Pharmacology of valsartan, an angiotensin II receptor antagonist.

Valsartan is the second orally-active, non-peptide angiotensin II receptor blocker to reach the market in Europe and the USA for the treatment of hypertension. Preclinical studies have demonstrated that this blocker is specific for the AT(1) receptor and has no affinity for the angiotensin II AT(2) receptor. Experimentally, valsartan dose-dependently inhibits the vasoconstriction induced by angiotensin II and lowers blood pressure in renin-dependent models of hypertension. Pharmacologically, oral valsartan is characterised by a low bioavailability but a rapid absorption and distribution with a half-life in keeping with once-daily administration. Thus, after oral administration, the maximal plasma concentration is reached 2 h after dosing and the elimination half-life is about 6 h. Clinically, several dose-finding and comparative studies have demonstrated that valsartan is an effective and well-tolerated antihypertensive drug in patients with mild to moderate hypertension. Valsartan has also been shown to be effective in severe hypertension. Valsartan is at least as effective as ACE inhibitors, diuretics, beta-blockers and calcium antagonists. However, none of the side-effects observed with these latter agents, including cough and lower limb oedema, has been observed with the administration of valsartan. Three large clinical trials are now underway to demonstrate whether valsartan can reduce morbidity and mortality: one in hypertensives with a high cardiovascular risk profile (VALUE), one in patients with heart failure previously treated with an angiotensin-converting enzyme inhibitor (VAL-HeFT) and one in post-myocardial infarct patients (VALIANT). These studies will further define the place of valsartan beyond the treatment of hypertension.

Cardiovascular actions of angiotensin-(1-7)

Angiotensin-(1-7) (Ang-(1-7)) is now considered to be a biologically active member of the renin-angiotensin system. The functions of Ang-(1-7) are often opposite to those attributed to the main effector component of the renin-angiotensin system, Ang II. Chronic administration of angiotensin-converting enzyme inhibitors (ACEI) increases 10- to 25-fold the plasma levels of this peptide, suggesting that part of the beneficial effects of ACEI could be mediated by Ang-(1-7). Ang-(1-7) can be formed from Ang II or directly from Ang I. Other enzymatic pathways for Ang-(1-7) generation have been recently described involving the novel ACE homologue ACE2. This enzyme can form Ang-(1-7) from Ang II or less efficiently by the hydrolysis of Ang I to Ang-(1-9) with subsequent Ang-(1-7) formation. The biological relevance of Ang-(1-7) has been recently reinforced by the identification of its receptor, the G-protein-coupled receptor Mas. Heart and blood vessels are important targets for the formation and actions of Ang-(1-7). In this review we will discuss recent findings concerning the biological role of Ang-(1-7) in the heart and blood vessels, taking into account aspects related to its formation and effects on these tissues. In addition, we will discuss the potential of Ang-(1-7) and its receptor as a target for the development of new cardiovascular drugs.

ACE-I inhibitory properties of hydrolysates from germinated and ungerminated Phaseolus lunatus proteins

Phaseolus lunatus protein concentrates and the proteases Alcalase(R) and Pepsin-Pancreatin were used for the production of protein hydrolysates that inhibit angiotensin-I converting enzyme (ACE). Protein concentrate obtained from germinated and ungerminated seeds flour was hydrolyzed with Alcalase(R) at enzyme/substrate ratio (E/S) 1/10 and during 0.5 and 2.0 h, respectively. On the other hand, protein concentrate obtained from ungerminated (E/S: 1/10) and germinated (E/S: 1/50) seeds flour was sequentially hydrolyzed with Pepsin-Pancreatin during 1.0 and 3.0 h, respectively. Peptide fractions with ACE inhibitory activity in a range of 0.9 to 3.8 µg/mL were obtained by G-50 gel filtration chromatography and high- performance liquid chromatography C18 reverse phase chromatography. The observed amino acid composition suggests a substantial contribution of hydrophobic residues to the peptides’ inhibitory potency, which potentially acts via blocking of angiotensin II production. These results show that P. lunatus seed proteins are a potential source of ACE inhibitory peptides when hydrolyzed with Alcalase(R) and Pepsin-Pancreatin.

