TLR2 and TLR4 agonists stimulate unique repertoires of host resistance genes in murine macrophages: interferon-beta-dependent signaling in TLR4-mediated responses.

That TLRs share a common MyD88-dependent signaling pathway which results in the generation of nuclear DNA-binding proteins, such as NF-kappaB, is a well-accepted paradigm. However, studies from our laboratories and others suggested that TLR4 agonists elicit a more diverse pattern of gene expression in murine macrophages than TLR2 agonists. The data presented show that activation of TLR4 by Escherichia coli LPS results in an MyD88-independent, TIRAP/Mal-dependent signaling pathway that, in turn, leads to early induction of interferon-beta (IFN-beta). IFN-beta, in turn, acts in an autocrine/paracrine fashion on the macrophage to activate STAT1-containing DNA binding complexes that participate in the induction of genes not expressed in response to natural or synthetic TLR2 agonists. These data support the hypothesis that the host response to microbes is controlled by TLRs at two levels: (i) the "sensing" of differences in microbial structures through the TLR extracellular domain; and (ii) signaling pathways that are initiated via interactions through unique intracytoplasmic regions of different TLRs with adaptor proteins.

Mice deficient in the CXCR2 ligand, CXCL1 (KC/GRO-alpha), exhibit increased susceptibility to dextran sodium sulfate (DSS)-induced colitis.

The role of TLRs and MyD88 in the maintenance of gut integrity in response to dextran sodium sulfate (DSS)-induced colitis was demonstrated recently and led to the conclusion that the innate immune response to luminal commensal flora provides necessary signals that facilitate epithelial repair and permits a return to homeostasis after colonic injury. In this report, we demonstrate that a deficit in a single neutrophil chemokine, CXCL1/KC, also results in a greatly exaggerated response to DSS. Mice with a targeted mutation in the gene that encodes this chemokine responded to 2.5% DSS in their drinking water with significant weight loss, bloody stools, and a complete loss of gut integrity in the proximal and distal colon, accompanied by a predominantly mononuclear infiltrate, with few detectable neutrophils. In contrast, CXCL1/KC(- /-) and wild-type C57BL/6J mice provided water showed no signs of inflammation and, at this concentration of DSS, wild-type mice showed only minimal histopathology, but significantly more infiltrating neutrophils. This finding implies that neutrophil infiltration induced by CXCL1/KC is an essential component of the intestinal response to inflammatory stimuli as well as the ability of the intestine to restore mucosal barrier integrity.

TLR4, but not TLR2, mediates IFN-beta-induced STAT1alpha/beta-dependent gene expression in macrophages.

Toll-like receptor 2 (TLR2) agonists induce a subset of TLR4-inducible proinflammatory genes, which suggests the use of differential signaling pathways. Murine macrophages stimulated with the TLR4 agonist Escherichia coli lipopolysaccharide (LPS), but not with TLR2 agonists, induced phosphorylation of signal transducer and activator of transcription 1alpha (STAT1alpha) and STAT1beta, which was blocked by antibodies to interferon beta (IFN-beta) but not IFN-alpha. All TLR2 agonists poorly induced IFN-beta, which is encoded by an immediate early LPS-inducible gene. Thus, the failure of TLR2 agonists to induce STAT1-dependent genes resulted, in part, from their inability to express IFN-beta. TLR4-induced IFN-beta mRNA was MyD88- and PKR (double-stranded RNA-dependent protein kinase)-independent, but TIRAP (Toll-interleukin 1 receptor domain-containing adapter protein)-dependent. Together, these findings provide the first mechanistic basis for differential patterns of gene expression activated by TLR4 and TLR2 agonists.