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Overexpression of CD14, TLR4, and MD-2 in HEK 293T cells does not prevent induction of in vitro endotoxin tolerance.
- Source :
-
Journal of endotoxin research [J Endotoxin Res] 2003; Vol. 9 (1), pp. 60-4. - Publication Year :
- 2003
-
Abstract
- TLR4 and MD-2 are necessary for conferring cellular responsiveness to LPS. Prior exposure to LPS induces a transient state of cell refractoriness to subsequent LPS re-stimulation, known as 'endotoxin tolerance'. While induction of LPS tolerance has been reported to correlate with down-regulation of cell surface expression of TLR4/MD-2, other mechanisms of LPS tolerance have been revealed that target intracellular intermediates downstream of the TLR4/MD-2 complex. In this study, we sought to examine whether endotoxin tolerance could be induced under conditions where expression of TLR4 and MD-2 proteins is not affected by LPS. Human HEK 293T cells are completely unresponsive to LPS, but acquire high LPS sensitivity following transient transfection with CD14, TLR4, and MD-2 (293T/CD14/TLR4/MD-2 cells), as judged by NF-kappaB activation, ERK 1/2 phosphorylation, and TNF-alpha gene expression. Prior exposure of 293T/CD14/TLR4/MD-2 cells to LPS resulted in a significant decrease of LPS-mediated responses, yet failed to affect expression levels of TLR4 and MD-2. Thus, altered expression and/or function of intracellular mediators downstream of the TLR4/MD-2 complex play an important role in mediating LPS tolerance.
- Subjects :
- Blotting, Western
Cell Line drug effects
Cell Line immunology
Cell Line metabolism
Dose-Response Relationship, Drug
Down-Regulation immunology
Drug Tolerance
Escherichia coli immunology
Gene Expression
Humans
Kidney
Lipopolysaccharides pharmacology
Lymphocyte Antigen 96
Mitogen-Activated Protein Kinase 1 genetics
Mitogen-Activated Protein Kinase 1 metabolism
Mitogen-Activated Protein Kinase 3
Mitogen-Activated Protein Kinases genetics
Mitogen-Activated Protein Kinases metabolism
NF-kappa B genetics
NF-kappa B metabolism
Phosphorylation
RNA, Messenger metabolism
Reverse Transcriptase Polymerase Chain Reaction
Toll-Like Receptor 4
Toll-Like Receptors
Transfection
Tumor Necrosis Factor-alpha genetics
Tumor Necrosis Factor-alpha metabolism
Antigens, Surface metabolism
Immune Tolerance drug effects
Immune Tolerance immunology
Lipopolysaccharide Receptors metabolism
Membrane Glycoproteins metabolism
Receptors, Cell Surface metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0968-0519
- Volume :
- 9
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- Journal of endotoxin research
- Publication Type :
- Academic Journal
- Accession number :
- 12691621
- Full Text :
- https://doi.org/10.1179/096805103125001360