Volume 4, Number 2, June 2003

PAPERNo involvement of the AT₂-receptor in angiotensin II-enhanced sympathetic transmission in vitro
Alexander Nap, Jippe C Balt, Martin Pfaffendorf, Pieter A van Zwieten

Angiotensin II (Ang II) enhances sympathetic neurotransmission via AT₁-receptors located on sympathetic nerve terminals. We recently demonstrated that inhibition of Ang II-mediated facilitation in the pithed rat by irbesartan resulted in a U-shaped dose response curve, which was not observed when PD 123319, at a concentration that selectively blocks the AT₂-receptor, was co-administered. Hence, the irbesartan-mediated ‘upstroke’ might be explained by the involvement of the AT₂-receptor after AT₁ blockade with high-dose irbesartan. In the present study, we further investigated the possible role of the AT₂-receptor in Ang II-mediated facilitation in vitro.
We studied the effect of the AT₂-receptor antagonist PD 123319 (10 nM) on Ang II-enhanced sympathetic outflow evoked by electrical field stimulation (EFS) in the rat isolated inferior vena cava. Additionally, we investigated the effect of the AT₁-receptor blocker irbesartan (0.1 nM–1 µM) on the sequelae of Ang II-enhanced, EFS-evoked sympathetic nerve traffic in the presence or absence of PD 123319 (10 nM). PD 123319 did not influence Ang II-enhanced sympathetic outflow. Irbesartan dose-dependently attenuate Ang II-augmented transmitter release (pIC50 7.99+0.03), whereas no U-shaped concentration-response relationship for irbesartan was observed. Co-administration of PD 123319 with irbesartan proved unable to influence Ang II-mediated facilitation differently compared with irbesartan alone. The experimental observations indicate that the AT₂-receptor is not involved in Ang II-mediated enhancement of sympathetic nerve traffic in the present in vitro study.

JRAAS 2003;4:100-105.

PAPERInvolvement of renal ACE activity in proteinuria-associated renal damage in untreated and treated adriamycin nephrotic rats
Hendrik Bos, Gozewijn D Laverman, Robert H Henning, Anton TMG Tiebosch, Paul E de Jong, Dick de Zeeuw, Gerjan Navis

Proteinuria is assumed to play a pathogenetic role in progressive renal damage. Angiotensin-converting enzyme (ACE) inhibition reduces proteinuria and provides renoprotection. This suggests that ACE activity might play a pathogenetic role in the development of proteinuria-induced renal structural damage. We investigated this hypothesis in untreated and treated established adriamycin nephrosis, a model of proteinuria-induced renal damage.
In a time-course experiment, the development of renal structural damage in untreated adriamycin nephrotic rats was paralleled by a significant rise in renal ACE activity. Moreover, on cross-sectional analysis, a consistent positive correlation between renal, but not plasma, ACE activity and proteinuria, focal glomerulosclerosis and interstitial injury was present. Notably, these associations were present, not only in the untreated condition, but also during intervention with either ACE inhibition or AT₁-receptor antagonism. Interestingly, we found that higher renal ACE activity is associated with more severe renal damage for a given amount of proteinuria, suggesting that renal ACE activity may be either a permissive or a promoting factor in the processes by which proteinuria eventually leads to renal structural damage. This relationship was abolished by renin-angiotensin system (RAS)-blockade, suggesting that RAS-mediated effects are involved in the relationship between renal ACE activity and proteinuria-induced renal damage.
In conclusion, in untreated as well as treated adriamycin nephrotic rats, renal ACE activity is closely associated with renal outcome. This association appears to be independent of the specific mode of blockade of the RAS. Renal ACE activity is a consistent marker of individual differences in proteinuria-associated renal damage: further studies are needed to investigate a possible pathogenetic role in renal damage.

JRAAS 2003;4:106-112.

