2-Furoyl-LIGRLO-amide

BACKGROUND AND PURPOSE: ;In non-obese diabetic animals, protease-activated receptor-2 (PAR2) agonists are more effective vasodilators, which is attributed to increased COX-2 and endothelial NOS (eNOS) activities. Under conditions of diabetes and obesity, the effectiveness of PAR2 agonists is unknown. We compared the vasodilator responses of small calibre mesenteric arteries from obese diabetic B6.BKS(D)-Lepr(db) /J (db/db) induced by PAR2-activating agonists 2-furoyl-LIGRLO-amide (2fly) and trypsin to those obtained in controls [C57BL/6J (C57)], and assessed the contributions of COX, NOS and calcium-activated potassium channels (K(Ca)) to these responses.;EXPERIMENTAL APPROACH: ;Arteries mounted in wire myographs under isometric tension conditions were contracted submaximally by U46619 then exposed to vasodilators. mRNA and protein expression of PAR2, eNOS and soluble GC (sGC) were determined by real-time PCR and Western blots.;KEY RESULTS: ;ACh- and nitroprusside-induced relaxations were attenuated in db/db compared with C57. In contrast, 2fly- and trypsin-induced relaxations were largely retained in db/db. A NOS inhibitor partly inhibited ACh- and 2fly-induced relaxations in C57, but not those in db/db.

This study investigated relaxation of vascular smooth muscle by acetylcholine, bradykinin and protease-activated receptor 2 (PAR-2) to characterise endothelial dysfunction in spontaneously hypertensive mice (BPH/2). We hypothesised that PAR-2 induced vasodilation would be preserved in BPH/2 despite the presence of hypertension and impaired vasodilator responses to acetylcholine and bradykinin. Mean arterial blood pressure (MAP), heart rate and locomotor activity were assessed in conscious mice over 24-h periods by radiotelemetry. Relaxation responses of small mesenteric arteries to acetylcholine, bradykinin and the PAR-2 agonist, 2-furoyl-LIGRLO-amide (2fly), were assessed using wire myographs. MAP and heart rate of BPH/2 were 15 and 18%, respectively, higher than in controls (BPN/3). BPH/2 also exhibited increased locomotor activity. Maximal relaxations of arteries by acetylcholine and bradykinin in BPH/2 were reduced by 25-50% relative to BPN/3. In contrast, relaxation responses to 2fly were only slightly (6%), albeit significantly, reduced. Sodium nitroprusside-induced relaxations were not different between strains. Treatment of BPH/2 arteries with inhibitors of calcium-activated K(+) channels was sufficient to block persistent 2fly- and residual ACh- and bradykinin-induced relaxations, whereas NO synthase inhibitor failed to inhibit these relaxations.

A peptide corresponding to a proteinase-activated receptor 2 (PAR(2))-activating peptide with an N-terminal furoyl group modification, 2-furoyl-LIGRLO-NH(2), was assessed for PAR(2)-dependent and -independent biological activities. 2-Furoyl-LIGRLO-NH(2) was equally effective to and 10 to 25 times more potent than SLIGRLNH(2) for increasing intracellular calcium in cultured human and rat PAR(2)-expressing cells, respectively. In bioassays of tissue PAR(2) activity, measured as arterial vasodilation and hyperpolarization, 2-furoyl-LIGRLO-NH(2) was 10 to 300 times more potent than SLIGRL-NH(2). Unlike trans-cinnamoyl-LIGRLO-NH(2), 2-furoyl-LI-GRLO-NH(2) did not cause a prominent non-PAR(2)-mediated contraction of murine femoral arteries. In conclusion, 2-furoyl-LI-GRLO-NH(2) represents the most potent and selective activator of PAR(2) in biological systems described to date.