endothelin-1 and acetovanillone

The effects of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase inhibitor apocynin on the enhanced hypoxia induced factor-1α (HIF-1α) and endothelin-1 (ET-1) expression, elevated systolic blood pressure under chronic intermittent hypoxia (CIH) condition and its action mechanism were investigated. Thirty healthy 8-week old Sprague-Dawley (SD) male rats were randomly divided into three groups (n=10 each): sham group, CIH group, and apocynin-treated CIH group. Tail artery systolic blood pressure was measured by tail-cuff method. Real-time fluorescence quantitative polymerase chain reaction (PCR) was used to detect the mRNA expression of HIF-1α and ET-1 in the carotid body, and the HIF-1α protein expression was examined by using Western blotting. The levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were determined by using colorimetric method. In addition, the plasma ET-1 and HIF-1α levels were measured by using enzyme-linked immunosorbent assay. It was found that CIH exposure was associated with increased MDA levels, and apocynin-treated CIH animals showed reduction in MDA levels. Apocynin treatment prevented CIH-induced hypertension as well as CIH-induced decrease in SOD. The increases of HIF-1α and ET-1 mRNA along with HIF-1α protein expression in the carotid body, and elevated circulating HIF-1α and ET-1 levels were observed in CIH-exposed animals. Treatment with apocynin significantly decreased the ET-1 mRNA, HIF-1α protein expression and circulating HIF-1α level in CIH-exposed animals, and there was no statistically significant difference in the HIF-1α mRNA expression between CIH group and apocynin-treated group. These results indicated that apocynin alleviated CIH-induced hypertension by inhibiting NADPH oxidase, further leading to the reduced vasoconstrictor ET-1 level and oxidative stress. HIF-1α/ET-1 system signal pathway may interact with CIH-induced NADPH oxidase-dependent oxidative stress. Inhibition of NADPH oxidase activity may hopefully serve as a useful strategy for prevention and treatment of obstructive sleep apnea hypopnea syndrome-induced hypertension.

Topics: Acetophenones; Animals; Antioxidants; Carotid Body; Endothelin-1; Hypoxia; Hypoxia-Inducible Factor 1, alpha Subunit; Male; NADP; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Treatment Outcome

The aim of this study was to assess whether apocynin, an nicotinamide adenine dinucleotide phosphate (NADPH) oxidase blocker, influences lipid peroxidation TBARS, hydrogen peroxide (H2O2) content, protein level, heart edema, tumor necrosis factor α (TNF-α) concentration or the glutathione redox system in heart homogenates obtained from endothelin 1 (ET-1)-induced oxidative stress rats.. Experiments were carried out on adult male Wistar-Kyoto rats. The animals were divided into 4 groups: Group I: saline-treated control; Group II: saline followed by ET-1 (3 μg/kg b.w., iv); Group III: apocynin (5 mg/kg b.w., iv) administered half an hour before saline; Group IV: apocynin (5 mg/kg b.w., iv) administered half an hour before ET-1 (3 μg/kg b.w., iv).. Injection of ET-1 alone showed a significant (p < 0.001) increase in thiobarbituric acid reactive substances (TBARS) and the hydrogen peroxide level (p < 0.01) vs. control, as well as a decrease (p < 0.001) in the GSH level. Apocynin significantly decreased TBARS (p < 0.001) and H2O2 (p < 0.05) level (vs. control) as well as improved protein level (p < 0.001) in the heart. Apocynin also prevented ET-1-induced heart edema (p < 0.05). The presence of ET-1 increased the concentration of TNF-α (p < 0.05) while apocynin decreased it (p < 0.05). Our results indicate that ET-1 may induce oxidative stress in heart tissue by reducing the GSH/GSSG ratio, stimulating lipid peroxidation and increasing TNF-α concentration. Apocynin diminished these measures of oxidative stress and TNF-α.. ET-1-induced formation of ROS in the heart is at least partially regulated via NADPH oxidase.

