| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
1 Department of Cardiovascular Medicine, University of Oxford, Wellcome Trust Centre for Human Genetics, Roosevelt Drive, Oxford OX3 9DU, UK 2 Kobe University of Medicine, Japan
 |
Abstract |
|---|
 Top Abstract IntroductionMethodsResultsDiscussionReferences |
|---|
(Received 17 July 2006;
accepted after revision 18 September 2006; first published online 28 September 2006)
Corresponding author K. M. Channon: Department of Cardiovascular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, OX3 8DU, UK. Email: keith.channon{at}cardiov.ox.ac.uk
|
Introduction |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
While these observations establish the basis for a relationship between eNOS protein levels, NO-mediated endothelial function and blood pressure, recent evidence suggests that physiological regulation of eNOS is more complex. The pteridine cofactor tetrahydrobiopterin (BH4) is critical for eNOS enzymatic activity, hence NO production. In the absence of adequate levels of BH4, eNOS becomes ‘uncoupled’ from L-arginine oxidation and instead molecular oxygen is reduced to form superoxide (Vasquez-Vivar et al. 1998, 2003; Landmesser et al. 2003). Although transgenic mice with endothelium-targeted eNOS overexpression have increased NO production and low blood pressure (Ohashi et al. 1998; van Haperen et al. 2002), the greatly increased levels of eNOS in these animals leads to eNOS uncoupling that is related to eNOS–BH4 stoichiometry in the endothelium (Bendall et al. 2005. Specifically, transgenic mice with endothelial eNOS overexpression have a marked increase in eNOS-derived superoxide production, owing to eNOS uncoupling, that is normalized by augmentiion of the endothelial BH4 levels by crossing with transgenic mice with endothelial expression of GTP cyclohydrolase 1 (GCH), the rate limiting enzyme in BH4 biosythesis (Bendall & Channon, 2005). However, it is not known whether the effects of coupled versus uncoupled eNOS on local NO and superoxide production have important effects on haemodynamic regulation. In particular, it remains unclear whether NO production alone, or other functionally related radicals such as superoxide, is the key determinant of vasorelaxation responses in isolated vessels, and how the changes in vasorelaxation remain quantitatively related to changes in blood pressure in vivo.
In this paper, we compare changes in blood pressure, NOS activity and vascular relaxation responses in endothelium-targeted transgenic mice with overexpression of either eNOS or GTP cyclohydrolase 1, and in double transgenic mice with both eNOS and GCH overexpression. We demonstrate that blood pressure is determined principally by NO production rather than eNOS coupling, and that chronic increases in vascular NO production lead to desensitization of downstream cGMP-mediated signalling, independent of eNOS uncoupling.
|
Methods |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Studies were performed in accordance with the UK Home Office Animals (Scientific Procedures) Act 1986. Mice were provided with standard chow and water ad libitum and housed singly at 24°C in individually ventilated cages (Techniplast Inc., Buguggiate, Italy). All mice were exposed to a regular 12 h–12 h light–dark cycle and were 11–18 weeks old at the time of study.
Mouse lines had previously been fully back-crossed onto C57bL/6 strain. Endothelial nitric oxide synthase transgenic mice (eNOS-Tg) were generated by targeting bovine eNOS overexpression to vascular endothelium under the control of the murine preproendothelin-1 promoter, as previously described (Ohashi et al. 1998). These animals have been shown to have an eightfold elevation in eNOS protein levels in lung and aortic tissue, with transgene expression confined to the endothelium (Ohashi et al. 1998; Bendall & Channon, 2005). Guanosine tripohosphate cyclohydrolase 1 transgenic mice (GCH-Tg) were generated by endothelium-targeted overexpression of human GCH under the control of the murine Tie-2 promoter, as previously described (Alp et al. 2003, 2004). These animals have been shown to have a threefold increase in tissue BH4 levels in lung, heart and aortic tissue (Alp et al. 2003; Bendall & Channon, 2005). eNOS-Tg and GCH-Tg heterozygote mice were crossbred to produce eNOS/GCH double Tg, eNOS-Tg, GCH-Tg and wild-type (WT) littermates in a 1:1:1:1 ratio for study. All animals were genotyped by polymerase chain reactions (PCR) on DNA prepared by phenol–chloroform extraction of ear-notch biopsies. Genotypes were double-checked using DNA prepared from lung tissue snap frozen at the time of killing.
