Dysregulation of LPS-induced Toll-like receptor 4-MyD88 complex formation and IL-1 receptor-associated kinase 1 activation in endotoxin-tolerant cells.
Prior exposure to LPS induces a transient state of cell refractoriness to subsequent LPS restimulation, known as endotoxin tolerance. Induction of LPS tolerance has been reported to correlate with decreased cell surface expression of the LPS receptor complex, Toll-like receptor 4 (TLR4)/MD-2. However, other results have underscored the existence of mechanisms of LPS tolerance that operate downstream of TLR4/MD-2. In the present study we sought to delineate further the molecular basis of LPS tolerance by examining the TLR4 signaling pathway in endotoxin-tolerant cells. Pretreatment of human monocytes with LPS decreased LPS-mediated NF-kappaB activation, p38 mitogen-activated protein kinase phosphorylation, and TNF-alpha gene expression, documenting the induction of endotoxin tolerance. FACS and Western blot analyses of LPS-tolerant monocytes showed increased TLR2 expression, whereas TLR4 expression levels were not affected. Comparable levels of mRNA and protein for myeloid differentiation factor 88 (MyD88), IL-1R-associated kinase 1 (IRAK-1), and TNFR-associated factor-6 were found in normal and LPS-tolerant monocytes, while MD-2 mRNA expression was slightly increased in LPS-tolerant cells. LPS induced the association of MyD88 with TLR4 and increased IRAK-1 activity in medium-pretreated cells. In LPS-tolerant monocytes, however, MyD88 failed to be recruited to TLR4, and IRAK-1 was not activated in response to LPS stimulation. Moreover, endotoxin-tolerant CHO cells that overexpress human TLR4 and MD-2 also showed decreased IRAK-1 kinase activity in response to LPS despite the failure of LPS to inhibit cell surface expression of transfected TLR4 and MD-2 proteins. Thus, decreased TLR4-MyD88 complex formation with subsequent impairment of IRAK-1 activity may underlie the LPS-tolerant phenotype.
Medvedev
Andrei E
AE
Department of Microbiology and Immunology, University of Maryland, Baltimore 21201, USA.
Lentschat
Arnd
A
Wahl
Larry M
LM
Golenbock
Douglas T
DT
Vogel
Stefanie N
SN
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000943:Antigens, Differentiation / Q000378:metabolism; D016466:CHO Cells; D006224:Cricetinae; D015536:Down-Regulation / Q000276:immunology*; D029721:Drosophila Proteins; D004789:Enzyme Activation / Q000276:immunology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007108:Immune Tolerance; D053592:Interleukin-1 Receptor-Associated Kinases; D007424:Intracellular Fluid / Q000276:immunology / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology; D009000:Monocytes / Q000201:enzymology / Q000276:immunology / Q000378:metabolism; D053594:Myeloid Differentiation Factor 88; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors* / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology; D011971:Receptors, Immunologic / Q000037:antagonists & inhibitors* / Q000378:metabolism; D017472:Receptors, Interleukin-1 / Q000378:metabolism*; D015398:Signal Transduction / Q000276:immunology; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors
D048868:Adaptor Proteins, Signal Transducing; D000943:Antigens, Differentiation; D029721:Drosophila Proteins; D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; C507779:MYD88 protein, human; D046911:Macromolecular Substances; D008562:Membrane Glycoproteins; D053594:Myeloid Differentiation Factor 88; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D011971:Receptors, Immunologic; D017472:Receptors, Interleukin-1; C495342:TLR2 protein, human; C495345:TLR4 protein, human; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D011494:Protein Kinases; D053592:Interleukin-1 Receptor-Associated Kinases