Topics: Acetophenones; Animals; Antioxidants; Edema; Endothelin-1; Glutathione; Lipid Peroxidation; Male; Myocardium; NADPH Oxidases; Oxidative Stress; Rats; Tumor Necrosis Factor-alpha

The mechanisms by which angiotensin-(1-7) [Ang-(1-7)] exerts its beneficial effects on end-organ damage associated with diabetes and hypertension are not well understood. The purpose of this study was A) to compare the effects of apocynin with Ang-(1-7) on renal vascular dysfunction and NADPH oxidase activity in a combined model of diabetes and hypertension and B) to further determine whether chronic treatment with Ang-(1-7) can modulate renal catalase, and peroxisome proliferator activated receptor- gamma (PPAR-gamma) levels in streptozotocin-induced diabetes in both normotensive Wistar Kyoto rats (WKY) and in spontaneously hypertensive rats (SHR). Apocynin or Ang-(1-7) treatment for one month starting at the onset of diabetes similarly attenuated elevation of renal NADPH oxidase activity in the diabetic SHR kidney and reduced the degree of proteinuria and hyperglycemia, but had little or modest effect on reducing mean arterial pressure. Both drugs also attenuated the diabetes-induced increase in renal vascular responsiveness to endothelin-1. Induction of diabetes in WKY and SHR animals resulted in significantly reduced renal catalase activity and in PPAR-gamma mRNA and protein levels. Treatment with Ang-(1-7) significantly prevented diabetes-induced reduction in catalase activity and the reduction in PPAR-gamma mRNA and protein levels in both animal models. Taken together, these data suggest that activation of Ang-(1-7)-mediated signaling could be an effective way to prevent the elevation of NADPH oxidase activity and inhibition of PPAR-gamma and catalase activities in diabetes and/or hypertension.

Topics: Acetophenones; Angiotensin I; Animals; Antihypertensive Agents; Antioxidants; Blood Glucose; Blood Pressure; Catalase; Diabetes Mellitus, Experimental; Endothelin-1; Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating); Hyperglycemia; Hypertension; Kidney; Male; NADPH Oxidases; Peptide Fragments; PPAR gamma; Proteinuria; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Renal Artery

As recently demonstrated, angiotensin II (Ang II) induces an increase in myocardial distensibility. Although endothelin-1 and the endocardial endothelium (EE) also modulate myocardial diastolic properties, their interaction with Ang II at this level has not yet been investigated. Increasing concentrations of Ang II (from 10(-8) to 10(-5) M) were studied in rabbit right papillary muscles in the following conditions: (1) baseline; (2) after selective removal of EE with Triton X-100; and (3) with intact EE in presence of a non-selective endothelin receptor antagonist (PD-145065), a selective endothelin type A receptor antagonist (BQ-123), an inhibitor of nitric oxide synthesis (N(G)-nitro-L-arginine (L-NA) or an inhibitor of the NAD(P)H oxidase (apocynin). At baseline, Ang II induced a concentration-dependent positive inotropic effect and an increase in passive muscle length (L) up to 1.020 +/- 0.004 L/L(max). After restoring muscle length to maximal physiological length (L(max)), passive tension decreased by 46.1 +/- 4.0%. When the EE was removed, the effect on myocardial distensibility was abolished. With intact EE in presence of PD-145065, BQ-123 or L-NA, the effects of Ang II on myocardial distensibility were attenuated, with a maximal increase in passive muscle length of 1.0087 +/- 0.0012, 1.0068 +/- 0.0022 and 1.0066 +/- 0.0020 L/L(max) and a decrease in resting tension of 22.6 +/- 3.6, 16.1 +/- 6.0 and 20.4 +/- 5.6%, respectively. In the presence of apocynin, the effect on myocardial distensibility was abolished. In conclusion, the Ang II-dependent acute increase in myocardial distensibility is abolished by the selective removal of the EE and attenuated in the presence of endothelin-1 receptor antagonists, an inhibitor of nitric oxide synthesis or an inhibitor of NAD(P)H oxidase.

