2-furoyl-ligrlo-amide and Diabetes-Mellitus--Type-2

Protease-activated receptor-2 (PAR2) is expressed in endothelial cells and mediates endothelium-dependent vasodilation. We hypothesized that PAR2 regulates tumor necrosis factor-alpha (TNF-α)-induced coronary arteriolar dysfunction in type 2 diabetic (db/db) mice. To test this, coronary arterioles from WT control, db/db, db/db mice treated with PAR2 antagonist FSLLRY-NH₂ (db/db+FSLLRY-NH₂) and db/db mice null for TNF (db(TNF-)/db(TNF-)) were isolated and pressurized (60 cmH₂O) without flow. Although vasodilation to the endothelium-independent vasodilator sodium nitroprusside (SNP) was not different among WT, db/db, db/db+FSLLRY-NH₂ and db(TNF-)/db(TNF-), endothelium-dependent acetylcholine (ACh)- and flow-mediated vasodilation were impaired in db/db mice but were enhanced in db(TNF-)/db(TNF-) mice and db/db mice treated with PAR2 antagonist. NOS inhibitor N (G)-nitro-L-arginine-methyl ester (L-NAME) significantly reduced ACh-induced dilation in WT, db(TNF-)/db(TNF-) and db/db+FSLLRY-NH₂, but did not alter the vasodilation in db/db mice. In contrast, cyclooxygenase (COX) inhibitor indomethacin (Indo) did not alter ACh-induced vasodilation in these four groups of mice. PAR2-activating peptide (PAR2-AP, 2-Furoyl-LIGRLO-am)-induced dilation was higher in db/db mice than that in WT, db(TNF-)/db(TNF-) and db/db mice treated with PAR2 antagonist. These effects were abolished by denudation, or in the presence of L-NAME or Indo. Protein expressions of TNF-α, PAR2, gp91(phox) and p47(phox) in the heart and isolated coronary arterioles were higher in db/db mice compared to WT mice. Administration of PAR2 antagonist to db/db mice reduced protein expression of TNF-α, gp91(phox) and PAR2. Protein expression of gp91(phox) and p47(phox) was lower in db(TNF-)/db(TNF-) compared to db/db mice. These results indicate that PAR2 plays a pivotal role in endothelial dysfunction in type 2 diabetes by up-regulating the expression/production of TNF-α and activating NAD(P)H oxidase subunit p47(phox).

Topics: Animals; Arterioles; Cells, Cultured; Coronary Vessels; Diabetes Mellitus, Type 2; Endothelium, Vascular; Female; Gene Expression Regulation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; NADPH Oxidases; Oligopeptides; Receptor, PAR-2; Tumor Necrosis Factor-alpha; Vasodilation

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.. 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.. 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. Inhibitors of the COX-cAMP pathway (FR122044, SC560, NS398, SC58125, SQ22536, CAY10441) did not affect these relaxation responses in either strain. Charybdotoxin (BK(Ca), SK3.1 blocker), but not iberiotoxin (BK(Ca) blocker), inhibited responses to the PAR2 agonists in db/db. In db/db protein levels of eNOS were higher, whereas those of sGC were lower than in C57. PAR2 mRNA expression in db/db was higher than in C57.. PAR2-mediated vasodilatation is protected against the negative effects of obesity and diabetes in mice. In diabetic vascular dysfunction, preserved PAR2 vasodilatation was linked to activation of SK3.1.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Acetylcholinesterase; Alcohol Oxidoreductases; Animals; Arteries; Blood Glucose; Diabetes Mellitus, Type 2; Dose-Response Relationship, Drug; Endothelium, Vascular; Gene Expression Regulation, Enzymologic; Glucose; Glycosuria; Male; Mice; Mice, Inbred C57BL; Nitroprusside; Obesity; Oligopeptides; Phenylephrine; Potassium; Receptor, PAR-2; Receptors, Leptin; RNA, Messenger; Vasodilation