Potentiation of des-Arg9-Kallidin-Induced Vasoconstrictor Responses by Metallopeptidase Inhibition in Isolated Human Umbilical Artery

Abstract

Several metallopeptidases have been reported to be involved in bradykinin (BK) B 1 receptor agonist metabolism. Our goal was to evaluate in vitro roles of metallopeptidases [e.g., neutral endopeptidase (NEP), aminopeptidase M (APM), and angiotensin-converting enzyme (ACE)] as functional inactivators of the selective BKB 1 receptor agonist Lys-des-Arg 9 -BK (DAKD) in isolated human umbilical artery (HUA) rings. Concentrationresponse curves (CRCs) to DAKD were performed after a 5-h incubation period. Treatment with 10 M phosphoramidon (NEP inhibitor) or 10 M amastatin (APM inhibitor) potentiated DAKD-elicited responses, whereas 1 M captopril (ACE inhibitor) had no significant effects. However, when the three enzymes were simultaneously inhibited, a significant potentiation over responses obtained under concurrent NEP and aminopeptidase M inhibition was observed. In contrast, responses induced by the peptidase resistant BKB 1 receptor agonist Sar-D-Phe 8 -des-Arg 9 -BK were not modified by triple peptidase inhibition. In addition, endothelial denudation failed to alter DAKD-induced responses in HUA. Finally, in the presence of NEP, ACE, and APM inhibition, Lys-des-Arg 9 -[Leu 8 ]-BK, the potent BKB 1 receptor antagonist, produced a parallel, concentration-dependent, rightward shift of DAKD CRCs. The obtained pK B (8.57) and the Schild slope not different from unity are in agreement with an interaction at a single homogeneous BKB 1 receptor population. In summary, this work constitutes the first pharmacological evidence that metallopeptidases NEP, APM, and ACE represent a relevant inactivation mechanism of the endogenous BKB 1 receptor agonist DAKD in isolated HUA.

The Kallikrein-Kinin System: Current and Future Pharmacological Targets

“…Indeed, the reaction product des-Arg 9 -BK has a much lower affinity at the human, rabbit, porcine, and bovine B1R. APN mediates the major inactivation pathway for des-Arg 10 -KD in the human isolated umbilical artery (127) and rabbit aorta (F. Marceau and A. Adam, unpublished observations). APN blockade with amastatin potentiates the hypotensive effect of the B1R agonist des-Arg 10 -KD in the LPS-pretreated rabbit (128).…”

Neutral Endopeptidase Up-Regulation in Isolated Human Umbilical Artery: Involvement in Desensitization of Bradykinin-Induced Vasoconstrictor Effects

“…Whenever necessary, effective inhibitory doses of peptidase inhibitors were used: 1 M captopril (ACE inhibitor, IC 50 38 nM) (Miyamoto et al, 2002), 10 M phosphoramidon (NEP inhibitor, IC 50 10 nM) (Loffler, 2000), 10 M thiorphan (NEP inhibitor, IC 50 1.4 nM) (Miyamoto et al, 2002), and 10 M amastatin [aminopeptidase M (APM) inhibitor, IC 50 50 nM] (Proud et al, 1987) were applied 30 min before the addition of BKB 2 and BKB 1 receptor agonists. The concentrations of amastatin, captopril, and phosphoramidon used in this study were previously shown to produce the inhibition of des-Arg 10 -kallidin inactivation as assessed by functional contractility studies in HUA (Pelorosso et al, 2005). Phosphoramidon is also able to inhibit ACE (IC 50 78 M) (Kukkola et al, 1995) and endothelin-converting enzyme (ECE) (IC 50 2.5 M) (Hoang and Turner, 1997).…”

“…In addition, Gafford et al (1983) have shown that neutral endopeptidase (NEP) is able to hydrolyze BK, yielding inactive fragments. Interestingly, we have recently shown that ACE and NEP are present in HUA smooth muscle after prolonged in vitro incubation (Pelorosso et al, 2005).…”

Vasopeptidase-activated latent ligands of the histamine receptor-1

“…This vein also expresses ACE [2,15]. The ectopeptidase APN is present in the vascular smooth muscle cells of the umbilical artery [3,4] and at least in the endothelium of the umbilical vein [16,17].…”

“…However, we reported lately that the peptide Met-Lys-bradykinin-Ser-Ser is paradoxically activated by its reaction with ACE, that frees the BK C-terminal sequence needed to activate the cognate B 2 receptor (B 2 R) [2]. The modulatory role of vascular aminopeptidase N (APN) has also been illustrated, as the blockade of this ectopeptidase, expressed in vascular smooth muscle, potentiates such peptides as Lys-des-Arg 9 -BK (the optimal agonist of human and rabbit BK B 1 receptor), some peptide antagonists of B 1 receptors and angiotensin III [3,4].…”