THE INVOLVEMENT OF MACROPHAGE-DERIVED TUMOR-NECROSIS-FACTOR AND LIPOXYGENASE PRODUCTS ON THE NEUTROPHIL RECRUITMENT INDUCED BY CLOSTRIDIUM-DIFFICILE TOXIN-B

Clostridium difficile (Cd) toxins appear to mediate the inflammatory r esponse in pseudomembranous colitis and/or colitis associated with the use of antibiotics. In contrast to Cd Toxin A (TxA), Cd Toxin B (TxB) has been reported not to promote fluid secretion or morphological dam age in rabbits and hamsters and also does not induce neutrophil chemot axis in vitro. However, TxB is about 1000 times more patent than TxA i n stimulating the release of tumour necrosis factor-alpha (TNF-alpha) by cultured monocytes. In the present study, we investigated the abili ty of TxB to promote neutrophil migration into peritoneal cavities and subcutaneous air-pouches of rats. We also examined the role of reside nt peritoneal cells in this process as well as the inflammatory mediat ors involved. TxB caused a significant and dose-dependent neutrophil i nflux with a maximal response at 0.1 mu g/cavity after 4 hr. Depleting the peritoneal resident cell population by washing the peritoneal cav ity or increasing this population by pretreating the animals with thio glycollate blocked and amplified the TxB-induced neutrophil migration, respectively. Pretreating the animals with MK886 (a lipoxygenase inhi bitor), NDGA (a dual cyclo- and lipoxygenase inhibitor) or the glucoco rticoid, dexamethasone, but not with indomethacin (a cyclo-oxygenase i nhibitor), or BN52021 (a platelet-activating factor antagonist), inhib ited the neutrophil migration evoked by TxB. Pretreatment with dexamet hasone or the administration of anti-TNF-alpha serum into the air-pouc hes also significantly reduced the TxB-induced neutrophil migration. S upernatants from TxB-stimulated macrophages induced neutrophil migrati on when injected into the rat peritoneal cavity. This effect was atten uated by the addition of either MK886 or dexamethasone to the macropha ge monolayer and by preincubating the supernatants with anti-TNF-alpha serum. TxB also stimulated the release of TNF-alpha by macrophages. O verall, these results suggest that TxB induces an intense neutrophil m igration which is mediated by macrophage-derived TNF-alpha and lipoxyg enase products.