Intracellular third loops in AT1 and AT2 receptors determine subtype specificity.

The recently cloned angiotensin II type 2 (AT2) receptor is a member of the seven transmembrane G-protein coupled receptor superfamily with a relatively low sequence homology with the angiotensin II type 1 (AT1) receptor subtype and counteracts the growth action of AT1 receptor. Intracellular third loops are known to be involved in interactions with various G proteins. We hypothesized that the intracellular third loop plays critical roles in determining the specificity of opposite functions of AT1 and AT2 receptor subtypes and examined this possibility using chimeric AT1 receptor, of which intracellular third loop is replaced with that of AT2 receptor. We transfected this chimeric receptor into PC 12 cells and observed that stimulation of this receptor inhibited extracellular signal-regulated kinase (ERK) activation and induces apoptosis, whereas the binding characteristics of this receptor remained those of ATI receptor. Taken together, these results support the notion that intracellular third loop is the critical determinant for mutually antagonistic AT1 and AT2 receptors' signaling pathways.

Alterations of load-induced p38 MAP kinase activation in failing rat hearts.

Hemodynamic load-induced cardiac p38 mitogen-activated protein kinase (MAPK) activation was studied in normotensive control Dahl rats (n = 10) and hypertensive Dahl rats with heart failure (n = 16). The isolated heart from each animal was stretched on a Langendorff apparatus at an equivalent diastolic wall stress, and the p38-MAPK activity of the left ventricular (LV) myocardium was analyzed by immunoprecipitation-kinase assay. Compared to the control hearts, the stretch-induced p38-MAPK activities were significantly decreased, and inversely correlated with the LV diameter (r = -0.73, P < 0.01). Chronic treatment with an angiotensin II AT1-receptor antagonist, valsartan (10 mg/kg/day), ameliorated cardiac function and remodeling process in the failing hearts, which was associated with an improvement of the p38-MAPK activities. Thus, the mechano-signal transduction of p38-MAPK pathway is downregulated in the failing hearts, along with progressive ventricular remodeling. The data also suggest that the beneficial effects of the AT1-receptor antagonists are potentially mediated by the restoration of cardiac growth-related signal transduction.

Stage-specific differential activation of mitogen-activated protein kinases in hypertrophied and failing rat hearts.

Mitogen-activated protein kinases (MAPKs) are involved in the early development of cardiac hypertrophy, but their roles in chronic left ventricular hypertrophy (LVH) are unclear. We studied the angiotensin (Ang) II-induced cardiac MAPK activation of the hypertensive Dahl salt-sensitive (DS) rats in the subacute developing LVH stage, the chronic compensated LVH stage, and the congestive heart failure (CHF) stage. In the isolated, coronary-perfused heart preparation, Ang II infusion (1x10(-6)mol/l) activated extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38-MAPK in the LV myocardium. No substantial differences were observed in the Ang II-induced ERK activation between the normotensive control DS rats and the hypertensive DS rats in either stage. In contrast, the Ang II-induced activation of JNK and p38-MAPK was augmented in the subacute LVH stage of the hypertensive DS rats, but then progressively attenuated in the chronic LVH and CHF stages. Chronic treatment with an angiotensin converting enzyme inhibitor, temocapril (20 mg/kg/day), ameliorated the responsiveness of the JNK/p38-MAPK activation, suggesting that the decreased JNK/p38-MAPK activation is a consequence of negative feedback regulation for the activated cardiac renin-angiotensin system in chronic LVH and CHF. Thus, the Ang II-induced activation of multiple cardiac MAPK pathways are differentially regulated, depending on the stages of chronic hypertrophic process. The JNK and p38-MAPK activation may be involved in the early development of adaptive LVH. However, the responsiveness of the cardiac JNK/p38-MAPK pathways progressively decreased in chronic LVH and CHF under the chronic activation of tissue renin-angiotensin system.

Calcium handling and sarcoplasmic-reticular protein functions during heart-failure transition in ventricular myocardium from rats with hypertension.

