Exercise testing criteria to diagnose lower extremity peripheral artery disease assessed by computed-tomography angiography.

BACKGROUND: The sensitivity and specificity of exercise testing have never been studied simultaneously against an objective quantification of arterial stenosis. Aims were to define the sensitivity and specificity of several exercise tests to detect peripheral artery disease (PAD), and to assess whether or not defined criteria defined in patients suspected of having a PAD show a difference dependent on the resting ABI. METHODS: In this prospective study, consecutive patients with exertional limb pain referred to our vascular center were included. All patients had an ABI, a treadmill exercise-oximetry test, a second treadmill test (both 10% slope; 3.2km/h speed) with post-exercise pressures, and a computed-tomography-angiography (CTA). The receiver-operating-characteristic curve was used to define a cut-off point corresponding to the best area under the curve (AUC; [CI95%]) to detect arterial stenosis ≥50% as determined by the CTA. RESULTS: Sixty-three patients (61+/-11 years-old) were included. Similar AUCs from 0.72[0.63-0.79] to 0.83[0.75-0.89] were found for the different tests in the overall population. To detect arterial stenosis ≥50%, cut-off values of ABI, post-exercise ABI, post-exercise ABI decrease, post-exercise ankle pressure decrease, and distal delta from rest oxygen pressure (DROP) index were ≤0.91, ≤0.52, ≥43%, ≥20mmHg and ≤-15mmHg, respectively (p<0.01). In the subset of patients with an ABI >0.91, cut-off values of post-exercise ABI decrease (AUC = 0.67[0.53-0.78]), and DROP (AUC = 0.67[0.53-0.78]) were ≥18.5%, and ≤-15mmHg respectively (p<0.05). CONCLUSION: Resting ABI is as accurate as exercise testing in patients with exertional limb pain. Specific exercise testing cut-off values should be used in patients with normal ABI to diagnose PAD.

Modulation of functionally active endothelin-converting enzyme by chronic neutral endopeptidase inhibition in experimental atherosclerosis.

BACKGROUND: Endothelin-converting enzyme-1 (ECE-1) processes big endothelin-1 (ET-1) to ET-1, a peptide implicated in atheroma formation. ECE-1 exists in 2 isoforms (ECE-1alpha and ECE-1beta), the result of alternative splicing of a common gene. Neutral endopeptidase (NEP) is a genetically distinct metallopeptidase that degrades the natriuretic peptides. These peptides mediate antiproliferative and vasodilating actions. We sought to demonstrate the distribution of the 2 ECE-1 isoforms in experimental atherosclerosis, to determine the effects of chronic NEP-I on plasma cGMP concentrations, vascular wall ECE-1 activity, and ET-1 concentration, and to correlate these actions with atheroma formation. Our hypothesis was that chronic NEP-I, in association with augmented cGMP, would inhibit ECE-1 conversion of big ET-1 to active ET-1, thus reducing tissue ET-1 concentrations and associated atheroma formation. METHODS AND RESULTS: Cholesterol-fed New Zealand White rabbits (n=8, 1% cholesterol diet) and NEP-I-treated cholesterol-fed New Zealand White rabbits (n=8; candoxatril, 30 mg/kg per day, Pfizer) were euthanized after 8 weeks of feeding. ECE-1alpha and ECE-1beta immunoreactivity was present in the aortas of both groups. Compared with control values, plasma cGMP concentrations were increased (2.8+/-0.6 versus 8.4+/-1.2 pmol/mL, P<0.05), ECE-1 activity was attenuated (68+/-3% versus 32+/-8%, P<0. 05), aortic tissue ET-1 concentrations were reduced (4.6+/-0.5 versus 3.2+/-0.3 pg/mg protein, P<0.05), and atheroma formation was attenuated (62+/-6% versus 34+/-5%, P<0.01) by NEP-I. CONCLUSIONS: These studies suggest that ECE-1 is present and functionally active in the vascular wall in atherosclerosis. Inhibition of ECE-1 by NEP-I represents a novel approach to interruption of the endothelin system in this cardiovascular disease state.

Vascular actions of brain natriuretic peptide: modulation by atherosclerosis and neutral endopeptidase inhibition.

