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The Journal of Immunology, 2000, 164: 5564-5574.
Copyright © 2000 by The American Association of Immunologists

Inhibition of Lipopolysaccharide-Induced Signal Transduction in Endotoxin-Tolerized Mouse Macrophages: Dysregulation of Cytokine, Chemokine, and Toll-Like Receptor 2 and 4 Gene Expression1

Andrei E. Medvedev, Karen M. Kopydlowski and Stefanie N. Vogel2

Department of Microbiology and Immunology, Uniformed Services University of the Health Sciences, Bethesda, MD 20814

In this study, the effect of in vitro endotoxin tolerance on LPS-induced mitogen-activated protein kinase activation, transcription factor induction, and cytokine, chemokine, and Toll-like receptor (TLR) 2 and 4 gene expression, as well as the involvement of TNF and IL-1 signaling pathways in tolerance, were examined. Pretreatment of mouse macrophages with LPS inhibited phosphorylation of the extracellular signal-regulated kinases, c-Jun NH2-terminal kinases, and p38 kinase; degradation of I-{kappa}B{alpha} (inhibitory protein that dissociates from NF-{kappa}B) and I-{kappa}Bß; and activation of the transcription factors NF-{kappa}B and AP-1 in response to subsequent LPS stimulation. These changes were accompanied by suppression of LPS-induced expression of mRNA for GM-CSF, IFN-{gamma}-inducible protein-10, KC, JE/monocyte chemoattractant protein-1, macrophage-inflammatory protein-1ß, and macrophage-inflammatory protein-2, with concurrent inhibition of chemokine secretion. In contrast to control cells, endotoxin-tolerant macrophages exhibited an increased basal level of TLR2 mRNA, and failed to increase levels of TLR2 mRNA or to down-regulate TLR4 gene expression upon restimulation with LPS. As judged by transcription factor activation, LPS and IL-1 were found to induce a state of cross-tolerance against each other, while no such reciprocal effect was seen for LPS and TNF-{alpha}. In addition, macrophages from TNFR I/II double knockout mice were LPS tolerizable, and blocking of endogenous TNF-{alpha} with TNFR-Fc fusion protein did not affect the capacity of LPS to tolerize macrophages. These data extend our understanding of LPS-signaling mechanisms that are inhibited in endotoxin-tolerized macrophages and suggest that endotoxin tolerance might result from impaired expression and/or functions of common signaling intermediates involved in LPS and IL-1 signaling.




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