The objective of this study was to determine the primary event that occurs in Ca2+-regulatory sarcoplasmic-reticular (SR) proteins during subacute transition from concentric/mechanically-compensated left ventricular (LV) hypertrophy to eccentric/decompensated hypertrophy. Using Dahl salt-sensitive rats with hypertension, changes of myocardial contraction, intracellular Ca2+ transients, SR Ca2+ uptake, protein levels of SR Ca2+ ATPase (SERCA2), phospholamban, and calsequestrin (CSQ), and mRNA levels of SERCA2 and CSQ were serially determined and compared between the established stage of LV hypertrophy (LVH) and the subsequent stage of overt LV dysfunction (CHF). In LVH, isolated LV papillary muscle preparations showed an equal peak-tension level and a mild prolongation of the isometric tension decay compared to those of age-matched controls. The Ca2+ transients as measured by aequorin were unchanged. The Ca2+ uptake of isolated SR vesicles and the protein/mRNA levels of SR proteins were also equivalent to those of the controls. In contrast, in CHF, the failing myocardium showed a further prolongation of the contraction time course and a 39% reduction of the peak-tension development. The Ca2+ transients showed changes consisting of a decrease in the peak level and a prolongation of the time course. In addition, the SR Ca2+ uptake was decreased by 41%. Despite these functional changes, the protein and mRNA levels of the SR components remained equivalent to those of the age-matched controls. Thus, in this hypertensive animal, 1) at the LVH stage, myocardial contractility and intracellular capability to regulate Ca2+ remained normal; 2) at the CHF stage, impaired SR Ca2+ handling and the subsequent reduction of myocardial contraction were in progress; and 3) impairments of SR function occurred at the post-translational protein level rather than at the transcriptional/translational levels. Our findings support the role of SR proteins as the primary determinant of the contractile dysfunction that occurs during the heart-failure transition; however, post-translational modulators of these SR elements may also be critical.

Cardiac calcineurin during transition from hypertrophy to heart failure in rats.

We studied an alteration of calcineurin expression in the heart and its modification by cyclosporin A and an ACE inhibitor, temocapril, using Dahl salt-sensitive (DS) rats with hypertensive left ventricular hypertrophy (LVH) and congestive heart failure (CHF). Calcineurin protein expression in the LV myocardium was increased in the LVH stage, but then decreased during CHF transition. Chronic cyclosporin A treatment (10 mg/kg/day), which inhibits calcineurin activity, could not block the increases of LV weight and dimensions and did not improve the LV systolic function during the CHF transition. In contrast, chronic temocapril treatment (20 mg/kg/day) restored the downregulation of calcineurin expression, but progression of the hypertrophic process was inhibited. Therefore, cardiac calcineurin is increased in the hypertensive LVH and may be involved in the development of the adaptive hypertrophic process. However, calcineurin expression is downregulated during CHF transition and may no longer play a major role in the pathogenesis of myocardial hypertrophy in the failing hearts.

Interferon-gamma induces AT(2) receptor expression in fibroblasts by Jak/STAT pathway and interferon regulatory factor-1.

The expression of angiotensin II type 2 (AT(2)) receptor is closely associated with cell growth, differentiation, and/or injury. We examined the effect of interferon (IFN)-gamma on AT(2) receptor expression in mouse fibroblast R3T3 cells and demonstrated that IFN-gamma treatment increased the expression of AT(2) receptor mRNA as well as its binding. Interferon regulatory factor (IRF)-1 was induced in mouse fibroblast R3T3 cells after IFN-gamma stimulation, and electrophoretic mobility shift assay showed an increase in IRF-1 binding with the IRF-specific binding sequence in the AT(2) receptor gene promoter region after IFN-gamma stimulation. The IRF-1 gene promoter contains an IFN-gamma-activated sequence (GAS) motif for possible binding of signal transducer(s) and activator(s) of transcription (STAT). Indeed, in R3T3 cells, IFN-gamma treatment resulted in rapid activation of Janus kinase (Jak) 1, Jak2, and STAT1 via tyrosine phosphorylation. Electrophoretic mobility shift assay with the GAS probe revealed increased STAT1 binding to the IRF-1 gene promoter in response to IFN-gamma stimulation. Transfection of GAS-binding oligonucleotides inhibited the effect of IFN-gamma on IRF-1 production, resulting in the AT(2) receptor trans-activation. Taken together, our data show that IFN-gamma upregulates AT(2) receptor expression in R3T3 cells via the activation of the intracellular Jak/STAT pathway and production of IRF-1.

Anti-ischemic effect of a novel cardioprotective agent, JTV519, is mediated through specific activation of delta-isoform of protein kinase C in rat ventricular myocardium.

