Mechanism of thrombin mediated eNOS phosphorylation in endothelial cells is dependent on ATP levels after stimulation.

2.50
Hdl Handle:
http://hdl.handle.net/2336/42233
Title:
Mechanism of thrombin mediated eNOS phosphorylation in endothelial cells is dependent on ATP levels after stimulation.
Authors:
Thors, Brynhildur; Halldorsson, Haraldur; Jonsdottir, Gudbjorg; Thorgeirsson, Gudmundur
Citation:
Biochim. Biophys. Acta. 2008, 1783(10):1893-902
Issue Date:
1-Oct-2008
Abstract:
Conflicting results have been reported concerning the role of AMP-activated protein kinase (AMPK) in mediating thrombin stimulation of endothelial NO-synthase (eNOS). We examined the involvement of two upstream kinases in AMPK activation in cultured human umbilical endothelial cells, LKB1 stimulated by a rise in intracellular AMP/ATP ratio, and Ca(+2)/CaM kinase kinase (CaMKK) responding to elevation of intracellular Ca(+2). We also studied the effects of AMPK activation on the downstream target eNOS. In culture medium 1640 the level of intracellular ATP was unchanged after thrombin stimulation and the CaMKK inhibitor STO-609 totally inhibited phosphorylation of AMPK and acetyl coenzyme A carboxylase (ACC) but not eNOS. In Morgan's medium 199 thrombin caused a significant lowering of intracellular ATP and STO-609 only partially inhibited the phosphorylation of AMPK, ACC and eNOS. Inhibition of AMPK by Compound C or AMPK downregulation using siRNA partially inhibited the phosphorylation of eNOS in medium 199 but not in 1640, underscoring a clear difference in the pathways mediating thrombin-stimulated eNOS phosphorylation in different culture media. Thus, conditions subjecting endothelial cells to a fall in ATP after thrombin stimulation facilitate activation of pathways partly dependent on AMPK causing downstream phosphorylation of eNOS. In contrast, under culture conditions that do not facilitate a fall in ATP after stimulation, AMPK activation is exclusively mediated by CaMKK and does not contribute to the phosphorylation of eNOS.
Description:
To access publisher full text version of this article. Please click on the hyperlink in Additional Links field
Additional Links:
http://www.sciencedirect.com/science/article/B6T20-4T13CDP-1/2/52f74882df808b44c80eac1cc4a56625

Full metadata record

DC FieldValue Language
dc.contributor.authorThors, Brynhildur-
dc.contributor.authorHalldorsson, Haraldur-
dc.contributor.authorJonsdottir, Gudbjorg-
dc.contributor.authorThorgeirsson, Gudmundur-
dc.date.accessioned2008-12-11T09:15:33Z-
dc.date.available2008-12-11T09:15:33Z-
dc.date.issued2008-10-01-
dc.date.submitted2008-12-11-
dc.identifier.citationBiochim. Biophys. Acta. 2008, 1783(10):1893-902en
dc.identifier.issn0006-3002-
dc.identifier.pmid18687367-
dc.identifier.doi10.1016/j.bbamcr.2008.07.003-
dc.identifier.urihttp://hdl.handle.net/2336/42233-
dc.descriptionTo access publisher full text version of this article. Please click on the hyperlink in Additional Links fielden
dc.description.abstractConflicting results have been reported concerning the role of AMP-activated protein kinase (AMPK) in mediating thrombin stimulation of endothelial NO-synthase (eNOS). We examined the involvement of two upstream kinases in AMPK activation in cultured human umbilical endothelial cells, LKB1 stimulated by a rise in intracellular AMP/ATP ratio, and Ca(+2)/CaM kinase kinase (CaMKK) responding to elevation of intracellular Ca(+2). We also studied the effects of AMPK activation on the downstream target eNOS. In culture medium 1640 the level of intracellular ATP was unchanged after thrombin stimulation and the CaMKK inhibitor STO-609 totally inhibited phosphorylation of AMPK and acetyl coenzyme A carboxylase (ACC) but not eNOS. In Morgan's medium 199 thrombin caused a significant lowering of intracellular ATP and STO-609 only partially inhibited the phosphorylation of AMPK, ACC and eNOS. Inhibition of AMPK by Compound C or AMPK downregulation using siRNA partially inhibited the phosphorylation of eNOS in medium 199 but not in 1640, underscoring a clear difference in the pathways mediating thrombin-stimulated eNOS phosphorylation in different culture media. Thus, conditions subjecting endothelial cells to a fall in ATP after thrombin stimulation facilitate activation of pathways partly dependent on AMPK causing downstream phosphorylation of eNOS. In contrast, under culture conditions that do not facilitate a fall in ATP after stimulation, AMPK activation is exclusively mediated by CaMKK and does not contribute to the phosphorylation of eNOS.en
dc.language.isoenen
dc.publisherElsevier Pub. Co.en
dc.relation.urlhttp://www.sciencedirect.com/science/article/B6T20-4T13CDP-1/2/52f74882df808b44c80eac1cc4a56625en
dc.subject.meshAcetyl-CoA Carboxylaseen
dc.subject.meshAdenosine Triphosphateen
dc.subject.meshCalcimycinen
dc.subject.meshCells, Cultureden
dc.subject.meshCulture Media, Conditioneden
dc.subject.meshEndothelial Cellsen
dc.subject.meshHumansen
dc.subject.meshMultienzyme Complexesen
dc.subject.meshNitric Oxideen
dc.subject.meshNitric Oxide Synthase Type IIIen
dc.subject.meshPhosphorylationen
dc.subject.meshProtein-Serine-Threonine Kinasesen
dc.subject.meshThrombinen
dc.titleMechanism of thrombin mediated eNOS phosphorylation in endothelial cells is dependent on ATP levels after stimulation.en
dc.typeArticleen
dc.contributor.departmentInstitute of Pharmacy, Pharmacology and Toxicology, University of Iceland, Hagi Hofsvallagotu 53, Reykjavik, Iceland.en
dc.identifier.journalBiochimica et biophysica actaen
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