Angiotensin-(1-7) is a modulator of the human renin-angiotensin system.

The renin-angiotensin system is important for cardiovascular homeostasis. Currently, therapies for different cardiovascular diseases are based on inhibition of angiotensin-converting enzyme (ACE) or angiotensin II receptor blockade. Inhibition of ACE blocks metabolism of angiotensin-(1-7) to angiotensin-(1-5) and can lead to elevation of angiotensin-(1-7) levels in plasma and tissue. In animal models, angiotensin-(1-7) itself causes or enhances vasodilation and inhibits vascular contractions to angiotensin II. The function of angiotensin-(1-5) is unknown. We investigated whether angiotensin-(1-7) and angiotensin-(1-5) inhibit ACE or antagonize angiotensin-induced vasoconstrictions in humans. ACE activity in plasma and atrial tissue was inhibited by angiotensin-(1-7) up to 100%, with an IC(50) of 3.0 and 4.0 micromol/L, respectively. In human internal mammary arteries, contractions induced by angiotensin I and II and the non-ACE-specific substrate [Pro(11),D-Ala(12)]-angiotensin I were antagonized by angiotensin-(1-7) (10(-5) mol/L) in a noncompetitive way, with a 60% inhibition of the maximal response to angiotensin II. Contractions to ACE-specific substrate [Pro(10)]-angiotensin I were also inhibited, an effect only partly accounted for by antagonism of angiotensin II. Angiotensin-(1-5) inhibited plasma ACE activity with a potency equal to that of angiotensin I but had no effect on arterial contractions. In conclusion, angiotensin-(1-7) blocks angiotensin II-induced vasoconstriction and inhibits ACE in human cardiovascular tissues. Angiotensin-(1-5) only inhibits ACE. These results show that angiotensin-(1-7) may be an important modulator of the human renin-angiotensin system.     

Evidence against a major role for angiotensin converting enzyme-related carboxypeptidase (ACE2) in angiotensin peptide metabolism in the human coronary circulation

OBJECTIVE: To investigate the role of angiotensin-converting enzyme-related carboxypeptidase (ACE2) in angiotensin peptide metabolism in the human coronary circulation. METHODS: Angiotensin I and angiotensin II, and their respective carboxypeptidase metabolites, angiotensin-(1-9) and angiotensin-(1-7), were measured in arterial and coronary sinus blood of heart failure subjects receiving angiotensin-converting enzyme (ACE) inhibitor therapy and in normal subjects not receiving ACE inhibitor therapy. In addition, angiotensin I, angiotensin II and angiotensin-(1-7) were measured in arterial and coronary sinus blood of subjects with coronary artery disease before, and at 2, 5 and 10 min after, intravenous administration of ACE inhibitor. RESULTS: In comparison with normal subjects, heart failure subjects receiving ACE inhibitor therapy had a greater than 40-fold increase in angiotensin I levels, but angiotensin-(1-9) levels were low (1-2 fmol/ml), and similar to those of normal subjects. Moreover, angiotensin-(1-7) levels increased in parallel with angiotensin I levels and the angiotensin-(1-7)/angiotensin II ratio increased by 7.5-fold in coronary sinus blood. Intravenous administration of ACE inhibitor to subjects with coronary artery disease rapidly decreased angiotensin II levels by 54-58% and increased angiotensin I levels by 2.4- to 2.8-fold, but did not alter angiotensin-(1-7) levels or net angiotensin-(1-7) production across the myocardial vascular bed. CONCLUSIONS: The failure of angiotensin-(1-9) levels to increase in response to increased angiotensin I levels indicated little role for ACE2 in angiotensin I metabolism. Additionally, the levels of angiotensin-(1-7) were more linked to those of angiotensin I than angiotensin II, consistent with its formation by endopeptidase-mediated metabolism of angiotensin I, rather than by ACE2-mediated metabolism of angiotensin II.     

Differential regulation of angiotensin peptide levels in plasma and kidney of the rat.