PAPERTransforming growth factor-?₁-mediated collagen gel contraction by cardiac fibroblasts
Paul Lijnen, Victor Petrov, Robert Fagard

Objective: Myofibroblasts and transforming growth factor-β₁ (TGF-β₁) are key elements of cardiac tissue fibrosis development. The aim of this study was to determine whether the ability of TGF-β₁ to affect the contractile activity of cardiac fibroblasts depends on their differentiation into myofibroblasts.
Methods: Cardiac fibroblasts (from male adult Wistar rats) from passage two were cultured to confluency and incubated on a hydrated collagen gel with and without TGF-β₁ (0, 20, 40, 100, 200, 400 or 600 pmol/L) for one, two and three days in a Dulbecco's Modified Eagle's Medium without foetal bovine serum.
Results: TGF-β₁ dose-dependently increased the contraction of collagen gel mediated by cardiac fibroblasts, reaching a maximal effect at 100 pmol/L TGF-β₁. TGF-β₁ also stimulated 3^H-thymidine incorporation and total protein content in cardiac fibroblasts in the collagen gel lattice. TGF-β₁ dose-dependently induced an increase in β-smooth muscle actin, a marker of myofibroblasts. The TGF-β₁-induced reduction of area of the collagen gel was negatively correlated to the TGF-β₁-evoked appearance of a-smooth muscle actin in the collagen gel matrix.
Conclusion: Our data demonstrate that TGF-β₁ increased the contractile activity of adult rat cardiac fibroblasts and their ability to differentiate into myofibroblasts. Because contractile activity was correlated with differentiation, the influence of TGF-β₁ on cardiac fibroblast-induced collagen gel contraction might depend on the promotion of myofibroblast differentiation.

JRAAS 2003;4:113-118.

POPULAR
TOPICPAPEREffect of angiotensin-converting enzyme inhibition on endothelial function and insulin sensitivity in hypertensive patients
Hakan Tezcan, Dilek Yavuz, Ahmet Toprak, Ihsan Akpinar, Mehmet Koç, Oguzhan Deyneli, Sema Akalin

Introduction: Evidence suggests an association between insulin resistance, hypertension and impaired endothelial function. Studies have shown that insulin resistance precedes the development of hypertension. By improving insulin sensitivity, it may be possible to improve hypertension and the subsequent damage to vessel walls. Some data indicates beneficial effects of angiotensin-converting enzyme (ACE) inhibitors on insulin sensitivity and endothelial function. We aimed to investigate these effects of ACE inhibition in the same group of patients with essential hypertension.
Materials and methods: Nine non-smoking, untreated hypertensive patients (38.3+9 years, 4/5 male/female) and 12 age-matched healthy subjects (35.2+6.7 years, 5/7 male/female) were included in the study. Hypertensive patients were given enalapril maleate (5–40 mg/day) for six months. The following parameters were studied at baseline and at the end of treatment period. Whole body insulin sensitivity was measured by a formula derived from an oral glucose tolerance test and named as the insulin sensitivity index (ISI). Insulin was measured by chemiluminescence and glucose by a glucose oxidase method. Endothelial function was evaluated as flow-mediated dilatation (FMD) of the brachial artery by ultrasonography and expressed as a percentage change relative to baseline diameter. Endothelial- independent vasodilatation was measured after sublingual nitroglycerine.
Results: FMD was impaired in the hypertensive group compared with healthy subjects (7.3+3.1% vs. 15.3+4.8%, p<0.0005), and ISI values were 1.18+0.6 vs. 4.4+0.9 (p<0.0001) respectively. Both insulin sensitivity and FMD improved after the treatment period compared with baseline values, FMD increased from 7.3+3.1% to 16.0+2.9% (p<0.0005) and ISI from 1.18+0.6 to 4.2+1.0 (p<0.0001). FMD and ISI showed a significant positive correlation (r=0.67, p<0.001) in the hypertensive group.
Conclusions: Patients with essential hypertension have impaired endothelial function and decreased whole body insulin sensitivity compared with healthy subjects. Treatment for six months with enalapril maleate seems to improve both FMD and ISI. This study confirms the beneficial effects of ACE inhibition on both endothelial function and insulin sensitivity tested in the same group of essential hypertensive patients. The mechanism of these favourable effects of ACE inhibition needs to be clarified.

JRAAS 2003;4:119-123.

