ClC-2 is required for rapid restoration of epithelial tight junctions in ischemic-injured murine jejunum |
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Authors: | Nighot Prashant K Moeser Adam J Ryan Kathleen A Ghashghaei Troy Blikslager Anthony T |
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Affiliation: | a Department of Clinical Science, College of Veterinary Medicine, North Carolina State University, 4700 Hillsborough Street Raleigh, NC 27606, USA b Department of Population Health and Pathobiology, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, USA c Department of Molecular Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, USA |
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Abstract: | Background and aimsInvolvement of the epithelial chloride channel ClC-2 has been implicated in barrier recovery following ischemic injury, possibly via a mechanism involving ClC-2 localization to the tight junction. The present study investigated mechanisms of intestinal barrier repair following ischemic injury in ClC-2−/− mice.MethodsWild type, ClC-2 heterozygous and ClC-2−/− murine jejunal mucosa was subjected to complete ischemia, after which recovery of barrier function was monitored by measuring in vivo blood-to-lumen clearance of 3H-mannitol. Tissues were examined by light and electron microscopy. The role of ClC-2 in re-assembly of the tight junction during barrier recovery was studied by immunoblotting, immunolocalization and immunoprecipitation.ResultsFollowing ischemic injury, ClC-2−/− mice had impaired barrier recovery compared to wild type mice, defined by increases in epithelial paracellular permeability independent of epithelial restitution. The recovering ClC-2−/− mucosa also had evidence of ultrastructural paracellular defects. The tight junction proteins occludin and claudin-1 shifted significantly to the detergent soluble membrane fraction during post-ischemic recovery in ClC-2−/− mice whereas wild type mice had a greater proportion of junctional proteins in the detergent insoluble fraction. Occludin was co-immunoprecipitated with ClC-2 in uninjured wild type mucosa, and the association between occludin and ClC-2 was re-established during ischemic recovery. Based on immunofluorescence studies, re-localization of occludin from diffuse sub-apical areas to apical tight junctions was impaired in ClC-2−/− mice.ConclusionsThese data demonstrate a pivotal role of ClC-2 in recovery of the intestinal epithelium barrier by anchoring assembly of tight junctions following ischemic injury. |
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Keywords: | Tight junction Intestinal permeability Ischemia ClC-2 |
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