<em>Angiotensin</em>-(<em>1</em>-<em>7</em>), an active fragment of both <em>angiotensins</em> I and II, generally opposes the vascular and proliferative actions of <em>angiotensin</em> II. Here we evaluated effects of the <em>angiotensin</em>-(<em>1</em>-<em>7</em>) receptor Mas on renal physiology and morphology using Mas-knockout mice. Compared to the wild-type animals, Mas knockout mice had significant reductions in urine volume and fractional sodium excretion without any significant change in free-water clearance. A significantly higher inulin clearance and microalbuminuria concomitant with a reduced renal blood flow suggest that glomerular hyperfiltration occurs in the knockout mice. Histological analysis found reduced glomerular tuft diameter and increased expression of collagen IV and fibronectin in the both the mesangium and interstitium, along with increased collagen III in the interstitium. These fibrogenic changes and the renal dysfunction of the knockout mice were associated with an upregulation of <em>angiotensin</em> II AT<em>1</em> receptor and transforming growth factor-beta mRNA. Our study suggests that Mas acts as a critical regulator of renal fibrogenesis by controlling effects transduced through <em>angiotensin</em> II AT<em>1</em> receptors in the kidney.

Recent studies have linked fetal exposure to a suboptimal intrauterine environment with adult hypertension. The aims of the present study were to see whether prenatal dexamethasone administered intravenously to the ewe between 26 to 28 days of gestation (<em>1</em>) resulted in high blood pressure in male and female offspring and whether hypertension in males was modulated by testosterone status, and (2) altered gene expression for <em>angiotensin</em>ogen and <em>angiotensin</em> type <em>1</em> (AT<em>1</em>) receptors in the brain in late gestation and in the adult. Basal mean arterial pressure (MAP) at 2 years of age was significantly higher in wethers exposed to prenatal dexamethasone (group D; <em>1</em>06+/-5 mm Hg, n=9) compared with the control group (group S; 9<em>1</em>+/-3 mm Hg, n=8; P<0.0<em>1</em>). Infusion of testosterone for 3 weeks had no effect on MAP in either treatment group. At <em>1</em>30 days of gestation, dexamethasone administered between 26 to 28 days of gestation (group DF; n=8), resulted in an increased expression of <em>angiotensin</em>ogen in hypothalamus (in arbitrary units: 2.5+/-0.3 versus <em>1</em>.3+/-0.3 in the saline group [group SF], n=<em>1</em>0; P<0.05). In addition, there was higher expression of the AT<em>1</em> receptors in medulla oblongata in group DF (2.6+/-0.6 versus <em>1</em>.<em>1</em>+/-0.2 in group SF; P<0.0<em>1</em>). This effect of prenatal dexamethasone treatment was still evident in females at <em>7</em> years of age (group DA; n=5; 2.6+/-0.5 versus <em>1</em>.<em>1</em>+/-0.2 in group SA; n=6, P<0.05). In conclusion, brief prenatal exposure of the pregnant ewe to dexamethasone leads to hypertension in adult animals of both sexes. Most interestingly, the mechanism leading to programming of hypertension might be linked with the brain <em>angiotensin</em> system.

Beyond its antidiabetic activity justifying its use in the treatment of the type 2 diabetes, metformin (MET [dimethylguanidine, Glucophage]) has been shown to exhibit antioxidant properties in vitro, which could contribute to limit the deleterious vascular complications of diabetes. We investigated whether MET, at the pharmacological level of <em>1</em>0 -5 mol/L, was able to modulate intracellular production of reactive oxygen species (ROS) both in quiescent bovine aortic endothelial cells (BAECs) and in BAECs stimulated by a short incubation with high levels of glucose (30 mmol/L, 2 hours) or <em>angiotensin</em> II (<em>1</em>0 -<em>7</em> mol/L, <em>1</em> hour). Intracellular ROS production was measured by fluorescence of the DCF (2,<em>7</em>-dichlorodihydrofluorescein) probe. Our results showed that MET was able to reduce the intracellular production of ROS in both nonstimulated BAECs (-20%, P < .05) and BAEC stimulated by high levels of glucose or <em>angiotensin</em> II (-28% and -<em>7</em>2%, respectively, P < .0<em>1</em>). Experiments performed in the presence of the nicotinamide adenine dinucleotide phosphate [NAD(P)H] oxidase inhibitor apocynin or the respiratory mitochondrial chain inhibitor rotenone indicated that MET exerted its effect partly through an inhibition of the formation of ROS produced mainly by NAD(P)H oxidase and also, to a lesser extent, by the respiratory mitochondrial chain.