BACKGROUND: A new 1,4-benzothiazepine derivative, JTV519, has a strong protective effect against Ca(2+) overload-induced myocardial injury. We investigated the effect of JTV519 on ischemia/reperfusion injury in isolated rat hearts. METHODS AND RESULTS: At 30 minutes of reperfusion after 30-minute global ischemia, the percent recovery of left ventricular developed pressure was improved, and the creatine phosphokinase and lactate dehydrogenase leakage was reduced in a concentration-dependent manner when JTV519 was administered in the coronary perfusate both at 5 minutes before the induction of ischemia and at the time of reperfusion. The myocardial protective effect of JTV519 was completely blocked by pretreatment of the heart with GF109203X, a specific protein kinase C (PKC) inhibitor. In contrast, the effect of JTV519 was not affected by alpha(1)-, A(1)-, and B(2)-receptor blockers that couple with PKC in the cardiomyocyte. Both immunofluorescence images and immunoblots of JTV519-treated left ventricular myocardium and isolated ventricular myocytes demonstrated that this agent induced concentration-dependent translocation of the delta-isoform but not the other isoforms of PKC to the plasma membrane. CONCLUSIONS: The mechanism of cardioprotection by JTV519 against ischemia/reperfusion injury involves isozyme-specific PKC activation through a receptor-independent mechanism. This agent may provide a novel pharmacological approach for the treatment of patients with acute coronary diseases via a subcellular mechanism mimicking ischemic preconditioning.

Stimulation of different subtypes of angiotensin II receptors, AT1 and AT2 receptors, regulates STAT activation by negative crosstalk.

Angiotensin II type 2 (AT2) receptor exerts an inhibitory action on cell growth. In the present study, we report that the stimulation of AT2 receptor in AT2 receptor cDNA-transfected rat adult vascular smooth muscle cells (VSMCs) inhibited angiotensin II type 1 (AT1) receptor-mediated tyrosine phosphorylation of STAT (signal transducers and activators of transcription) 1alpha/beta, STAT2, and STAT3 without influence on Janus kinase. AT2 receptor activation also inhibited the tyrosine phosphorylation of STAT1alpha/beta induced by interferon-gamma, epidermal growth factor, and platelet-derived growth factor. Similar effects of AT2 receptor were observed in R3T3 fibroblast and mouse fetal VSMCs, which express endogenous AT2 receptor. Moreover, AT2 receptor inhibited serine phosphorylation of STAT1alpha and STAT3 via the inhibition of extracellular signal-regulated kinase (ERK) activation. Stimulation of AT2 receptor inhibited the binding of STATs with sis-inducing element in c-fos promoter, resulting in decreased c-fos expression. Taken together, our results suggest that AT2 receptor can crosstalk negatively with multiple families of growth receptors by inhibiting ERK and STAT activation.

Load-sensitive diastolic relaxation in hypertrophied left ventricles.

We studied effects of enalaprilat and L-158,809, an angiotensin II type-1 receptor antagonist, on left ventricular (LV) diastolic relaxation in 11 normal control dogs and 16 LV hypertrophied (LVH) dogs with perinephritic hypertension. At baseline, LV systolic and end-diastolic pressures and end-systolic elastance were increased in the LVH group (all P < 0.01 vs. the control group). LV relaxation time constant was also prolonged (P < 0.01), suggesting impaired LV diastolic relaxation in this model of LVH. Before and after the administration of enalaprilat (0.25 mg/kg) and L-158,809 (0.30 mg/kg), LV relaxation was assessed over a wide range of LV loading conditions during vena caval occlusion. LV relaxation time constant was insensitive to load reduction in the control group, which was not affected by enalaprilat or L-158,809. In contrast, LV unloading caused a significant prolongation of the relaxation time constant in the LVH group. This load-sensitive LV relaxation abnormality was significantly improved by enalaprilat or L-158,809. These results support the concept that angiotensin II is involved in the pathogenesis of diastolic dysfunction in pressure-overloaded LVH and also suggest that angiotensin-converting enzyme inhibitors and angiotensin II type-1 receptor antagonists are potentially beneficial in the treatment of the hypertrophied heart.

Angiotensin type 2 receptor dephosphorylates Bcl-2 by activating mitogen-activated protein kinase phosphatase-1 and induces apoptosis.