Haemodynamic studies
Blood pressure was measured by direct invasive methods under general anaesthesia using the Millar® catheter system. Animals were anaesthetized using inhalational isofluorane vaporised on oxygen and maintained at a temperature of 36.5°C using a warming blanket. Surgical anaesthesia was determined by loss of the pedal withdrawal reflex. A mid-line incision was made in the neck, and the left carotid artery isolated. The cranial end of the artery was tied using 3.0 mersilk, and a stay suture looped around the caudal artery. A 1.5 F Millar® catheter was then introduced and advanced until a good blood pressure waveform trace could be detected. The proximal suture was then tied to provide haemostasis. Experimental anaesthesia was lightened to obtain a respiratory rate of 80 min–1. Following equilibration for 15 min, intra-arterial blood pressure was recorded for 10 min. In some animals, pharmacological studies were performed by intraperitoneal (I.P.) injections of the NOS inhibitor N
-nitro-L-arginine methyl ester hydrochloride (L-NAME, 100 mg kg–1) or the adrenergic agonist phenylephrine (PE, 3 mg kg–1).
Isometric tension vasomotor studies
Aortic vasomotor function was assessed using isometric tension studies in a wire myograph (Multi-Myograph 610M, Danish Myo Technology, Aarhus, Denmark). Mice were killed by cervical dislocation. Freshly harvested and cleaned thoracic aortas (n
= 5–10 per group) were cut into two 2 mm aortic rings, which were mounted in organ bath chambers containing 5 ml of Krebs–Henseleit buffer (KHB [in mmol l–1]: NaCl, 120; KCl, 4.7; MgSO4, 1.2; KH2PO4, 1.2; CaCl2, 2.5; NaHCO3, 25; and glucose, 5.5) at 37°C, gassed with 95% O2–5% CO2. All experiments were performed in the presence of 10 µM indomethacin to inhibit endogenous prostaglandin release. Rings were first allowed to equilibrate for 30 min and then gradually stretched to a passive tension of 15 mN. Rings were constricted with 60 mM KCl for 5 min to assess vessel viability. Dose–response contraction curves were performed using cumulative half-log concentrations of phenylephrine (10–9–10–5
M). Vessels were then washed repeatedly with fresh KHB for 30 min, and then precontracted to approximately 90% of maximal tension with PE (typically 3 x 10–6
M). Dose–response relaxation curves for increasing cumulative concentrations of acetylcholine (ACh, 10–9–10–5
M) were then established to assess endothelium-dependent relaxation mediated by endothelial NO release. Responses were expressed as a percentage of the precontracted tension. Vessels were washed and precontracted again to 90% maximal contraction with PE. N-Nitro-L-arginine methyl ester (10–4
M; Sigma-Aldrich, UK) was then added to inhibit endogenous NO release from eNOS. Finally, the NO donor sodium nitroprusside (SNP, 10–10–10–6
M) was added in increasing cumulative concentrations to test endothelium-independent smooth muscle relaxation in response to exogenous NO.