The renin-<em>angiotensin</em>-aldosterone system contributes to cardiac remodeling, hypertrophy, and left ventricular dysfunction. <em>Angiotensin</em> II and aldosterone (corticosterone in rodents) together generate reactive oxygen species (ROS) via reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, which likely facilitate this hypertrophy and remodeling. This investigation sought to determine whether cardiac oxidative stress and cellular remodeling could be attenuated by in vivo mineralocorticoid receptor (MR) blockade in a rodent model of the chronically elevated tissue renin-<em>angiotensin</em>-aldosterone system, the transgenic TG (mRen2) 2<em>7</em> rat (Ren2). The Ren2 overexpresses the mouse renin transgene with resultant hypertension, insulin resistance, proteinuria, and cardiovascular damage. Young (6- to <em>7</em>-wk-old) male Ren2 and age-matched Sprague-Dawley rats were treated with spironolactone or placebo for 3 wk. Heart tissue ROS, immunohistochemical analysis of 3-nitrotyrosine, and NADPH oxidase (NOX) subunits (gp9<em>1</em>(phox) recently renamed NOX2, p22(phox), Rac<em>1</em>, NOX<em>1</em>, and NOX4) were measured. Structural changes were assessed with cine-magnetic resonance imaging, transmission electron microscopy, and light microscopy. Significant increases in Ren2 septal wall thickness (cine-magnetic resonance imaging) were accompanied by perivascular fibrosis, increased mitochondria, and other ultrastructural changes visible by light microscopy and transmission electron microscopy. Although there was no significant reduction in systolic blood pressure, significant improvements were seen with MR blockade on ROS formation and NOX subunits (each P < 0.05). Collectively, these data suggest that MR blockade, independent of systolic blood pressure reduction, improves cardiac oxidative stress-induced structural and functional changes, which are driven, in part, by <em>angiotensin</em> type <em>1</em> receptor-mediated increases in NOX.

Although renin, the rate-limiting enzyme of the renin-<em>angiotensin</em> system (RAS), was first discovered by Robert Tigerstedt and Bergman more than a century ago, the research on the RAS still remains stronger than ever. The RAS, once considered to be an endocrine system, is now widely recognized as dual (circulating and local/tissue) or multiple hormonal systems (endocrine, paracrine and intracrine). In addition to the classical renin/<em>angiotensin</em> I-converting enzyme (ACE)/<em>angiotensin</em> II (Ang II)/Ang II receptor (AT₁/AT₂) axis, the prorenin/(Pro)renin receptor (PRR)/MAP kinase axis, the ACE2/Ang (<em>1</em>-<em>7</em>)/Mas receptor axis, and the Ang IV/AT₄/insulin-regulated aminopeptidase (IRAP) axis have recently been discovered. Furthermore, the roles of the evolving RAS have been extended far beyond blood pressure control, aldosterone synthesis, and body fluid and electrolyte homeostasis. Indeed, novel actions and underlying signaling mechanisms for each member of the RAS in physiology and diseases are continuously uncovered. However, many challenges still remain in the RAS research field despite of more than one century's research effort. It is expected that the research on the expanded RAS will continue to play a prominent role in cardiovascular, renal and hypertension research. The purpose of this article is to review the progress recently being made in the RAS research, with special emphasis on the local RAS in the kidney and the newly discovered prorenin/PRR/MAP kinase axis, the ACE2/Ang (<em>1</em>-<em>7</em>)/Mas receptor axis, the Ang IV/AT₄/IRAP axis, and intracrine/intracellular Ang II. The improved knowledge of the expanded RAS will help us better understand how the classical renin/ACE/Ang II/AT₁ receptor axis, extracellular and/or intracellular origin, interacts with other novel RAS axes to regulate blood pressure and cardiovascular and kidney function in both physiological and diseased states.