Caractérisation du gène de l'enzyme de conversion de l'angiotensine-2 dans le rein diabétique et implication dans le développement de la néphropathie diabétique et de l'hypertension

De nombreuses études ont bien démontré que l’activation du système rénine-angiotensine (RAS) joue un rôle important dans le développement de l’hypertension et de la néphropathie diabétique (DN). La découverte de l’enzyme de conversion de l’angiotensine-2 (ACE2) et l’identification du récepteur MAS, spécifique pour l’angiotensine 1-7 (Ang 1-7), ont permis d’identifier deux nouveaux membres du RAS. L’axe ACE2/Ang 1-7/MAS contrebalance les effets de l’axe ACE/Ang II/AT1. Plusieurs évidences impliquent la contribution du RAS intrarénal dans la DN. Des études réalisées dans notre laboratoire avec des souris transgéniques surexprimant l’angiotensinogène de rat dans les cellules de leurs tubules proximaux rénaux (RPTCs) ont permis de démontrer l’importance du RAS intrarénal dans l’induction de l’hypertension et les dommages rénaux. Nous avons également observé que l’expression rénale de l’ACE2 et les niveaux urinaires d’ANG 1-7 sont plus faibles chez les souris Akita (diabète de type 1) et qu’un traitement avec des bloqueurs du RAS permet de normaliser l’expression de l’ACE2 et de prévenir le développement de l’hypertension dans le modèle des souris Akita. Dans un milieu diabétique, à la fois la glycémie et l’angiotensine II (Ang II) peuvent induire la génération des espèces réactives de l’oxygène (ROS), contribuant ainsi aux dommages rénaux. Afin d’explorer la relation entre les ROS, ACE2 et la DN, nous avons créé des souris Akita transgéniques surexprimant la catalase (Cat) dans les RPTCs, en croisant des souris Akita diabétique de type 1 à notre modèle de souris transgéniques surexprimant la Cat de rat dans les RPTCs. Dans une seconde étude, des souris Akita ont été traitées avec l’Ang 1-7 ou une combinaison d’Ang 1-7 et de son antagoniste, A779, afin d’étudier la relation entre l’action de l’Ang 1-7, l’hypertension systolique (sHTN), le stress oxydatif, les dommages rénaux, ACE2 et l’expression du récepteur Mas. Nos résultats ont montré que la surexpression de Cat atténue le stress oxydatif rénal; prévient l’hypertension, améliore le taux de filtration glomérulaire, l’albuminurie, l’hypertrophie rénale, la fibrose tubulo-interstitielle et l’apoptose tubulaire; et supprime l’expression des gènes profibrotiques et proapoptotiques dans les RPTCs des souris Akita Cat-Tg lorsque comparées aux souris Akita. De plus, la surexpression de Cat dans les RPTC des souris Akita normalise l’expression rénale de l’ACE2 et les niveaux urinaires d’Ang 1-7. D’autre part, l’administration d’Ang 1-7 prévient l’hypertension systémique, normalise le ratio albumine/créatinine urinaire et atténue l’hyperfiltration glomérulaire des souris Akita, sans affecter la glycémie sanguine. De plus, le traitement avec l’Ang 1-7 atténue aussi le stress oxydatif et l’expression de la NADPH oxydase, Agt, ACE, TGF-β1 (transforming growth factor-β1) et collagène IV, tout en augmentant l’expression de l’ACE2 et du récepteur Mas dans les reins des souris Akita. Ces effets sont renversés par la co-admininstration d’A779. Ces résultats démontrent que la surexpression de Cat prévient l’hypertension et la progression de la néphropathie, en plus de mettre en lumière l’importance du stress oxydatif intrarénal et l’expression de l’ACE2 comme facteurs contribuant à l’hypertension et les dommages rénaux observés dans le diabète. En outre, nos données suggèrent que l’Ang 1-7 joue un rôle protecteur dans l’hypertension et les dommages aux RPTC dans le diabète, principalement en réduisant les voies de signalisations du stress oxydatif dans les reins et en normalisant l’expression de l’ACE2 et du récepteur Mas. Nos résultats indiquent aussi que l’Ang 1-7 pourrait agir comme un agent thérapeutique potentiel dans le traitement de l’hypertension systémique et les dommages rénaux observés dans le diabète. En conséquence, l’Ang 1-7 est responsable du rôle protecteur de l’ACE2 dans l’hypertension et la DN.

