Superinduction of TNF-alpha and IL-6 in macrophages by vomitoxin (deoxynivalenol) modulated by mRNA stabilization.

ABSTRACT

Vomitoxin (VT or deoxynivalenol), a trichothecene, superinduces proinflammatory cytokine gene expression in vitro and in vivo. To better understand the underlying molecular mechanisms for this observation, post-transcriptional effects of VT on TNF-alpha and IL-6 gene expression were studied in lipopolysaccharide (LPS)-stimulated macrophage RAW 264.7 cells. VT was found to enhance both TNF-alpha and IL-6 protein secretion in the presence of LPS. Upon addition of the transcriptional inhibitor, 5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB), secretion of both cytokines was inhibited. Using Northern analysis, the mRNA stabilities of TNF-alpha and IL-6 were studied in DRB-treated cells exposed to VT and LPS in both asynchronous and delayed synchronous modes. In the asynchronous model, cells were first incubated with LPS for 2 h, and then the medium was removed and replaced with medium containing DRB and VT. In the delayed synchronous model, cells were pretreated with LPS for 2 h and then DRB and VT were added to the culture. TNF-alpha and IL-6 mRNA were rapidly stabilized by VT (100 and 250 ng/ml) in both asynchronous and delayed synchronous models. In the asynchronous model, TNF-alpha mRNA half-life was 25 min but this was extended in the presence of 100 and 250 ng/ml of VT to >3 h. VT also extended half-lives of IL-6 mRNA from 60 min to >3 h. In the delayed synchronous model, the half-lives for TNF-alpha and IL-6 mRNA of 1.3 and 1.5 h, respectively, were extended to >3 h upon incubation with 100 and 250 ng/ml VT. These results suggest that post-transcriptional control via enhancement of mRNA stability is likely to contribute to proinflammatory cytokine superinduction in macrophages by VT and other trichothecenes.