Abstract
Previous work has suggested that the peptide-carrier, heat-shock protein (hsp)70, could directly activate APC. Here we show that this ability is related to endotoxin contamination of the human rhsp70 produced in Escherichia coli. Hence, the ability of 1-3 microg/ml of rhsp70 to induce the maturation of human monocyte-derived DC is abrogated in the presence of the LPS-antagonist polymyxin B or when the rhsp70 contains less than 60 IU/mg endotoxin. Such a level of contamination of the rhsp70 is, however, sufficient - in the presence of soluble rCD14, the LPS co-receptor - to induce cytokine secretion from monocytes and DC, despite the presence of polymyxin B. However, when endotoxin contamination is below 10 IU/mg, rhsp70 does not induce cytokine secretion - even in the presence of soluble rCD14 - or activate p38 mitogen-activated protein kinase signaling pathways, thus showing that an "endotoxin free" hsp70 does not activate APC.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Amino Acid Sequence
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Antigen-Presenting Cells / cytology
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Antigen-Presenting Cells / drug effects*
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Antigen-Presenting Cells / enzymology
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Antigen-Presenting Cells / immunology*
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Cell Differentiation / drug effects
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Cytokines / biosynthesis
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Dendritic Cells / cytology
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Dendritic Cells / drug effects
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Dendritic Cells / enzymology
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Dendritic Cells / immunology
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Drug Contamination
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Endotoxins / antagonists & inhibitors
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Endotoxins / isolation & purification
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Endotoxins / pharmacology
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Enzyme Activation / drug effects
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HSP70 Heat-Shock Proteins / genetics
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HSP70 Heat-Shock Proteins / isolation & purification
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HSP70 Heat-Shock Proteins / pharmacology*
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Humans
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In Vitro Techniques
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Mitogen-Activated Protein Kinases / metabolism
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Molecular Sequence Data
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Monocytes / cytology
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Monocytes / drug effects
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Monocytes / enzymology
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Monocytes / immunology
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Polymyxin B / pharmacology
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Recombinant Proteins / genetics
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Recombinant Proteins / isolation & purification
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Recombinant Proteins / pharmacology
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Signal Transduction / drug effects
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p38 Mitogen-Activated Protein Kinases
Substances
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Cytokines
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Endotoxins
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HSP70 Heat-Shock Proteins
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Recombinant Proteins
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Mitogen-Activated Protein Kinases
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p38 Mitogen-Activated Protein Kinases
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Polymyxin B