We compared the effects of the converting enzyme inhibitor perindopril on components of the renin-angiotensin system in plasma and kidney of male Sprague-Dawley rats administered perindopril in their drinking water at two doses (1.4 and 4.2 mg/kg) over 7 days. Eight angiotensin peptides were measured in plasma and kidney: angiotensin-(1-7), angiotensin II, angiotensin-(1-9), angiotensin I, angiotensin-(2-7), angiotensin III, angiotensin-(2-9), and angiotensin-(2-10). In addition, angiotensin converting enzyme activity, renin, and angiotensinogen were measured in plasma, and renin, angiotensinogen, and their respective messenger RNAs were measured in kidney; angiotensinogen messenger RNA was also measured in liver. In plasma, the highest dose of perindopril reduced angiotensin converting enzyme activity to 11% of control, increased renin 200-fold, reduced angiotensinogen to 11% of control, increased angiotensin-(1-7), angiotensin I, angiotensin-(2-7), and angiotensin-(2-10) levels 25-, 9-, 10-, and 13-fold, respectively; angiotensin II levels were not significantly different from control. By contrast, for the kidney, angiotensin-(1-7), angiotensin I, angiotensin-(2-7), and angiotensin-(2-10) levels did not increase; angiotensin II levels fell to 14% of control, and angiotensinogen fell to 12% of control. Kidney renin messenger RNA levels increased 12-fold, but renal renin content and angiotensinogen messenger RNA levels in kidney and liver were not influenced by perindopril treatment. These results demonstrate a differential regulation of angiotensin peptides in plasma and kidney and provide direct support for the proposal that the cardiovascular effects of converting enzyme inhibitors depend on modulation of tissue angiotensin systems. Moreover, the failure of kidney angiotensin I levels to increase with perindopril treatment, taken together with the fall in kidney angiotensinogen levels, suggests that angiotensinogen may be a major rate-limiting determinant of angiotensin peptide levels in the kidney.     

Attenuated responses to angiotensin II in follitropin receptor knockout mice,a model of menopause-associated hypertension

Activation of the renin-angiotensin system has been implicated in the development of hypertension in menopausal women. We investigated whether blood pressure is elevated and whether angiotensin II (Ang II)-induced vascular reactivity is increased in follitropin receptor knockout (FORKO) female mice. These mice are estrogen-deficient and have characteristics similar to postmenopausal women. Serum estradiol levels were significantly reduced in FORKO versus wild-type mice (1.4+/-0.2 versus 15+/-3 pg/mL, P<0.01). Blood pressure, measured by telemetry, was significantly increased in FORKO (120+/-2/92+/-2 mm Hg) compared with wild-type counterparts (110+/-1/85+/-2 mm Hg, P<0.05). Vascular dose responses to acetylcholine (endothelium-dependent dilation) and sodium nitroprusside (endothelium-independent dilation) were not different. Ang II-induced vasoconstriction was blunted in FORKO compared with wild-type mice (P<0.05). Media-to-lumen ratio was significantly increased in FORKO (6.2+/-0.5%) versus control mice (5.2+/-0.3%), indicating vascular remodeling. Aortic*O2- levels, NADH-inducible.O2- generation, and plasma levels of thiobarbituric acid reactive substances (TBARS), indexes of oxidative stress, were not significantly different between wild-type and FORKO mice. Vascular AT1 receptor content, assessed by immunoblotting, was reduced by 40% in FORKO compared with wild-type mice (P<0.01). This was associated with decreased circulating Ang II levels in FORKO versus control mice. These data indicate that FORKO mice have increased blood pressure, vascular remodeling, and attenuated vascular responses to Ang II. Our findings suggest that vascular Ang II signaling is downregulated in female FORKO mice and that Ang II may not play an important role in blood pressure elevation in this model of menopause-associated hypertension.     

Differential regulation of elevated renal angiotensin II in chronic renal ischemia

The present study was undertaken to clarify the role of intrarenal angiotensin (Ang) II and its generating pathways in clipped and nonclipped kidneys of 4-week unilateral renal artery stenosis in anesthetized dogs. After 4 weeks, renal plasma flow (RPF) decreased in clipped and nonclipped kidneys (baseline, 59+/-3; clipped, 16+/-1; nonclipped, 44+/-2 mL/min; P<0.01, n=22). Renal Ang I levels increased only in clipped, whereas intrarenal Ang II contents were elevated in both clipped (from 0.7+/-0.1 to 2.0+/-0.2 pg/mg tissue) and nonclipped kidneys (from 0.6+/-0.1 to 2.5+/-0.3 pg/mg tissue). Intrarenal ACE activity was increased in nonclipped kidneys but was unaltered in clipped kidneys. An angiotensin receptor antagonist (olmesartan medoxomil) given into the renal artery markedly restored RPF, and dilated both afferent and efferent arterioles (using intravital videomicroscopy). Furthermore, in clipped kidneys, the elevated Ang II was suppressed by a chymase inhibitor, chymostatin (from 2.1+/-0.6 to 0.8+/-0.1 pg/mg tissue; P<0.05), but not by cilazaprilat. In nonclipped kidneys, in contrast, cilazaprilat, but not chymostatin, potently inhibited the intrarenal Ang II generation (from 2.4+/-0.3 to 1.5+/-0.2 pg/mg tissue; P<0.05). Finally, [Pro11-D-Ala12]Ang I (an inactive precursor that yields Ang II by chymase but not by ACE; 1 to 50 nmol/kg) markedly elevated intrarenal Ang II in clipped, but not in nonclipped, kidneys. In conclusion, renal Ang II contents were elevated in both clipped and nonclipped kidneys, which contributed to the altered renal hemodynamics and microvascular tone. Furthermore, the mechanisms for intrarenal Ang II generation differ, and chymase activity is enhanced in clipped kidneys, whereas ACE-mediated Ang II generation is possibly responsible for elevated Ang II contents in nonclipped kidneys.     