The renin-<em>angiotensin</em>-system (RAS) constitutes an important hormonal system in the physiological regulation of blood pressure. Indeed, dysregulation of the RAS may lead to the development of cardiovascular pathologies including kidney injury. Moreover, the blockade of this system by the inhibition of <em>angiotensin</em> converting enzyme (ACE) or antagonism of the <em>angiotensin</em> type <em>1</em> receptor (AT<em>1</em>R) constitutes an effective therapeutic regimen. It is now apparent with the identification of multiple components of the RAS that the system is comprised of different <em>angiotensin</em> peptides with diverse biological actions mediated by distinct receptor subtypes. The classic RAS can be defined as the ACE-Ang II-AT<em>1</em>R axis that promotes vasoconstriction, sodium retention, and other mechanisms to maintain blood pressure, as well as increased oxidative stress, fibrosis, cellular growth, and inflammation in pathological conditions. In contrast, the non-classical RAS composed of the ACE2-Ang-(<em>1</em>-<em>7</em>)-Mas receptor axis generally opposes the actions of a stimulated Ang II-AT<em>1</em>R axis through an increase in nitric oxide and prostaglandins and mediates vasodilation, natriuresis, diuresis, and oxidative stress. Thus, a reduced tone of the Ang-(<em>1</em>-<em>7</em>) system may contribute to these pathologies as well. Moreover, the non-classical RAS components may contribute to the effects of therapeutic blockade of the classical system to reduce blood pressure and attenuate various indices of renal injury. The review considers recent studies on the ACE2-Ang-(<em>1</em>-<em>7</em>)-Mas receptor axis regarding the precursor for Ang-(<em>1</em>-<em>7</em>), the intracellular expression and sex differences of this system, as well as an emerging role of the Ang<em>1</em>-(<em>1</em>-<em>7</em>) pathway in fetal programing events and cardiovascular dysfunction.

The aim of the present study was to test the hypothesis that the activation of the <em>angiotensin</em>-converting enzyme (ACE)2/<em>angiotensin</em>-(<em>1</em>-<em>7</em>)/Mas receptor axis by use of a novel ACE2 activator (XNT) would protect against thrombosis. Thrombi were induced in the vena cava of spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) rats, and ACE2 and ACE activity in the thrombus was determined. Real-time thrombus formation was viewed through intravital microscopy of vessels in nude mice. Thrombus weight was 40% greater in the SHR (4.99 +/- 0.39 versus <em>7</em>.04 +/- 0.66 mg). This weight increase was associated with a 20% decrease in ACE2 activity in the thrombus. In contrast, there were no differences between the WKY and SHR in ACE2 protein and ACE activity in the thrombi. ACE2 inhibition (DX600; 0.<em>1</em> micromol/L/kg) increased thrombus weight by 30% and XNT treatment (<em>1</em>0 mg/kg) resulted in a 30% attenuation of thrombus formation in the SHR. Moreover, XNT reduced platelet attachment to injured vessels, reduced thrombus size, and prolonged the time for complete vessel occlusion in mice. Thus, a decrease in thrombus ACE2 activity is associated with increased thrombus formation in SHR. Furthermore, ACE2 activation attenuates thrombus formation and reduces platelet attachment to vessels. These results suggest that ACE2 could be a novel target for the treatment of thrombogenic diseases.