Topics: Acetophenones; Angiotensin II; Animals; Diastole; Dose-Response Relationship, Drug; Endocardium; Endothelin Receptor Antagonists; Endothelin-1; Endothelium; Enzyme Inhibitors; In Vitro Techniques; Male; NADPH Oxidases; Nitric Oxide; Nitric Oxide Synthase; Nitroarginine; Oligopeptides; Papillary Muscles; Peptides, Cyclic; Rabbits; Receptors, Endothelin

The endothelial nitric oxide synthase (eNOS) has been implicated in the rapid (Frank-Starling) and slow (Anrep) cardiac response to stretch. Our work and that of others have demonstrated that a neuronal nitric oxide synthase (nNOS) localized to the myocardium plays an important role in the regulation of cardiac function and calcium handling. However, the effect of nNOS on the myocardial response to stretch has yet to be investigated. Recent evidence suggests that the stretch-induced release of angiotensin II (Ang II) and endothelin 1 (ET-1) stimulates myocardial superoxide production from NADPH oxidases which, in turn, contributes to the Anrep effect. nNOS has also been shown to regulate the production of myocardial superoxide, suggesting that this isoform may influence the cardiac response to stretch or ET-1 by altering the NO-redox balance in the myocardium. Here we show that the increase in left ventricular (LV) myocyte shortening in response to the application of ET-1 (10 nM, 5 min) did not differ between nNOS(-/-) mice and their wild type littermates (nNOS(+/+)). Pre-incubating LV myocytes with the NADPH oxidase inhibitor, apocynin (100 microM, 30 min), reduced cell shortening in nNOS(-/-) myocytes only but prevented the positive inotropic effects of ET-1 in both groups. Superoxide production (O(2)(-)) was enhanced in nNOS(-/-) myocytes compared to nNOS(+/+); however, this difference was abolished by pre-incubation with apocynin. There was no detectable increase in O(2)(-) production in ET-1 pre-treated LV myocytes. Inhibition of protein kinase C (chelerythrine, 1 microM) did not affect cell shortening in either group, however, protein kinase A inhibitor, PKI (2 microM), significantly reduced the positive inotropic effects of ET-1 in both nNOS(+/+) and nNOS(-/-) myocytes. Taken together, our findings show that the positive inotropic effect of ET-1 in murine LV myocytes is independent of nNOS but requires NADPH oxidases and protein kinase A (PKA)-dependent signaling. These results may further our understanding of the signaling pathways involved in the myocardial inotropic response to stretch.

Topics: Acetophenones; Animals; Benzophenanthridines; Cell Separation; Cyclic AMP-Dependent Protein Kinases; Endothelin-1; Heart Ventricles; Mice; Mice, Inbred Strains; Myocardial Contraction; Myocardium; Myocytes, Cardiac; NADPH Oxidases; Nitric Oxide; Nitric Oxide Synthase Type I; Protein Kinase C; Reactive Oxygen Species; Signal Transduction; Stimulation, Chemical; Superoxides; Time Factors

The role of endothelial dysfunction in coronary vasospasm is controversial. We hypothesized that reactive oxygen species (ROS) and endothelin-1 (ET-1) are plausible candidates as the mediator of vasospasm is linked to endothelial dysfunction. In a pig model with repetitive endothelial injury in coronary arteries, intracoronary administration of serotonin induced a vasospasm at the endothelial injury site. The level of endothelin-converting enzyme was upregulated at that site where, upon exposure to serotonin, there were also increases in p47(phox), ROS, and ET-1 fluorescence intensities, and myosin light chain phosphorylation and RhoA activation were detected. The nicotinamide adenine dinucleotide phosphate oxidase inhibitor, apocynin, had the effect of extinguishing not only ROS but also the appearance of ET-1. The chronic blockade of endothelin type-A receptor prevented a serotonin-triggered vasospasm along with the inhibition of ROS generation and myosin light chain phosphorylation. Under the coronary artery endothelial dysfunction, ET-1 is essential for an ROS-dependent coronary vasospasm. Our findings suggest that endothelial dysfunction plays a critical role in clinically defined human Prinzmetal angina.

