title: A role for proteinase-activated receptor 2 and PKC-epsilon in thrombin-mediated induction of decay-accelerating factor on human endothelial cells.
abstract: Thrombin, an important mediator of thrombosis and inflammation, may also enhance vascular cytoprotection. Thus thrombin induces expression of the complement-inhibitory protein decay-accelerating factor (DAF) in human umbilical vein endothelial cells (HUVECs), thus increasing protection against complement-mediated injury. Using PKC isozyme-specific peptide antagonists and adenoviral constructs, we have shown in the present study that PKC-epsilon is the primary isozyme involved in DAF induction by thrombin. Experiments with proteinase-activated receptor-1 (PAR1) and PAR2 activating peptides (APs) showed that DAF expression induced by PAR1-AP was PKC-alpha-dependent; in contrast, PAR2-AP induction of DAF required activation of PKC-epsilon. PAR1-AP and PAR2-AP in combination exerted an additive effect on DAF protein expression, which was equivalent to that observed with thrombin alone. These data implied a specific role for PAR2 in DAF induction, which was supported by the observation that upregulation of endothelial cell (EC) PAR2-enhanced DAF induction by thrombin. ERK1/2, p38, and JNK MAPK were also involved in thrombin-induced DAF upregulation, with evidence of interdependence between ERK1/2 and JNK. A role for transactivation of PAR2 by PAR1 was suggested by partial inhibition of thrombin-induced DAF expression by the PAR1 signaling antagonists BMS-200261 and SCH79797, whereas inhibition of thrombin-induced cleavage of PAR1 by specific MAbs or hirudin completely abrogated the response. Together, these data imply that the predominant pathway for thrombin-induced DAF expression involves transactivation of PAR2 by PAR1 and signaling via PKC-epsilon/MAPK. This may represent an important, novel pathway for endothelial cytoprotection during inflammation and angiogenesis and suggests that PAR2 may play a central role in some thrombin-induced responses.
authors.affiliation: Cardiovascular Medicine Unit, Imperial College, Hammersmith Hospital, DuCane Road, London W12 ONN, UK.
authors.forename: Elaine A
authors.initials: EA
authors.lastname: Lidington
authors.forename: Rivka
authors.initials: R
authors.lastname: Steinberg
authors.forename: Anne R
authors.initials: AR
authors.lastname: Kinderlerer
authors.forename: R Clive
authors.initials: RC
authors.lastname: Landis
authors.forename: Motoi
authors.initials: M
authors.lastname: Ohba
authors.forename: Allen
authors.initials: A
authors.lastname: Samarel
authors.forename: Dorian O
authors.initials: DO
authors.lastname: Haskard
authors.forename: Justin C
authors.initials: JC
authors.lastname: Mason
mesh_terms: D018958:CD55 Antigens / Q000096:biosynthesis*; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D006146:Guanidines / Q000494:pharmacology; D006629:Hirudins / Q000494:pharmacology; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000502:physiology; D009842:Oligopeptides / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology; D010766:Phosphorylation; D051744:Protein Kinase C-epsilon / Q000037:antagonists & inhibitors / Q000378:metabolism*; D044463:Receptor, PAR-1 / Q000235:genetics / Q000378:metabolism; D044464:Receptor, PAR-2 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013917:Thrombin / Q000378:metabolism*; D015533:Transcriptional Activation; D015854:Up-Regulation; D010975:Platelet Aggregation Inhibitors
chemical_list: D018958:CD55 Antigens; D006146:Guanidines; D006629:Hirudins; D007527:Isoenzymes; C479887:N-cinnamoyl-4-fluorophenylalanyl-4-guanidinophenylalanyl-leucyl-arginyl-argininamide; D009842:Oligopeptides; D010455:Peptides; D044463:Receptor, PAR-1; D044464:Receptor, PAR-2; C016977:amastatin; D051744:Protein Kinase C-epsilon; D013917:Thrombin; D010975:Platelet Aggregation Inhibitors