Therapeutic potential of S-nitrosothiols as nitric oxide donor drugs.
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Animals↗
Biomedical subjects
D J Webb
Publications and source records attributed to D J Webb.
The endothelin system in cardiovascular physiology and pathophysiology.
Endothelin-1, a member of a novel family of regulatory peptides, is the most potent vasoconstrictor and pressor substance known. Endothelin-1 is a 21-amino-acid endothelium-derived peptide causing uniquely sustained vasoconstriction. In addition, endothelin-1 has pronounced effects on the coronary, renal and cerebral circulations, enhances responses to other vasoconstrictors, and is comitogenic. Recent studies have shown that the endothelins are essential for normal fetal development, and that endothelin-1 plays an important physiological role in the regulation of basal vascular tone and blood pressure in healthy humans. There is now also a wealth of evidence suggesting that endothelin-1 is a key mediator in a range of cardiovascular diseases associated with sustained vasoconstriction, such as chronic heart failure, and with vasospasm, such as subarachnoid haemorrhage. In addition, endothelin-1 appears to act in opposition to nitric oxide to promote the atherosclerotic process. There are a large number of oral and intravenously active endothelin antagonists entering clinical development and a number of clinical studies, particularly with endothelin receptor antagonists, are now under way. Such studies are beginning to define the role of the endothelins in cardiovascular disease and to confirm the potential of the endothelin system as an important new therapeutic target.
Amino Acid Sequence↗
Seasonal variation in glucocorticoid activity in healthy men.
Many endocrine systems are subject to seasonal variation. However, studies of the hypothalamic-pituitary-adrenal axis in man have been limited to patients with psychiatric illness with conflicting results. We studied 105 healthy men, age 24-33 yr, during a 15-month period that included two winters. We measured cortisol and its metabolites by gas chromatography/mass spectrometry in plasma and urine and the intensity of dermal blanching after overnight topical application of beclomethasone dipropionate. There were no differences between subjects studied during the two winter periods, but marked differences between subjects studied in winter and summer. In winter, 0900-h plasma cortisol concentrations were higher (73 +/- 10 ng/mL, n = 41 vs. 35 +/- 4, n = 25 in summer; P < 0.01), total cortisol metabolite excretion was lower (678 +/- 67 micrograms/mmol creatinine vs. 900 +/- 98; P < 0.05), the ratio of metabolites of cortisol to those of cortisone was higher (3.0 +/- 0.2 vs. 2.1 +/- 0.1; P < 0.01), and dermal glucocorticoid sensitivity was higher (7.2 +/- 0.4 arbitrary units vs. 5.6 +/- 0.5; P < 0.02). Although blood pressure and fasting insulin/glucose relationships were not measurably different between seasons, these correlated with dermal vasoconstriction and cortisol metabolite excretion rate. We conclude that plasma cortisol and tissue sensitivity to glucocorticoids are higher in winter, but cortisol production rate is reduced. This could be explained by a reduction in cortisol clearance rate: urinary free cortisol/cortisone ratios were not different but A-ring-reduced metabolites of cortisol were higher in winter, suggesting that conversion of cortisone to cortisol by hepatic 11 beta-hydroxysteroid dehydrogenase 1 is enhanced. It is an intriguing possibility that increased glucocorticoid activity contributes to the increased prevalence of disease during the winter.
Adult↗
Endothelial dysfunction and hypertension.
