, Hendricks DF, Keenan GF, et al. Infliximab (REMICADE) therapy in the treatment of pediatric Crohn’s disease. Am J Gastroenterol. 2003;98:833–8.
de Jong NSH, Leach ST, Day AS. Polymeric formula has direct anti-inflammatory effects on enterocytes in an in vitro model of intestinal inflammation. Dig Dis Sci. 2007;52:2029–36.
Gaillard JL, Finlay BB. Effect of cell polarization and differentiation on entry of Listeria monocytogenes into the enterocyte-like Caco-2 cell line. Infect Immun. 1996;64:1299–308.
Wang F, Graham WV, Wang Y, Witkowski ED, Schwarz BT, Turner JR. Interferon-gamma and tumor necrosis factor-alpha synergize to induce intestinal epithelial barrier dysfunction by up-regulating myosin light chain kinase expression. Am J Pathol. 2005;166:409–19.
Feighery LM, Cochrane SW, Quinn T, Baird AW, O’Toole D, Owens SE, et al. Myosin light chain kinase inhibition: correction of increased intestinal epithelial permeability in vitro. Pharm Res. 2008;25:1377–86.
Wroblewski LE, Shen L, Ogden S, Romero-Gallo J, Lapierre LA, Israel DA, et al. Helicobacter pylori dysregulation of gastric epithelial tight junctions by urease-mediated myosin II activation. Gastroenterology. 2009;136:236–46.
Fries W, Muja C, Crisafulli C, Cuzzocrea S, Mazzon E. Dynamics of enterocyte tight junctions: effect of experimental colitis and two different anti-TNF strategies. Am J Physiol Gastrointest Liver Physiol. 2008;294:G938–47.
Stevenson BR, Anderson JM, Bullivant S. The epithelial tight junction: structure, function and preliminary biochemical characterization. Mol Cell Biochem. 1988;83:129–45.
Gibson PR. Increased gut permeability in Crohn’s disease: is TNF the link? Gut. 2004;53:1724–5.
Schulzke J-D, Bojarski C, Zeissig S, Heller F, Gitter AH, Fromm M. Disrupted barrier function through epithelial cell apoptosis. Ann N Y Acad Sci. 2006;1072:288–99.
Zeissig S, Bojarski C, Buergel N, Mankertz J, Zeitz M, Fromm M, et al. Downregulation of epithelial apoptosis and barrier repair in active Crohn’s disease by tumour necrosis factor alpha antibody treatment. Gut. 2004;53:1295–302.
Watson AJ, Chu S, Sieck L, Gerasimenko O, Bullen T, Campbell F, et al. Epithelial barrier function in vivo is sustained despite gaps in epithelial layers. Gastroenterology. 2005;129:902–12.
Florian P, Schoneberg T, Schulzke JD, Fromm M, Gitter AH. Single-cell epithelial defects close rapidly by an actinomyosin purse string mechanism with functional tight junctions. J Physiol. 2002;545:485–99.
Graham WV, Wang F, Clayburgh DR, Cheng JX, Yoon B, Wang Y, et al. Tumor necrosis factor-induced long myosin light chain kinase transcription is regulated by differentiation-dependent signaling events. Characterization of the human long myosin light chain kinase promoter. J Biol Chem. 2006;281:26205–15.
Al-Sadi R, Ye D, Dokladny K, Ma TY. Mechanism of Il-1β-induced increase in intestinal epithelial tight junction permeability. J Immunol. 2008;180:5653–61.
Clayburgh DR, Rosen S, Witkowski ED, Wang F, Blair S, Dudek S, et al. A differentiation-dependent splice variant of myosin light chain kinase, MLCK1, regulates epithelial tight junction permeability. J Biol Chem. 2004;279:55506–13.
Weber CR, Turner JR. Inflammatory bowel disease: is it really just another break in the wall? Gut. 2007;56:6–8.
Ma TY, Tran D, Hoa N, Nguyen D, Merryfield M, Tarnawski A. Mechanism of extracellular calcium regulation of intestinal epithelial tight junction permeability: role of cytoskeletal involvement. Microsc Res Tech. 2000;51:156–68.
Vieira AT, Pinho V, Lepsch LB, Scavone C, Ribeiro IM, Tomassini T, et al. Mechanisms of the anti-inflammatory effects of the natural secosteroids physalins in a model of intestinal ischaemia and reperfusion injury. Br J Pharmacol. 2005;146:244–51.
Cury DHB, Costa JE, Irika K, Mijji L, Garcez A, Buchiguel C, et al. Protective effect of octreotide and infliximab in an experimental model of indomethacin-induced inflammatory bowel disease. Dig Dis Sci. 2008;53:2516–20.
Zhong Y, Cai D, Cai W, Geng S, Chen L, Han T. Protective effect of galactooligosaccharide-supplemented enteral nutrition on intestinal barrier function in rats with severe acute pancreatitis. Clin Nutr. 2009;28:575–80.
Halász J, Holczbauer A, Páska C, Kovács M, Benyó G, Verebély T, et al. Claudin-1 and claudin-2 differentiate fetal and embryonal components in human hepatoblastoma. Hum Pathol. 2006;37:555–61.
Orbán E, Szabó E, Lotz G, Kupcsulik P, Páska C, et al. Different expression of occludin and ZO-1 in primary and metastatic liver tumors. Pathol Oncol Res. 2008;14:299–306.
Ma TY, Boivin MA, Ye D, Pedram A, Said HM. Mechanism of TNF-{alpha} modulation of Caco-2 intestinal epithelial tight junction barrier: role of myosin light-chain kinase protein expression. Am J Physiol Gastrointest Liver Physiol. 2005;288:G422–30.
Acknowledgments
This work was made possible by the National Health and Medical Research Council, Australia (NHMRC, grant number 510230). Laboratory investigations were accomplished in the Westfield Research Laboratories. Osmolite was provided by Abbott Nutrition Australia.
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The authors declare that they have no conflict of interest.
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Nahidi, L., Day, A.S., Lemberg, D.A. et al. Differential effects of nutritional and non-nutritional therapies on intestinal barrier function in an in vitro model. J Gastroenterol 47, 107–117 (2012). https://doi.org/10.1007/s00535-011-0471-1
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DOI: https://doi.org/10.1007/s00535-011-0471-1