OBJECTIVES: We sought to define the vascular actions of the cardiac hormone brain natriuretic peptide (BNP) on cellular proliferation and cyclic guanosine monophosphate (cGMP) in human aortic vascular smooth muscle cells (HAVSMCs). Secondly, we investigated BNP and acetylcholine (ACh) vasorelaxations in aortic rings from normal and atherosclerotic rabbits in the presence and absence of long-term oral inhibition of neutral endopeptidase (NEP). BACKGROUND: The vascular actions of BNP are not well defined, despite the presence of its receptor in vascular smooth muscle and the upregulation of NEP, the ectoenzyme that degrades BNP, in the vascular wall in atherosclerosis. METHODS: HAVSMCs stimulated with fetal calf serum (FCS) were pulsed with bromodeoxyuridine (BrdU) with and without BNP. The HAVSMCs were incubated in the presence and absence of BNP to assess cGMP. Vasorelaxations to BNP and ACh were assessed in rings in normal and atherosclerotic rabbits in the presence and absence of long-term oral inhibition of NEP, together with assessment of atheroma formation. RESULTS: FCS-stimulated BrdU uptake in HAVSMCs was suppressed with BNP. BNP potentiated cGMP in HAVSMCs. BNP resulted in potent vasorelaxation in normal isolated aortic rings, which were impaired in atherosclerotic versus normal rabbits and preserved with NEP inhibition, which also decreased atheroma formation. Relaxations to ACh, which were also impaired in atherosclerosis, were preserved with inhibition of NEP. CONCLUSIONS: We conclude that BNP potently inhibits vascular smooth muscle cell proliferation and potentiates the generation of cGMP. BNP potently relaxes the normal rabbit aorta, and this response is impaired in atherosclerosis but preserved with inhibition of NEP, together with a reduction in atheroma formation and preservation of relaxations to ACh.

Presence of Dendroaspis natriuretic peptide-like immunoreactivity in human plasma and its increase during human heart failure.

OBJECTIVE: To determine whether Dendroaspis natriuretic peptide (DNP), a novel peptide isolated from the venom of the Dendroaspis angusticeps snake that contains a 17-amino acid disulfide ring structure similar to that in atrial, brain, and C-type natriuretic peptides, is present in normal human plasma and myocardium and whether, like the other natriuretic peptides, DNP-like immunoreactivity (DNP-LI) is activated in human congestive heart failure (CHF). MATERIAL AND METHODS: Circulating DNP-LI was assessed in 19 normal human subjects and 19 patients with CHF (New York Heart Association class III or IV) with a specific and sensitive radioimmunoassay for DNP with no cross-reactivity with the other natriuretic peptides. Immunohistochemical studies that used polyclonal rabbit anti-DNP antiserum were performed on human atrial myocardial tissue obtained from four patients with end-stage CHF who were undergoing cardiac transplantation and from three donor hearts at the time of transplantation. RESULTS: We report that DNP-LI circulates in normal human plasma and is present in the normal atrial myocardium. In addition, DNP-LI is increased in the plasma of patients with CHF. CONCLUSION: This study demonstrates, for the first time, the presence of a DNP-like peptide in normal human plasma and in the atrial myocardium. Additionally, these studies demonstrate increased plasma DNP-LI in human CHF. These results support the possible existence of an additional new natriuretic peptide in humans, which may have a role in the neurohumoral activation that characterizes human CHF.

Predicting superficial venous incompetence with strain gauge plethysmography.

BACKGROUND: With currently available effective interventional methods to treat superficial venous insufficiency, it becomes particularly important to have a simple and reliable method to evaluate the location and severity of venous reflux. To date, there are few studies that evaluated plethysmography with and without tourniquet application to differentiate superficial from deep venous incompetence. OBJECTIVES: To determine if strain gauge plethysmography (SGP) with and without tourniquet application can be used to distinguish between the superficial and deep venous components of venous incompetence. METHODS: We conducted a prospective study using SGP with and without tourniquet application and duplex ultrasound (duplex US) to assess the severity and location of venous incompetence in 62 patients (85 limbs, 42 women, with an age range of 32-81 years) referred to our vascular laboratory for haemodynamic evaluation. Based on duplex US results, patients were diagnosed with superficial (SVI), deep and superficial (mixed) and deep vein incompetence (DVI). RESULTS: Mixed incompetence was the most common type. Twenty-three out of 33 limbs in the SVI group normalized their refill rate (RR) with tourniquet application (69.6%). Normalization of the RR with tourniquet application was less common in the mixed (n: 17 out of 40, 42.5%) and DVI (n: 2 out of 6, 33.3%) groups. CONCLUSION: SGP with tourniquet application is a simple and fast technique that can identify patients with SVI, based on RR improvement, who probably would benefit more from ablation procedures. Further studies evaluating impact of SGP with tourniquet test results on clinical outcome of SVI invasive treatment are warranted.

Aging and vasoreactivity: in vivo responses in the beagle hindlimb.