Angiotensin-(1-7). A member of circulating angiotensin peptides.

We measured the concentrations of three principal products of the renin-angiotensin system and seven of their metabolites in the plasma of anesthetized normal dogs and in dogs 24 hours after bilateral nephrectomy. The levels of the angiotensin peptides were measured by high-performance liquid chromatography combined with radioimmunoassay using three specific antibodies that recognized different epitotes in the sequences of angiotensin I, angiotensin II, and angiotensin-(1-7). The analysis revealed that angiotensin-(1-7) is present in the plasma of intact (4.9 +/- 2.2 fmol/ml) and nephrectomized (0.5 +/- 0.5 fmol/ml) dogs. An intravenous injection of purified hog renin (0.01 Goldblatt unit/kg) increased plasma levels of angiotensin I, angiotensin II, and angiotensin-(1-7) both before and after nephrectomy. These changes were associated with parallel increases in the concentrations of fragments of the three parent peptides. Administration of MK-422 led to the disappearance of circulating angiotensin II and its fragments both before and after a second injection of the same dose of renin. In contrast, MK-422 augmented the plasma levels of both angiotensin I and angiotensin-(1-7). The concentrations of these two peptides, but not the blood pressure, were again augmented by a second injection of renin given after blockade of converting enzyme. These effects were observed both before and after bilateral nephrectomy. These findings show that angiotensin-(1-7) circulates in the blood of normal and nephrectomized dogs. In addition, we found that angiotensin-(1-7) is generated in the blood from the cleavage of angiotensin I through a pathway independent of converting enzyme (EC 3.4.15.1).     

Angiotensin-(1-9) attenuates cardiac fibrosis in the stroke-prone spontaneously hypertensive rat via the angiotensin type 2 receptor

The renin-angiotensin system regulates cardiovascular physiology via angiotensin II engaging the angiotensin type 1 or type 2 receptors. Classic actions are type 1 receptor mediated, whereas the type 2 receptor may counteract type 1 receptor activity. Angiotensin-converting enzyme 2 metabolizes angiotensin II to angiotensin-(1-7) and angiotensin I to angiotensin-(1-9). Angiotensin-(1-7) antagonizes angiotensin II actions via the receptor Mas. Angiotensin-(1-9) was shown recently to block cardiomyocyte hypertrophy via the angiotensin type 2 receptor. Here, we investigated in vivo effects of angiotensin-(1-9) via the angiotensin type 2 receptor. Angiotensin-(1-9) (100 ng/kg per minute) with or without the angiotensin type 2 receptor antagonist PD123 319 (100 ng/kg per minute) or PD123 319 alone was infused via osmotic minipump for 4 weeks into stroke-prone spontaneously hypertensive rats. We measured blood pressure by radiotelemetry and cardiac structure and function by echocardiography. Angiotensin-(1-9) did not affect blood pressure or left ventricular mass index but reduced cardiac fibrosis by 50% (P<0.01) through modulating collagen I expression, reversed by PD123 319 coinfusion. In addition, angiotensin-(1-9) inhibited fibroblast proliferation in vitro in a PD123 319-sensitive manner. Aortic myography revealed that angiotensin-(1-9) significantly increased contraction to phenylephrine compared with controls after N-nitro-l-arginine methyl ester treatment, an effect abolished by PD123 319 coinfusion (area under the curve: angiotensin-(1-9) N-nitro-l-arginine methyl ester=98.9±11.8%; control+N-nitro-l-arginine methyl ester=74.0±10.4%; P<0.01), suggesting that angiotensin-(1-9) improved basal NO bioavailability in an angiotensin type 2 receptor-sensitive manner. In summary, angiotensin-(1-9) reduced cardiac fibrosis and altered aortic contraction via the angiotensin type 2 receptor supporting a direct role for angiotensin-(1-9) in the renin-angiotensin system.     

