Superoxide dismutase gene transfer reduces portal pressure in CCl4 cirrhotic rats with portal hypertension.

BACKGROUND: Increased intrahepatic vascular tone in cirrhosis has been attributed to a decrease of hepatic nitric oxide (NO) secondary to disturbances in the post-translational regulation of the enzyme eNOS. NO scavenging by superoxide (O(2)(-)) further contributes to a reduction of NO bioavailability in cirrhotic livers. AIM: To investigate whether removing increased O(2)(-) levels could be a new therapeutic strategy to increase intrahepatic NO, improve endothelial dysfunction and reduce portal pressure in cirrhotic rats with portal hypertension. METHODS: Adenoviral vectors expressing extracellular superoxide dismutase (SOD) (AdECSOD) or beta-galactosidase (Adbetagal) were injected intravenously in control and CCl(4)-induced cirrhotic rats. After 3 days, liver O(2)(-) levels were determined by dihydroethidium staining, NO bioavailability by hepatic cGMP levels, nitrotyrosinated proteins by immunohistochemistry and western blot, and endothelial function by responses to acetylcholine in perfused rat livers. Mean arterial pressure (MAP) and portal pressure were evaluated in vivo. RESULTS: Transfection of cirrhotic livers with AdECSOD produced a significant reduction in O(2)(-) levels, a significant increase in hepatic cGMP, and a decrease in liver nitrotyrosinated proteins which were associated with a significant improvement in the endothelium-dependent vasodilatation to acetylcholine. In addition, in cirrhotic livers AdECSOD transfection produced a significant reduction in portal pressure (17.3 (SD 2) mm Hg vs 15 (SD 1.6) mm Hg; p<0.05) without significant changes in MAP. In control rats, AdECSOD transfection prevents the increase in portal perfusion pressure promoted by an ROS-generating system. CONCLUSIONS: In cirrhotic rats, reduction of O(2)(-) by AdECSOD increases NO bioavailability, improves intrahepatic endothelial function and reduces portal pressure. These findings suggest that scavenging of O(2)(-) might be a new therapeutic strategy in the management of portal hypertension.

Gene transfer after subarachnoid hemorrhage: a tool and potential therapy.

This mini-review describes steps towards gene therapy to prevent vasospasm after subarachnoid hemorrhage, and summarizes some remaining obstacles. With recombinant adenoviruses, it is now possible to prevent vasospasm in experimental animals. If an adenoviral or other effective vector is demonstrated to be safe, it is likely that gene therapy will be used in patients to prevent vasospasm.

Effect of acute hypoxia on vascular responsiveness in man. I. Responsiveness to lower body negative pressure and ice on the forehead. II. Responses to norepinephrine and angiotensin. 3. Effect of hypoxia and hypocapnia.

An effect of hypoxemia on vascular responsiveness and blood pressure regulation has not been demonstrated in man. The response of forearm resistance vessels to several vasoconstrictor stimuli was compared during normoxia and acute hypoxia. Forearm vasoconstrictor responses to lower body negative pressure and to the application of ice to the forehead, which are neurogenic stimuli, were decreased during acute hypoxia. Lower body negative pressure caused a decrease in mean arterial pressure during hypoxia, but not during normoxia. Because norepinephrine is the neurotransmitter released during reflex vasoconstriction, we considered the possibility that decreased responsiveness to norepinephrine might be one mechanism for diminished responses to lower body negative pressure and ice on the forehead during hypoxia. Hypoxia decreased the response of forearm resistance vessels to infusions of norepinephrine and angiotensin into the brachial artery. In addition, the effectiveness of intravenous infusions of norepinephrine in elevating mean arterial pressure was decreased during hypoxia. Since exposure to acute hypoxia stimulates hyperventilation and hypocapnia, experiments were done to determine the contribution of hypocapnia during hypoxia to the decreased vasoconstriction. The results indicate that hypocapnia may diminish the vascular response to some stimuli, but the reduction in oxygen appears to be the primary mechanism for decreased vasoconstrictor responses during acute hypoxia.