The purpose of this investigation was to determine whether vasodilator responses are attenuated and whether vasoconstriction is augmented with age in resistance vessels in the hindlimb of the dog. We examined blood flow (FAF) and pressure (FAP) responses in the femoral arterial system in older (109 +/- 8-mo-old) and younger mature (31 +/- 3-mo-old) female beagles during pentobarbital anesthesia. Vasodilator responses were evaluated during the intra-arterial administration of acetylcholine (ACh), which produces endothelium-dependent vasodilation, and albuterol, which mediates relaxation in vascular smooth muscle via beta-adrenoceptors. The vasoconstrictor response to phenylephrine (PE), an alpha-adrenergic agonist, was also examined. ACh and albuterol each induced dose-dependent vasodilation in the older and in the younger dogs. Resultant changes in neither FAF nor FAP were affected by age in response to either of these vasodilator substances. Likewise, reductions in femoral vascular resistance (FVR) in response to ACh or to albuterol were not age dependent. Vasodilation following induced hindlimb ischemia resulted in similar increases in FAF in both groups, but produced a greater reduction in FAP in older vs. younger dogs (P = 0.05). Similarly, FVR decreased more in the older beagles (P = 0.02). Vasoconstriction mediated by PE resulted in similar reductions in FAF in both age groups, but the increase in FAP was less at several PE doses in older vs. younger dogs (P < 0.05). However, increases in FVR in response to PE were not statistically different in the younger and older beagles.(ABSTRACT TRUNCATED AT 250 WORDS)

Further attenuation of endothelium-dependent relaxation imparted by natriuretic peptide receptor antagonism.

Nitric oxide (NO) is an important endothelium-derived relaxing factor that functions via activation of soluble guanylyl cyclase and cGMP generation in vascular smooth muscle. Recently, studies have described the synthesis and secretion of C-type natriuretic peptide (CNP) from endothelial cells. This peptide also mediates relaxation via cGMP but through activation of particulate guanylyl cyclase. We tested the hypothesis that endothelium-dependent relaxations to acetylcholine or bradykinin in isolated canine coronary arteries involve both releases of NO and CNP. Rings of canine coronary arteries were incubated with either inhibitors of NO production (N(G)-monomethyl-L-arginine, L-NMMA) or the natriuretic peptide receptor antagonist HS-142-1. CNP caused concentration-dependent relaxations of rings with and without endothelium. These relaxations were attenuated by HS-142-1. Relaxations to acetylcholine and bradykinin were attenuated by L-NMMA alone but not attenuated by HS-142-1 alone. Coinhibition with L-NMMA and HS-142-1 significantly inhibited acetylcholine- and bradykinin-induced relaxation to a magnitude greater than either inhibitor alone. In summary, a novel interaction between the NO and the natriuretic peptide system is demonstrated by increased attenuation of endothelium-dependent relaxations to acetylcholine and bradykinin when both NO synthase and natriuretic peptide receptors are inhibited. These investigations support the concept of release of multiple endothelium-derived factors in response to acetylcholine- and bradykinin-receptor stimulation in endothelial cells, which may include CNP, as well as NO.

Effects of aging per se on arterial stiffness: systemic and regional compliance in beagles.

Although previous studies have suggested that aging results in an increase in vascular stiffness, diseases that increase in prevalence with advanced age may have confounded the results of some of this past research. The purpose of this investigation was to determine whether aging per se results in reduced arterial compliance by using animals that are resistant to atherosclerosis and do not develop hypertension or hyperlipidemias with advanced age. We evaluated systemic and regional (femoral) arterial compliance in older (110 +/- 8 months old) and in younger (27 +/- 2 months old) female beagle dogs by using a computer-based assessment of the diastolic decay of arterial pressure waveforms and a modified Windkessel model of the circulation. Although systemic arterial pressure was very similar in both age groups, cardiac output was 29% lower (p = 0.03) and systemic vascular resistance was 24% higher (p = 0.02) in the older dogs. Moreover, there was an age-related reduction in systemic arterial compliance, derived both from the exponential decay in the arterial pulse (C1) (p = 0.05) and that derived from the oscillatory component of the diastolic pulse wave (C2) (p = 0.04). By contrast, although femoral vascular resistance was 25% higher in the older dogs (p = 0.04), regional (femoral) vascular compliance measured after femoral arterial occlusion was also 25% reduced but was not significantly changed with age (p = 0.14). These results demonstrate that systemic arterial compliance is reduced with age in dogs, extending this finding to animals without age-related diseases that frequently occur in older human beings. Regional compliance, evaluated in the isolated femoral vascular bed, also tends to be reduced with age, but variability in this parameter in dogs reduces the significance of this finding.

Role for the endogenous natriuretic peptide system in the control of basal coronary vascular tone in dogs.