Activation of <em>angiotensin</em>-converting enzyme 2 (ACE2), production of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] and stimulation of the Ang-(<em>1</em>-<em>7</em>) receptor Mas exert beneficial actions in various peripheral cardiovascular diseases, largely through opposition of the deleterious effects of <em>angiotensin</em> II via its type <em>1</em> receptor. Here we considered the possibility that Ang-(<em>1</em>-<em>7</em>) may exert beneficial effects against CNS damage and neurological deficits produced by cerebral ischaemic stroke. We determined the effects of central administration of Ang-(<em>1</em>-<em>7</em>) or pharmacological activation of ACE2 on the cerebral damage and behavioural deficits elicited by endothelin-<em>1</em> (ET-<em>1</em>)-induced middle cerebral artery occlusion (MCAO), a model of cerebral ischaemia. The results of the present study demonstrated that intracerebroventricular infusion of either Ang-(<em>1</em>-<em>7</em>) or an ACE2 activator, diminazine aceturate (DIZE), prior to and following ET-<em>1</em>-induced MCAO significantly attenuated the cerebral infarct size and neurological deficits measured <em>7</em>2 h after the insult. These beneficial actions of Ang-(<em>1</em>-<em>7</em>) and DIZE were reversed by co-intracerebroventricular administration of the Mas receptor inhibitor, A-<em>7</em><em>7</em>9. Neither the Ang-(<em>1</em>-<em>7</em>) nor the DIZE treatments altered the reduction in cerebral blood flow elicited by ET-<em>1</em>. Lastly, intracerebroventricular administration of Ang-(<em>1</em>-<em>7</em>) significantly reduced the increase in inducible nitric oxide synthase mRNA expression within the cerebral infarct that occurs following ET-<em>1</em>-induced MCAO. This is the first demonstration of cerebroprotective properties of the ACE2-Ang-(<em>1</em>-<em>7</em>)-Mas axis during ischaemic stroke, and suggests that the mechanism of the Ang-(<em>1</em>-<em>7</em>) protective action includes blunting of inducible nitric oxide synthase expression.

We tested the hypothesis that the renin inhibitor aliskiren ameliorates organ damage in rats transgenic for human renin and <em>angiotensin</em>ogen genes (double transgenic rat [dTGR]). Six-week-old dTGR were matched by albuminuria (2 mg per day) and divided into 5 groups. Untreated dTGR were compared with aliskiren (3 and 0.3 mg/kg per day)-treated and valsartan (Val; <em>1</em>0 and <em>1</em> mg/kg per day)-treated rats. Treatment was from week 6 through week 9. At week 6, all groups had elevated systolic blood pressure (BP). Untreated dTGR showed increased BP (202+/-4 mm Hg), serum creatinine, and albuminuria (34+/-5.<em>7</em> mg per day) at week <em>7</em>. At week 9, both doses of aliskiren lowered BP (<em>1</em><em>1</em>5+/-6 and <em>1</em>39+/-5 mm Hg) and albuminuria (0.4+/-0.<em>1</em> and <em>1</em>.6+/-0.6 mg per day) and normalized serum creatinine. Although high-dose Val lowered BP (<em>1</em>48+/-4 mm Hg) and albuminuria (2.<em>1</em>+/-0.<em>7</em> mg per day), low-dose Val reduced BP (<em>1</em>82+/-3 mm Hg) and albuminuria (24+/-3.8 mg per day) to a lesser extent. Mortality was <em>1</em>00% in untreated dTGR and 26% in Val (<em>1</em> mg/kg per day) treated rats, whereas in all other groups, survival was <em>1</em>00%. dTGR treated with low-dose Val had cardiac hypertrophy (4.4+/-0.<em>1</em> mg/g), increased left ventricular (LV) wall thickness, and diastolic dysfunction. LV atrial natriuretic peptide and beta-myosin heavy chain mRNA, albuminuria, fibrosis, and cell infiltration were also increased. In contrast, both aliskiren doses and the high-dose Val lowered BP to a similar extent and more effectively than low-dose Val. We conclude that in dTGR, equieffective antihypertensive doses of Val or aliskiren attenuated end-organ damage. Thus, renin inhibition compares favorably to <em>angiotensin</em> receptor blockade in reversing organ damage in dTGR.