Relationship between plasma sodium concentration and vascular reactivity in man.

Experiments were done to study the effects of acute changes in plasma sodium concentration [Na] on vasoconstrictor responses to norepinephrine and to reflex sympathetic stimulation in man. [Na] in the venous return from the forearm of each of 21 normal subjects was reduced (to an average of 118 mEq/liter), increased (147 mEq/liter), and maintained within the normal range (140 mEq/liter) by means of infusions into the brachial artery of three solutions containing different [Na]. Mannitol or sucrose and disodium sulfate were substituted for sodium chloride to produce the desired changes in [Na] without changing blood osmolarity. Blood flow to the forearm (plethysmograph) increased during infusion of the three solutions, but the increase in flow was not related to [Na]. Vasoconstrictor responses to injections of norepinephrine and angiotensin into the brachial artery were reduced at low [Na] and augmented at high [Na]. Reflex vasoconstriction, activated by lower body negative pressure, was similarly affected by changes in [Na]. In the isolated, perfused gracilis muscle of dog vasoconstrictor responses to norepinephrine and to nerve stimulation were attenuated, and the extraction of norepinephrine by this muscle was smaller when plasma [Na] was reduced.

Role of large arteries in regulation of cerebral blood flow in dogs.

Previous studies have demonstrated a significant pressure gradient from carotid artery to pial or middle cerebral arteries. This pressure gradient suggests that large cerebral arteries contribute to cerebral resistance. We have tested the hypothesis that large cerebral arteries contribute to regulation of cerebral blood flow during changes in blood gases and arterial pressure. Microspheres were used to measure brain blood flow in anesthetized dogs. Resistance of large cerebral arteries was estimated by determining the pressure gradient between common carotid and wedged vertebral artery catheters. Systemic hypercapnia and hypoxia dilated large cerebral arteries, and hypocapnia constricted large cerebral arteries. Resistance of large arteries was 0.6+/-0.1 (mean +/- SE) mm Hg per ml/min per 100 g during normocapnia. During hypercapnia and hypoxia, large artery resistance decreased significantly to 0.2 +/- 0.03 and 0.3 +/- 0.05, respectively. During hypocapnia large artery resistance increased significantly to 1.0 +/- 0.1. In other experiments, we found that large cerebral arteries participate in auto-regulatory responses to hemorrhagic hypotension. When arterial pressure was reduced from 110 to 58 mm Hg, autoregulation maintained cerebral blood flow constant, and resistance of large cerebral arteries decreased significantly from 1.0 +/- 0.2 to 0.6 +/- 0.1 mm Hg per ml/min per 100 g. In absolute terms, we calculated that 20-45% of the change in total cerebral resistance during these interventions was accounted for by changes in large artery resistance. These studies indicate that large cerebral arteries, as well as arterioles, participate actively in regulation of cerebral blood flow during changes in arterial blood gases and during autoregulatory responses to hemorrhagic hypotension.

Depression of ventilation by dopamine in man. Evidence for an effect on the chemoreceptor reflex.

Dopamine is present in the carotid body and has been postulated to be an inhibitory neurotransmitter. The purpose of this study was to determine the effects of dopamine on ventilation in man and to examine its mechanism of action. Dopamine (0.5-10 mug/kg per min) was infused in eight normal men at different levels of arterial chemoreceptor activity, produced by varying the inspired Po(2). During normoxia dopamine produced a small decrease in minute ventilation (Ve) and an increase in arterial Pco(2). When arterial chemoreceptors were stimulated by hypoxia, infusion of dopamine produced a marked initial depression of Ve followed by a sustained although less pronounced decrease in Ve. An increase in Pa(co) (2) and a decrease in Pao(2) were also observed. When arterial chemoreceptor activity was suppressed by hyperoxia, infusion of dopamine did not affect ventilation. Subjects also breathed a hypercarbic, hyperoxic gas mixture. The hypercarbia produces hyperventilation by stimulating central chemoreceptors, whereas the hyperoxia suppresses peripheral chemoreceptors. Dopamine did not alter ventilation while the subjects were breathing this gas mixture. These studies suggest that dopamine suppresses ventilation in man through an action on the arterial chemoreceptor reflex. These findings support the hypothesis that dopamine is an inhibitory neurotransmitter in the carotid body, and that release of dopamine may modulate the sensitivity of peripheral arterial chemoreceptors.