Production of novel ACE inhibitory peptides from beta-lactoglobulin using Protease N Amano

Potent angiotensin l-converting enzyme (ACE) inhibitory peptide mixtures were obtained from the hydrolysis of beta-lactoglobulin (beta Lg) using Protease N Amano, a food-grade commercial proteolytic preparation. Hydrolysis experiments were carried out for 8 h at two different temperatures and neutral pH. Based on their ACE inhibitory activity, samples of 6 h of digestion were chosen for further analysis. The temperature used for the hydrolysis had a marked influence on the type of peptides produced and their concentration in the hydrolysate. Protease N Amano was found to produce very complex peptide mixtures; however, the partially fractionated hydrolysates had already very potent ACE inhibitory activity. The novel heptapeptide SAPLRVY was isolated and characterised. It corresponded to beta Lg f(36-42) and had an IC50 value of 8 mu m, which is considerably lower than the most potent ACE inhibitory peptides derived from bovine beta Lg reported so far. (C) 2008 Elsevier Ltd. All rights reserved.

Endothelin-converting enzyme-1 degrades internalized somatostatin-14

Agonist-induced internalization of somatostatin receptors (ssts) determines subsequent cellular responsiveness to peptide agonists and influences sst receptor scintigraphy. To investigate sst2A trafficking, rat sst2A tagged with epitope was expressed in human embryonic kidney cells and tracked by antibody labeling. Confocal microscopical analysis revealed that stimulation with sst and octreotide induced internalization of sst2A. Internalized sst2A remained sequestrated within early endosomes, and 60 min after stimulation, internalized sst2A still colocalized with beta-arrestin1-enhanced green fluorescence protein (EGFP), endothelin-converting enzyme-1 (ECE-1), and rab5a. Internalized (125)I-Tyr(11)-SST-14 was rapidly hydrolyzed by endosomal endopeptidases, with radioactive metabolites being released from the cell. Internalized (125)I-Tyr(1)-octreotide accumulated as an intact peptide and was released from the cell as an intact peptide ligand. We have identified ECE-1 as one of the endopeptidases responsible for inactivation of internalized SST-14. ECE-1-mediated cleavage of SST-14 was inhibited by the specific ECE-1 inhibitor, SM-19712, and by preventing acidification of endosomes using bafilomycin A(1). ECE-1 cleaved SST-14 but not octreotide in an acidic environment. The metallopeptidases angiotensin-1 converting enzyme and ECE-2 did not hydrolyze SST-14 or octreotide. Our results show for the first time that stimulation with SST-14 and octreotide induced sequestration of sst2A into early endosomes and that endocytosed SST-14 is degraded by endopeptidases located in early endosomes. Furthermore, octreotide was not degraded by endosomal peptidases and was released as an intact peptide. This mechanism may explain functional differences between octreotide and SST-14 after sst2A stimulation. Moreover, further investigation of endopeptidase-regulated trafficking of neuropeptides may result in novel concepts of neuropeptide receptor inactivation in cancer diagnosis.

Endothelin-converting enzyme-1 regulates endosomal sorting of calcitonin receptor-like receptor and beta-arrestins

Although cell surface metalloendopeptidases degrade neuropeptides in the extracellular fluid to terminate signaling, the function of peptidases in endosomes is unclear. We report that isoforms of endothelin-converting enzyme-1 (ECE-1a-d) are present in early endosomes, where they degrade neuropeptides and regulate post-endocytic sorting of receptors. Calcitonin gene-related peptide (CGRP) co-internalizes with calcitonin receptor-like receptor (CLR), receptor activity-modifying protein 1 (RAMP1), beta-arrestin2, and ECE-1 to early endosomes, where ECE-1 degrades CGRP. CGRP degradation promotes CLR/RAMP1 recycling and beta-arrestin2 redistribution to the cytosol. ECE-1 inhibition or knockdown traps CLR/RAMP1 and beta-arrestin2 in endosomes and inhibits CLR/RAMP1 recycling and resensitization, whereas ECE-1 overexpression has the opposite effect. ECE-1 does not regulate either the resensitization of receptors for peptides that are not ECE-1 substrates (e.g., angiotensin II), or the recycling of the bradykinin B(2) receptor, which transiently interacts with beta-arrestins. We propose a mechanism by which endosomal ECE-1 degrades neuropeptides in endosomes to disrupt the peptide/receptor/beta-arrestin complex, freeing internalized receptors from beta-arrestins and promoting recycling and resensitization.

