Essential Involvement of IFN-γ in Clostridium difficile Toxin A-Induced Enteritis

Abstract

Clostridium difficile has emerged as the important causative agent of antibiotics-associated pesudomembranous colitis; especially its toxin A is presumed to be responsible for the colitis. We examined the pathophysiological roles of IFN-γ in toxin A-induced enteritis using IFN-γ knockout (KO) mice. When toxin A of C. difficile was injected into the ileal loops of BALB/c wild-type (WT) mice, massive fluid secretion, disruption of intestinal epithelial structure, and massive neutrophil infiltration developed within 4 h after the injection. IFN-γ protein was faintly detected in some CD3-positive lymphocytes in the lamina propria and submucosa of the ileum of untreated WT mice. On the contrary, at 2 and 4 h after toxin A injection, IFN-γ protein was detected in infiltrating neutrophils and to a lesser degree in CD3-positive lymphocytes. In the ileum of WT mice, toxin A treatment markedly enhanced the gene expression of TNF-α, macrophage inflammatory protein-1α and -2, KC, and ICAM-1 >2 h after treatment. In contrast, the histopathological changes were marginal, without enhanced fluid secretion in the ileum of toxin A-treated IFN-γ KO mice. Moreover, toxin A-induced gene expression of TNF-α, neutrophil chemotactic chemokines, and ICMA-1 was remarkably attenuated in IFN-γ KO mice. Furthermore, pretreatment of WT mice with a neutralizing anti-IFN-γ Ab prevented toxin A-induced enteritis. These observations indicate that IFN-γ is the crucial mediator of toxin A-induced acute enteritis and suggest that IFN-γ is an important molecular target for the control of C. difficile-associated pseudomembranous colitis.

Chemokine Receptor CX3CR1 Mediates Skin Wound Healing by Promoting Macrophage and Fibroblast Accumulation and Function

“…Wound sections were also analyzed by double-or triple-color immunofluorescence microscopy to determine the localization of CX3CL1, CX3CR1, and CCR2 proteins as described previously (30,31). Briefly, deparaffinized sections were incubated with PBS containing 1% normal donkey serum and 1% BSA to reduce nonspecific reactions.…”

Clostridium difficile Toxin A Regulates Inducible Cyclooxygenase-2 and Prostaglandin E2 Synthesis in Colonocytes via Reactive Oxygen Species and Activation of p38 MAPK

“…tory cytokines that mediate a profound and rapid inflammatory response (5)(6)(7). The induction of fluid secretion and inflammation by toxin A involves extensive signaling cross-talk between epithelial cells, mast cells, sensory neurons, and inflammatory cells of the intestinal lamina propria (4).…”

Protective Roles of CX3CR1-Mediated Signals in Toxin A-Induced Enteritis through the Induction of Heme Oxygenase-1 Expression

“…Deparaffinized 6-mm-thick sections were stained with H&E solution, and histological changes were graded by an examiner without any prior knowledge of the experimental procedures as described previously (8). In parallel, immunohistochemical analysis with anti-Gr-1 mAb, anti-F4/80 mAb, anti-CX3CL1 pAbs, anti-CX3CR1 pAbs, and anti-HO-1 pAbs was conducted as described previously (8).…”

“…Toxin A-induced enteritis was induced as described previously (8). Briefly, mice were anesthetized by i.p.…”

“…C. difficile colitis is characterized by diarrhea, epithelial cell destruction, pseudomembrane formation, and increased production of proinflammatory mediators (7). C. difficile produces two exotoxins, toxins A and B. Toxin A challenge increases fluid secretion and causes mucosal injury with massive neutrophil recruitment in animals (8,9).…”