Measurement of NO production
Nitric oxide production was measured using an L-arginine to citruilline conversion assay in the presence of the specific arginase inhibitor N-hydroxy-nor-L-arginine (nor-NOHA), as previously described (de Bono et al. 2006). Aortas were opened longitudinally, and incubated in 250 µl of KHB containing 1 µM calcium ionophore and 5 µl of 1.85 MBq ml–1 ubiquitously labelled [14C]L-arginine for 90 min at 37°C, prior to removing the supernatant. Endothelium lysis was induced by three cycles of freeze–thawing in 250 µl of water added to this supernatant. Sixty microlitres of 10% trichloroacetic acid was then added to deproteinate the samples, prior to centrifugation. Five hundred microlitres of this supernatant was then collected and added to 360 µl distilled water with 140 µl 10% trichloroacetic acid. Citruilline was resolved from arginine by HPLC, as previously described (de Bono et al. 2006), using a 250 mm x 4.6 mm Supelcosil LC-SCX 5 cation-exchange column (Sigma-Aldrich), a DG-980-50 degasser, two PU-2080 Plus pumps, a MX-2080-32 dynamic mixer (all from Jasco Ltd, Great Dunmow, UK) and a Lablogic ß-RAM Model 3 continuous flow liquid scintillation detector (Lablogic Systems Ltd, Sheffield, UK). Products of arginine metabolism were eluted over 30 min using the following buffer (rate, 1 ml min–1): 0–5 min 100% distilled water, 5–15 min linear gradient from 100% distilled water to 100% 200 mM sodium citrate (pH 3.0), and 15–30 min 100% sodium citrate (pH 3.0). Scintillant fluid (Lablogic) was mixed in-line at a ratio of 0.5:1.0 after elution from the column, before passage through the liquid scintillation detector. The integrals of citruilline peaks were expressed as a percentage of the total 14C count.
Measurement of cyclic GMP levels
Cyclic GMP levels in aortas were measured as previously described (Ohashi et al. 1998). Mice were killed by cervical dislocation. Briefly, aortas (n = 4–7 per group) were opened longitudinally and pre-incubated in oxygenated Krebs–Hepes solution with 0.1 mmol l–1 3-isobutyl-1-methylxanthine (IBMX; Sigma-Aldrich) at 37°C for 15 min and then stimulated with 1 µM acetylcholine for 3 min. Vessels were immediately snap-frozen in liquid nitrogen before homogenization in ice-cold 5% trichloroacetic acid containing 0.5 mM IBMX. Homogenates were centrifuged at 2000g, and trichloroacetic acid in the supernatant fractions was extracted with water-saturated ether. Cyclic GMP levels were measured in these fractions using a cGMP enzyme immunoassay kit (Cayman Chemical Co., Nottingham, UK), and results expressed as picomoles per milligram of trichloroacetic acid-precipitable protein solubilized with 1 M sodium hydroxide.
Statistics
Mean data for haemodynamic parameters, NO production and cGMP formation were analysed by one-way ANOVA with Bonferroni correction for repeated measures. For vasomotor studies, the mean responses of two arotic rings from each animal were combined to produce an n of 1. Dose–response curves from each group were compared using ANOVA for repeated measures followed by Bonferroni post hoc correction.
|
Results |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
The haemodynamic findings by direct intra-arterial Millar® catheter measurements revealed no differences in systolic, mean or diastolic blood pressure between GCH-Tg and WT littermate mice. There was a significant reduction in systolic blood pressure in eNOS-Tg compared with their WT littermates (P < 0.001), with a non-significant trend towards a reduction in mean and diastolic pressure. This was reflected by a significant reduction in pulse pressure in mice carrying the eNOS transgene (P < 0.001). However, the addition of the GCH transgene in eNOS/GCH double transgenic mice did not confer any additional effect on blood pressure when compared to eNOS-Tg alone. There were no significant differences in heart rate between the transgenic mice and their WT littermates (Fig. 1). This baseline haemodynamic phenotype was also confirmed by prior tail-cuff assessment in the same cohort (data not shown).
|
|
Vasomotor function studies
We determined the functional significance of altering eNOS protein and BH4 bioavailability in vitro using isometric tension studies on a wire myograph (Fig. 3). The contractile responses to PE were unchanged in GCH-Tg mice, but were significantly reduced in aortic rings from both eNOS-Tg and eNOS/GCH-Tg mice compared with wild-type littermates. Endothelium-dependent relaxations in reponse to the receptor-mediated eNOS agonist ACh were unchanged in GCH-Tg mice but were significantly attenuated in eNOS-Tg mice compared with wild-type littermates. However, the presence of the GCH transgene on the eNOS-Tg background in eNOS/GCH-Tg animals had no effect, since relaxations in response to ACh were not different between eNOS-Tg and eNOS/GCH-Tg mice. Endothelium-independent relaxations in response to the NO donor SNP, unchanged in GCH-Tg mice, were significantly attenuated in both eNOS-Tg and eNOS/GCH-Tg mice, indicating a reduction in vascular smooth muscle responsiveness to NO (Fig. 3).