We tested the hypothesis that superoxide anion (O(2)(-).) generated in the kidney by prolonged <em>angiotensin</em> II (ANG II) reduces renal cortical Po(2) and the use of O(2) for tubular sodium transport (T(Na):Q(O(2))). Groups (n = 8-<em>1</em><em>1</em>) of rats received <em>angiotensin</em> II (ANG II, 200 ng.kg(-<em>1</em>).min(-<em>1</em>) sc) or vehicle for 2 wk with concurrent infusions of a permeant nitroxide SOD mimetic 4-hydroxy-2,2,6,6-tetramethylpiperidine <em>1</em>-oxyl (Tempol, 200 nmol.kg(-<em>1</em>).min(-<em>1</em>)) or vehicle. Rats were studied under anesthesia with measurements of renal oxygen usage and Po(2) in the cortex and tubules with a glass electrode. Compared with vehicle, ANG II increased mean arterial pressure (<em>1</em>0<em>7</em> +/- 4 vs. <em>1</em>46 +/- 6 mmHg; P < 0.00<em>1</em>), renal vascular resistance (42 +/- 3 vs. 65 +/- <em>7</em> mmHg.ml(-<em>1</em>).min(-<em>1</em>).<em>1</em>00 g(-<em>1</em>); P < 0.00<em>1</em>), renal cortical NADPH oxidase activity (2.3 +/- 0.2 vs. 3.6 +/- 0.4 nmol O(2)(-)..min(-<em>1</em>).mg(-<em>1</em>) protein; P < 0.05), mRNA and protein expression for p22(phox) (2.<em>1</em>- and <em>1</em>.8-fold respectively; P < 0.05) and reduced the mRNA for extracellular (EC)-SOD (-<em>1</em>.8 fold; P < 0.05). ANG II reduced the Po(2) in the proximal tubule (39 +/- <em>1</em> vs. 34 +/- 2 mmHg; P < 0.05) and throughout the cortex and reduced the T(Na):Q(O(2)) (<em>1</em><em>7</em> +/- <em>1</em> vs. 9 +/- 2 mumol/mumol; P < 0.00<em>1</em>). Tempol blunted or prevented all these effects of ANG II. The effects of prolonged ANG II to cause hypertension, renal vasoconstriction, renal cortical hypoxia, and reduced efficiency of O(2) usage for Na(+) transport, activation of NADPH oxidase, increased expression of p22(phox), and reduced expression of EC-SOD can be ascribed to O(2)(-). generation because they are prevented by an SOD mimetic.

The discovery of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] as a bioactive Ang II fragment of the renin-<em>angiotensin</em> system (RAS) alters the current understanding of the enzymatic components that comprise the RAS cascade. Two neutral endopeptidases, prolyl endopeptidase (E.C. 3.4.2<em>1</em>.26) and neutral endopeptidase 24.<em>1</em><em>1</em> (E.C. 3.4.24.<em>1</em><em>1</em>), are capable of forming Ang-(<em>1</em>-<em>7</em>) from Ang I and have been implicated in the in vivo processing of Ang I. This makes them putative Ang processing enzymes and part of the RAS cascade. This review summarizes the physical characteristics and distribution of <em>angiotensin</em> converting enzyme (E.C. 3.4.<em>1</em>5.<em>1</em>), a known Ang I processing enzyme, and compares its features to what is known of prolyl endopeptidase and neutral endopeptidase 24.<em>1</em><em>1</em>.

Premenopausal women are protected to some extent from cardiovascular and kidney diseases. Because this protection weakens after menopause, sex hormones are believed to play an important role in the pathogenesis of cardiovascular and kidney diseases. The cardiovascular system and the kidneys are regulated by the renin-<em>angiotensin</em>-aldosterone system (RAAS), which in turn, appears to be regulated by sex hormones. In general, oestrogen increases <em>angiotensin</em>ogen levels and decreases renin levels, <em>angiotensin</em>-converting enzyme (ACE) activity, AT(<em>1</em>) receptor density, and aldosterone production. Oestrogen also activates counterparts of the RAAS such as natriuretic peptides, AT(2) receptor density, and <em>angiotensin</em>ogen (<em>1</em>-<em>7</em>). Progesterone competes with aldosterone for mineralocorticoid receptor. Less is known about androgens, but testosterone seems to increase renin levels and ACE activity. These effects of sex hormones on the RAAS can explain at least some of the gender differences in cardiovascular and kidney diseases.

