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Article: Gi proteins and the response to 5-hydroxytryptamine in porcine cultured endothelial cells with impaired release of EDRF

TitleGi proteins and the response to 5-hydroxytryptamine in porcine cultured endothelial cells with impaired release of EDRF
Authors
Issue Date1995
PublisherJohn Wiley & Sons Ltd. The Journal's web site is located at http://www.wiley.com/bw/journal.asp?ref=0007-1188&site=1
Citation
British Journal Of Pharmacology, 1995, v. 115 n. 5, p. 822-827 How to Cite?
Abstract1. The receptor-mediated release of endothelium-derived relaxing factor(s) (EDRF) requires the presence of different functional G proteins in endothelial cells. Release of EDRF in response to 5-hydroxytryptamine (5-HT), which involves activation of pertussis toxin-sensitive Gi proteins, is impaired in both regenerated endothelium of the coronary artery following balloon catheterization and in porcine cultured endothelial cells. This study used porcine cultered endothelial cells as a model of regenerated endothelium to determine if the abnormal release of EDRF in response to 5-HT may be associated with the loss of functional pertussis toxin-sensitive Gi proteins. 2. Binding studies on porcine cultured endothelial cells demonstrated specific binding sites for [3H]-5-HT. Scatchard analyses revealed a single binding site for [3H]-5-HT with K(d) of 7.2 ± 3.5 nM and maximal binding (B(max)) of 121.4 ± 51.3 fmol mg-1 protein. Binding of [3H]-5-HT was displaced by methiothepin (5-HT1 and 5-HT2 antagonist; K(i) = 6.2 ± 1.2 nM), but not by ketanserin (preferential 5-HT2 antagonist). 3. Giα1 protein was expressed in cultured but not in native endothelial cells. Giα2 and Giα3 proteins were expressed to significant levels in porcine native and cultured endothelial cells, as defected by Northern and Western blot analysis. 4. In membranes from cultured endothelial cells, two bands of 40 and 41 kDa. which corresponded to the Giα2 and the combination of Giα3-Giα1 proteins, respectively, were ADP-ribosylated by pertussis toxin. The labelling intensity was Giα2 > Giα3-Giα1 and the amount of ADP-ribosylation was not different between porcine native and cultured endothelial cells. Stimulation of the cultured cells with 5-HT (3 x 10-6 M; 4 min) decreased significantly further ADP-ribosylation of Giα2 by pertussis toxin, but not that of Giα3 and/or Giα1. 5. The present results suggest that porcine endothelial cell culture may lead to the abnormal expression of Giα1 protein and that the dysfunctional release of EDRF from cultured porcine endothelial cells in response to 5-HT is not associated with the loss of Giα proteins or the absence of 5-HT binding sites.
Persistent Identifierhttp://hdl.handle.net/10722/171159
ISSN
2015 Impact Factor: 5.259
2015 SCImago Journal Rankings: 2.368
ISI Accession Number ID

 