Production of novel ACE inhibitory peptides from β-lactoglobulin using Protease N Amano

Potent angiotensin I-converting enzyme (ACE) inhibitory peptide mixtures were obtained from the hydrolysis of β-lactoglobulin (βLg) using Protease N Amano, a food-grade commercial proteolytic preparation. Hydrolysis experiments were carried out for 8 h at two different temperatures and neutral pH. Based on their ACE inhibitory activity, samples of 6 h of digestion were chosen for further analysis. The temperature used for the hydrolysis had a marked influence on the type of peptides produced and their concentration in the hydrolysate. Protease N Amano was found to produce very complex peptide mixtures; however, the partially fractionated hydrolysates had already very potent ACE inhibitory activity. The novel heptapeptide SAPLRVY was isolated and characterised. It corresponded to βLg f(36–42) and had an IC50 value of 8 μm, which is considerably lower than the most potent ACE inhibitory peptides derived from bovine βLg reported so far.

The renin-angiotensin system plays a major role in voiding dysfunction of ovariectomized rats

Aims: The renin-angiotensin system (RAS) plays a major role in cardiovascular diseases in postmenopausal women, but little is known about its importance to lower urinary tract symptoms. In this study we have used the model of ovariectomized (OVX) estrogen-deficient rats to investigate the role of RAS in functional and molecular alterations in the urethra and bladder. Main methods: Responses to contractile and relaxant agents in isolated urethra and bladder, as well as cystometry were evaluated in 4-month OVX Sprague-Dawley rats. Angiotensin-converting enzyme activity and Western blotting for AT1/AT2 receptors were examined. Key findings: Cystometric evaluations in OVX rats showed increases in basal pressure, capacity and micturition frequency, as well as decreased voiding pressure. Angiotensin II and phenylephrine produced greater urethral contractions in OVX compared with Sham group. Carbachol-induced bladder contractions were significantly reduced in OVX group. Relaxations of urethra and bladder to sodium nitroprusside and BAY 41-2272 were unaffected by OVX. Angiotensin-converting enzyme activity was 2.6-fold greater (p < 0.05) in urethral tissue of OVX group, whereas enzyme activity in plasma and bladder remained unchanged. Expressions of AT1 and AT2 receptors in the urethra were markedly higher in OVX group. In bladder, AT1 receptors were not detected, whereas AT2 receptor expression was unchanged between groups. 17β-Estradiol replacement (0.1 mg/kg, weekly) or losartan (30 mg/kg/day) largely attenuated most of the alterations seen in OVX group. Significance: Prolonged estrogen deprivation leads to voiding dysfunction and urethral hypercontractility that are associated with increased ACE activity and up-regulation of angiotensin AT1/AT2 receptor in the urethral tissue. © 2013 Elsevier Inc. All rights reserved.

Effects of aerobic exercise training on cardiac renin-angiotensin system in an obese Zucker rat strain