Vascular endothelial cells play a key role in cardiovascular regulation by producing a number of potent vasoactive agents, including the vasodilator molecule nitric oxide (NO) and the vasoconstrictor peptide endothelin (ET)-1. A dysfunction of the vascular endothelium has been implicated in the pathophysiology of a number of cardiovascular diseases, important among which is essential hypertension. Impairment of NO synthesis, or increased inactivation of NO by superoxide radicals, may account for the increased peripheral vascular tone associated with hypertension, as well as contribute to the clinical consequences of this condition, which include vascular hypertrophy, increased platelet and monocyte adhesion to the endothelium, atherosclerosis, myocardial infarction and stroke. Similarly, increased ET-1 synthesis, or increased smooth muscle sensitivity to ET-1, could account for many of the features of hypertension, including increased peripheral vascular tone and vascular hypertrophy. Modulation of endothelial function is, therefore, an attractive therapeutic option in the treatment of hypertension. Calcium antagonists have been shown to enhance the effects of NO, and inhibit those of ET-1, on vascular smooth muscle cells. In addition, calcium antagonists have antiatherogenic and antioxidant properties and could, therefore, prove to be useful therapeutic agents in preventing some of the important complications of hypertension. The long term effects on cardiovascular morbidity and mortality of the long-acting nifedipine gastrointestinal therapeutic system (nifedipine GITS) used in the treatment of essential hypertension are currently being investigated in the first multinational outcome study (INSIGHT) of an antihypertensive agent since the major studies of beta-adrenoceptor blockers or thiazide diuretics. The results of this study are awaited with considerable interest.
Calcium Channel Blockers↗
An in vivo model for the assessment of acute fibrinolytic capacity of the endothelium.
The effects on blood flow and plasma fibrinolytic and coagulation parameters of intraarterial substance P, an endothelium dependent vasodilator, and sodium nitroprusside, a control endothelium independent vasodilator, were studied in the human forearm circulation. At subsystemic locally active doses, both substance P (2-8 pmol/min) and sodium nitroprusside (2-8 microg/min) caused dose-dependent vasodilatation (p <0.001 for both) without affecting plasma concentrations of PAI-1, von Willebrand factor antigen or factor VIII:C activity. Substance P caused local increases in t-PA antigen and activity (p <0.001) in the infused arm while sodium nitroprusside did not. At higher doses, substance P increased blood flow and t-PA concentrations in the noninfused arm. We conclude that brief, locally active and subsystemic infusions of intraarterial substance P cause a rapid and substantial local release of t-PA which appear to act via a flow and nitric oxide independent mechanism. This model should provide a useful and selective method of assessing the in vivo capacity of the forearm endothelium to release t-PA acutely.
Adult↗
Proteinases are isoform-specific regulators of the binding of transforming growth factor beta to alpha 2-macroglobulin.
alpha 2-Macroglobulin (alpha 2M) regulates growth and gene expression in many cell types by binding and neutralizing transforming growth factor beta (TGF-beta). In this study we characterized the effects of the serine proteinase, plasmin, on the interaction of alpha 2M with TGF-beta 1 and TGF-beta 2. Binding of both TGF-beta isoforms to purified alpha 2M-plasmin complex was primarily non-covalent and reversible. The binding affinity of alpha 2M for TGF-beta 1 was increased by plasmin; the Kd values were 320 and 84 nM for native alpha 2M and alpha 2M-plasmin respectively. In contrast the affinity of alpha 2M for TGF-beta 2 was decreased by plasmin; the Kd values were 14 and 80 nM for native alpha 2M and alpha 2M-plasmin respectively. Thrombin decreased the affinity of alpha 2M for TGF-beta 2 in a similar manner to plasmin. In assays of DNA synthesis in fetal bovine heart endothelial cells, native alpha 2M neutralized the activity of exogenously added TGF-beta 2, whereas alpha 2M-plasmin, at equivalent concentrations, had almost no effect. Native alpha 2M and methylamine-modified alpha 2M increased platelet-derived growth factor alpha-receptor expression in vascular smooth-muscle cells, an activity attributed to the neutralization of autocrine TGF-beta activity, whereas alpha 2M-plasmin was less effective at the same concentration. These studies demonstrate that the effects of proteinases on the cytokine-binding and cytokine-neutralizing activities of alpha 2M are cytokine-dependent. By reacting with alpha 2M, proteinases might regulate not only the availability of cytokines in the extracellular spaces but also the composition of the cytokine milieu.
