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Canadian Journal of Infectious Diseases
Volume 15, Issue 2, Pages 83-88
http://dx.doi.org/10.1155/2004/292580
Original Article

Translocation of Clostridium difficile Toxin B across Polarized Caco-2 Cell Monolayers Is Enhanced by Toxin A

Tim Du1 and Michelle J Alfa2

1Department of Medical Microbiology, University of Manitoba, Winnipeg, Manitoba, Canada
2St Boniface General Hospital, Winnipeg, Manitoba, Canada

Received 3 October 2003; Accepted 10 October 2003

Copyright © 2004 Hindawi Publishing Corporation. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Clostridium difficile is the etiological agent of antibiotic-associated diarrhea; the most common form of nosocomial infectious diarrhea. The basis for the shock-like systemic symptoms observed in severe cases of this infection are not known. It is hypothesized that the invasion of C difficile toxins A and/or B from the gut mucosa may contribute to these symptoms.

A polarized tissue culture model employing Caco-2 cells grown on transwell inserts was established to study the translocation of purified C difficile toxins A and B. C difficile toxins were 125I labelled and inoculated onto confluent polarized Caco-2 cell monolayers to study translocation dynamics. Electrical resistance measurements were used to monitor monolayer confluence and tight junction integrity. Samples were taken from the apical and basal sides of the insert, as well as the insert itself, and tested using the human foreskin fibroblasts cell cytotoxicity assay to monitor partitioning of the radiolabelled toxins.

Toxin A produced a 50% reduction in electrical resistance in 3 h whereas the same concentration of toxin B required at least 7 h to achieve the same effect. Both toxins A and B were able to translocate across confluent monolayers of Caco-2 cells. The combination of toxin A and B together was synergistic with respect to promoting the translocation of toxin B. Although the addition of toxin A resulted in a 100% increase in the amount of toxin B able to translocate, no increases in toxin A translocation were observed. These findings suggest a model of pathogenesis in which C difficile toxin A facilitates the translocation of toxin B from the gut into submucosal areas where it may play a role in inflammatory damage.