alexa Toll-like receptor responses of normal human urothelial cells to bacterial flagellin and lipopolysaccharide.
General Science

General Science

Journal of Bioterrorism & Biodefense

Author(s): Smith NJ, Varley CL, Eardley I, Feather S, Trejdosiewicz LK,

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Abstract PURPOSE: We determined toll-like receptor expression in normal human urothelium and functional responses in normal human urothelial cell cultures to bacterial lipopolysaccharide via toll-like receptor-4 and to flagellin via toll-like receptor-5. MATERIALS AND METHODS: Toll-like receptor protein expression was examined immunohistochemically. Toll-like receptor transcript expression was determined in freshly isolated urothelium, and in proliferating and differentiated normal human urothelial cultured cells. Lipopolysaccharide binding was assessed by flow cytometry. Functional responses of proliferating and differentiated normal human urothelial cells to lipopolysaccharide and flagellin were determined by interleukin-6 and 8 secretion, and transcription factor activation. Polymyxin B and siRNA were used to confirm the specificity of toll-like receptor-4 and 5 responses, respectively. Western blot detection of phosphorylated IκB was used to confirm toll-like receptor-4 results. RESULTS: Human urothelium expressed transcripts for toll-like receptor-4 and 5. Although bladder cancer derived T24 cells responded to lipopolysaccharide, there was no lipopolysaccharide binding to normal human urothelial cells and no functional response of proliferative or differentiated normal human urothelial cells even in the presence of exogenous CD14 and MD-2 accessory proteins. In contrast, flagellin evoked a toll-like receptor-5 mediated response in proliferating but not in differentiated normal human urothelial cells, which was abrogated by toll-like receptor-5 specific siRNA. CONCLUSIONS: Results suggest that human urothelium may mediate a host response to uropathogenic Escherichia coli through the detection of flagellin. The absent constitutive toll-like receptor-4 response may reflect an adaptation of urothelium toward sustaining barrier function and limiting inflammation to soluble bacterial products. Copyright © 2011 American Urological Association Education and Research, Inc. Published by Elsevier Inc. All rights reserved. This article was published in J Urol and referenced in Journal of Bioterrorism & Biodefense

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