Possible involvement of angiotensin-converting enzyme 2 and Mas activation in inhibitory effects of angiotensin II Type 1 receptor blockade on vascular remodeling

We explored the roles of angiotensin-converting enzyme 2 (ACE2), angiotensin-(1-7), and Mas activation in angiotensin II type 1 receptor blockade-mediated attenuation of vascular remodeling. Vascular injury was induced by polyethylene-cuff placement around the mouse femoral artery. After cuff placement, the mRNA level of both ACE2 and Mas was markedly decreased in wild-type mice, whereas ACE mRNA was not changed. Immunostaining of ACE2 and Mas was observed mainly in the media and was reduced in the injured artery. Administration of angiotensin-(1-7) decreased neointimal formation after cuff placement, whereas administration of [D-Ala(7)] angiotensin-(1-7), a Mas antagonist, increased it. Consistent with these results, we also demonstrated that neointimal formation induced by cuff placement was further increased in ACE2 knockout mice. In angiotensin II type 1a receptor knockout mice, mRNA expression and immunostaining of ACE2 and Mas in the injured artery were greater, with less neointimal formation than in wild-type mice. Increased ACE2 expression in the injured artery was also observed by treatment of wild-type mice with an angiotensin II type 1 receptor blocker, olmesartan. These results suggested that activation of the ACE2-angiotensin-(1-7)-Mas axis is at least partly involved in the beneficial effects of angiotensin II type 1 receptor blockade on vascular remodeling.     

Evidence that prolyl endopeptidase participates in the processing of brain angiotensin.

In order to understand angiotensin metabolism in the canine brain, we determined the molecular forms of angiotensin peptides present in the hypothalamus of the dog and carried out measurements of the metabolism of 125I-angiotensin I in homogenates of that tissue. Angiotensin peptides were extracted from canine hypothalamic tissue and quantified by specific radioimmunoassays combined with high-performance liquid chromatography. The major angiotensin peptides detected were angiotensin-(2-7) (391.2 +/- 16.8 pg/g tissue) and angiotensin-(3-7) (864.8 +/- 128.1 pg/g). Angiotensin II immunoreactivity was mainly composed of angiotensin-(3-8) (117.5 +/- 64 pg/g) and trace amounts of angiotensin II and angiotensin III. Angiotensin I immunoreactivity was composed of angiotensin I (52.3 +/- 5.8 pg/g). In separate experiments, addition of 125I-angiotensin I into supernatants (18,000 g for 2 min) of canine hypothalamic homogenates resulted in the accumulation of 125I-angiotensin-(1-7) as the major peptide product (14% of the total 125I-radioactivity) at 2 min. Incubation of the homogenate supernatants with enalaprilat (1 mumol/l), phosphoramidon (10 mumol/l), or ethylenediamine tetraacetic acid (1 mmol/l) did not inhibit the production of 125I-angiotensin-(1-7). In contrast, addition of Z-Pro-Prolinal (1 mumol/l), a specific inhibitor of prolyl endopeptidase, prevented the generation of 125I-angiotensin-(1-7) from 125I-angiotensin I by 47.0 +/- 8.0% (n = 6).(ABSTRACT TRUNCATED AT 250 WORDS)     

Modulation of reflex function by endogenous angiotensins in older transgenic rats with low glial angiotensinogen

Age-related impairments in baroreflex sensitivity in Sprague-Dawley rats are associated with low solitary tract nucleus content of angiotensin-(1-7). However, transgenic rats with low-brain angiotensinogen resulting from glial overexpression of an antisense oligonucleotide to angiotensinogen (ASrAOGEN) are spared age-related declines in cardiovascular function characteristic of Sprague-Dawley rats. We examine whether cardiovascular and reflex actions of angiotensin-(1-7) persist in the solitary tract nucleus of older (16 to 22 months) ASrAOGEN rats. Baroreflex sensitivity for control of heart rate and chemosensitive vagal afferent activation in response to phenylbiguanide were measured before and after bilateral microinjection of the angiotensin II type 1 receptor antagonist candesartan and angiotensin-(1-7) receptor antagonist (D-Ala(7))-angiotensin-(1-7) in urethane/chloralose-anesthetized rats. In older anesthetized ASrAOGEN rats, candesartan had no effect, whereas (D-Ala(7))-angiotensin-(1-7) significantly reduced baroreflex sensitivity (1.80+/-0.43 versus 0.50+/-0.17 ms/mm Hg). Phenylbiguanide responses were attenuated by injection of candesartan (-79+/-6 versus -55+/-12 mm Hg and -277+/-12 versus -156+/-27 bpm; P<0.05). In addition, resting blood pressure was reduced by injection of candesartan or (D-Ala(7))-angiotensin-(1-7). Within the solitary tract nucleus of older ASrAOGEN rats, it appears that glial angiotensinogen is the main source of angiotensin II attenuation of baroreflex sensitivity; endogenous angiotensin-(1-7) from nonglial sources enhances baroreflex sensitivity; nonglial sources of angiotensin II contribute to chemosensitive vagal afferent activation; and receptors for both peptides modulate resting arterial pressure under anesthesia. These results suggest a novel mechanism for the preservation of baroreflex sensitivity during aging.     