|
We examined the effect of endothelial eNOS and GCH overexpression on NO production from aortas by measurement of L-arginine to citruilline conversion in the presence of the specific arginase inhibitor N-hydroxy-nor-L-arginine (nor-NOHA). Citruilline production was markedly increased in the aortas of both eNOS-Tg and eNOS/GCH-Tg animals (P < 0.0001) compared with wild-type littermates. However, there was no further significant increase in NO production in eNOS/GCH-Tg aortas compared with eNOS-Tg alone (Fig. 4B).
|
Since aortic relaxations in response to exogenous, as well as endothelium-derived, NO were significantly attenuated in eNOS-Tg and eNOS/GCH-Tg mice, we hypothesized that this may result from a desensitization in their downstream NO–cGMP pathway as a result of chronically enhanced NO production. In accordance with the vasomotor studies, aortic ACh-stimulated cGMP formation was found to be significantly reduced in both eNOS-Tg and eNOS/GCH-Tg mice compared with wild-type littermates (Fig. 4A).
|
Discussion |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
The importance of BH4 for the coupling of eNOS enzymatic activity to arginine oxidation and NO production is now well recognized (Vasquez-Vivar et al. 1998, 2003, 2004). In situations where BH4 is limiting, eNOS uncoupling results in increased superoxide production. This results in reduced bioavailability of NO both as a result of a reduction in NO production and as a result of increased NO scavenging by reactive oxygen species, leading to the production of peroxynitrite. We have previously shown that eNOS-Tg mice have increased aortic NO but also increased L-NAME-inhibitable superoxide production owing to uncoupling of eNOS as a result of a discordance between eNOS protein and its essential cofactor, BH4 (Bendall & Channon, 2005). This increase in aortic superoxide production was reversed by increasing vascular BH4 production in eNOS/GCH-Tg mice, which overexpress both endothelial eNOS and GTP cyclohydrolase 1 (the rate-limiting enzyme in endothelial BH4 synthesis; Bendall & Channon, 2005). In this study, we describe the effect of endothelial upregulation of eNOS and its cofactor BH4 on haemodynamic regulation in mice in vivo and compare it to measurements of vasomotor function in vitro.
Endothelium-specific eNOS-Tg mice were significantly hypotensive compared with WT littermates, a difference abolished by the NOS inhibitor L-NAME. This conforms with previously reported data from the same animal model (Ohashi et al. 1998). This finding is consistent with the expected effect of endothelial eNOS overexpression on enhanced endothelial NO production, leading to a reduction in smooth muscle tone and reduced blood pressure. It might be expected that this model would be associated with improved vascular responsiveness to activators of endothelial NO release, such as acetylcholine. However, we observed discordance between in vivo hypotension versus a significant impairment of endothelium-dependant relaxations in vitro, measured using wire myography. Furthermore, in vitro, aortic contraction in response to phenylephrine and relaxation in response to an exogenous NO donor (SNP) were found to be significantly impaired in eNOS-Tg and eNOS/GCH-Tg animals. This was reflected by an impaired pressor response to a phenylephrine bolus in vivo. These findings are consistent with those reported in the eNOS knockout mouse model, in which the opposite effect has been reported (Kojda et al. 2001), and suggest that chronic alterations in endothelial free radical signalling also have secondary effects on endothelium-independent vascular smooth muscle reactivity.
The GCH-Tg mouse was normotensive and demonstrated no change in vasomotor function at myography compared with WT littermates. We have previously shown that the GCH-Tg preserves vasomotor function in the context of a disease model such as diabetes where it is otherwise impaired (Alp et al. 2003). The addition of the GCH transgene to eNOS-Tg mice in the eNOS/GCH-Tg animal had no additional effect on blood pressure compared with eNOS-Tg littermates. In accordance, vasorelaxations in response to endothelium-derived and exogenous NO were no different between eNOS/GCH-Tg and their eNOS-Tg littermates. This suggests that the known difference in superoxide production between the aortas of eNOS-Tg and eNOS/GCH-Tg animals is not an important determinant of blood pressure regulation or vascular tone. Rather, both blood pressure and vasomotor function appear to be more closely related to NO production, which is increased equally in both eNOS-Tg and eNOS/GCH-Tg mice.
