%0 Journal Article
%T Macrophages exposed continuously to lipopolysaccharide and other agonists that act via toll-like receptors exhibit a sustained and additive activation state
%A David A Hume
%A David M Underhill
%A Matthew J Sweet
%A Adrian O Ozinsky
%A Foo Y Liew
%A Alan Aderem
%J BMC Immunology
%D 2001
%I BioMed Central
%R 10.1186/1471-2172-2-11
%X RAW264 macrophage cells were doubly-transfected with reporter genes in which the IL-12p40, ELAM or IL-6 promoter controls firefly luciferase, and the human IL-1¦Â promoter drives renilla luciferase. The resultant stable lines provide robust assays of macrophage activation by TLR stimuli including LPS [TLR4], lipopeptide [TLR2], and bacterial DNA [TLR9], with each promoter demonstrating its own intrinsic characteristics. With each of the promoters, luciferase activity was induced over an 8 hr period, and thereafter reached a new steady state. Elevated expression required the continued presence of agonist. Sustained responses to different classes of agonist were perfectly additive. This pattern was confirmed by measuring inducible cytokine production in the same cells. While homodimerization of TLR4 mediates responses to LPS, TLR2 appears to require heterodimerization with another receptor such as TLR6. Transient expression of constitutively active forms of TLR4 or TLR2 plus TLR6 stimulated IL-12 promoter activity. The effect of LPS, a TLR4 agonist, was additive with that of TLR2/6 but not TLR4, whilst that of lipopeptide, a TLR2 agonist, was additive with TLR4 but not TLR2/6. Actions of bacterial DNA were additive with either TLR4 or TLR2/6.These findings indicate that maximal activation by any one TLR pathway does not preclude further activation by another, suggesting that common downstream regulatory components are not limiting. Upon exposure to a TLR agonist, macrophages enter a state of sustained activation in which they continuously sense the presence of a microbial challenge.Mammalian macrophages respond to a wide range of microbial products with induction of genes required for host defence. Amongst these genes is a suite of inducible cytokines, including IL-1, TNF-¦Á, IL-6 and IL-12, which are required for protective innate and acquired immunity but also mediate much of the pathology of disseminated infections including toxic shock [1]. The archetypal macrophage
%U http://www.biomedcentral.com/1471-2172/2/11