Angiotensin-converting enzyme 2 and angiotensin-(1-7): an evolving story in cardiovascular regulation

This lecture summarizes the chronology and rationale that led to the discovery of angiotensin-(1-7) as a hormone that, in its own right, opposes the vasoconstrictor and proliferative actions of angiotensin II. The work discussed here additionally analyzes the newest findings on angiotensin-converting enzyme 2, the angiotensin-converting enzyme homologue that efficiently hydrolyzes angiotensin II into angiotensin-(1-7). Both components of this system may significantly influence our future perspective of the role of the renin-angiotensin system, not just in terms of its role in the regulation of cardiovascular and renal function but, moreover, as regulators of a vast array of disease processes in which inflammation and immune mechanisms play a role.     

Angiotensin-(1-7) attenuates neointimal formation after stent implantation in the rat

Angiotensin-(1-7) is an endogenous, biologically active peptide of the renin-angiotensin system with vasodilatory, antithrombotic, and antiproliferative properties. This study examined the effects of angiotensin-(1-7) infusion on neointimal formation after stent placement in the rat. Male Wistar rats underwent stent implantation in the abdominal aorta or sham surgery. Subsequently, an osmotic minipump was placed for angiotensin-(1-7) (24 microg/kg per hour) or saline administration. After 4 weeks, histomorphometric and histological analyses were performed, and the endothelial function was measured in isolated thoracic aortic rings. Stent implantation resulted in equal mean injury scores within the groups. The angiotensin-(1-7)-treated group displayed a significant reduction in neointimal thickness (112+/-8 versus 141+/-11 microm; P<0.05), neointimal area (0.51+/-0.05 versus 0.70+/-0.07 mm2; P<0.05), and percentage stenosis (10.4+/-1.0 versus 14.0+/-1.3%; P<0.05) compared with the saline-treated group. Furthermore, angiotensin-(1-7) infusion attenuated the stenting-induced impairment in endothelium-dependent relaxation (42.6+/-3.0 versus 64.5+/-6.0% of phenylephrine maximal contraction; P<0.001). In conclusion, angiotensin-(1-7) treatment attenuates neointimal formation after stent implantation in the rat, combined with an improvement of endothelial function.     

Angiotensin-converting enzyme 2 deficiency is associated with impaired gestational weight gain and fetal growth restriction

Angiotensin-converting enzyme 2 (ACE2) is a key enzyme of the renin-angiotensin system that influences the relative expression of angiotensin II (Ang II) and Ang-(1-7). Although ACE2 expression increases in normal pregnancy, the impact of ACE2 deficiency in pregnancy has not been elucidated. We determined the influence of ACE2 deficiency on circulating and tissue renin-angiotensin system components, fetal and maternal growth characteristics, and maternal hemodynamics (mean blood pressure and cardiac output) at day 18 of gestation. Gestational body weight gain was lower in the ACE2 knockout (KO) versus C57BL/6 (wild-type) mice (30.3±4.7 versus 38.2±1.0 g; P<0.001). Fetal weight (0.94±0.1 versus 1.24±0.01 g; P<0.01) and length (19.6±0.2 versus 22.2±0.2 mm; P<0.001) were less in KO. Mean blood pressure was significantly reduced in C57BL/6 with pregnancy; it was elevated (P<0.05) in the KO virgin and pregnant mice, and this was associated with an increased cardiac output in both C57BL/6 and KO pregnant mice (P<0.05). Plasma Ang-(1-7) was reduced in pregnant KO mice (P<0.05). Placenta Ang II levels were higher in KO mice (52.9±6.0 versus 22.0±3.3 fmol/mg of protein; P<0.001). Renal Ang II levels were greater in KO virgin mice (30.0±1.7 versus 23.7±1.1 fmol/mg of protein; P<0.001). There was no change in the Ang-(1-7) levels in the KO placenta and virgin kidney. These results suggest that ACE2 deficiency and associated elevated placenta Ang II levels impact pregnancy by impairing gestational weight gain and restricting fetal growth.     