We examined the cellular and signaling mechanism of angiotensin II (Ang II) type 2 (AT2) receptor-induced apoptosis in PC12W (rat pheochromocytoma cell line) cells that express abundant AT2 receptor but not Ang II type 1 receptor. In these cells, nerve growth factor (NGF) inhibited the internucleosomal DNA fragmentation induced by serum depletion, whereas Ang II antagonized this NGF cell survival action and induced apoptosis. We studied the mechanism of NGF and AT2 receptor interaction on apoptosis by examining their effects on the survival factor Bcl-2. AT2 receptor activation did affect intracellular Bcl-2 protein levels. Bcl-2 phosphorylation was stimulated by NGF, whereas AT2 receptor activation blocked this NGF effect. Pretreatment with antisense oligonucleotide of mitogen-activated protein (MAP) kinase phosphatase-1 enhanced the effects of NGF on MAP kinase activation and Bcl-2 phosphorylation but attenuated the inhibitory effects of AT2 receptor on MAP kinase, Bcl-2 phosphorylation, and apoptosis. Taken together, these results suggest that MAP kinase plays a critical role in inhibiting apoptosis by phosphorylating Bcl-2. The AT2 receptor inhibits MAP kinase activation, resulting in the inactivation of Bcl-2 and the induction of apoptosis.

Interferon regulatory factor-1 up-regulates angiotensin II type 2 receptor and induces apoptosis.

The expression of the angiotensin II type 2 (AT2) receptor is developmentally and growth regulated. In cultured R3T3 cells, expression of this receptor is markedly induced at the confluent state and with serum deprivation. In this study we demonstrated that the removal of serum from culture media resulted in the induction of apoptosis in these cells and the addition of angiotensin II further enhanced apoptosis. We have previously identified an interferon regulatory factor (IRF) binding motif in the mouse AT2 receptor gene promoter region. In this report, we observed that serum removal increased IRF-1 expression, with a rapid and transient decrease of IRF-2. To prove that the changes in IRFs after serum removal mediated apoptosis and up-regulated AT2 receptor, we transfected antisense oligonucleotides for IRF-1 or IRF-2 into R3T3 cells and observed that IRF-1 antisense oligonucleotide attenuated apoptosis and abolished the up-regulation of AT2 receptor. IRF-2 antisense oligonucleotide pretreatment did not affect the onset of apoptosis after serum removal; instead, it increased AT2 receptor binding and enhanced angiotensin II-mediated apoptosis. Taken together, these results suggest that increased IRF-1 after serum starvation contributes to the induction of apoptosis and that increased IRF-1 up-regulates the AT2 receptor expression after serum starvation, resulting in enhanced angiotensin II-mediated apoptosis.

Diastolic properties in canine hypertensive left ventricular hypertrophy: effects of angiotensin converting enzyme inhibition and angiotensin II type-1 receptor blockade.

OBJECTIVE: Angiotensin II has been suggested to be involved in the pathogenesis of diastolic dysfunction in left ventricular hypertrophy (LVH). The purpose of this study was to asses the effects of enalaprilat and L-158,809, an angiotensin II type-1 receptor antagonist, on LV diastolic function in 16 normal control dogs and 20 LVH dogs with perinephritic hypertension. METHODS: LV hemodynamics was studied before and after intravenous injection of enalaprilat (0.25 mg/kg) or L-158,809 (0.3 mg/kg). The hemodynamic data were analyzed in relation to the changes in myocardial blood flow (measured by radioactive microspheres) and in the circulating angiotensin II and norepinephrine levels. RESULTS AND CONCLUSIONS: At baseline, significant increases were observed for LV/body weight ratio as well as LV systolic and end-diastolic pressure in the LVH dogs (all P < 0.01 vs. the control group). In addition, LV relaxation time constant was prolonged and the chamber and myocardial stiffness constants were increased (P < 0.01) in the LVH dogs, suggesting an impairment of LV diastolic function. Administration of enalaprilat or L-158,809 improved LV stiffness constants in the LVH dogs (P < 0.05). The diastolic LV pressure-diameter relation shifted downwards in the LVH dogs whereas diastolic distensibility was not altered in the control dogs. Although the circulating angiotensin II levels were significantly decreased by enalaprilat in the LVH dogs, they did not correlate with the changes in the stiffness constants. Furthermore, the alterations of LV diastolic properties in the LVH group could not be attributed to myocardial perfusion, which was rather decreased by administration of enalaprilat and L-158,809. These results suggest that angiotensin II, particularly at the local level, is involved in the pathogenesis of diastolic dysfunction in pressure-overload LVH. The data also support the concept that ACE inhibitors and angiotensin II receptor blockers are potentially beneficial in the treatment of the hypertrophied heart.