Restoration of endothelium-dependent relaxation by dietary treatment of atherosclerosis.

Atherosclerosis results in impaired relaxation to acetylcholine, thrombin, and the calcium ionophore A23187, all agents that require the presence of endothelium. We now report that dietary treatment of atherosclerosis in monkeys not only produces morphological improvement of the atherosclerotic lesion but restores endothelium-dependent vascular relaxation to normal. Because the intima remains thickened after regression of atherosclerosis, these studies suggest that intimal thickening which is present in both atherosclerotic vessels and after regression of atherosclerosis does not prevent the endothelium-derived relaxing factor from reaching the underlying vascular smooth muscle.

Impaired reflex vasoconstriction in chronically hypoxemic patients.

Acute hypoxia impairs vasoconstrictor responses in normal men. The present study was done to determine whether reflex vasoconstriction is impaired in chronically hypoxemic patients and whether correction of hypoxemia in these patients improves their cardiovascular reflexes. In eight chronically hypoxemic patients, arterial P(O2) was increased from an average of 45 mm Hg while breathing room air to 161 mm Hg while breathing 40-100% oxygen, with minimal changes in arterial P(CO2) or pH. Correction of hypoxemia did not cause changes in resting arterial pressure or in forearm vascular resistance, but it caused a small increase in resting heart rate. Reflex responses to lower body negative pressure, which causes pooling of blood in the lower part of the body, were observed. When the patients were hypoxemic, lower body negative pressure caused a fall in arterial pressure, slight constriction of forearm vessels, and a small increase in heart rate. When hypoxemia was corrected, the same intervention caused marked vasoconstriction and a greater increase in heart rate, and there was no decrease in arterial pressure. The results indicate that reflex vasoconstrictor responses are depressed in chronic hypoxemia, indicating that adaptive mechanisms which occur in chronic hypoxemia do not include preservation of sympathetic reflexes.

Reflex vascular responses to left ventricular outflow obstruction and activation of ventricular baroreceptors in dogs.

Reflex vascular responses to acute left ventricular outflow obstruction were studied in anesthetized dogs. The studies were done to compare the effects of activation of ventricular baroreceptors on vascular resistance in skeletal muscle (gracilis muscle) and skin (hindpaw); to identify afferent and efferent pathways which mediate the reflex vasodilatation; and to assess the relative contribution of ventricular baroreceptors and baroreceptors in left atrium and pulmonary vessels in responses to left ventricular outflow obstruction. The gracilis artery and the cranial tibial artery to the paw were perfused separately at constant flow. Changes in perfusion pressure to each bed reflected changes in vascular resistance. Outflow obstruction was produced by inflating a balloon in the left ventricular outflow tract for 15 s while pressures in the left ventricle and aortic arch were measured. Inflation of the balloon increased left ventricular pressure and decreased pressure in the aortic arch. Low and high levels of obstruction produced dilator responses averaging -5+/-3 (SE) and -42+/-11 mm Hg in muscle and -1+/-1 and -3+/-2 mm Hg in paw. Denervation, phentolamine, and glyceryltrinitrate caused greater dilatation in paw than did left ventricular outflow obstruction. This indicates that dilator responses in the paw were not limited by a low level of resting neurogenic constrictor tone or by a negligible dilator capacity of these vessels. Obstruction to left ventricular inflow increased left atrial pressure, but did not cause reflex vasodilatation. This suggests that low pressure baroreceptors in atria or pulmonary vessels did not contribute to vasodilator responses to left ventricular outflow obstruction. Vasodilator responses to outflow obstruction were blocked by bilateral vagotomy, sectioning the sciatic and obturator nerves, and administration of phentolamine, but were not decreased by atropine or tripelennamine. The results indicate that activation of left ventricular baroreceptors produces striking vasodilatation in skeletal muscle, but only slight vasodilatation in skin. The data suggest that the difference in dilator responses in the two beds results from greater withdrawal of adrenergic constrictor tone to skeletal muscle than to skin. Activation of sympathetic cholinergic or histaminergic dilator pathways does not contribute to the dilatation.