Topics: Acetophenones; Angina Pectoris, Variant; Animals; Aspartic Acid Endopeptidases; Coronary Angiography; Coronary Vasospasm; Disease Models, Animal; Endothelin A Receptor Antagonists; Endothelin-1; Endothelin-Converting Enzymes; Endothelium, Vascular; Enzyme Inhibitors; Metalloendopeptidases; Myosin Light Chains; NADPH Oxidases; Phosphorylation; Pyrimidines; Reactive Oxygen Species; Receptor, Endothelin A; rhoA GTP-Binding Protein; Serotonin; Signal Transduction; Sulfonamides; Swine; Time Factors; Vasoconstriction

Reactive oxygen species (ROS) play important roles in renal vasoconstrictor responses to acute and chronic stimulation by angiotensin II and norepinephrine, as well as in long-term effects of endothelin-1 (ET-1). Little is known about participation of ROS in acute vasoconstriction produced by ET-1. We tested the influence of NAD(P)H oxidase inhibition by apocynin [4 mg.kg(-1).min(-1), infused into the renal artery (ira)] on ET(A) and ET(B) receptor signaling in the renal microcirculation. Both receptors were stimulated by ET-1, ET(A) receptors by ET-1 during ET(B) antagonist BQ-788, and ET(B) by ET(B) agonist sarafotoxin 6C. ET-1 (1.5 pmol injected ira) reduced renal blood flow (RBF) 17 +/- 4%. Apocynin raised baseline RBF (+10 +/- 1%, P < 0.001) and attenuated the ET-1 response to 10 +/- 2%, i.e., 35 +/- 9% inhibition (P < 0.05). Apocynin reduced ET(A)-induced vasoconstriction by 42 +/- 12% (P < 0.05) and that of ET(B) stimulation by 50 +/- 8% (P < 0.001). During nitric oxide (NO) synthase inhibition (N(omega)-nitro-l-arginine methyl ester), apocynin blunted ET(A)-mediated vasoconstriction by 60 +/- 8% (P < 0.01), whereas its effect on the ET(B) response (by 87 +/- 8%, P < 0.001) was even larger without than with NO present (P < 0.05). The cell-permeable superoxide dismutase mimetic tempol (5 mg.kg(-1).min(-1) ira), which reduces O(2)(-) and may elevate H(2)O(2), attenuated ET-1 responses similar to apocynin (by 38 +/- 6%, P < 0.01). We conclude that ROS, O(2)(-) rather than H(2)O(2), contribute substantially to acute renal vasoconstriction elicited by both ET(A) and ET(B) receptors and to basal renal vasomotor tone in vivo. This physiological constrictor action of ROS does not depend on scavenging of NO. In contrast, scavenging of O(2)(-) by NO seems to be more important during ET(B) stimulation.

Topics: Acetophenones; Animals; Blood Pressure; Cyclic N-Oxides; Endothelin-1; Femoral Artery; Male; NG-Nitroarginine Methyl Ester; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Receptor, Endothelin A; Receptor, Endothelin B; Renal Circulation; Spin Labels; Vasoconstriction; Vasodilation

