Common and distinct signalling cascades in the production of tumour necrosis factor-α and interleukin-13 induced by lipopolysaccharide in RBL-2H3 cells.

Activation of mast cells by lipopolysaccharide (LPS) results in the production of TNF-α and IL-13. TNF-α and IL-13 are key mediators in the development of neutrophilic and allergic inflammation, respectively. LPS-induced TNF-α and IL-13 production in mast cells has been reported to be mediated by Toll-like receptor 4 (TLR4) signalling, but differences in signal transduction mechanisms leading to the production of these cytokines are not clearly defined.We investigated the molecular mechanisms responsible for LPS-induced TNF-α and IL-13 production in mast cells.TNF-α and IL-13 production by LPS was assessed by transfecting RBL-2H3 cells with dominant-negative (DN) expression vectors.Transfection of RBL-2H3 cells with plasmids encoding DN mutants of myeloid differentiation protein (MyD88) and TNFR-associated factor (TRAF6) inhibited both LPS-induced TNF-α and IL-13 production. IκBα-DN inhibited LPS-induced production of TNF-α, but not IL-13. We also found that inhibition of p38 kinase suppressed both TNF-α and IL-13 induction by LPS, and inhibition of JNK reduced IL-13 production, but not TNF-α. Furthermore, we found that protein kinase R (PKR) was activated by LPS in these cells. Treatment with 2-aminopurine, a PKR inhibitor, attenuated LPS-induced nuclear factor-κB activation and TNF-α production, whereas inhibition of PKR had little effect on IL-13 production.These findings indicate that the production of TNF-α and IL-13 by LPS required TLR4/MyD88/TRAF6 signalling as a common pathway of mast cell-mediated inflammation. We furthermore found that TNF-α and IL-13 production were differentially regulated by signalling cascades through PKR and mitogen-activated protein kinases downstream of TRAF6 in mast cells. [ABSTRACT FROM AUTHOR]