Since aortic responses to exogenous as well as endothelium-dependent NO were attenuated in eNOS-Tg and eNOS/GCH-Tg mice, we hypothesized that this may result from downstream desensitization of their NO–cGMP signalling pathway as a result of chronic elevation of NO levels rather than owing to endothelial dysfunction. Acetylcholine-stimulated cGMP production was found to be significantly reduced in both eNOS-Tg and eNOS/GCH-Tg aortas, the two groups with attenuated vasorelaxations to ACh and SNP. It remains possible, however, that a component of the desensitization of the NO–cGMP pathway may result from a reduction in endothelial NO production in response to exogenous stimulation with ACh. Hence, whilst maximal NO production measured by arginine to citruilline conversion (in the presence of calcium ionophore) is increased in eNOS-Tg and eNOS/GCH-Tg aortas, it cannot be said with certainty that NO production in response to ACh is likewise increased.
There is therefore an apparent discordance between hypotension in mice expressing the eNOS transgene and vascular hyporeactivity, although both appear to be mediated directly or indirectly by increased NO production and are independent of superoxide. One explanation might be that the mechanisms which regulate the set-point of vascular tone and therefore blood pressure are independent of those which regulate vascular reactivity. It would then be possible to have lower vascular resistance in the context of impaired vascular responsiveness. Thus, whilst we have shown that vascular reactivity is greatly influenced by the NO–cGMP pathway, perhaps vascular tone is mediated by a different NO-dependent pathway. This could be the case if the mechanism of action of NO were different in resistance arterioles from that in conduit vessels. This is suggested by the redundancy of vasoreactive mechanisms previously described in resistance vessels which are not mediators in conduit vessels, including endothelium-derived hyperpolarization factor, products of cyclo-oxygenase and neuronal NOS-derived NO (Popp et al. 1998; Chataigneau et al. 1999; Sun et al. 1999; Ding et al. 2000; Huang et al. 2001; Scotland et al. 2001). An alternative explanation is that endothelial NO production may mediate effects on chronic blood pressure independantly of vascular smooth muscle by effecting renal regulation of endovascular salt and volume.
The results of this study have implications for our understanding of the role of NO signalling in vascular function and haemodynamic regulation. Our data show discordance between systemic blood pressure and vasomotor function by aortic wire myography. This suggests that vasorelaxation responses in models of altered NO signalling should not be interpreted as direct measures of NOS enzymatic activity. Our data support the hypothesis that endothelial NO signalling is an important regulator of systemic blood pressure but suggest that therapeutic strategies targeting upregulation of eNOS should also consider the potential impact of chronic elevation of NO on desensitization of downstream signalling pathways. Furthermore, even correction of eNOS uncoupling and BH4 supplementation in vivo seems unlikely to alter desensitization of downstream NO signalling. This may have clinical implications for patients receiving therapeutic exogenous nitrate donors, since chronic administration in the context of disease states associated with increased oxidative stress in the vascular wall might lead to desensitization of the NO–cGMP pathway and exacerbate impairment of vascular reactivity and endothelial function.
|
Footnotes |
|---|
|
References |
|---|
|
TopAbstractIntroductionMethodsResultsDiscussionReferences |
|---|
Alp NJ, Mussa S, Khoo J, Guzik TJ, Cai S, Jefferson A, Rockett KA & Channon KM (2003). Tetrahydrobiopterin-dependent preservation of nitric oxide-mediated endothelial function in diabetes by targeted transgenic GTP-cyclohydrolase I overexpression. J Clin Invest 112, 725–735.[CrossRef][Medline]
Bendall JK, Alp NJ, Warrick N, Cai S, Adlam D, Rockett K, Yokoyama M, Kawashima S & Channon KM (2005). Stoichiometric relationships between endothelial tetrahydrobiopterin, eNOS activity and eNOS coupling in vivo: insights from transgenic mice with endothelial-targeted GTP cyclohydrolase 1 and eNOS over-expression. Circ Res 97, 864–871.