Hemodynamic and cardiac effects of the selective T-type and L-type calcium channel blocking agent mibefradil in patients with varying degrees of left ventricular systolic dysfunction.

OBJECTIVES: This study sought to assess the hemodynamic and cardiac effects of two dose levels of mibefradil in patients with varying degrees of ischemic left ventricular dysfunction. BACKGROUND: Mibefradil is a new, selective T-type and L-type calcium channel blocking agent. Because L-type channel blockade may depress myocardial performance, an invasive hemodynamic study was performed to assess the safety of this agent. METHODS: We performed an open label study, examining the effects of two intravenous doses of mibefradil, selected to produce plasma levels comparable to those measured after oral administration of 50 mg (dose 1: 400 ng/ml) or 100 mg (dose 2: 800 ng/ml) of the drug. Variables studied included the indexes of left ventricular function and neurohormone levels. Patients were stratified according to ejection fraction (EF) (> or = 40%, n = 26; < 40%, n = 24) and the presence (n = 15) or absence (n = 35) of heart failure. RESULTS: In patients with preserved systolic function, dose 1 had no clinically significant hemodynamic effects, but dose 2 decreased mean aortic pressure and systemic vascular resistance (-8.5 mm Hg, -12%, both p < 0.01) and also reduced end-systolic stress and volume, thus improving EF (52% to 58%, p < 0.01). Heart rate tended to decrease. In patients with depressed EF, heart rate decreased significantly with both doses. The effects of dose 1 mimicked those observed after dose 2 in patients with preserved EF. Dose 2 (plasma levels 1,052 +/- 284 ng/ml) still decreased left ventricular systolic wall stress and improved EF (24.0% to 28.5%, p < 0.05) but also significantly depressed the maximal first derivative of left ventricular pressure. Examination of individual pressure-volume loops in two patients with heart failure showed a clear rightward shift of the loop despite a decrease in systolic pressure, suggesting negative inotropy. Neurohormone levels were unchanged at both dose levels and in all subgroups. CONCLUSIONS: Intravenous mibefradil was well tolerated and produced an overall favorable cardiovascular response. However, high plasma concentrations might produce myocardial depression in patients with heart failure, and caution should be exerted in this setting.

Intracellular third loop domain of angiotensin II type-2 receptor. Role in mediating signal transduction and cellular function.

The present study tests the hypothesis that the unique intracellular third loop domain of angiotensin II type-2 (AT2) receptor is essential for the subsequent intracellular signaling and plays an important role in mediating receptor function. Synthetic intracellular third loop peptide of the AT2 receptor (AT2-3LP, 22 amino acids) and control peptide consisting of the same amino acid composition in random sequence were delivered into adult rat aortic vascular smooth muscle cells by cationic liposome-mediated transfection. Successful intracellular peptide delivery was confirmed by microscopic localization of the fluorescein-labeled AT2-3LP within the cells and also by co-immunoprecipitation of the 125I-labeled 3LP complexed with Gi protein using anti-Gialpha antibody. The AT2-3LP-transfected cells showed reduction of serum-stimulated DNA synthesis and cell proliferation as well as a decrease in mitogen-activated protein kinase activity, simulating the effects of AT2 receptor stimulation. The antagonistic effect of the AT2-3LP on mitogen-activated protein kinase activity and DNA synthesis were reversed by pertussis toxin and sodium orthovanadate. Thus, our data suggest that the intracellular third loop domain of the AT2 receptor is closely linked with the cellular signaling pathways of vascular smooth muscle cells in which Gi and protein-phosphotyrosine phosphatase are involved, resulting in the alteration of mitogen-activated protein kinase activity and in growth inhibition.

Change in aortic end-systolic pressure by alterations in loading sequence and its relation to left ventricular isovolumic relaxation.