Interaction of baroreceptor and chemoreceptor reflexes. Modulation of the chemoreceptor reflex by changes in baroreceptor activity.

The purpose of this study was to determine whether the level of arterial pressure and degree of baroreceptor activation affect responses to stimulation of chemoreceptors. Chemoreceptors were stimulated by injecting nicotine into the common carotid artery of anesthetized and paralyzed dogs. Responses were observed in the innervated gracilis muscle, perfused at constant flow while perfusion pressure was measured. Arterial pressure was lowered by bleeding the animals and raised by transient occlusion of the descending aorta. Vasoconstrictor responses to stimulation of chemoreceptors were enhanced by hypotension and inhibited by elevation of arterial pressure. Potentiation of the chemoreceptor reflex by hemorrhagic hypotension was not the result of altered vascular resistance in the gracilis muscle, sensitization of chemoreceptors by catecholamines or acidosis, or changes in cerebral perfusion pressure. Additional studies were done in which we excluded the possibility that the changes resulted from direct effects of changes in arterial pressure on chemoreceptors. Both carotid bifurcations were isolated and perfused. On one side, pressure was raised to stimulate the carotid sinus baroreceptors. On the other side, the carotid body chemoreceptors were stimulated by nicotine or by hypoxic and hypercapnic blood. Activation of baroreceptors on one side attenuated the vasoconstrictor response to chemoreceptor stimulation on the other side. This excludes a direct effect of changes in arterial pressure on the chemoreceptors and suggests a central interaction of these reflexes. We conclude that vasoconstrictor responses to stimulation of chemoreceptors are potentiated by hypotension and inhibited by transient hypertension. These effects appear to result at least in part from a central interaction of chemoreceptor and baroreceptor reflexes.

Effects of adrenergic stimulation on ventilation in man.

The mechanism by which catecholamines affect ventilation in man is not known. Ventilatory responses to catecholamines were observed in normal subjects before and after adrenergic receptor blockade. Intravenous infusions of norepinephrine and isoproterenol caused significant increases in minute volume and decreases in end-tidal P(Co2) which were blocked by the administration of propranolol, a beta adrenergic receptor blocker. The hyperventilatory response to hypoxia was not altered by propranolol. Intravenous infusion of phenylephrine caused a small but significant decrease in minute volume which was antagonized by phentolamine, an alpha adrenergic receptor blocker. Angiotensin, a nonadrenergic pressor agent, also decreased minute volume significantly.100% oxygen was administered to suppress arterial chemoreceptors. Increases in minute volume and decreases in arterial P(Co2) in response to norepinephrine and isoproterenol were blocked by breathing 100% oxygen. The decrease in minute volume during phenylephrine was not altered by 100% oxygen. THE RESULTS INDICATE THAT: (a) beta adrenergic receptors mediate the hyperventilatory response to norepinephrine and isoproterenol but not to hypoxia. (b) the pressor agents phenylephrine and angiotensin decrease ventilation, and (c) suppression of chemoreceptors blocks the ventilatory response to norepinephrine and isoproterenol but not to phenylephrine. Implications concerning the interaction of adrenergic receptors and chemoreceptors with respect to the hyperventilatory response to catecholamines are discussed.

Differences in direct effects of adrenergic stimuli on coronary, cutaneous, and muscular vessels.