Chataigneau T, Feletou M, Huang PL, Fishman MC, Duhault J & Vanhoutte PM (1999). Acetylcholine-induced relaxation in blood vessels from endothelial nitric oxide synthase knockout mice. Br J Pharmacol 126, 219–226.[CrossRef][Medline]
de Bono J, Warrick N, Bendall J, Channon KM & Alp N (2006). Radiochemical HPLC detection of arginine metabolism: Measurement of nitric oxide synthesis and arginase activity in vascular tissue. Nitric Oxide, doi: 10.1016/j.niox.2006.03.008
Ding H, Kubes P & Triggle C (2000). Potassium- and acetylcholine-induced vasorelaxation in mice lacking endothelial nitric oxide synthase. Br J Pharmacol 129, 1194–1200.[CrossRef][Medline]
Furchgott RF & Zawadzki JV (1980). The obligatory role of endothelial cells in the relaxation of arterial smooth muscle by acetylcholine. Nature 288, 373–376.[CrossRef][Medline]
Heitzer T, Schlinzig T, Krohn K, Meinertz T & Munzel T (2001). Endothelial dysfunction, oxidative stress, and risk of cardiovascular events in patients with coronary artery disease. Circulation 104, 2673–2678.
Henry RM, Ferreira I, Kostense PJ, Dekker JM, Nijpels G, Heine RJ, Kamp O, Bouter LM & Stehouwer CD (2004). Type 2 diabetes is associated with impaired endothelium-dependent, flow-mediated dilation, but impaired glucose metabolism is not; The Hoorn Study. Atherosclerosis 174, 49–56.[CrossRef][Medline]
Huang A, Sun D, Carroll MA, Jiang H, Smith CJ, Connetta JA, Falck JR, Shesely EG, Koller A & Kaley G (2001). EDHF mediates flow-induced dilation in skeletal muscle arterioles of female eNOS-KO mice. Am J Physiol Heart Circ Physiol 280, H2462–H2469.
Huang PL, Huang ZH, Mashimo H, Bloch KD, Moskowitz MA, Bevan JA & Fishman MC (1995). Hypertension in mice lacking the gene for endothelial nitric oxide synthase. Nature 377, 239–242.[CrossRef][Medline]
Ignarro LJ (2002). Nitric oxide as a unique signaling molecule in the vascular system: a historical overview. J Physiol Pharmacol 53, 503–514.[Medline]
Kiff RJ, Gardiner SM, Compton AM & Bennett T (1991). Selective impairment of hindquarters vasodilator responses to bradykinin in conscious Wistar rats with streptozotocin-induced diabetes mellitus. Br J Pharmacol 103, 1357–1362.[Medline]
Kojda G, Cheng YC, Burchfield J & Harrison DG (2001). Dysfunctional regulation of endothelial nitric oxide synthase (eNOS) expression in response to exercise in mice lacking one eNOS gene. Circulation 103, 2839–2844.
Kojda G, Laursen JB, Ramasamy S, Kent JD, Kurz S, Burchfield J, Shesely EG & Harrison DG (1999). Protein expression, vascular reactivity and soluble guanylate cyclase activity in mice lacking the endothelial cell nitric oxide synthase: contributions of NOS isoforms to blood pressure and heart rate control. Cardiovasc Res 42, 206–213.
Landmesser U, Dikalov S, Price SR, McCann L, Fukai T, Holland SM, Mitch WE & Harrison DG (2003). Oxidation of tetrahydrobiopterin leads to uncoupling of endothelial cell nitric oxide synthase in hypertension. J Clin Invest 111, 1201–1209.[CrossRef][Medline]
Laursen JB, Somers M, Kurz S, McCann L, Warnholtz A, Freeman BA, Tarpey M, Fukai T & Harrison DG (2001). Endothelial regulation of vasomotion in apoE-deficient mice: implications for interactions between peroxynitrite and tetrahydrobiopterin. Circulation 103, 1282–1288.