We have recently reported that adventitial fibroblasts are able to express endothelin-1 (ET-1) in response to angiotensin II (Ang II) stimulation. However, the mechanism by which this occurs in the adventitia remains unclear. As Ang II has been reported to increase oxidant production by NADPH oxidase, we examined the role of this complex in Ang II stimulated ET-1 expression in vascular adventitial fibroblasts.. Adventitial fibroblasts were isolated and cultured from mouse aorta. Cells were treated with Ang II (100 nmol/L) in the presence or absence of NADPH oxidase inhibitors, apocynin (100 micromol/L) and diphenyleneiodonium (10 micromol/L), superoxide scavengers, SOD (350 units/mL), tempol (100 micromol/L), tiron (100 micromol/L), and ET-receptor antagonists (10 microM), BQ123 (for ET(A)-) and BQ788 (for ET(B)-). PreproET-1 mRNA and ET-1 level were determined by relative RT-PCR and ELISA, respectively. Type I procollagen-alpha-I (collagen) level was detected by Western blot. Superoxide anion (superoxide) production was determined by coelenterazine or lucigenin chemiluminescence. Ang II-induced collagen expression was inhibited by BQ123, suggesting that adventitial ET-1 plays a significant role in regulating the extracellular matrix. NADPH oxidase inhibitors and superoxide scavengers significantly decreased Ang II-induced ET-1 mRNA and peptide expression, superoxide production as well as collagen expression. Furthermore, deletion of gp91(phox) (a key subunit of NADPH oxidase) and overexpression of SOD1 attenuated Ang II-induced responses.. Ang II-evoked expression of ET-1 in adventitial fibroblasts appears to be mediated, at least in part, by NADPH oxidase. Functionally, this mechanism stimulates collagen expression thereby implicating the adventitia as a potential contributor to the vascular pathophysiology associated with oxidative stress and vascular remodeling.

Topics: 1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; Acetophenones; Adenoviridae; Angiotensin II; Animals; Antioxidants; Aorta; Biomarkers; Catecholamines; Cells, Cultured; Collagen Type I; Collagen Type I, alpha 1 Chain; Connective Tissue; Cyclic N-Oxides; Dopamine Agonists; Endothelin A Receptor Antagonists; Endothelin B Receptor Antagonists; Endothelin-1; Fibroblasts; Gene Expression; Genetic Vectors; Image Processing, Computer-Assisted; Imidazolines; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth, Vascular; NADPH Oxidase 2; NADPH Oxidases; Oligopeptides; Peptides, Cyclic; Piperidines; Protein Precursors; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spin Labels; Superoxide Dismutase; Transduction, Genetic

Red wine polyphenols (RWPs) have been reported to exert beneficial effects in preventing cardiovascular diseases, such as hypertension. We studied the effects of chronic treatment with RWPs and apocynin, an inhibitor of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, on blood pressure, endothelial function, and oxidative status in deoxycorticosterone acetate (DOCA)-salt-induced hypertension. Rats were administered RWPs (40 mg/kg) or apocynin (33 microg/kg) daily by gavage for 5 weeks. Plasma catechin levels were detected only after RWP treatment. RWPs and apocynin prevented both the increase in systolic blood pressure and the proteinuria induced by DOCA-salt. Plasma malonyldialdehyde levels, urinary iso-prostaglandin F(2alpha) excretion, aortic superoxide production, and aortic NADPH oxidase activity were found to be increased in animals of the DOCA group. RWP and apocynin treatments reduced these parameters in DOCA-salt rats, having no effect on control rats. However, only RWPs reduced the increase in plasma endothelin-1 (ET-1) levels and aortic p22(phox) gene overexpression found in DOCA-salt animals. RWPs and apocynin also improved the blunted endothelium-dependent relaxation response to acetylcholine in noradrenaline-precontracted aortic rings. All these results suggest that chronic treatment with RWPs prevents hypertension and vascular dysfunction. RWPs prevent vascular oxidative stress by inhibiting NADPH oxidase activity and/or by reducing ET-1 release.