Direct effects of adrenergic stimuli on coronary vessels in dogs were compared with effects on vessels to skin (hind paw) and skeletal muscle (gracilis muscle) after intravenous administration of practolol (2 mg/kg), a selective myocardial beta receptor blocker which minimized indirect effects of myocardial stimulation on coronary vascular resistance. The left circumflex coronary, cranial tibial, and gracilis arteries were perfused separately but simultaneously at constant flow. Perfusion pressures, left ventricular pressure and dP/dt. and heart rate were recorded. Changes in perfusion pressure to each bed reflected changes in vascular resistance. The direct constrictor effects of sympathetic nerve stimulation, norepinephrine and phenylephrine on coronary vessels were minimal compared with effects on cutaneous and muscular vessels. Subsequent blockade of vascular beta receptors did not augment the constrictor responses. Angiotensin, a nonadrenergic stimulus, produced striking coronary vasoconstriction which exceeded that in skin and approximated that in muscle. These results suggests that there is a paucity of alpha adrenergic receptors in coronary vessels compared to cutaneous and muscular vessels. Direct dilator responses to isoproterenol were similar in coronary and cutaneous vessels, but were greater in muscular vessels. Responses to glyceryl trinitrate, a nonadrenergic dilator, also were greater in skeletal muscle. Therefore, differences in effects of isoproterenol on the three beds may reflect differences in reactivity to dilator stimuli rather than differences in the density of beta receptors. In contrast to norepinephrine, the predominant direct effect of epinephrine on coronary vessels was dilatation mediated through activation of vascular beta receptors. A constrictor effect caused by stimulation of alpha receptors was unmasked by propranolol.Finally, the order of potency of agonists in stimulating coronary vascular beta receptors and the demonstration of selective beta receptor blockade with practolol suggest that beta receptors in coronary vessels resemble those in peripheral vessels more than those in myocardium.

Hemodynamic sequelae of regression of experimental atherosclerosis.

Regression of experimental atherosclerosis is characterized by decreased intimal thickness and luminal enlargement, but intimal fibrosis becomes more dense. We tested the hypothesis that fibrosis of arteries during regression might limit vasodilator capacity and restrict hemodynamic improvement despite luminal improvement. We studied limb, coronary, and cerebral hemodynamics in 11 normal cynomolgus monkeys, 10 monkeys given an atherogenic diet for 20 mo and 8 monkeys given a regression diet for an additional 18 mo. The atherogenic diet induced lesions of moderate severity (50-60% stenosis); owing to characteristic vessel growth during the atherogenic period, luminal size did not decrease correspondingly. Regression monkeys showed typical changes of regression with luminal enlargement but increased fibrosis. The iliac artery was perfused at constant blood flow and maximal vasodilatation was produced with papaverine. Blood flow was measured with microspheres during maximal vasodilatation in the coronary bed (adenosine) and cerebral bed (hypercapnia). In normal monkeys, minimal vascular resistances were 1.95 +/- 0.19 mm Hg/ml/min X 100 g (mean +/- SE) (limb), 0.13 +/- 0.01 (coronary), and 0.44 +/- 0.02 (cerebral). In atherosclerotic monkeys minimal resistance increased (P less than 0.05) 108, 62, and 166% in the limb, coronary, and cerebral beds, respectively. In regression monkeys, minimal resistance increased from values found in atherosclerotic animals in the limb (+22%), decreased inconsistently in the coronary bed (-19%), and decreased significantly in the cerebral bed (-44%, P less than 0.05). Thus morphologic regression was accompanied by significant hemodynamic improvement during maximal dilatation only in cerebral vessels. We conclude that increases in luminal size during regression of atherosclerotic lesions may not be associated with increases in vasodilator capacity, as intimal fibrosis may limit physiologically important hemodynamic improvement.

Regulation of blood flow to the aortic media in dogs.