Plasma angiotensin II and the antihypertensive action of angiotensin-converting enzyme inhibition

The measurement of immunoreactive "angiotensin II" in plasma cannot provide an accurate reflection of the efficacy of angiotensin-converting enzyme (ACE) inhibition because different angiotensin fragments interfere in all radioimmunoassays available so far. More complex methods are necessary in order to measure specifically angiotensin-(1-8)octapeptide. With such methodology it can be shown that no tolerance develops to the angiotensin II-reducing effect of ACE inhibitors after prolonged administration. Marked reduction of angiotensin II levels can be shown even in patients with primary aldosteronism. At peak blockade, the level of plasma angiotensin II is still related to circulating active renin and angiotensin I. Accordingly, because ACE inhibitors raise circulating angiotensin I in a dose-dependent fashion, this should be taken into account when dosing ACE inhibitors. The hypothesis that tissue renin-angiotensin systems play an important independent role in determining vasomotor tone is very interesting. However, any discussion on whether tissue or plasma renin determines the pharmacological effect of ACE inhibitors should be based on the simultaneous measurement of true angiotensin II in tissue and plasma under steady-state conditions.     

Evidence for the role of angiotensin II biosynthesis in the rat internal anal sphincter tone

BACKGROUND & AIMS: The internal anal sphincter tone is important for anorectal continence. This study examined the role of angiotensin II as a neurohumoral signal for the myogenic tone in the internal anal sphincter. METHODS: We determined the effect of angiotensin I, II, III, and IV and angiotensin-(1-7) on the basal tone of the rat internal anal sphincter smooth muscle before and after selective receptor antagonists and biosynthesis inhibitors. Selective pharmacological tools used were losartan (for the AT(1) receptor), PD123,319 (for AT(2)), A-779 [for angiotensin-(1-7)], captopril (for angiotensin-converting enzyme), and amastatin (for aminopeptidases A and N). Angiotensins were measured by using high-performance liquid chromatography/UV. Western blot studies were used to determine AT(1) and AT(2) receptors, ACE, and aminopeptidases A and N. RESULTS: Angiotensin I, II, and III produced concentration-dependent contraction in the internal anal sphincter mediated by AT(1) receptors. However, in the higher concentrations (from 100 nM to 10 microM), angiotensin II showed an inhibitory effect via AT(2) receptors. Captopril (1 microM) inhibited the biosynthesis of angiotensin II in the internal anal sphincter, antagonized the contractile effects of angiotensin I, and, importantly, caused a decrease in the basal tone. Amastatin inhibited the effects of angiotensin II while augmenting those of angiotensin III. In contrast, angiotensin-(1-7) and angiotensin IV had only minor effects in the internal anal sphincter. Angiotensin I, II, and III; angiotensin-converting enzyme; aminopeptidase A and aminopeptidase n; at(1); and at(2) receptors were shown to be present in the internal anal sphincter. CONCLUSIONS: Locally produced angiotensin II may partially regulate basal tone in the internal anal sphincter.     

The renal antihypertensive effect of angiotensin I converting enzyme inhibitors

Intrarenal administration of angiotensin I converting enzyme (ACE) inhibitors carried out in norepinephrine- (NE; 2-4 micrograms/kg per min) or in angiotensin II- (ANG II; 60-90 ng/kg per min) induced acute hypertension in conscious unrestrained rabbits. Intrarenal administration of captopril (5 mg/kg) and MK-422 (1 mg/kg) caused no significant effect when injected intravenously. However, it showed a prompt and marked depressor effect in NE- but not in ANG II-induced hypertension. This effect was not observed after intrarenal infusion of saralasin (2 and 10 micrograms/kg per min) in NE-induced hypertension. While pretreatment with aprotinin or indomethacin failed to inhibit the depressor action, 2-bromoethylamine hydrobromide (BEA), which is known to induce necrosis of the renal papilla, produced complete abolition of the depressor effect of an intrarenal injection of MK-422 in NE-induced hypertension. These results indicate that the kidney plays an important role in the depressor action of ACE inhibitors in NE- but not in ANG II-induced acute hypertension, and that this effect may be related to the potentiation of antihypertensive renomedullary lipids rather than the inhibition of the renin-angiotensin system or the potentiation of bradykinin or prostaglandins.     