1. While the natriuretic peptides (atrial, brain and C-type) mediate potent endothelium-independent vasorelaxing actions in vitro, the role of the endogenous natriuretic peptide system in vascular regulation in vivo remains unclear. 2. HS-142-1 is a novel natriuretic peptide receptor antagonist derived from a fungus named Aureobasidium sp. which selectively blocks particulate guanylate cyclase-linked natriuretic peptide A and B receptors that bind atrial, brain and C-type natriuretic peptide, and thus attenuates the generation of cGMP. 3. To characterize the vascular actions of the endogenous natriuretic peptide system in the control of basal coronary and systemic haemodynamics, six normal male mongrel anaesthetized dogs were studied while a second group of five dogs served as a control. HS-142-1 was given as an intravenous bolus at 3 mg/kg and was studied over five 20 min periods. 4. No significant difference after HS-142-1 was observed in mean arterial pressure, heart rate, cardiac output, right atrial pressure, pulmonary capillary wedge pressure or systemic vascular resistance compared with control. In contrast, a significant increase in coronary vascular resistance and decrease in coronary blood flow were observed which were different from the baseline values and the responses of the control group. 5. These studies demonstrate that HS-142-1 produces vasoconstriction in the coronary circulation. We conclude that the endogenous natriuretic peptide system, which is of cardiac and endothelial cell origin, is an important regulator of basal coronary vascular tone.

Enhanced endothelin-converting enzyme immunoreactivity in early atherosclerosis.

Endothelin-1 (ET-1) is a 21-amino-acid local and circulating factor whose plasma concentrations are increased in advanced atherosclerosis. ET-1 is cleaved from a prohormone (big ET-1) by endothelin-converting enzymes (ECEs) into the biologically active mature form which mediates vasoconstriction and cell proliferation. This study was designed to test by immunohistochemistry the hypothesis that ECE is present locally in the neointima of atherosclerotic vessels. Two groups of rabbits, control (n = 6) and cholesterol-fed (1% cholesterol diet for 8 weeks; n = 6) were sacrificed. Aortas were excised and divided for determination of tissue ET-1 concentration by RIA and immunohistochemical analysis of ECE. Vascular wall ET-1 was increased in the atherosclerotic aorta (6.1 +/- 0.8 vs. 9.8 +/- 0.9 pg/mg protein; p < 0.05), whereas circulating ET-1 concentrations were similar in the two groups (3.8 +/- 0.4 vs. 2.4 +/- 1.4 pg/ml). Immunostaining revealed the presence of ECE in endothelial and vascular smooth-muscle cells of the control group. Enhanced ECE immunoreactivity was present in atherosclerotic aortas, particularly in the neointimal macrophages and smooth-muscle cells. We conclude that local vascular wall, but not circulating ET-1, is increased in early atherosclerosis. In addition, ECE immunoreactivity is increased in early atherosclerosis and may therefore contribute to the generation of local ET-1 in early experimental atherosclerosis. These studies provide important insights into the regulation of ET-1 in early atherosclerosis, which may contribute to the elucidation of factors involved in the progression of atherosclerosis.

Renal actions of synthetic dendroaspis natriuretic peptide.

BACKGROUND: Dendroaspis natriuretic peptide (DNP), recently isolated from the venom of the green Mamba snake Dendroaspis angusticeps, is a 38 amino acid peptide containing a 17 amino acid disulfide ring structure similar to that of atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP), and C-type natriuretic peptide (CNP). DNP-like immunoreactivity (DNP-LI) was reported to be present in human plasma and atrial myocardium and to be elevated in human congestive heart failure. Although previously named DNP, it remains unknown if DNP is natriuretic or if is it present in canine plasma, urine, and atrial myocardium. METHOD: Studies were performed in vivo in anesthetized dogs (N = 6) using intravenous infusion of synthetic DNP at 10 and 50 ng/kg/min. Employing a sensitive and specific radioimmunoassay for DNP, the presence of DNP-like peptide was assessed in the canine plasma and urine before, during, and following the administration of exogenous synthetic DNP. Additionally, we performed immunohistochemical studies using the indirect immunoperoxidase method with polyclonal DNP antiserum in normal atrial myocardium (N = 10). Atrial concentrations of DNP-LI were also assessed. RESULTS: We report that DNP is markedly natriuretic and diuretic, which, like ANP and BNP, is associated with the increase in urinary and plasma cGMP. DNP-like peptide is also detected in canine plasma, urine, and atrial myocardium. CONCLUSION: These studies establish that DNP is a potent natriuretic and diuretic peptide with tubular actions linked to cGMP and that DNP may play a physiological role in the regulation of sodium excretion.