Morphologic observations suggest that the inner layers of the thoracic aorta in man and dog are avascular and the outer layers have vasa vasorum. It appears that vasa vasorum are essential in the thoracic aorta because their interruption produces medial necrosis. These experiments provide the first measurements of blood flow through aortic vasa vasorum and examine physiologic regulation of that flow. During control conditions the outer two-thirds of the media of the thoracic aorta received 10 ml/min per 100 g blood flow through vasa vasorum. Flow to the inner third of the aorta was 1 ml/min per 100 g. Flow to both the inner and outer media of the abdominal aorta was less than 1 ml/min per 100 g. Adenosine increased blood flow to vasa vasorum in the outer media of the thoracic aorta from 7 to 18 ml/min per 100 g, but did not increase flow to the inner layers of the aorta. Hemorrhagic hypotension decreased flow in the outer media of the thoracic aorta from 14 to 2 ml/min per 100 g. Acute hypertension failed to increase blood flow through vasa vasorum, as conductance decreased significantly. These studies indicate that vasa vasorum provide a considerable amount of blood flow to the outer layers of the thoracic aorta. The vessels are responsive to physiologic stimuli because they dilate during infusion of adenosine and constrict during both hemorrhagic hypotension and acute hypertension. We speculate that the failure of blood flow to the aortic wall to increase during acute hypertension might, if it were sustained, contribute to aortic medial necrosis.

Regulation of blood flow in Paget's disease of bone.

Previous studies have demonstrated an increase in blood flow to extremities involved by Paget's disease of bone. It has been assumed that the increase in blood flow is through bone, but warmth of skin overlying Pagetic bone suggests that cutaneous blood flow might be increased. In three patients with Paget's disease involving one extremity, we compared blood flow in "Pagetic" extremities with flow in the contralateral normal extremities. Resting blood flow (measured with water plethysmographs) was 14.2plus or minus2.9 (meanplus or minusSE) ml/min times 100 ml extremity in the Pagetic limbs. The contribution of cutaneous flow to the increase in extremity blood flow was evaluated with epinephrine iontophoresis, which suppresses flow to skin but not to underlying tissue. Epinephrine iontophoresis of the Pagetic extremities decreased blood flow from 14.2plus or minus2.9 to 3.6plus or minus1.5 ml/min. Local heating (which increases cutaneous flow only) failed to increase blood flow in the Pagetic extremities as much as it did in the normal extremities. This suggests that cutaneous vessels in the Pagetic extremities were already dilated. During heating, blood flow in the normal extremities was similar to resting flow in the Pagetic extremities; this indicates that increases in cutaneous flow could account for most of the increase in total blood flow in the Pagetic extremities. Adrenergic control of blood flow to the Pagetic extremities was compared with that of the normal extremities. Vasoconstrictor responses to reflex stimuli in the Pagetic extremities were reduced; when vasoconstriction occurred it was gradual and sustained after termination of the stimuli, which suggests an exaggerated humoral response but reduced neural response to the stimuli. Intravenous epinephrine produced vasoconstriction in the Pagetic extremities and vasodilatation in the normal extremities. In summary, responses to epinephrine iontophoresis and heating suggest that the increase in blood flow in Pagetic extremities is primarily the result of cutaneous vasodilatation.

The role of low pressure baroreceptors in reflex vasoconstrictor responses in man.

