title: Macrophage-dependent stimulation of T cell-depleted spleen cells by Clostridium difficile toxin A and calcium ionophore.
abstract: Clostridium difficile toxin A causes severe intestinal inflammation and fluid secretion in rabbit ileum and is chemotactic for neutrophils in vitro. The mechanism of intestinal injury produced by toxin A appears to involve direct epithelial cell damage as well as recruitment of an inflammatory cell response. The current study was undertaken to determine if toxin A can directly stimulate a proliferative response in lymphocytes. Highly purified toxin A, in the presence of the calcium ionophore, ionomycin, stimulated substantial [3H]thymidine incorporation by murine splenic lymphocytes, which was maximal at 10(-9) M toxin A and 800 ng/ml ionomycin. Removal of T cells with anti-Thy-1.2 antibody plus complement had no effect on the proliferative response induced by toxin A. However, [3H]thymidine incorporation in response to toxin A was significantly inhibited (P less than 0.001) by the removal of macrophages from splenocyte suspensions and was restored by the addition of peritoneal macrophages or cell-free supernatant from toxin A-treated macrophage cultures. Analysis of the toxin A-treated macrophage supernatants showed high levels of IL-1, but not IL-2 or IL-4. The combination of recombinant IL-1 plus ionomycin was found to stimulate [3H]thymidine incorporation by T cell-depleted splenic lymphocytes. These results suggest that toxin A stimulates the release of IL-1, and possibly other factors, from macrophages which can costimulate murine B lymphocytes.
authors.affiliation: Evans Memorial Department of Clinical Research, Boston University Medical Center, Massachusetts 02118.
authors.forename: P D
authors.initials: PD
authors.lastname: Miller
authors.forename: C
authors.initials: C
authors.lastname: Pothoulakis
authors.forename: T R
authors.initials: TR
authors.lastname: Baeker
authors.forename: J T
authors.initials: JT
authors.lastname: LaMont
authors.forename: T L
authors.initials: TL
authors.lastname: Rothstein
mesh_terms: D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology*; D002478:Cells, Cultured; D003013:Clostridium / Q000472:pathogenicity*; D004768:Enterotoxins / Q000494:pharmacology*; D007375:Interleukin-1 / Q000378:metabolism / Q000494:pharmacology; D015759:Ionomycin / Q000494:pharmacology*; D008213:Lymphocyte Activation / Q000187:drug effects*; D008212:Lymphocyte Depletion; D008264:Macrophages / Q000187:drug effects / Q000502:physiology*; D008297:Male; D051379:Mice; D013601:T-Lymphocytes / Q000502:physiology*
chemical_list: D001427:Bacterial Toxins; D004768:Enterotoxins; D007375:Interleukin-1; C041348:tcdA protein, Clostridium difficile; D015759:Ionomycin