Abstract
Diarrhea-associated hemolytic uremic syndrome (D(+)HUS) is caused by the ingestion of Escherichia coli that produce Shiga toxin (Stx), which is composed of a cytotoxic A subunit and pentameric B subunits that bind globotriaosylceramide on susceptible cells. Stx occurs in 2 types, Stx1 and Stx2. B subunits of either type stimulate von Willebrand factor (VWF) secretion from human umbilical vein endothelial cells (HUVECs), and Stx2B can cause thrombotic microangiopathy in Adamts13(-/-) mice. We have now determined that Stx1B and Stx2B activate different signaling pathways in HUVECs. VWF secretion induced by Stx1B is associated with a transient rise in intracellular Ca(2+) level that is blocked by chelation with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester, removal of extracellular Ca(2+), the phospholipase C inhibitor U73122, the protein kinase inhibitor staurosporine, or small interfering RNA knockdown of protein kinase Cα. In contrast, Stx2B-induced VWF secretion is associated with activation of protein kinase A (PKA) and is blocked by the PKA inhibitor H89 or small interfering RNA knockdown of PKA. Stx2B does not increase cAMP levels and may activate PKA by a cAMP-independent mechanism. The activation of distinct signaling pathways may be relevant to understanding why E coli that express Stx2 are more likely to cause D(+)HUS than are E coli expressing only Stx1.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Animals
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Calcium / metabolism
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Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
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Cyclic AMP-Dependent Protein Kinases / metabolism
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Diarrhea / metabolism*
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Diarrhea / microbiology
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Diarrhea / pathology
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Egtazic Acid / analogs & derivatives
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Egtazic Acid / metabolism
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Endothelial Cells / cytology
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Endothelial Cells / drug effects
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Endothelial Cells / metabolism*
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Escherichia coli / chemistry
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Escherichia coli / metabolism
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Escherichia coli Infections / metabolism*
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Escherichia coli Infections / microbiology
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Escherichia coli Infections / pathology
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Gene Silencing / drug effects
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Hemolytic-Uremic Syndrome / metabolism*
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Hemolytic-Uremic Syndrome / microbiology
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Hemolytic-Uremic Syndrome / pathology
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Humans
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Mice
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Protein Binding
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Protein Kinase C-alpha / antagonists & inhibitors
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Protein Kinase C-alpha / metabolism
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Protein Kinase Inhibitors / pharmacology
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RNA, Small Interfering / pharmacology
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Shiga Toxin 1* / adverse effects
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Shiga Toxin 1* / pharmacology
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Shiga Toxin 2* / adverse effects
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Shiga Toxin 2* / pharmacology
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Signal Transduction* / drug effects
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Trihexosylceramides / metabolism
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Umbilical Veins / cytology
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von Willebrand Factor / metabolism*
Substances
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Protein Kinase Inhibitors
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RNA, Small Interfering
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Shiga Toxin 1
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Shiga Toxin 2
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Trihexosylceramides
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von Willebrand Factor
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Egtazic Acid
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globotriaosylceramide
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Cyclic AMP-Dependent Protein Kinases
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Protein Kinase C-alpha
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1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid
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Calcium