BACKGROUND

Overactivity of the brain renin-<em>angiotensin</em> system is a major contributor to neurogenic hypertension. Although overexpression of <em>angiotensin</em>-converting enzyme type 2 (ACE2) has been shown to be beneficial in reducing hypertension by transforming <em>angiotensin</em> II into <em>angiotensin</em>-(<em>1</em>-<em>7</em>), several groups have reported decreased brain ACE2 expression and activity during the development of hypertension.

OBJECTIVE

We hypothesized that ADAM<em>1</em><em>7</em>-mediated ACE2 shedding results in decreased membrane-bound ACE2 in the brain, thus promoting the development of neurogenic hypertension.

RESULTS

To test this hypothesis, we used the deoxycorticosterone acetate-salt model of neurogenic hypertension in nontransgenic and syn-hACE2 mice overexpressing ACE2 in neurons. Deoxycorticosterone acetate-salt treatment in nontransgenic mice led to significant increases in blood pressure, hypothalamic <em>angiotensin</em> II levels, inflammation, impaired baroreflex sensitivity, and autonomic dysfunction, as well as decreased hypothalamic ACE2 activity and expression, although these changes were blunted or prevented in syn-hACE2 mice. In addition, reduction of ACE2 expression and activity in the brain paralleled an increase in ACE2 activity in the cerebrospinal fluid of nontransgenic mice after deoxycorticosterone acetate-salt treatment and were accompanied by enhanced ADAM<em>1</em><em>7</em> expression and activity in the hypothalamus. Chronic knockdown of ADAM<em>1</em><em>7</em> in the brain blunted the development of hypertension and restored ACE2 activity and baroreflex function.

CONCLUSIONS

Our data provide the first evidence that ADAM<em>1</em><em>7</em>-mediated shedding impairs brain ACE2 compensatory activity, thus contributing to the development of neurogenic hypertension.

OBJECTIVE

The benefit of using a renin-angiotensin-aldosterone system blocker such as an angiotensin-converting enzyme inhibitor (ACEI) or an angiotensin II receptor blocker (ARB) for patients with advanced chronic kidney disease (CKD) remains undetermined.

OBJECTIVE

To assess the effectiveness and safety of ACEI/ARB use for advanced predialysis CKD in patients with hypertension and anemia. DESIGN Prospective cohort study.

METHODS

Taiwan.

METHODS

From January 1, 2000, through June 30, 2009, we selected 28 497 hypertensive adult patients with CKD. Serum creatinine levels were greater than 6 mg/dL, hematocrit levels were less than 28%, and patients were treated with erythropoiesis-stimulating agents.

METHODS

Users (n = 14,117) and nonusers (n = 14,380) of ACEIs/ARBs.

METHODS

We used Cox proportional hazards regression models to estimate hazard ratios (HRs) for commencement of long-term dialysis and all-cause mortality for ACRI/ARB users vs nonusers.

RESULTS

In a median follow-up of 7 months, 20,152 patients (70.7%) required long-term dialysis and 5696 (20.0%) died before progression to end-stage renal disease requiring dialysis. Use of ACEIs/ARBs was associated with a lower risk for long-term dialysis (HR, 0.94 [95% CI, 0.91-0.97]) and the composite outcome of long-term dialysis or death (0.94 [0.92-0.97]). The renal benefit of ACEI/ARB use was consistent across most patient subgroups, as was that of ACEI or ARB monotherapy. Compared with nonusers, the ACEI/ARB users had a higher hyperkalemia-associated hospitalization rate, but the risk of predialysis mortality caused by hyperkalemia was not significantly increased (HR, 1.03 [95% CI, 0.92-1.16]; P = .30).

CONCLUSIONS

Patients with stable hypertension and advanced CKD who receive therapy with ACEIs/ARBs exhibit an association with lower risk for long-term dialysis or death by 6%. This benefit does not increase the risk of all-cause mortality.

BACKGROUND

Thioredoxin (Trx)<em>1</em> inhibits pathological cardiac hypertrophy. MicroRNAs (miRNAs) are small noncoding RNAs that downregulate posttranscriptional expression of target molecules.

OBJECTIVE

We investigated the role of miRNAs in mediating the antihypertrophic effect of Trx<em>1</em> on angiotensin II (Ang II)-induced cardiac hypertrophy.