PAPERBiphasic effects of angiotensin (1-7) and its interactions with angiotensin II in rat aorta
Ion Haulica, Walther Bild, Christian N Mihaila, Teodor Ionita, Corneliu P Boisteanu, Bogdan Neagu

Using isolated rat aortic rings perfused with Krebs-Henseleit saline, the vascular effects of angiotensin (1-7) (Ang [1-7]) and its interactions with angiotensin II (Ang II) were investigated.
Ang (1-7) induced endothelium-dependent relaxation and vasodilating effects in preparations precontracted with phenylephrine. Without preconstriction, Ang (1-7) at high doses (10^-6–10^-5 M) produced either a significant inhibition of Ang II-induced vasoconstriction or a non-tachyphylactic vasopressor response. While losartan inhibited the vasoconstriction induced by Ang (1-7), A779 blocked only its relaxation. Unlike losartan, blockade of AT₂-receptors with PD 123319 had no effect. Taking into account the biphasic effects of angiotensin (1-7), we propose that it is one of the active components of the renin-angiotensin system, which is involved as a modulator both in the counter-regulatory actions of Ang II and in the self-regulation of its own vasodilating effects.

JRAAS 2003;4:124-128.

POPULAR
TOPICEDITORIAL REVIEWInteractions of angiotensin II with NAD(P)H oxidase, oxidant stress and cardiovascular disease
David G Harrison, Hua Cai, Ulf Landmesser, Kathy K Griendling

An elevation in angiotensin II (Ang II) levels is a common occurrence in a diverse number of cardiovascular diseases including hypertension, hypercholesterolaemia, atherosclerotic coronary artery disease, left ventricular hypertrophy (LVH), heart failure and diabetes. An important effect of Ang II is activation of the NAD(P)H oxidase, a major source of reactive oxygen species (ROS) production by vascular cells. This increase in cellular ROS contributes to the pathogenesis of vascular disease by altering endothelial cell function, enhancing smooth muscle cell growth and proliferation, stimulating inflammatory proteins, including macrophage chemoattractant agents, growth factors and cytokines, and modulating matrix remodelling. Studies of genetically-altered mice have unequivocally shown that activation of the NAD(P)H oxidase by Ang II contributes to hypertension, LVH and atherosclerosis. Furthermore, increasing evidence suggest that the NAD(P)H oxidase contributes to human disease, suggesting that it is a potential target for future therapeutic intervention.

JRAAS 2003;4:51-61.

EDITORIAL REVIEWAngiotensin II receptor blockers and cardiovascular outcomes: what does the future hold?
Michael A Weber

The ability of angiotensin-converting enzyme (ACE) inhibitors and angiotensin II receptor blockers (ARBs) to lower blood pressure (BP) is well established. ACE inhibitors (ACE-Is) have also been shown to improve the prognosis of a broad range of patients at high cardiovascular risk, including those with heart failure, post-myocardial infarction (MI), and nephropathy. These benefits suggest that interrupting the renin-angiotensin-aldosterone system (RAAS) with ACE-Is has a widespread vasculoprotective effect, provided that BP is also adequately controlled. Evidence that RAAS blockade by ARBs also improves long-term clinical outcomes in patients with cardiovascular disease has started to accumulate, and will be tested further during the coming years as a number of large-scale, prospective trials are completed. These trials are investigating the long-term protective effects of ARBs on morbidity and mortality in patients with hypertension, heart failure, diabetes mellitus, acute MI, or established vascular disease. The results should establish the extent to which ARBs exhibit the vasculoprotective properties demonstrated by ACE-Is in patients at high cardiovascular risk. If ARBs are found to provide benefits that are similar to, or even greater than ACE-Is, it may have important implications for drug selection, given the excellent tolerability of ARBs. Some studies are also investigating whether more extensive RAAS blockade using a combination of an ARB and an ACE-I will offer even greater protection than either agent alone.

JRAAS 2003;4:62-73.

EDITORIAL REVIEWACE inhibition or angiotensin receptor blockade: which should we use in diabetic patients?
Luis M Ruilope, Julian Segura, Ernesto L Schiffrin

Blockade of the effects of angiotensin II (Ang II) by using an angiotensin-converting enzyme (ACE) inhibitor has been proven to be of value in Type 1 diabetic nephropathy and in non-diabetic renal disease. Evidence in favour of Ang II blockade in Type 2 diabetic patients with renal damage is still lacking for ACE inhibitors (ACE-Is), while recent data indicate that angiotensin receptor blockers (ARBs) could be the drugs of choice in this situation. On the other hand, renal damage from the onset of disease is accompanied by a very significant increment in global cardiovascular risk. This fact, as well as that of simultaneous renal and cardiovascular protection, have to be considered for drug selection. In this sense, ACE-Is have been shown to be the drugs of choice when secondary cardiovascular prevention is required, while the evidence in primary prevention in hypertensive patients has been shown with losartan in the Losartan Intervention For Endpoint reduction in hypertension (LIFE) study. All these facts led to the conclusion that both ACE-Is and ARBs can be considered when both renal and cardiovascular protection are aimed for in Type 2 diabetic patients.