BACKGROUND: A brief, sustained constriction of the descending and the ascending aortas produces systolic loads at different times during ejection, and descending intervention prolongs left ventricular (LV) relaxation more than ascending intervention. Although alterations in the sequence of loading the ventricle have been suggested as a cause of such load-induced relaxation abnormalities, the relation of the loading system to relaxation has been unclear. METHODS AND RESULTS: LV peak systolic pressure was elevated by approximately 40 mm Hg by constricting the descending and ascending aortas in seven anesthetized dogs. The descending intervention increased aortic end-systolic pressure (AoESP, 110.4 +/- 9.3 to 150.8 +/- 11.5 mm Hg; P < .05), reduced aortic flow (P < .05), and prolonged LV relaxation (time constant [T], 31.9 +/- 4.4 to 69.8 +/- 12.8 ms; P < .05). LV ejection time was reduced, but the systolic time interval was unchanged. In contrast, ascending intervention decreased AoESP (111.9 +/- 11.4 to 101.5 +/- 10.3 mm Hg; P < .05), reduced aortic flow (P < .05), and prolonged T (31.2 +/- 5.4 to 42.2 +/- 8.3 ms; P < .05), whereas ejection time and systolic time interval increased (both P < .01). Prolongation of T was significantly greater during descending intervention (P < .05) and was associated with an increase in AoESP during descending intervention but a decrease in AoESP during ascending intervention. CONCLUSIONS: Descending intervention induced greater prolongation of T than ascending intervention. Prolongation of T was closely related to an increase in AoESP in the descending intervention but a decrease in AoESP in the ascending intervention. These data suggest that not only the loading sequence but also the pressure level at the onset of isovolumic relaxation determines LV relaxation.

Early diastolic regional function of the hypertrophied left ventricle.

We analyzed cardiac catheterization data from 7 patients with aortic stenosis and 10 patients with nonobstructive hypertrophic cardiomyopathy to compare left ventricular regional diastolic function. Left ventriculogram in the right anterior oblique projection was analyzed by the area method, and regional wall stress and regional area were computed for 4 regions in the mid-portion of the left ventricle. For each region, we assessed the percent area changes (normalized by end-diastolic regional area) and time constant for regional wall stress decrease during the isovolumic relaxation period. Regional non-uniformity during the isovolumic relaxation period was then evaluated by standard deviations for the percent area changes and for regional time constants of the 4 ventricular regions. In patients with hypertrophic cardiomyopathy, both the standard deviations for the percent area changes and the regional time constants were greater (P < 0.05) than those in patients with aortic stenosis, suggesting the presence of pronounced non-uniformity of regional relaxation in hypertrophic cardiomyopathy. The time constant of left ventricular pressure decrease during early relaxation phase was significantly greater (P < 0.01), and the early diastolic peak filling rate of the global left ventricle was significantly smaller (P < 0.05) in patients with hypertrophic cardiomyopathy. Thus, early diastolic left ventricular regional non-uniformity was more pronounced in hypertrophic cardiomyopathy than in aortic stenosis, which was associated with the impairment of relaxation and early filling of the global left ventricle. These findings suggest that different mechanisms are responsible for diastolic dysfunction in primary versus secondary myocardial hypertrophy.

The angiotensin II type 2 (AT2) receptor antagonizes the growth effects of the AT1 receptor: gain-of-function study using gene transfer.

The type 1 angiotensin II (AT1) receptor is well characterized but the type 2 (AT2) receptor remains an enigma. We tested the hypothesis that the AT2 receptor can modulate the growth of vascular smooth muscle cells by transfecting an AT2 receptor expression vector into the balloon-injured rat carotid artery and observed that overexpression of the AT2 receptor attenuated neointimal formation. In cultured smooth muscle cells, AT2 receptor transfection reduced proliferation and inhibited mitogen-activated protein kinase activity. Furthermore, we demonstrated that the AT2 receptor mediated the developmentally regulated decrease in aortic DNA synthesis at the latter stages of gestation. These results suggest that the AT2 receptor exerts an antiproliferative effect, counteracting the growth action of AT1 receptor.

Role of endogenous endothelin-1 in experimental renal hypertension in dogs.