RESULTS

Microarray analyses of mature rodent microRNAs and quantitative RT-PCR/Northern blot analyses showed that Trx<em>1</em> upregulates members of the let-7 family, including miR-98, in the heart and the cardiomyocytes therein. Adenovirus-mediated expression of miR-98 in cardiomyocytes reduced cell size both at baseline and in response to Ang II. Knockdown of miR-98, and of other members of the let-7 family, augmented Ang II-induced cardiac hypertrophy, and attenuated Trx<em>1</em>-mediated inhibition of Ang II-induced cardiac hypertrophy, suggesting that endogenous miR-98/let-7 mediates the antihypertrophic effect of Trx<em>1</em>. Cyclin D2 is one of the predicted targets of miR-98. Ang II significantly upregulated cyclin D2, which in turn plays an essential role in mediating Ang II-induced cardiac hypertrophy, whereas overexpression of Trx<em>1</em> inhibited Ang II-induced upregulation of cyclin D2. miR-98 decreased both expression of cyclin D2 and the activity of a cyclin D2 3'UTR luciferase reporter, suggesting that both Trx<em>1</em> and miR-98 negatively regulate cyclin D2. Overexpression of cyclin D2 attenuated the suppression of Ang II-induced cardiac hypertrophy by miR-98, suggesting that the antihypertrophic actions of miR-98 are mediated in part by downregulation of cyclin D2.

CONCLUSIONS

These results suggest that Trx<em>1</em> upregulates expression of the let-7 family, including miR-98, which in turn inhibits cardiac hypertrophy, in part through downregulation of cyclin D2.

Docosahexaenoic acid (DHA), a peroxisome proliferator-activated receptor-alpha (PPARalpha) activator, reduces blood pressure (BP) in some hypertensive models by unclear mechanisms. We tested the hypothesis that DHA would prevent BP elevation and improve vascular dysfunction in <em>angiotensin</em> (Ang) II-infused rats by modulating of NADPH oxidase activity and inflammation in vascular wall. Sprague-Dawley rats received Ang II (<em>1</em>20 ng/kg per minute SC) with or without DHA (2.5 mL of oil containing 40% DHA/d PO) for <em>7</em> days. Systolic BP (mm Hg), elevated in Ang II-infused rats (<em>1</em><em>7</em>2+/-3) versus controls (<em>1</em>08+/-2, P<0.0<em>1</em>), was reduced by DHA (<em>1</em><em>1</em>2+/-4). In mesenteric small arteries studied in a pressurized myograph, media/lumen ratio was increased (P<0.05) and acetylcholine-induced relaxation impaired in Ang II-infused rats (P<0.05); both were normalized by DHA. In blood vessels of Ang II-infused rats, NADPH oxidase activity measured by chemiluminescence and expression of adhesion molecules intercellular adhesion molecule and vascular cell adhesion molecule-<em>1</em> were significantly increased. These changes were abrogated by DHA. PPARalpha activator DHA attenuated the development of hypertension, corrected structural abnormalities, and improved endothelial dysfunction induced by Ang II. These effects are associated with decreased oxidative stress and inflammation in the vascular wall.

The renin-<em>angiotensin</em> system (RAS), critically involved in the control of blood pressure and volume homeostasis, is a dual system comprising a circulating component and a local tissue component. The rate limiting enzyme is renin, which in the circulating RAS derives from the kidney to generate Ang II, which in turn regulates cardiovascular function by binding to AT(<em>1</em>) and AT(2) receptors on cardiac, renal and vascular cells. The tissue RAS can operate independently of the circulating RAS and may be activated even when the circulating RAS is suppressed or normal. A functional tissue RAS has been identified in brain, kidney, heart, adipose tissue, hematopoietic tissue, gastrointestinal tract, liver, endocrine system and blood vessels. Whereas <em>angiotensin</em>sinogen, <em>angiotensin</em> converting enzyme (ACE), Ang I and Ang II are synthesized within these tissues, there is still controversy as to whether renin is produced locally or whether it is taken up from the circulation, possibly by the (pro)renin receptor. This is particularly true in the vascular wall, where expression of renin is very low. The exact function of the vascular RAS remains elusive, but may contribute to fine-tuning of vascular tone and arterial structure and may amplify vascular effects of the circulating RAS, particularly in pathological conditions, such as in hypertension, atherosclerosis and diabetes. New concepts relating to the vascular RAS have recently been elucidated including: (<em>1</em>) the presence of functionally active Ang-(<em>1</em>-<em>7</em>)-Mas axis in the vascular system, (2) the importance of the RAS in perivascular adipose tissue and cross talk with vessels, and (3) the contribution to vascular RAS of Ang II derived from immune and inflammatory cells within the vascular wall. The present review highlights recent progress in the RAS field, focusing on the tissue system and particularly on the vascular RAS.