Animals↗
Vasodilator effects of endothelin-converting enzyme inhibition and endothelin ETA receptor blockade in chronic heart failure patients treated with ACE inhibitors.
BACKGROUND: The importance of endothelin-1 in chronic heart failure (CHF) is unclear. We therefore investigated the effects of endothelin-converting enzyme (ECE) inhibition and endothelin ETA receptor blockade in CHF patients treated with ACE inhibitors. We also compared the function of ETA and ETB receptors in healthy subjects and patients with CHF. METHODS AND RESULTS: Locally active doses of study drugs were infused into the nondominant brachial artery while forearm blood flow (FBF was measured by venous occlusion plethysmography. In CHF patients (n = 10), phosphoramidon (a combined ECE and neutral endopeptidase inhibitor) and BQ-123 (an ETA receptor antagonist) increased FBF by 52 +/- 10% (P = .0006) and 31 +/- 6% (P = .002), respectively, and thiorphan (a selective neutral endopeptidase inhibitor) reduced FBF by 15 +/- 5% (P = .0007). Forearm vasoconstriction to endothelin-1 (an ETA and ETB receptor agonist) was significantly blunted in CHF patients compared with control subjects (both n = 10; CHF versus control subjects, P < .001), whereas vasoconstriction to sarafotoxin S6c (an ETB receptor agonist) was significantly enhanced in CHF patients compared with control subjects (both n = 10; CHF versus control subjects. P < .05). CONCLUSIONS: ECE inhibitors and ETA receptor antagonists may be useful as vasodilator agents in CHF patients already receiving treatment with an ACE inhibitor. Both ETA and ETB receptors can mediate agonist-induced vasoconstriction in healthy subjects and patients with CHF, but further studies are required to clarify the contribution of each receptor subtype in mediating the effects of endogenous endothelin-1.
Aged↗
Murine alpha-macroglobulins demonstrate divergent activities as neutralizers of transforming growth factor-beta and as inducers of nitric oxide synthesis. A possible mechanism for the endotoxin insensitivity of the alpha2-macroglobulin gene knock-out mouse.
alpha2-Macroglobulin null mice demonstrate increased resistance to endotoxin challenge (Umans, L., Serneels, L., Overbergh, L., Van Leuven, F., and Van den Berghe, H. (1995) J. Biol. Chem. 270, 19778-19785). We hypothesized that this phenotype might reflect the function of murine alpha2M (malpha2M) as a neutralizer of transforming growth factor-beta (TGF-beta) and inducer of nitric oxide synthesis in vivo. When incubated with wild-type mouse plasma, TGF-beta1 and TGF-beta2 bound only to malpha2M. Alternative TGF-beta-binding proteins were not detected in plasma from alpha2M(-/-) mice. Wild-type mouse plasma, but not plasma from alpha2M(-/-) mice, inhibited TGF-beta1 binding to TGF-beta receptors on fibroblasts. Purified malpha2M bound TGF-beta1 and TGF-beta2 with similar affinity; the KD values were 28 +/- 4 and 33 +/- 4 nM, respectively. Murinoglobulin, the second murine alpha-macroglobulin, bound both TGF-beta isoforms with 30-fold lower affinity. Malpha2M counteracted the activities of TGF-beta1 and TGF-beta2 in an endothelial cell growth assay. Malpha2M also induced NO synthesis when incubated with RAW 264.7 cells, an activity which probably results from the neutralization of autocrine TGF-beta activity. Human alpha2M induced NO synthesis comparably to malpha2M; however, MUG had no effect. These studies demonstrate that the ability to neutralize TGF-beta is a property of malpha2M, which is not redundant in the murine alpha-macroglobulin family or in murine plasma. Malpha2M is the only murine alpha-macroglobulin that promotes NO synthesis. The absence of malpha2M, in alpha2M(-/-) mice, may allow TGF-beta to more efficiently suppress excessive iNOS expression following endotoxin challenge.
Animals↗
Systemic endothelin receptor blockade decreases peripheral vascular resistance and blood pressure in humans.
