Christian Barmeyer1,2, Irene Erko1, Karem Awad2, Anja Fromm2, Christian Bojarski1, Svenja Meissner1, Christoph Loddenkemper3,4, Martin Kerick5, Britta Siegmund1, Michael Fromm2, Michal R Schweiger5,6, Jörg-Dieter Schulzke7,8. 1. Department of Gastroenterology, Infectious Diseases and Rheumatology, Charité, Campus Benjamin Franklin, Berlin, Germany. 2. Institute of Clinical Physiology, Charité, Campus Benjamin Franklin, Hindenburgdamm 30, 12203, Berlin, Germany. 3. Institute of Pathology, Charité, Campus Benjamin Franklin, Berlin, Germany. 4. Institute of Pathology PathoTres, Berlin, Germany. 5. Max Planck Institute for Molecular Genetics, Berlin, Germany. 6. Cologne Center for Genomics, University of Cologne, Cologne, Germany. 7. Department of Gastroenterology, Infectious Diseases and Rheumatology, Charité, Campus Benjamin Franklin, Berlin, Germany. joerg.schulzke@charite.de. 8. Institute of Clinical Physiology, Charité, Campus Benjamin Franklin, Hindenburgdamm 30, 12203, Berlin, Germany. joerg.schulzke@charite.de.
Abstract
BACKGROUND: Watery diarrhea is the cardinal symptom of lymphocytic colitis (LC). We have previously shown that colonic Na malabsorption is one of the major pathologic alterations of LC and found evidence for an epithelial barrier defect. On these grounds, this study aimed to identify the inherent mechanisms of this epithelial barrier dysfunction and its regulatory features. METHODS: Epithelial resistance (R epi) was determined by one-path impedance spectroscopy and 3H-mannitol fluxes were performed on biopsies from sigmoid colon in miniaturized Ussing chambers. Tight junction proteins were analyzed by Western blot and confocal microscopy. Inflammatory signaling was characterized in HT-29/B6 cells. Apoptosis and mucosal surface parameters were quantified morphologically. RESULTS: R epi was reduced to 53% and 3H-mannitol fluxes increased 1.7-fold in LC due to lower expression of claudin-4, -5, and -8 and altered subcellular claudin-5 and -8 distributions off the tight junction. TNFα and IFNγ could mimic subcellular redistribution in HT-29/B6 cells, a process which was independent on MLCK activation. Epithelial apoptosis did not contribute to barrier dysfunction in LC and mucosal surface area was unchanged. CONCLUSIONS: Epithelial barrier dysfunction in LC occurs through downregulation of claudin-4, -5, and -8, and redistribution of claudin-5 and -8 off the tight junction, which contributes to diarrhea by a leak-flux mechanism. The key effector cytokines TNFα and IFNγ turned out to be the trigger for redistribution of claudin-5 and -8. Thus, alongside sodium malabsorption, leak-flux is yet another important diarrheal mechanism in LC.
BACKGROUND: Watery diarrhea is the cardinal symptom of lymphocytic colitis (LC). We have previously shown that colonic Na malabsorption is one of the major pathologic alterations of LC and found evidence for an epithelial barrier defect. On these grounds, this study aimed to identify the inherent mechanisms of this epithelial barrier dysfunction and its regulatory features. METHODS: Epithelial resistance (R epi) was determined by one-path impedance spectroscopy and 3H-mannitol fluxes were performed on biopsies from sigmoid colon in miniaturized Ussing chambers. Tight junction proteins were analyzed by Western blot and confocal microscopy. Inflammatory signaling was characterized in HT-29/B6 cells. Apoptosis and mucosal surface parameters were quantified morphologically. RESULTS: R epi was reduced to 53% and 3H-mannitol fluxes increased 1.7-fold in LC due to lower expression of claudin-4, -5, and -8 and altered subcellular claudin-5 and -8 distributions off the tight junction. TNFα and IFNγ could mimic subcellular redistribution in HT-29/B6 cells, a process which was independent on MLCK activation. Epithelial apoptosis did not contribute to barrier dysfunction in LC and mucosal surface area was unchanged. CONCLUSIONS: Epithelial barrier dysfunction in LC occurs through downregulation of claudin-4, -5, and -8, and redistribution of claudin-5 and -8 off the tight junction, which contributes to diarrhea by a leak-flux mechanism. The key effector cytokines TNFα and IFNγ turned out to be the trigger for redistribution of claudin-5 and -8. Thus, alongside sodium malabsorption, leak-flux is yet another important diarrheal mechanism in LC.
Authors: Christian Barmeyer; Irene Erko; Anja Fromm; Christian Bojarski; Christoph Loddenkemper; Petra Dames; Martin Kerick; Britta Siegmund; Michael Fromm; Michal R Schweiger; Jörg-Dieter Schulzke Journal: Inflamm Bowel Dis Date: 2016-03 Impact factor: 5.325
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