Topics: Acetophenones; Animals; Blood Pressure; Desoxycorticosterone; Endothelin-1; Endothelium, Vascular; Flavonoids; Gene Expression; Hypertension; Male; NADPH Oxidases; Phenols; Polyphenols; Proteinuria; Rats; Rats, Wistar; Reactive Oxygen Species; Sodium Chloride

Experiments were conducted to test the hypothesis that hypertension produced by chronic ET-1 infusion is mediated by NADPH oxidase-dependent superoxide production. Mean arterial pressure (MAP) was continuously monitored in male Sprague Dawley rats by telemetry. After baseline measurements, rats were placed on a high-salt diet (8% NaCl) and osmotic minipumps were implanted to infuse ET-1 (5 pmol/kg per minute intravenous) for 12 days. Control rats were maintained on the high-salt diet only. Separate groups of rats were also infused with ET-1 and given the superoxide dismutase mimetic, tempol (1 mmol/L), or the NADPH oxidase inhibitor, apocynin (1.5 mmol/L), in the drinking water. Infusion of ET-1 significantly increased MAP when compared with baseline values (132+/-3 versus 114+/-2 mm Hg, P<0.05). Neither tempol nor apocynin treatment had any effect on the increase in MAP produced by ET-1 when compared with baseline values (127+/-5 versus 113+/-2 and 130+/-3 versus 115+/-2 mm Hg, respectively). Plasma 8-isoprostane, an indicator of oxidative stress, was significantly increased in ET-1-infused rats compared with rats on a high-salt diet alone (128+/-33 versus 51+/-5 pg/mL; P<0.05). Both tempol and apocynin treatment significantly attenuated the ET-1-induced increase in plasma 8-isoprostane (72+/-10 and 61+/-6 pg/mL, respectively). Similarly, ET-1 infusion also significantly increased aortic superoxide production (chemiluminescence and dihydroethidium staining techniques), which was prevented by both tempol and apocynin. These data provide evidence that chronic ET-1 infusion increases vascular NADPH oxidase-dependent superoxide production but does not account for chronic ET-1-induced hypertension.

Topics: Acetophenones; Animals; Antioxidants; Cyclic N-Oxides; Drug Administration Schedule; Endothelin-1; Enzyme Inhibitors; Hypertension; Infusion Pumps; Male; NADPH Oxidases; Oxidative Stress; Rats; Rats, Sprague-Dawley; Spin Labels; Superoxides

Experiments were designed to test the hypothesis that elevated levels of endothelin 1 (ET-1) in the vasculature activate NADPH oxidase and/or uncoupled nitric-oxide synthase (NOS), resulting in O2-* production, and mediate increased constriction. Rat aortic rings were incubated with ET-1 or vehicle in the presence and absence of superoxide dismutase (SOD), ebselen (glutathione peroxidase mimetic), apocynin (NADPH oxidase inhibitor), L-NAME (Nomega-nitro-L-arginine methyl ester) (NOS inhibitor), tetrahydrobiopterin (BH4) (NOS cofactor), or selective ETA and ETB receptor antagonists (BQ-123 [cyclo(D-Asp-Pro-D-Val-Leu-D-Trp)] and A-192621 [[2R-(4-propoxyphenyl)-4S-(1,3-benzodioxol-5-yl)-1-(N-(2,6-diethylphenyl)aminocarbonyl-methyl)-pyrrolidine-3R-carboxylic acid]], respectively). O2-* production was monitored by oxidized dihydroethidine staining and/or lucigenin chemiluminescence. ET-1 significantly increased O2-* production compared with vehicle. SOD, ebselen, and apocynin inhibited the ET-1-induced increase in O2-* in intact and endothelium-denuded aorta. L-NAME and BH4 inhibited the ET-1-induced increase in O2-* in intact tissue, whereas these two compounds had no effect on ET-1-induced O2-* in endothelium-denuded aorta. Preincubation with BQ-123 or A-192621, individually, had no effect on ET-1-induced O2-*; however combining both antagonists inhibited the ET-1-stimulated increase in O2-*. Rat aortic rings were incubated with ET-1 or vehicle in the presence or absence of sepiapterin (BH4 synthesis substrate) or apocynin and mounted on wire myographs to determine isometric force generation in response to increasing KCl concentrations. ET-1 increased the contractile response to KCl compared with vehicle. Treatment with either sepiapterin or apocynin attenuated the ET-1-mediated increase with no effect of sepiapterin or apocynin alone. These data support the hypothesis that ET-1 increases vascular tone, in part, through ETA/ETB receptor activation of O2-* production from NADPH oxidase and NOS uncoupling.