BACKGROUND: Although local inhibition of the generation or actions of endothelin-1 has been shown to cause forearm vasodilatation, the systemic effects of endothelin receptor blockade in healthy humans are unknown. We therefore investigated the cardiovascular effects of a potent peptide endothelin ETA/B receptor antagonist, TAK-044, in healthy men. METHODS AND RESULTS: Two randomized, placebo-controlled, crossover studies were performed. In nine subjects, TAK-044 (10 to 1000 mg IV over a 15-minute period) caused sustained dose-dependent peripheral vasodilatation and hypotension. Four hours after infusion of the highest dose (1000 mg), there were decreases in mean arterial pressure of 18 mm Hg and total peripheral resistance of 665 AU and increases in heart rate of 8 bpm and cardiac index of 0.9 L x min(-1) x m(-2) compared with placebo. TAK-044 caused a rapid, dose-dependent increase in plasma immunoreactive endothelin (from 3.3 to 35.7 pg/mL within 30 minutes after 1000 mg). In a second study in eight subjects, intravenous administration of TAK-044 at doses of 30, 250, and 750 mg also caused peripheral vasodilatation, and all three doses abolished local forearm vasoconstriction to brachial artery infusion of endothelin-1. Brachial artery infusion of TAK-044 caused local forearm vasodilation. CONCLUSIONS: The endothelin ETA/B receptor antagonist TAK-044 decreases peripheral vascular resistance and, to a lesser extent, blood pressure; increases circulating endothelin concentrations; and blocks forearm vasoconstriction to exogenous endothelin-1. These results suggest that endogenous generation of endothelin-1 plays a fundamental physiological role in maintenance of peripheral vascular tone and blood pressure. The vasodilator properties of endothelin receptor antagonists may prove valuable therapeutically.
Adult↗
Endothelial nitric oxide production and insulin sensitivity. A physiological link with implications for pathogenesis of cardiovascular disease.
BACKGROUND: Insulin sensitivity varies up to threefold in apparently healthy individuals, but the mechanism for this is unknown. We have examined the hypothesis that vascular endothelial nitric oxide production and insulin sensitivity are directly related in humans. METHODS AND RESULTS: Nineteen healthy male subjects were studied on 3 separate days 1 week apart during which time they underwent measurement of insulin sensitivity by the euglycemic hyperinsulinemic clamp technique (soluble insulin 1.5 mU . kg-1 . min-1) and measurement of in vivo basal and stimulated endothelial nitric oxide production by forearm venous occlusion plethysmography. There was a correlation between insulin sensitivity and forearm vasoconstrictor responses to NG-monomethyl-L-arginine, the substrate inhibitor of nitric oxide synthase (r = .52, P < .05). No correlations were observed between insulin sensitivity and noradrenaline, acetylcholine, or sodium nitroprusside responses. CONCLUSIONS: Endothelial nitric oxide synthesis and insulin sensitivity are positively related in healthy humans, which suggests a direct physiological link.
Acetylcholine↗
Local inhibition of nitric oxide generation in man reduces blood flow in finger pulp but not in hand dorsum skin.
1. Nitric oxide generation is important in the regulation of resistance vessel tone. Until now, however, there has been no evidence of such a role for basal generation of nitric oxide in the skin microcirculation of humans. 2. To investigate this, L-NG-monomethylarginine (L-NMMA), a competitive inhibitor of nitric oxide synthase, was administered at 1, 2 and 4 mumol min-1 (each for 10 min), via the brachial artery, in six healthy male subjects. 3. At each dose, using laser Doppler fluximetry, red blood cell flux was measured as an index of blood flow in the pulp of the thumb, an area rich in arteriovenous anastomoses, and on the dorsal surface of the hand, where arteriovenous anastomoses are rare. Finger nailfold capillary blood velocity was monitored at each dose using videomicroscopy. Forearm blood flow was measured by venous occlusion plethysmography, before, and 8 min after, completing infusion of L-NMMA. All data were obtained from both the infused and control arms. 4. L-NMMA reduced blood flow in the infused forearm by 37% (P = 0.005). In contrast, dorsum red cell flux, capillary blood velocity, and skin temperature were unchanged. There was, however, a significant reduction in thumb red cell flux (ANOVA, P = 0.0001), reaching a maximum reduction of 33% with 4 mumol min-1 L-NMMA. There were no effects apparent in the opposite arm. 5. These results suggest that endogenous nitric oxide production may be more important in regulating microvascular skin blood flow in regions rich in arteriovenous anastomoses than in areas containing mainly nutritive vessels.
