Abstract
In this work, we investigated the potential role of the small G protein RhoA in ethanol-induced tight junction (TJ) protein disassembly and increased intestinal epithelial barrier (IEB) permeability. Our study used Caco-2 cells as an in vitro IEB model and RhoA short hairpin RNA (shRNA) interference to establish whether RhoA plays a role in ethanol-induced TJ opening. RhoA shRNA interference partially inhibited epithelial leakage and restored normal transepithelial electrical resistance (TEER) values in the IEB. Moreover, RhoA shRNA interference prevented a shift in occludin distribution from insoluble to soluble fractions. Additionally, RhoA shRNA interference inhibited the ethanol-induced expression of zonula occludens-1 (ZO-1). Finally, RhoA shRNA interference inhibited an ethanol-induced increase in RhoA activity. The contributions of RhoA to an ethanol-induced increase in IEB permeability are associated with TJ disassembly.
Highlights
The barrier function of intestinal epithelium is provided by tight junctions (TJ), the highly specialized junctional complexes located at the apical end of epithelial cells
Claudins and junction adhesion molecules are the major transmembrane proteins that interact with intracellular plaque proteins, such as zonula occludens (ZOs), including zonula occludens-1 (ZO-1), ZO-2 and ZO-3, which in turn interact with the actin cytoskeleton to anchor occludin and other transmembrane proteins at the apical end of the lateral membrane
The results showed that cell viability, evaluated by the MTT assay, was not altered at ethanol concentrations of less than
Summary
The barrier function of intestinal epithelium is provided by tight junctions (TJ), the highly specialized junctional complexes located at the apical end of epithelial cells. A disruption of tight junctions may lead to increased permeability to allergens, toxins and pathogens, which appears to be a common mechanism involved in the pathogenesis of a number of gastrointestinal diseases, such as inflammatory bowel disease, celiac disease and alcoholic liver disease (ALD). Most recent studies have addressed the possible effect of ethanol on paracellular permeability and the disruption of epithelial tight junctions using a cell culture model of intestinal epithelium, the Caco-2 cell monolayer [2,3,4,5]. The aforementioned study suggests that a possible mechanism exists that drives the ethanol-induced opening of the intestinal epithelial barrier (IEB). An IEB model system was used to evaluate the relationship among TJ protein, IEB permeability and RhoA activation
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