Objective: Obesity and renin angiotensin system (RAS) hyperactivity are profoundly involved in cardiovascular diseases, however aerobic exercise training (EXT) can prevent obesity and cardiac RAS activation. The study hypothesis was to investigate whether obesity and its association with EXT alter the systemic and cardiac RAS components in an obese Zucker rat strain. Methods: The rats were divided into the following groups: Lean Zucker rats (LZR); lean Zucker rats plus EXT (LZR+EXT); obese Zucker rats (OZR) and obese Zucker rats plus EXT (OZR+EXT). EXT consisted of 10 weeks of 60-min swimming sessions, 5 days/week. At the end of the training protocol heart rate (HR), systolic blood pressure (SBP), cardiac hypertrophy (CH) and function, local and systemic components of RAS were evaluated. Also, systemic glucose, triglycerides, total cholesterol and its LDL and HDL fractions were measured. Results: The resting HR decreased (, 12%) for both LZR+EXT and OZR+EXT. However, only the LZR+EXT reached significance (p, 0.05), while a tendency was found for OZR versus OZR+EXT (p = 0.07). In addition, exercise reduced (57%) triglycerides and (61%) LDL in the OZR+EXT. The systemic angiotensin I-converting enzyme (ACE) activity did not differ regardless of obesity and EXT, however, the OZR and OZR+EXT showed (66%) and (42%), respectively, less angiotensin II (Ang II) plasma concentration when compared with LZR. Furthermore, the results showed that EXT in the OZR prevented increase in CH, cardiac ACE activity, Ang II and AT2 receptor caused by obesity. In addition, exercise augmented cardiac ACE2 in both training groups. Conclusion: Despite the unchanged ACE and lower systemic Ang II levels in obesity, the cardiac RAS was increased in OZR and EXT in obese Zucker rats reduced some of the cardiac RAS components and prevented obesity-related CH. These results show that EXT prevented the heart RAS hyperactivity and cardiac maladaptive morphological alterations in obese Zucker rats.

Sirolimus and everolimus reduce albumin endocytosis in proximal tubule cells via an angiotensin II-dependent pathway

The proliferation signal inhibitors (PSIs) sirolimus (SRL) and everolimus (ERL) often induce proteinuria due to glomerular but also tubular dysfunction in transplant patients. The beneficial effect of angiotensin converting enzyme inhibitors (ACE-I) and angiotensin II (Ang II) type 1 receptor blockers (ARB) has been reported.

Contribution of the genes of the renin-angiotensin system to interindividual differences in blood pressure levels in Caucasians from Rochester, Minnesota

The Renin-Angiotensin system (RAS) regulates blood pressure through its effects on vascular tone, renal hemodynamics, and renal sodium and fluid balance. The genes encoding the four major components of the RAS, angiotensinogen, renin, angiotensin I-converting enzyme (ACE), and angiotensin II receptor type 1 (AT1), have been investigated as candidate genes in the pathogenesis of essential hypertension. However, studies have primarily focused on small samples of diseased individuals, and, therefore, have provided little information about the determinants of interindividual variation in blood pressure (BP) in the general population.^ Using data from a large population-based sample from Rochester, MN, I have evaluated the contribution of variation in the region of the RAS genes to interindividual variation in systolic, diastolic, and mean arterial pressure in the population-at-large. Marker genotype data from four polymorphisms located within or very near these genes were first collected on 3,974 individuals from 583 randomly ascertained three-generation pedigrees. Haseman-Elston regression and variance component methods of linkage analysis were then carried out to estimate the proportion of interindividual variance in BP attributable to the effects of variation at these four measured loci.^ A significant effect of the ACE locus on interindividual variation in mean arterial pressure (MAP) was detected in a sample of siblings belonging to the youngest generation. After allowing for measured covariates, this effect accounted for 15-25% of the interindividual variance in MAP, and was even greater in a subset with a positive family history of hypertension. When gender-specific analyses were carried out, this effect was significant in males but not in females. Extended pedigree analyses also provided evidence for an effect of the ACE locus on interindividual variation in MAP, but no difference between males and females was observed. Circumstantial evidence suggests that the ACE gene itself may be responsible for the observed effects on BP, although the possibility that other genes in the region may be at play cannot be excluded.^ No definitive evidence for an effect of the renin, angiotensinogen, or AT1 loci on interindividual variation in BP was obtained in this study, suggesting that the impact of these genes on BP may not be great in the Caucasian population-at-large. However, this does not preclude a larger effect of these genes in some subsets of individuals, especially among those with clinically manifest hypertension or coronary heart disease, or in other populations. ^

Reduced growth, abnormal kidney structure, and type 2 (AT2) angiotensin receptor-mediated blood pressure regulation in mice lacking both AT1A and AT1B receptors for angiotensin II

