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Salmonella typhimurium translocates flagellin across intestinal epithelia, inducing a proinflammatory response
Andrew T. Gewirtz, … , Andrew S. Neish, James L. Madara
Andrew T. Gewirtz, … , Andrew S. Neish, James L. Madara
Published January 1, 2001
Citation Information: J Clin Invest. 2001;107(1):99-109. https://doi.org/10.1172/JCI10501.
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Article

Salmonella typhimurium translocates flagellin across intestinal epithelia, inducing a proinflammatory response

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Abstract

This study investigated whether soluble paracrine factors mediated Salmonella-induced IL-8 expression in polarized model intestinal epithelia. We found that the basolateral media of model epithelia that had been apically infected with Salmonella typhimurium for a short period (10 minutes) could activate IL-8 secretion in virgin model epithelia, demonstrating that a proinflammatory factor (PIF) was indeed present. Initial characterization found that PIF was a heat-stable protein with a molecular mass of about 50 kDa that acts on the basolateral, but not apical, surface of model intestinal epithelia to elicit IL-8 secretion. PIF was not present in the media of model epithelia stimulated with other inducers of IL-8 secretion (TNF-α or carbachol) but was present in S. typhimurium supernatants, indicating PIF is of bacterial origin. PIF was purified from bacterial culture supernatants by anion/cation exchange chromatography and SDS-PAGE and found by using microsequencing to be the protein flagellin. In support of this finding, flagellin-deficient S. typhimurium mutants did not secrete detectable levels of PIF (i.e., a bioactivity that induced IL-8 secretion when placed basolaterally on model epithelia). Furthermore, viable flagellin-deficient mutant organisms (fliC/fljB and flhD) failed to elicit IL-8 secretion when added apically to model intestinal epithelia. These findings indicate that translocation of flagellin across epithelia, subsequent to apical epithelial–S. typhimurium interaction, is likely a major means of activating a mucosal inflammatory response.

Authors

Andrew T. Gewirtz, Peter O. Simon Jr., Clare K. Schmitt, Laura J. Taylor, Curt H. Hagedorn, Alison D. O’Brien, Andrew S. Neish, James L. Madara

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Figure 1

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Basolaterally conditioned media of S. typhimurium–infected epithelia ind...
Basolaterally conditioned media of S. typhimurium–infected epithelia induce IL-8 secretion in virgin epithelia. Model intestinal epithelia were apically exposed to S. typhimurium. The epithelial media was then transferred to untreated model epithelia. IL-8 was assayed in basolateral media 5 hours later. (a) Media was transferred maintaining its polarity of origin (i.e., apical to apical, basolateral to basolateral) 1 hour after addition of bacteria. Live S. typhimurium (added apically for 5 hours) served as a positive control. (b) Basolateral media was transferred at the indicated time after addition of bacteria.

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ISSN: 0021-9738 (print), 1558-8238 (online)

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