Upregulation of hepatic angiotensin-converting enzyme 2 (ACE2) and angiotensin-(1-7) levels in experimental biliary fibrosis

BACKGROUND/AIMS: Angiotensin-converting enzyme 2 (ACE2), its product, angiotensin-(1-7) and its receptor, Mas, may moderate the adverse effects of angiotensin II in liver disease. We examined the expression of these novel components of the renin angiotensin system (RAS) and the production and vasoactive effects of angiotensin-(1-7) in the bile duct ligated (BDL) rat. METHODS: BDL or sham-operated rats were sacrificed at 1, 2, 3 and 4 weeks. Tissue and blood were collected for gene expression, enzyme activity and peptide measurements. In situ perfused livers were used to assess angiotensin peptide production and their effects on portal resistance. RESULTS: Hepatic ACE2 gene and activity (P<0.0005), plasma angiotensin-(1-7) (P<0.0005) and Mas receptor expression (P<0.01) were increased following BDL compared to shams. Perfusion experiments confirmed that BDL livers produced increased angiotensin-(1-7) (P<0.05) from angiotensin II and this was augmented (P<0.01) by ACE inhibition. Whilst angiotensin II increased vasoconstriction in cirrhotic livers, angiotensin-(1-7) had no effect on portal resistance. CONCLUSIONS: RAS activation in chronic liver injury is associated with upregulation of ACE2, Mas and hepatic conversion of angiotensin II to angiotensin-(1-7) leading to increased circulating angiotensin-(1-7). These results support the presence of an ACE2-angiotensin-(1-7)-Mas axis in liver injury which may counteract the effects of angiotensin II.     

Differential effects of angiotensin II and angiotensin-(1-7) at the nucleus tractus solitarii of transgenic rats with low brain angiotensinogen

OBJECTIVES : In this study, we investigated the effects of angiotensin II and angiotensin-(1-7) at the nucleus tractus solitarii (nTS) in transgenic rats with a severe deficit in brain angiotensinogen production, TGR(ASrAOGEN) (TGR). METHODS : Angiotensin II (10 pmol), angiotensin-(1-7) (10 pmol) or NaCl (0.9%/50 nl) were microinjected into the nTS of urethane-anaesthetized TGR (n = 36) and Sprague Dawley (SD) (n = 34) rats. Mean arterial pressure (MAP) and heart rate were measured via a femoral artery catheter and the baroreflex control of heart rate was evaluated after increases in MAP induced by phenylephrine (baroreflex bradycardia). RESULTS : Angiotensin II microinjections into the nTS of the TGR induced a higher decrease in MAP and heart rate (-37 +/- 5 mmHg and -69 +/- 12 b.p.m., respectively) in comparison to SD rats (-18 +/- 1 mmHg and -43 +/- 5 b.p.m., respectively). In contrast, changes after angiotensin-(1-7) microinjections into the nTS of TGR (-6 +/- 1 mmHg and -13 +/- 4 b.p.m.) were significantly smaller than that induced in SD (-11 +/- 2 mmHg and -24 +/- 6 b.p.m.). The baseline baroreflex sensitivity to phenylephrine of TGR was accentuated in comparison to SD rats (0.70 +/- 0.06 versus 0.44 +/- 0.03 ms/mmHg). Angiotensin II microinjection into the nTS produced similar attenuation in the baroreflex bradycardia in both SD (0.28 +/- 0.07 versus 0.5 +/- 0.07 ms/mmHg, before injection) and TGR (0.44 +/- 0.1 versus 0.82 +/- 0.1 ms/mmHg, before injection). In contrast, angiotensin-(1-7) microinjection elicited a facilitation of the baroreflex bradycardia in SD (0.68 +/- 0.12 versus 0.41 +/- 0.03 ms/mmHg, before injection), while in TGR, angiotensin-(1-7) induced an attenuation of baroreflex bradycardia (0.34 +/- 0.07 ms/mmHg versus 0.55 +/- 0.05 ms/mmHg, before injection). CONCLUSIONS : These results indicate that a permanent inhibition of angiotensinogen synthesis in the brain can lead to an increase in the sensitivity of the baroreflex control of heart rate (baroreflex bradycardia) and an increase in angiotensin II responsiveness at the nTS. However, the nTS effect of angiotensin-(1-7) was significantly attenuated in these transgenic rats. These data further indicate that the decrease in brain angiotensins in the transgenic rats may be functionally relevant and support the concept of differential regulatory mechanisms for the effects of the two angiotensin peptides.     