Adult↗
Endothelin production in sepsis and the adult respiratory distress syndrome.
OBJECTIVE: Septic shock is characterised by a decrease in systemic vascular resistance. Nevertheless, regional increases in vascular resistance can occur which may predispose to organ dysfunction, including the adult respiratory distress syndrome (ARDS). Because endothelial damage is a major feature of acute lung injury, we examined whether the potent endothelial vasoconstrictor peptide endothelin-1 plays a pathophysiological role in sepsis or ARDS. DESIGN: Plasma endothelin was measured in mixed venous, pulmonary capillary and arterial blood, and the relationship with outcome measures was determined. SETTING: The intensive care unit of a university teaching hospital. PATIENTS AND PARTICIPANTS: A consecutive series of well-characterised patients with sepsis syndrome, both with (n = 11) and without (n = 15) ARDS, and ventilated controls without sepsis or ARDS (n = 7). MEASUREMENTS AND RESULTS: Plasma endothelin was significantly elevated in patients with sepsis alone and in patients with sepsis and ARDS. Plasma endothelin did not differ among mixed venous, pulmonary capillary and systemic arterial blood. On multiple regression analysis, plasma endothelin correlated positively with organ failure score and with oxygen consumption, and negatively with the PaO2 : FiO2 ratio. There was no correlation with plasma creatinine, suggesting that decreased renal clearance did not account for the high plasma endothelin concentrations. CONCLUSIONS: Although the lung does not appear to be the major site of endothelin production in critically ill patients with sepsis, increased endothelin production may contribute to regional increases in vascular [correction of vacular] resistance, hyperfusion, and the development of organ failure, including ARDS, in patients with sepsis.
Case-Control Studies↗
Life as an academic clinical pharmacologist.
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Career Choice↗
The use of the Micral-Test strip to identify the presence of microalbuminuria in people with insulin dependent diabetes mellitus (IDDM) participating in the EUCLID study.
In IDDM, microalbuminuria (urinary albumin excretion rate (AER) of 20-200 micrograms/min) is a predictor of persistent proteinuria and diabetic nephropathy. Early intervention may prevent or reduce the rate of progression of renal complications. The Micral-Test strip can be used to establish a semi-quantitative estimate of AER. We assessed the field performance of the Micral-Test strip in detecting microalbuminuria in the EUCLID study, an European wide, 18 centre study of 530 IDDM participants, aged 20 to 59 years. People with macroalbuminuria were excluded. On entry, all participants had albumin concentrations from two overnight urine collections measured by a central laboratory, and the corresponding Micral-Test performed on the two collections locally. a cut off of > or = mg/l albumin from the first Micral-Test, to detect a centrally measured albumin concentration > or = 20 mg/l, yielded 29 (5.8%) false negative results and 58 (11.6%) false positive results (sensitivity 70%, specificity 87%). The mean AER, from two collections, was compared with the corresponding 'pooled' Micral-Test results (mean of the two readings). Receiver Operating Characteristic (ROC) curves were used to assess if there was a suitable 'pooled' Micral-Test result for screening microalbuminuria. A 'pooled' Micral-Test result (> or = 15 mg/l) was used to detect mean AER > or = 20 micrograms/min (sensitivity 78%, specificity 77%). This 'pooled cut-off' had already been used for screening on to the study and led to an over-estimate (154 vs. 77) of the true number of microalbuminuric participants on the study. In conclusion, our findings suggest that the Micral-Test strip is not an effective screening tool for microalbuminuria, using the 'pooled' result from two measurements did not improve the sensitivity of the test.