Ohashi Y, Kawashima S, Hirata K, Yamashita T, Ishida T, Inoue N, Sakoda T, Kurihara H, Yazaki Y & Yokoyama M (1998). Hypotension and reduced nitric oxide-elicited vasorelaxation in transgenic mice overexpressing endothelial nitric oxide synthase. J Clin Invest 102, 2061–2071.[Medline]
Panza JA, García CE, Kilcoyne CM, Quyyumi AA & Cannon RO III (1995). Impaired endothelium-dependent vasodilation in patients with essential hypertension: evidence that nitric oxide abnormality is not localized to a single signal transduction pathway. Circulation 91, 1732–1738.
Popp R, Fleming I & Busse R (1998). Pulsatile stretch in coronary arteries elicits release of endothelium-derived hyperpolarizing factor: a modulator of arterial compliance. Circ Res 82, 696–703.
Scotland RS, Chauhan S, Vallance PJ & Ahluwalia A (2001). An endothelium-derived hyperpolarizing factor-like factor moderates myogenic constriction of mesenteric resistance arteries in the absence of endothelial nitric oxide synthase-derived nitric oxide. Hypertension 38, 833–839.
Shesely EG, Maeda N, Kim HS, Desai KM, Krege JH, Laubach VE, Sherman PA, Sessa WC & Smithies O (1996). Elevated blood pressures in mice lacking endothelial nitric oxide synthase. Proc Natl Acad Sci U S A 93, 13176–13181.
Sun D, Huang A, Smith CJ, Stackpole CJ, Connetta JA, Shesely EG, Koller A & Kaley G (1999). Enhanced release of prostaglandins contributes to flow-induced arteriolar dilation in eNOS knockout mice. Circ Res 85, 288–293.
van Haperen R, de Waard M, van Deel E, Mees B, Kutryk M, van Aken T, Hamming J, Grosveld F, Duncker DJ & de Crom R (2002). Reduction of blood pressure, plasma cholesterol, and atherosclerosis by elevated endothelial nitric oxide. J Biol Chem 277, 48803–48807.
Vasquez-Vivar J, Kalyanaraman B & Martasek P (2003). The role of tetrahydrobiopterin in superoxide generation from eNOS: enzymology and physiological implications. Free Radic Res 37, 121–127.[CrossRef][Medline]
Vasquez-Vivar J, Kalyanaraman B, Martasek P, Hogg N & Masters BS, Karoui H, Tordo P & Pritchard KA Jr (1998). Superoxide generation by endothelial nitric oxide synthase: the influence of cofactors. Proc Natl Acad Sci U S A 95, 9220–9225.
Vasquez-Vivar J, Martasek P & Kalyanaraman B (2004). Superoxide generation from nitric oxide synthase: role of cofactors and protein interaction. In Biological Magnetic Resonance, pp.75–91. Kluger Academic Publishers.
This article has been cited by other articles:
|
|
 |
|
  L.-C. Kung, S. H. H. Chan, K. L. H. Wu, C.-C. Ou, M.-H. Tai, and J. Y. H. Chan Mitochondrial Respiratory Enzyme Complexes in Rostral Ventrolateral Medulla as Cellular Targets of Nitric Oxide and Superoxide Interaction in the Antagonism of Antihypertensive Action of eNOS Transgene Mol. Pharmacol., November 1, 2008; 74(5): 1319 - 1332. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. L. Moens, H. C. Champion, M. J. Claeys, B. Tavazzi, P. M. Kaminski, M. S. Wolin, D. J. Borgonjon, L. Van Nassauw, A. Haile, M. Zviman, et al. High-Dose Folic Acid Pretreatment Blunts Cardiac Dysfunction During Ischemia Coupled to Maintenance of High-Energy Phosphates and Reduces Postreperfusion Injury Circulation, April 8, 2008; 117(14): 1810 - 1819. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 T. M. Paravicini and R. M. Touyz NADPH Oxidases, Reactive Oxygen Species, and Hypertension: Clinical implications and therapeutic possibilities Diabetes Care, February 1, 2008; 31(Supplement_2): S170 - S180. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
-adrenergic receptor agonist phenylephrine (A), the endothelium-dependent NO agonist acetylcholine (B) and the exogenous NO donor sodium nitroprusside (C). *P < 0.05, **P < 0.01 compared with wild-type measured using repeated measures ANOVA (means ±
S.E.M.).