Role of nNOS in regulation of renal function in angiotensin II-induced hypertension

Previous studies have indicated that in normotensive rats, NO produced by neuronal NO synthase (nNOS) plays an important role in modulating tubuloglomerular feedback (TGF)-mediated afferent arteriolar constriction. It has also been shown that in angiotensin (Ang) II-infused hypertensive rats, there is a reduced ability of nNOS-derived NO to counteract this vasoconstriction. The present study was performed to (1) assess in vivo renal functional responses to intrarenal nNOS inhibition in control and Ang II-infused rats and (2) determine whether changes in renal function following nNOS inhibition are mediated by unopposed stimulation of Ang II receptor subtype 1 (AT(1)). Wistar rats were infused with either saline (SAL) or Ang II (80 ng/min) by osmotic minipumps implanted subcutaneously. Mean arterial blood pressure of SAL- and Ang II-infused rats on day 13 after implantation averaged 121+/-4 (n=28) and 151+/-5 (n=30), respectively (P<0.05). There were no differences in glomerular filtration rate (GFR) (0.68+/-0.09 versus 0.59+/-0.09 mL. min(-1). g(-1)), renal plasma flow (RPF) (2.66+/-0.31 versus 2.34+/-0.39 mL. min(-1). g(-1)), and absolute sodium excretion (0.37+/-0.07 versus 0.42+/-0.09 micromol. min(-1). g(-1)). Intrarenal infusion of SAL did not change GFR, RPF, and sodium excretion in either SAL-infused (n=7) or Ang II-infused rats (n=8). Acute intrarenal administration of the nNOS inhibitor S-methyl-L-thiocitrulline (L-SMTC; 0.3 mg/h) decreased GFR, RPF, and sodium excretion in SAL-infused rats (n=9) by 29+/-4%, 38+/-4%, and 70+/-4% compared with control values (P<0.05). The pretreatment by the AT(1) receptor antagonist candesartan (750 ng IR) in SAL-infused rats (n=7) effectively prevented the decrease in RPF (-3+/-3%) elicited by nNOS inhibition and resulted in an increase in GFR (+25+/-12, P<0.05) and a concomitant greater increase in sodium excretion (84+/-12%, P<0.05) compared with control values. In contrast, in Ang II-infused rats (n=10) intrarenal inhibition of nNOS by L-SMTC did not cause significant decreases in GFR, RPF and sodium excretion (-2+/-2%, -15+/-10%, and -14+/-10%, respectively). These results suggest that in normotensive rats nNOS-derived NO counteracts Ang II-mediated vasoconstriction in the pre- and postglomerular microcirculation. Furthermore, Ang II-infused rats exhibit an impaired ability to release NO by nNOS. Decreased nNOS activity is likely to account at least partially for the enhanced TGF responsiveness in Ang II-infused rats and thus may contribute to the maintenance of hypertension in this model.     

Angiotensin-(1-7) attenuates vasoconstriction evoked by angiotensin II but not by noradrenaline in man

Angiotensin-(1-7) has been suggested to be a novel vasodilating peptide. We investigated the direct vascular effect of angiotensin-(1-7) in human forearm resistant vessels, particularly with regard to the interaction with angiotensin II, in healthy normotensive men by strain-gauge venous occlusion plethysmography with intra-arterial infusions of peptides. Intra-arterial infusion of angiotensin-(1-7) at 0.1 to 2000 pmol/min did not cause vasodilatation but rather reduced forearm blood flow by approximately 10% at the highest dose. A placebo-controlled study showed that angiotensin-(1-7) at 0.5 to 40 nmol/min caused weak but significant vasoconstriction (P=0.0016 by ANOVA). Angiotensin-(1-7) at 100 pmol/min, but not at 10 pmol/min, significantly shifted the angiotensin II dose-response curve toward the right (mean+/-SD of percent changes in forearm blood flow: -19+/-17%, -33+/-22%, -55+/-12%, -63+/-10%, and -68+/-5% at 5, 10, 25, 50, and 100 pmol/min of angiotensin II, respectively, with saline; 5+/-13%, 0. 9+/-18%, -40+/-16%, -54+/-9%, and -61+/-6% with angiotensin-(1-7), P=0.0021 by ANOVA). Angiotensin-(1-7) did not affect the dose-response curve of noradrenaline [3+/-12%, 5+/-16%, -20+/-22%, -31+/-18%, and -40+/-12% at 25, 50, 100, 300, and 600 pmol/min of noradrenaline, respectively, with saline; -4+/-15%, -2+/-23%, -29+/-22%, -34+/-16%, and -42+/-9% with angiotensin-(1-7)]. Our results suggest that angiotensin-(1-7) antagonizes vasoconstriction by angiotensin II in human resistant vessels and might act as an endogenous angiotensin II antagonist.