Topics: Acetophenones; Animals; Aorta; Azoles; Biopterins; Endothelin-1; Endothelium, Vascular; Enzyme Inhibitors; Isoindoles; Luminescence; Luminescent Measurements; Male; NADPH Oxidases; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Organoselenium Compounds; Pyrrolidines; Rats; Rats, Sprague-Dawley; Superoxide Dismutase; Superoxides; Vasoconstriction

Deoxycorticosterone acetate (DOCA)-salt hypertension is characterized by low renin/angiotensin but increased arterial superoxide levels. We have recently reported that the arterial endothelin-1 (ET-1) level is increased, resulting in NADPH oxidase activation and superoxide generation. However, the effect of ET-1 on venous superoxide production and its relation to venoconstriction are unknown. The present study tested the hypotheses that ET-1 stimulates venous NADPH oxidase and superoxide via its ET(A) receptors, resulting in enhanced venoconstriction in DOCA-salt hypertensive rats. Treatment with ET-1 (0.01 to 1 nmol/L), but not the selective ET(B) receptor agonist sarafotoxin s6c, of vena cavas of normal rats concentration-dependently increased superoxide levels, an effect that was abolished by the selective ET(A) receptor antagonist ABT-627. Although the ET-1 level was not increased in the vena cava and plasma, both venous NADPH oxidase activity and superoxide levels were significantly higher in DOCA-salt compared with sham rats. Moreover, ET-1 treatment (10(-9) mol/L, 10 minutes) of isolated vena cavas further elevated superoxide levels in DOCA-salt rats only but not sham rats, an effect that was abrogated by the superoxide scavenger tempol. Similarly, ET-1-induced contractions of isolated vena cavas of DOCA-salt but not sham rats were significantly inhibited by tempol. The NADPH oxidase inhibitor apocynin significantly reduced superoxide levels in vena cavas of DOCA-salt rats and in ET-1-treated vena cavas of normal rats. Finally, in vivo ET(A) receptor blockade by ABT-627 significantly lowered venous superoxide levels and blood pressure in DOCA-salt but not sham rats. These results suggest that superoxide contributes to ET-1-induced venoconstriction through an elevated venous NADPH oxidase activity in mineralocorticoid hypertension.

Topics: Acetophenones; Allopurinol; Animals; Atrasentan; Desoxycorticosterone; Dose-Response Relationship, Drug; Endothelin A Receptor Antagonists; Endothelin Receptor Antagonists; Endothelin-1; Enzyme Inhibitors; Hypertension; In Vitro Techniques; Male; NADPH Oxidases; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Pyrrolidines; Rats; Rats, Sprague-Dawley; Receptors, Mineralocorticoid; Superoxides; Vasoconstriction; Vasoconstrictor Agents; Venae Cavae; Viper Venoms; Xanthine Oxidase

We recently reported that arterial superoxide (O2-) is augmented by increased endothelin-1 (ET-1) in deoxycorticosterone acetate (DOCA)-salt hypertension, a model of low renin hypertension. Tetrahydrobiopterin (BH4), a potent reducing molecule with antioxidant properties and an essential cofactor for endothelial nitric oxide synthase, protects against O2--induced vascular dysfunction. However, the interaction between O2- and BH4 on endothelial function and the underlying mechanisms are unknown.. The present study tested the hypothesis that BH4 deficiency due to ET-1-induced O2- leads to impaired endothelium-dependent relaxation and that gene transfer of human guanosine 5'-triphosphate (GTP) cyclohydrolase I (GTPCH I), the first and rate-limiting enzyme for BH4 biosynthesis, reverses such deficiency and endothelial dysfunction in carotid arteries of DOCA-salt rats. There were significantly increased arterial O2- levels and decreased GTPCH I activity and BH4 levels in DOCA-salt compared with sham rats. Treatment of arteries of DOCA-salt rats with the selective ETA receptor antagonist ABT-627, NADPH oxidase inhibitor apocynin, or superoxide dismutase (SOD) mimetic tempol abolished O2- and restored BH4 levels. Basal arterial NO release and endothelium-dependent relaxations were impaired in DOCA-salt rats, conditions that were improved by apocynin or tempol treatment. Gene transfer of GTPCH I restored arterial GTPCH I activity and BH4 levels, resulting in reduced O2- and improved endothelium-dependent relaxation and basal NO release in DOCA-salt rats.. These results indicate that a BH4 deficiency resulting from ET-1-induced O2- via an ETA/NADPH oxidase pathway leads to endothelial dysfunction, and gene transfer of GTPCH I reverses the BH4 deficiency and endothelial dysfunction by reducing O2- in low renin mineralocorticoid hypertension.

