title: Neutral endopeptidase up-regulation in isolated human umbilical artery: involvement in desensitization of bradykinin-induced vasoconstrictor effects.
abstract: Previous reports show that bradykinin B(2) receptors mediate contractile responses induced by bradykinin (BK) in human umbilical artery (HUA). However, although it has been reported that BK-induced responses can desensitize in several inflammatory models, the effects of prolonged in vitro incubation on BK-induced vasoconstriction in HUA have not been studied. In isolated HUA rings, BK-induced responses after a 5-h in vitro incubation showed a marked desensitization compared with responses at 2 h. Inhibition of either angiotensin-converting enzyme (ACE) or neutral endopeptidase (NEP), both BK-inactivating enzymes, failed to modify responses to BK at 2 h. After 5 h, ACE inhibition produced only a slight potentiation of BK-induced responses. In contrast, BK-induced vasoconstriction at 5 h was markedly potentiated by NEP inhibition. Moreover, NEP activity, measured by hydrolysis of its synthetic substrate (Z-Ala-Ala-Leu-p-nitroanilide), showed a 2.4-fold increase in 5-h incubated versus 2-h incubated tissues, which was completely reversed by cycloheximide (CHX) treatment. Furthermore, CHX significantly potentiated BK-induced responses, suggesting that NEP-mediated kininase activity increase at 5 h depends on de novo protein synthesis. In addition, under NEP inhibition, CHX treatment failed to produce an additional potentiation of BK-induced vasoconstriction. Still, NEP up-regulation was confirmed by Western blot, showing a 2.1-fold increase in immunoreactive NEP in 5-h incubated versus 2-h incubated HUA. In summary, the present study provides strong pharmacological evidence that NEP is up-regulated and plays a key role in desensitization of BK-induced vasoconstriction after prolonged in vitro incubation in HUA. Our results provide new insights into the possible mechanisms involved in BK-induced response desensitization during sustained inflammatory conditions.
authors.affiliation: Departamento de Farmacología, Universidad de Buenos Aires, Argentina.
authors.forename: Facundo Germán
authors.initials: FG
authors.lastname: Pelorosso
authors.forename: Ana Verónica
authors.initials: AV
authors.lastname: Halperin
authors.forename: Alejandro Martín
authors.initials: AM
authors.lastname: Palma
authors.forename: Wanda
authors.initials: W
authors.lastname: Nowak
authors.forename: Andrea Emilse
authors.initials: AE
authors.lastname: Errasti
authors.forename: Rodolfo Pedro
authors.initials: RP
authors.lastname: Rothlin
mesh_terms: D000806:Angiotensin-Converting Enzyme Inhibitors / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology*; D006020:Glycopeptides / Q000494:pharmacology; D006801:Humans; D015260:Neprilysin / Q000502:physiology*; D014176:Protein Biosynthesis; D043783:Receptor, Bradykinin B1 / Q000502:physiology; D043782:Receptor, Bradykinin B2 / Q000502:physiology; D013997:Time Factors; D014469:Umbilical Arteries / Q000187:drug effects* / Q000502:physiology; D015854:Up-Regulation; D014661:Vasoconstriction / Q000187:drug effects*
chemical_list: D000806:Angiotensin-Converting Enzyme Inhibitors; D006020:Glycopeptides; D043783:Receptor, Bradykinin B1; D043782:Receptor, Bradykinin B2; D015260:Neprilysin; D001920:Bradykinin; C008890:phosphoramidon