Bacterial DNA and lipopolysaccharide induce synergistic production of TNF-α through a post-transcriptional mechanism

LPS is well recognized for its potent capacity to activate mouse macrophages to produce TNF-α, an important inflammatory mediator in bacterial infection-related diseases such as septic shock. We demonstrate here that while inducing only Iow levels of TNF-α alone, DNA from both Gram-negative and Gram-positive bacteria synergizes with subthreshold concentrations of LPS (0.3 ng/ml) to induce TNF-α in the RAW 264.7 macrophage-like cell line. The bacterial DNA effects are mimicked by synthetic CpG-containing oligodeoxynucleotides, but not non-CpG-containing oligodeoxynucleotides. Pretreatment of macrophages with either DNA for 2-8 h inhibits macrophage TNF-α production in responses to DNA/LPS. However, when pretreatment was extended to 24 h, DNA/LPS synergy on TNF-α is further enhanced. RT-PCR analysis indicates that mRNA levels of the TNF-α gene, however, are not synergistically induced by bacterial DNA and LPS. Analyses of the half-life of TNF-α mRNA indicate that TNF-α message has a longer half-life in bacterial DNA- and LPS-treated macrophages than that in bacterial DNA- or LPS-treated macrophages. These findings indicate that the temporally controlled, synergistic induction of TNF-α by bacterial DNA and LPS is not mediated at the transcriptional level. Instead, this synergy may occur via a post-transcriptional mechanism.