Studies were performed on 11 healthy men to evaluate the role of low pressure baroreceptors in the reflex forearm vasoconstrictor responses (plethysmography) to venous pooling produced by lower body negative pressure. Lower body negative pressure (LBNP) at - 5, - 10, - 20, and - 40 mm Hg lowered central venous pressure by 42, 59, 74, and 93%, respectively, and decreased forearm vascular conductance by 24, 29, 34, and 40%, respectively. The decreases in forearm blood flow and conductance during the low levels of venous pooling (LBNP - 5 and - 10 mm Hg) occurred without significant changes in arterial pressure, arterial dP/dt. and heart rate. These results with the low levels indicate that maneuvers which decrease venous return and central venous pressure in man can influence forearm vascular tone without significant changes in the determinants of carotid and aortic baroreceptor activity. During high levels of venous pooling (LBNP - 20 and - 40 mm Hg), significant decreases in arterial pressure and dP/dt and significant increases in heart rate accompanied the further reductions in central venous pressure, forearm blood flow, and forearm vascular conductance. About 73% of the decrease in conductance during venous pooling at LBNP - 40 mm Hg, which was sufficient to decrease arterial pressure and activate high pressure baroreceptor reflexes, occurred during low levels of venous pooling at LBNP - 10 mm Hg without changes in arterial pressure. This suggests that much of the forearm vasoconstriction with the high levels of venous pooling, which were sufficient to decrease arterial pressure, may be accounted for by reflexes originating in areas other than high pressure baroreceptors. The results of these studies suggest that low pressure baroreceptors exert an important influence on forearm vascular tone during decreases in venous return and central venous pressure in man.

Abnormal vascular responses to exercise in patients with aortic stenosis.

We tested the hypothesis that the normal forearm vasoconstrictor response to leg exercise is inhibited or reversed in patients with aortic stenosis, possibly because of activation of left ventricular baroreceptors. Forearm vascular responses to supine leg exercise were measured in 10 patients with aortic stenosis and in 2 control groups of 6 patients with mitral stenosis and 5 patients without valvular heart disease.Forearm vasoconstriction occurred during exercise in the control groups. In contrast, forearm blood flow increased and forearm vascular resistance did not change in patients with aortic stenosis. In six patients with aortic stenosis and a history of exertional syncope, forearm vasodilatation occurred during the second minute of leg exercise. Inhibition or reversal of forearm vasoconstrictor responses in aortic stenosis was asscociated with significant increases in left ventricular pressure. In three patients with aortic stenosis and exertional syncope, forearm vasodilator responses to exercise changed to vasoconstrictor responses after aortic valve replacement. The results indicate that forearm vasoconstrictor responses to leg exercise are inhibited or reversed in patients with aortic stenosis, possibly because of activation of left ventricular baroreceptors. The observations suggest that reflex vasodilatation resulting from activation of left ventricular baroreceptors may contribute to exertional syncope in patients with aortic stenosis.

Vascular dysfunction in monkeys with diet-induced hyperhomocyst(e)inemia.

Elevated plasma homocyst(e)ine may predispose to complications of vascular disease. Homocysteine alters vasomotor regulatory and anticoagulant properties of cultured vascular endothelial cells, but little is known about effects of hyperhomocyst(e)inemia on vascular function in vivo. We tested the hypothesis that diet-induced moderate hyperhomocyst(e)inemia is associated with vascular dysfunction in cynomolgus monkeys. Plasma homocyst(e)ine increased from 4.O +/- O.2 microM when monkeys were fed normal diet to 10.6 +/- 2.6 microM when they were fed modified diet (mean +/- SE; P = 0.02). Vasomotor responses were assessed in vivo by quantitative angiography and Doppler measurement of blood flow velocity. In response to activation of platelets by intraarterial infusion of collagen, blood flow to the leg decreased by 42 +/- 9% in monkeys fed modified diet, compared with 14 +/- 11% in monkeys fed normal diet (P = 0.008), Responses of resistance vessels to the endothelium-dependent vasodilators acetylcholine and ADP were markedly impaired in hyperhomocyst(e)inemic monkeys, which suggests that increased vasoconstriction in response to collagen may be caused by decreased vasodilator responsiveness to platelet-generated ADP. Relaxation to acetylcholine and, to a lesser extent, nitroprusside, was impaired ex vivo in carotid arteries from monkeys fed modified diet. Thrombomodulin anticoagulant activity in aorta decreased by 34 +/- 15% in hyperhomocyst(e)inemic monkeys (P = 0.03). We conclude that diet-induced moderate hyperhomocyst(e)inemia is associated with altered vascular function.