DC FieldValueLanguage
dc.contributor.authorDay, NSen_US
dc.contributor.authorGe, Ten_US
dc.contributor.authorCodina, Jen_US
dc.contributor.authorBirnbaumer, Len_US
dc.contributor.authorVanhoutte, PMen_US
dc.contributor.authorBoulanger, CMen_US
dc.date.accessioned2012-10-30T06:12:27Z-
dc.date.available2012-10-30T06:12:27Z-
dc.date.issued1995en_US
dc.identifier.citationBritish Journal Of Pharmacology, 1995, v. 115 n. 5, p. 822-827en_US
dc.identifier.issn0007-1188en_US
dc.identifier.urihttp://hdl.handle.net/10722/171159-
dc.description.abstract1. The receptor-mediated release of endothelium-derived relaxing factor(s) (EDRF) requires the presence of different functional G proteins in endothelial cells. Release of EDRF in response to 5-hydroxytryptamine (5-HT), which involves activation of pertussis toxin-sensitive Gi proteins, is impaired in both regenerated endothelium of the coronary artery following balloon catheterization and in porcine cultured endothelial cells. This study used porcine cultered endothelial cells as a model of regenerated endothelium to determine if the abnormal release of EDRF in response to 5-HT may be associated with the loss of functional pertussis toxin-sensitive Gi proteins. 2. Binding studies on porcine cultured endothelial cells demonstrated specific binding sites for [3H]-5-HT. Scatchard analyses revealed a single binding site for [3H]-5-HT with K(d) of 7.2 ± 3.5 nM and maximal binding (B(max)) of 121.4 ± 51.3 fmol mg-1 protein. Binding of [3H]-5-HT was displaced by methiothepin (5-HT1 and 5-HT2 antagonist; K(i) = 6.2 ± 1.2 nM), but not by ketanserin (preferential 5-HT2 antagonist). 3. Giα1 protein was expressed in cultured but not in native endothelial cells. Giα2 and Giα3 proteins were expressed to significant levels in porcine native and cultured endothelial cells, as defected by Northern and Western blot analysis. 4. In membranes from cultured endothelial cells, two bands of 40 and 41 kDa. which corresponded to the Giα2 and the combination of Giα3-Giα1 proteins, respectively, were ADP-ribosylated by pertussis toxin. The labelling intensity was Giα2 > Giα3-Giα1 and the amount of ADP-ribosylation was not different between porcine native and cultured endothelial cells. Stimulation of the cultured cells with 5-HT (3 x 10-6 M; 4 min) decreased significantly further ADP-ribosylation of Giα2 by pertussis toxin, but not that of Giα3 and/or Giα1. 5. The present results suggest that porcine endothelial cell culture may lead to the abnormal expression of Giα1 protein and that the dysfunctional release of EDRF from cultured porcine endothelial cells in response to 5-HT is not associated with the loss of Giα proteins or the absence of 5-HT binding sites.en_US
dc.languageengen_US
dc.publisherJohn Wiley & Sons Ltd. The Journal's web site is located at http://www.wiley.com/bw/journal.asp?ref=0007-1188&site=1en_US
dc.relation.ispartofBritish Journal of Pharmacologyen_US
dc.subject.meshAdenosine Diphosphate Ribose - Metabolismen_US
dc.subject.meshAnimalsen_US
dc.subject.meshBase Sequenceen_US
dc.subject.meshBlotting, Northernen_US
dc.subject.meshBlotting, Westernen_US
dc.subject.meshCells, Cultureden_US
dc.subject.meshElectrophoresis, Polyacrylamide Gelen_US
dc.subject.meshEndothelium, Vascular - Drug Effects - Metabolismen_US
dc.subject.meshGtp-Binding Proteins - Metabolismen_US
dc.subject.meshMolecular Sequence Dataen_US
dc.subject.meshNitric Oxide - Metabolismen_US
dc.subject.meshPertussis Toxinen_US
dc.subject.meshPolymerase Chain Reactionen_US
dc.subject.meshRegeneration - Physiologyen_US
dc.subject.meshSerotonin - Metabolism - Pharmacologyen_US
dc.subject.meshSwineen_US
dc.subject.meshVirulence Factors, Bordetella - Pharmacologyen_US
dc.titleGi proteins and the response to 5-hydroxytryptamine in porcine cultured endothelial cells with impaired release of EDRFen_US
dc.typeArticleen_US
dc.identifier.emailVanhoutte, PM:vanhoutt@hku.hken_US
dc.identifier.authorityVanhoutte, PM=rp00238en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.doi10.1111/j.1476-5381.1995.tb15006.x-
dc.identifier.pmid8548182-
dc.identifier.scopuseid_2-s2.0-0028978989en_US
dc.identifier.volume115en_US
dc.identifier.issue5en_US
dc.identifier.spage822en_US
dc.identifier.epage827en_US
dc.identifier.isiWOS:A1995RF92300016-
dc.publisher.placeUnited Kingdomen_US
dc.identifier.scopusauthoridDay, NS=36854953000en_US
dc.identifier.scopusauthoridGe, T=7003328740en_US
dc.identifier.scopusauthoridCodina, J=7005011473en_US
dc.identifier.scopusauthoridBirnbaumer, L=35401284700en_US
dc.identifier.scopusauthoridVanhoutte, PM=7202304247en_US
dc.identifier.scopusauthoridBoulanger, CM=7006599024en_US

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