title: Enzymatic pathways involved in the generation of endothelin-1(1-31) from exogenous big endothelin-1 in the rabbit aorta.
abstract: We investigated whether blood vessels contribute to the production of ET-1(1-31) from exogenous big endothelin-1 (BigET-1) in the rabbit and assessed which enzymes are involved in this process. Vascular reactivity experiments, using standard muscle bath procedures, showed that BigET-1 induces contraction in endothelium-intact rabbit aortic rings. Preincubation of the rings with phosphoramidon, CGS35066 or thiorphan reduced BigET-1-induced contraction. Conversely, chymostatin did not affect BigET-1-induced contraction. Thiorphan and phosphoramidon, but not CGS35066 or chymostatin, reduced ET-1(1-31)-induced contraction. None of the enzymatic inhibitors affected the contraction afforded by ET-1.BQ123-, but not BQ788-, selective antagonists for ET(A) and ET(B) receptors, respectively, produced concentration-dependent rightward displacements of the ET-1(1-31) and ET-1 concentration-response curves. By the use of enzymatic assays, we found that the aorta, as well as the heart, lung, kidney and liver, possess a chymase-like activity. Enzyme immunoassays detected significant levels of Ir-ET-1(1-31) in bathing medium of aortas after the addition of BigET-1 (30 nM). Neither thiorphan nor chymostatin altered the levels of Ir-ET-1(1-31). Conversely, the levels of Ir-ET-1(1-31) were increased in the presence of phosphoramidon. This marked increase of the 31-amino-acid peptide was abolished when phosphoramidon and chymostatin were added simultaneously. The major new finding of the present work is that the rabbit aorta generates ET-1(1-31) from exogenously administered BigET-1. Additionally, by measuring the production of ET-1(1-31), we showed that a chymase-like enzyme is involved in this process when ECE and NEP are inhibited by phosphoramidon. Our results also suggest that ET-1(1-31) is an alternate intermediate in the production of ET-1 following BigET-1 administration. Finally, we showed that NEP is the predominant enzymatic pathway involved in the cleavage of ET-1(1-31) to a bioactive metabolite that will act on ET(A) receptors to induce contraction in the rabbit aorta.
authors.affiliation: Department of Pharmacology, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Québec, Canada.
authors.forename: Carlos R
authors.initials: CR
authors.lastname: Tirapelli
authors.forename: Marie-Helene
authors.initials: MH
authors.lastname: Fecteau
authors.forename: Jean-Claude
authors.initials: JC
authors.lastname: Honore
authors.forename: Eurode
authors.initials: E
authors.lastname: Legros
authors.forename: Fernand
authors.initials: F
authors.lastname: Gobeil
authors.forename: Pedro
authors.initials: P
authors.lastname: D'Orleans-Juste
mesh_terms: D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000378:metabolism*; D016282:Aspartic Acid Endopeptidases / Q000502:physiology; D053818:Chymases; D004305:Dose-Response Relationship, Drug; D019332:Endothelin-1 / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism* / Q000494:pharmacology; D000072396:Endothelin-Converting Enzymes; D005260:Female; D008297:Male; D008666:Metalloendopeptidases / Q000502:physiology; D015260:Neprilysin / Q000502:physiology; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000494:pharmacology; D010456:Peptides, Cyclic / Q000494:pharmacology; D010880:Piperidines / Q000494:pharmacology; D011817:Rabbits; D012697:Serine Endopeptidases / Q000502:physiology; D014661:Vasoconstriction / Q000187:drug effects
chemical_list: D019332:Endothelin-1; D009842:Oligopeptides; D010446:Peptide Fragments; D010456:Peptides, Cyclic; D010880:Piperidines; C114932:endothelin-1 (1-31); C086539:BQ 788; D012697:Serine Endopeptidases; D053818:Chymases; D016282:Aspartic Acid Endopeptidases; D008666:Metalloendopeptidases; D015260:Neprilysin; D000072396:Endothelin-Converting Enzymes; C072247:cyclo(Trp-Asp-Pro-Val-Leu)