The classically recognized functions of the renin–angiotensin system are mediated by type 1 (AT1) angiotensin receptors. Whereas man possesses a single AT1 receptor, there are two AT1 receptor isoforms in rodents (AT1A and AT1B) that are products of separate genes (Agtr1a and Agtr1b). We have generated mice lacking AT1B (Agtr1b −/−) and both AT1A and AT1B receptors (Agtr1a −/−Agtr1b −/−). Agtr1b −/− mice are healthy, without an abnormal phenotype. In contrast, Agtr1a −/−Agtr1b −/− mice have diminished growth, vascular thickening within the kidney, and atrophy of the inner renal medulla. This phenotype is virtually identical to that seen in angiotensinogen-deficient (Agt−/−) and angiotensin-converting enzyme-deficient (Ace −/−) mice that are unable to synthesize angiotensin II. Agtr1a −/−Agtr1b −/− mice have no systemic pressor response to infusions of angiotensin II, but they respond normally to another vasoconstrictor, epinephrine. Blood pressure is reduced substantially in the Agtr1a −/− Agtr1b −/− mice and following administration of an angiotensin converting enzyme inhibitor, their blood pressure increases paradoxically. We suggest that this is a result of interruption of AT2-receptor signaling. In summary, our studies suggest that both AT1 receptors promote somatic growth and maintenance of normal kidney structure. The absence of either of the AT1 receptor isoforms alone can be compensated in varying degrees by the other isoform. These studies reaffirm and extend the importance of AT1 receptors to mediate physiological functions of the renin–angiotensin system.

Benefits of ACE inhibitors with tissue-active levels in the cardiac-compromised patient

Angiotensin converting enzyme inhibitors (ACEI) have been proven beneficial to the cardiac-compromised patient, but whether there is an advantage associated with using a tissue-active or systemically-active ACEI is debatable. An investigation into the clinical benefits of tissue ACEI for veterinary patients was undertaken by comparing enalapril with ramipril. Results obtained concluded that although there is much evidence to prove that tissue ACEIs are superior over systemic ACEIs at the cellular level, this does not correlate in the clinical sense. Both enalapril and ramipril provided similar clinical benefits to the cardiac-compromised patient.

Blockade Of Renin-angiotensin System Prevents Micturition Dysfunction In Renovascular Hypertensive Rats.

Association between hypertension and bladder symptoms has been described. We hypothesized that micturition dysfunction may be associated with renin-angiotensin system (RAS) acting in urethra. The effects of the anti-hypertensive drugs losartan (AT1 antagonist) and captopril (angiotensin-converting enzyme inhibitor) in comparison with atenolol (β1-adrenoceptor antagonist independently of RAS blockade) have been investigated in bladder and urethral dysfunctions during renovascular hypertension in rats. Two kidney-1 clip (2K-1C) rats were treated with losartan (30 mg/kg/day), captopril (50mg/kg/day) or atenolol (90 mg/kg/day) for eight weeks. Cystometric study, bladder and urethra smooth muscle reactivities, measurement of cAMP levels and p38 MAPK phosphorylation in urinary tract were determined. Losartan and captopril markedly reduced blood pressure in 2K-1C rats. The increases in non-voiding contractions, voiding frequency and bladder capacity in 2K-1C rats were prevented by treatments with both drugs. Likewise, losartan and captopril prevented the enhanced bladder contractions to electrical-field stimulation (EFS) and carbachol, along with the impaired relaxations to β-adrenergic-cAMP stimulation. Enhanced neurogenic contractions and impaired nitrergic relaxations were observed in urethra from 2K-1C rats. Angiotensin II also produced greater urethral contractions that were accompanied by higher phosphorylation of p38 MAPK in urethral tissues of 2K-1C rats. Losartan and captopril normalized the urethral dysfunctions in 2K-1C rats. In contrast, atenolol treatment largely reduced the blood pressure in 2K-1C rats but failed to affect the urinary tract smooth muscle dysfunction. The urinary tract smooth muscle dysfunction in 2K-1C rats takes place by local RAS activation irrespective of levels of arterial blood pressure.