Topics: Acetophenones; Animals; Antioxidants; Atrasentan; Biopterins; Carotid Arteries; Cyclic N-Oxides; Desoxycorticosterone; Disease Models, Animal; Endothelin Receptor Antagonists; Endothelin-1; Endothelium, Vascular; Enzyme Inhibitors; Gene Transfer Techniques; Genetic Therapy; GTP Cyclohydrolase; Humans; Hypertension; In Vitro Techniques; Male; Nitric Oxide; Pyrrolidines; Rats; Rats, Sprague-Dawley; Receptor, Endothelin A; Sodium Chloride; Spin Labels; Superoxides; Vasodilation

To investigate whether superoxide mediates angiotensin (Ang) II-induced vasoconstriction.. Human coronary arteries (HCAs), porcine femoral arteries (PFA) and porcine coronary arteries (PCAs) were mounted in organ baths and concentration-response curves to Ang II, the nitric oxide (NO) donor S-nitroso-N-acetylpenicillamine (SNAP) and the NAD(P)H oxidase substrate NADH were constructed in the absence and presence of superoxide inhibiting and activating drugs. Extracellular superoxide was measured using cytochrome c reduction.. Ang II constricted both HCAs and PFAs. In HCAs, the NAD(P)H inhibitors diphenyleneiodonium (DPI) and apocynin, and the xanthine oxidase (XO) inhibitor allopurinol, but not the superoxide dismutase (SOD) mimetic tempol or the SOD inhibitor diethyldithiocarbamate (DETCA), reduced this constriction. Catalase potentiated Ang II in HCAs, indicating a vasodilator role for H2O2. DPI, tempol and SOD did not affect Ang II in PFAs. DPI, apocynin and allopurinol relaxed preconstricted HCAs. Although the relaxant effects of the NO donor SNAP in PCAs was reduced by DETCA, indicating that superoxide-induced constrictions depend on NO inactivation, the apocynin-induced relaxations were NO independent. Moreover, NADH relaxed all vessels, and this effect was blocked by KCl but not DPI or NO removal. Xanthine plus XO also relaxed HCAs and PCAs. Incubation of human or porcine arteries with Ang II or NADH did not result in detectable increases of extracellular superoxide within 1 h.. Acute vasoconstriction by Ang II is not mediated via superoxide generated through NAD(P)H oxidase and/or XO activation. Such activation, if occurring, rather results in the generation of the vasodilator H2O2.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Acetophenones; Adolescent; Adult; Angiotensin II; Animals; Catecholamines; Child; Child, Preschool; Coronary Vessels; Cyclic N-Oxides; Dopamine Agonists; Dose-Response Relationship, Drug; Endothelin-1; Enzyme Inhibitors; Female; Femoral Artery; Free Radical Scavengers; Humans; Imidazolines; Male; Myocardial Contraction; NAD; NADP; Nitroprusside; Oxidants; S-Nitroso-N-Acetylpenicillamine; Spin Labels; Superoxide Dismutase; Superoxides; Swine; Time Factors; Vasoconstriction; Vasoconstrictor Agents; Vasodilation; Vasodilator Agents; Xanthine Oxidase