Adult↗
The endothelin system and its potential as a therapeutic target in cardiovascular disease.
Endothelin (ET)-1, an endothelium-derived peptide, is the most potent vasoconstrictor agent described to date. ET-1 also has positive inotropic and chronotropic effects in the heart and is a co-mitogen in both cardiac and vascular myocytes. The major elements of the system involved in formation of ET-1 and its isopeptides, as well as the receptors mediating their effects, have been cloned and characterised. Antagonists of the ET receptors are now available, and selective inhibitors of the ET-converting enzymes are being developed. Early studies using receptor antagonists support the involvement of ET-1 in the pathophysiology of several cardiovascular diseases. The relative merits of ET-converting enzyme inhibitors and receptor antagonists for the treatment of cardiovascular disease are discussed.
Amino Acid Sequence↗
Activation of the endothelin B receptor causes a dose-dependent accumulation of cyclic GMP in human platelets.
Endothelins modulate in vitro aggregation of human platelets in a bi-directional manner. Thus endothelin-1 has been shown to act as a potentiator of primary aggregation and an inhibitor of secondary aggregation. The endothelin receptors and corresponding second messengers which cause these effects have not yet been characterised. This study investigated the effect of endothelin-1, an agonist at both the ETA and the ETB receptors and sarafotoxin (SRTX) S6c, a selective ETB agonist, on human platelet cyclic nucleotide levels. Neither endothelin-1 (10(-11) -10(-7) M) nor SRTX S6c (10(-11) -10(-7) M) significantly altered platelet cAMP levels. In contrast, both agonists produced a dose-dependent increase in platelet cGMP. From these data, we conclude that activation of the ETB receptor in human platelets is responsible for an increase in platelet cGMP and may contribute to the inhibition of platelet aggregation caused by the endothelins.
Adult↗
The increase in human plasma immunoreactive endothelin but not big endothelin-1 or its C-terminal fragment induced by systemic administration of the endothelin antagonist TAK-044.
1. We examined the effects of systemic infusion, in healthy human volunteers, of the endothelin antagonist TAK-044 on the plasma concentrations of mature endothelin, big endothelin-1 and the C-terminal fragment of big endothelin-1, by selective solid-phase extraction and specific radioimmunoassays. 2. Unlabelled TAK-044 competed with specific [125I]-endothelin-1 binding to human left ventricle tissue in a biphasic manner giving KD values of 0.11 nM and 26.8 nM at the ETA and ETB receptor subtypes, respectively, indicating a 244 fold selectivity for the ETA receptor subtype. 3. A 15 min intravenous infusion of placebo or 30 mg TAK-044 (giving a serum concentration of 2 nM, calculated to block > 95% of ETA but < 5% ETB receptors) had no effect on the immunoreactive plasma concentrations of the three peptides. 4. At the higher dose of 750 mg TAK-044 (giving a serum concentration of 80 nM, calculated to block > 99% of ETA and > 75% ETB receptors), the immunoreactive plasma endothelin concentrations were increased 3.3 fold over basal levels (P < 0.01). The concentrations of big endothelin-1 or C-terminal fragment of big endothelin-1 were unchanged. 5. At both doses of TAK-044, there were significant decreases in diastolic blood pressure, and peripheral vascular resistance, with corresponding increases in cardiac index and stroke index. There were no changes in systolic or mean arterial blood pressures or heart rate. 6. Since only the concentrations of the mature peptide were increased, we conclude that the most likely sources of endothelin contributing to the observed rise were displacement of receptor-bound peptide and reduction in plasma clearance rather than peptide synthesis.
Adult↗
Role of endothelin in the regulation of vascular tone and blood pressure.
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Blood Pressure↗