Gene transfer of calcitonin gene-related peptide prevents vasoconstriction after subarachnoid hemorrhage.

We sought to determine whether adenovirus-mediated gene transfer in vivo of calcitonin gene-related peptide (CGRP), a potent vasodilator, ameliorates cerebral vasoconstriction after experimental subarachnoid hemorrhage (SAH). Arterial blood was injected into the cisterna magna of rabbits to mimic SAH 5 days after injection of AdRSVCGRP (8x10(8) pfu), AdRSVbetagal (control virus), or vehicle. After injection of AdRSVCGRP, there was a 400-fold increase in CGRP in cerebrospinal fluid. Contraction of the basilar artery to serotonin in vitro was greater in rabbits after SAH than after injection of artificial cerebrospinal fluid (P<0.001). Contraction to serotonin was less in rabbits with SAH after AdRSVCGRP than after AdRSVbetagal or vehicle (P:<0.02). Basal diameter of the basilar artery before SAH (measured with digital subtraction angiogram) was 13% greater in rabbits treated with AdRSVCGRP than in rabbits treated with vehicle or AdRSVbetagal (P:<0.005). In rabbits treated with vehicle or AdRSVbetagal, arterial diameter after SAH was 25+/-3% smaller than before SAH (P<0.0005). In rabbits treated with AdRSVCGRP, arterial diameter was similar before and after SAH and was reduced by 19+/-3% (P<0.01) after intracisternal injection of CGRP-(8-37) (0.5 nmol/kg), a CGRP(1) receptor antagonist. To determine whether gene transfer of CGRP after SAH may prevent cerebral vasoconstriction, we constructed a virus with a cytomegalovirus (CMV) promoter, which results in rapid expression of the transgene product. Treatment of rabbits with AdCMVCGRP after experimental SAH prevented constriction of the basilar artery 2 days after SAH. Thus, gene transfer of CGRP prevents cerebral vasoconstriction in vivo after experimental SAH.

Endothelial dysfunction and elevation of S-adenosylhomocysteine in cystathionine beta-synthase-deficient mice.

Hyperhomocysteinemia is associated with increased risk for cardiovascular events, but it is not certain whether it is a mediator of vascular dysfunction or a marker for another risk factor. Homocysteine levels are regulated by folate bioavailability and also by the methyl donor S-adenosylmethionine (SAM) and its metabolite S-adenosylhomocysteine (SAH). We tested the hypotheses that endothelial dysfunction occurs in hyperhomocysteinemic mice in the absence of folate deficiency and that levels of SAM and SAH are altered in mice with dysfunction. Heterozygous cystathionine beta-synthase-deficient (CBS(+/-)) and wild-type (CBS(+/+)) mice were fed a folate-replete, methionine-enriched diet. Plasma levels of total homocysteine were elevated in CBS(+/-) mice compared with CBS(+/+) mice after 7 weeks (27.1+/-5.2 versus 8.8+/-1.1 micromol/L; P<0.001) and 15 weeks (23.9+/-3.0 versus 13.0+/-2.3 micromol/L; P<0.01). After 15 weeks, but not 7 weeks, relaxation of aortic rings to acetylcholine was selectively impaired by 35% (P<0.05) and thrombomodulin anticoagulant activity was decreased by 20% (P<0.05) in CBS(+/-) mice. Plasma levels of folate did not differ between groups. Levels of SAH were elevated approximately 2-fold in liver and brain of CBS(+/-) mice, and correlations were observed between plasma total homocysteine and SAH in liver (r=0.54; P<0.001) and brain (r=0.67; P<0.001). These results indicate that endothelial dysfunction occurs in hyperhomocysteinemic mice even in the absence of folate deficiency. Endothelial dysfunction in CBS(+/-) mice was associated with increased tissue levels of SAH, which suggests that altered SAM-dependent methylation may contribute to vascular dysfunction in hyperhomocysteinemia.