BACKGROUND: Endothelin-1, a vasoconstrictive peptide released by endothelium, may be involved in the pathophysiology of hypertension. The goal of the present study was to evaluate the role of endogenous endothelin-1 in renal hypertension in dogs. The model of hypertension consisted of silk tissue wrapping of the left kidney, which produced hypertension associated with perinephritis after 6 to 8 weeks. METHODS AND RESULTS: Thirty-two anesthetized open chest dogs were studied randomly: 8 dogs with perinephritic hypertension received the nonpeptidic ETA-ETB receptor antagonist bosentan (group 1); 8 other hypertensive dogs received the vehicle solution (group 2); 8 healthy dogs received bosentan (group 3); and 8 healthy dogs received the vehicle solution (group 4). Bosentan was injected as an intravenous bolus (3 mg/kg) followed by a 1-hour infusion at a rate of 7 mg.kg-1.h-1. In hypertensive dogs, bosentan produced a similar decrease (P = .0001) of both left ventricular systolic and mean aortic pressures, which averaged 38 mm Hg (-22% and -24%, respectively). These parameters remained unchanged with the vehicle solution. Left ventricular end-diastolic and left atrial pressures also declined significantly with bosentan (P = .0005 and P < .05, respectively). Left ventricular lengths tended to decrease. The other cardiovascular parameters (heart rate, peak [+]dP/dt, time constant of relaxation, and coronary vascular resistance) did not change significantly. In healthy dogs, bosentan decreased mean aortic pressure by 19 mm Hg (P = .004). Vehicle solution had no effect. Plasma endothelin-1 levels, similar under basal conditions in healthy and hypertensive dogs, increased 30-fold with bosentan (P = .0001). CONCLUSIONS: Specific endothelin-1 receptor antagonism markedly lowers blood pressure in experimental hypertension but is less effective on blood pressure of healthy animals. This suggests that endothelin-1 plays a role in the pathophysiology of hypertension but contributes to a lesser extent to the maintenance of normal blood pressure. This role of endothelin-1 is unrelated to its plasma levels. The increase of plasma endothelin-1 with bosentan, due either to a displacement of endothelin-1 from its receptor or to a feedback mechanism, does not prevent this blood pressure reduction.

Intracardiac angiotensin-converting enzyme inhibition improves diastolic function in patients with left ventricular hypertrophy due to aortic stenosis.

BACKGROUND: Cardiac hypertrophy is associated with elevated intracardiac angiotensin-converting enzyme activity, which may contribute to diastolic dysfunction. METHODS AND RESULTS: We infused enalaprilat (0.05 mg/min) for 15 minutes into the left coronary arteries of 20 adult patients with left ventricular (LV) hypertrophy due to aortic stenosis (mean aortic valve area, 0.7 +/- 0.2 cm2) and 10 patients with dilated cardiomyopathy (mean ejection fraction, 35 +/- 4%) and assessed (1) simultaneous changes in LV micromanometer pressure and dimensions, (2) LV regional wall motion analyzed by the area method, and (3) Doppler flow-velocity profiles. Systemic neurohormonal activation did not occur with the selective left coronary artery infusion; there were no changes in plasma renin activity, angiotensin-converting enzyme activity, or atrial natriuretic peptide. In patients with aortic stenosis, LV end-diastolic pressure declined from 25 +/- 2 to 20 +/- 2 mm Hg (P < .05). LV pressure-volume and LV pressure-dimension relations showed downward shifts by ventriculography and echocardiography, respectively, indicating improved diastolic distensibility. Regional area change during isovolumic relaxation increased in the anterior segments perfused with enalaprilat but decreased in the inferior segments, indicating acceleration of isovolumic relaxation in the anterior segments and reciprocal shortening in the inferior segments. Regional peak filling rate increased in the anterior segments but not in the inferior segments, and the regional area stiffness constant decreased in the anterior segments but not in the inferior segments. There were no changes in heart rate, cardiac output, or right atrial pressure, excluding alterations in right ventricular/pericardial constraint. In contrast, in the patients with dilated cardiomyopathy the decrease in LV end-diastolic pressure from 22 +/- 2 to 18 +/- 2 mm Hg (P < .05) was accompanied by a significant fall in right atrial pressure (9 +/- 1 to 6 +/- 1 mm Hg), implicating alterations in pericardial constraint. The patients with dilated cardiomyopathy showed no improvement in regional diastolic relaxation, filling, or distensibility. CONCLUSIONS: Intracoronary enalaprilat at a dosage that did not cause systemic neurohormonal activation improved LV diastolic chamber distensibility and regional relaxation and filling in patients with LV hypertrophy due to aortic stenosis. In contrast, these effects of intracoronary enalaprilat on diastolic function were not observed in patients with dilated cardiomyopathy who did not have concentric hypertrophy. These observations support the hypothesis that the cardiac renin-angiotensin system is activated in patients with concentric pressure-overload hypertrophy and that this activation may contribute to impaired diastolic function.