JRAAS 2003;4:74-79.

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TOPICEDITORIAL REVIEWThe renin-angiotensin-aldosterone system and Alzheimer’s disease?
Patrick G Kehoe

Epidemiological studies from the last decade have begun to produce evidence that the perceived joint occurrence of vascular disease and Alzheimer's disease (AD), both common elderly disorders more often believed to occur by chance due to their high prevalence, may now actually have a more pathological significance. The following review discusses some of this evidence and the implications for cognitive decline and the development of AD and how a well-known cardiovascular risk factor gene, the apolipoprotein E (APOE) gene, plays a significant role in the molecular genetics of AD. It also introduces and discusses recent and compelling evidence for the involvement of another well-known cardiovascular risk factor gene, the angiotensin-converting enzyme (ACE1) gene, in the pathogenesis of AD. This role is suggested in terms of recent molecular genetic association evidence implicating the ACE1 insertion/deletion (indel) polymorphism, a more recent large haplotype study that greatly extends the ACE1 indel evidence and incorporates knowledge accrued from previous cardiovascular disease-focused ACE1 haplotype studies. Finally, this paper discusses very recent biological evidence that further supports a role for ACE1 and hypothesises a number of readily testable mechanisms by which the ACE1 enzyme and other components of the renin-angiotensin-aldosterone system may be implicated in increased risk and/or the progression of AD.

JRAAS 2003;4:80-93.

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TOPICEDITORIAL REVIEWImprovement of impaired coronary vasodilator reserve in hypertensive patients by low-dose ACE inhibitor/diuretic therapy: a pilot PET study
Jean-Jacques Mourad, Olivier Hanon, Jean-Robert Deverre, Paolo G Camici, Philippe Sellier, Denis Duboc, Michel E Safar

JRAAS 2003;4:94-95.

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TOPICEDITORIAL REVIEWDual blockade with candesartan cilexetil and lisinopril in hypertensive patients with diabetes mellitus: rationale and design
Niels H Andersen, Søren T Knudsen, Per L Poulsen, Steen H Poulsen, Kjeld Helleberg, Hans Eiskjær, Klaus W Hansen, Toke Bek, Carl E Mogensen

Background: Blood pressure (BP) reduction is the key to risk reduction of cardiovascular disease or renal failure in hypertensive patients with diabetes mellitus. Inhibition of the renin-angiotensin system by an angiotensin-converting enzyme (ACE) inhibitor or an angiotensin II receptor blocker (ARB) provides efficient BP reduction and renal protection in hypertensive diabetes patients. But, despite this, the recommended BP levels can be difficult to achieve and dual blockade therapy might be a possible way of obtaining efficient BP reduction in hypertensive patients with diabetes.
Dual blockade treatment is based on a principle of obtaining the broadest and most efficient blockade of angiotensin II, by using the combination of an ACE-inhibitor and an ARB.
Methods: The Candesartan And Lisinopril Microalbuminuria (CALM II) study is a one centre, one observer, double-blind, randomised, active-controlled, parallel-group study, investigating the efficacy and tolerability of candesartan cilexetil in combination with lisinopril, compared with the maximum recommended dose of lisinopril in hypertensive patients with diabetes mellitus.
The study design consists of two treatment arms with either 16 mg candesartan cilexetil or 20 mg lisinopril added to concomitant treatment with 20 mg lisinopril. It comprises 80 patients with a minimum of 35 patients in each group and statistical power of 90% to detect a difference in systolic BP reduction of 6.5 mmHg.
Conclusion: The CALM II study aims to investigate the effects of dual blockade on systolic BP, albuminuria, left ventricular mass and function, and retinopathy in hypertensive patients with diabetes mellitus.

JRAAS 2003;4:96-99.