Cardiac remodeling is a hallmark hypertension-induced pathophysiology. In the current study, the role of the <em>angiotensin</em>-(<em>1</em>-<em>7</em>) fragment in modulating cardiac remodeling was examined. Sprague-Dawley rats underwent uninephrectomy surgery and were implanted with a deoxycorticosterone acetate (DOCA) pellet. DOCA animals had their drinking water replaced with 0.9% saline solution. A subgroup of DOCA-salt animals was implanted with osmotic minipumps, which delivered <em>angiotensin</em>-(<em>1</em>-<em>7</em>) chronically (<em>1</em>00 ng.kg(-<em>1</em>).min(-<em>1</em>)). Control animals underwent sham surgery and were maintained on normal drinking water. Blood pressure was measured weekly with the use of the tail-cuff method, and after 4 wk of treatment, blood pressure responses to graded doses of <em>angiotensin</em> II were determined by direct carotid artery cannulation. Ventricle size was measured, and cross sections of the heart ventricles were paraffin embedded and stained using Masson's Trichrome to measure interstitial and perivascular collagen deposition and myocyte diameter. DOCA-salt treatment caused significant increases in blood pressure, cardiac hypertrophy, and myocardial and perivascular fibrosis. <em>Angiotensin</em>-(<em>1</em>-<em>7</em>) infusion prevented the collagen deposition effects without any effect on blood pressure or cardiac hypertrophy. These results indicate that <em>angiotensin</em>-(<em>1</em>-<em>7</em>) selectively prevents cardiac fibrosis independent of blood pressure or cardiac hypertrophy in the DOCA-salt model of hypertension.

Since <em>angiotensin</em>-(<em>1</em>-<em>1</em>2) [Ang-(<em>1</em>-<em>1</em>2)] is a non-renin dependent alternate precursor for the generation of cardiac Ang peptides in rat tissue, we investigated the metabolism of Ang-(<em>1</em>-<em>1</em>2) by plasma membranes (PM) isolated from human atrial appendage tissue from nine patients undergoing cardiac surgery for primary control of atrial fibrillation (MAZE surgical procedure). PM was incubated with highly purified ¹²⁵I-Ang-(<em>1</em>-<em>1</em>2) at 3<em>7</em>°C for <em>1</em> h with or without renin-<em>angiotensin</em> system (RAS) inhibitors [lisinopril for <em>angiotensin</em> converting enzyme (ACE), SCH393<em>7</em>0 for neprilysin (NEP), MLN-4<em>7</em>60 for ACE2 and chymostatin for chymase; 50 µM each]. ¹²⁵I-Ang peptide fractions were identified by HPLC coupled to an inline γ-detector. In the absence of all RAS inhibitor, ¹²⁵I-Ang-(<em>1</em>-<em>1</em>2) was converted into Ang I (2±2%), Ang II (69±2<em>1</em>%), Ang-(<em>1</em>-<em>7</em>) (5±2%), and Ang-(<em>1</em>-4) (2±<em>1</em>%). In the absence of all RAS inhibitor, only 22±<em>1</em>0% of ¹²⁵I-Ang-(<em>1</em>-<em>1</em>2) was unmetabolized, whereas, in the presence of the all RAS inhibitors, 98±<em>7</em>% of ¹²⁵I-Ang-(<em>1</em>-<em>1</em>2) remained intact. The relative contribution of selective inhibition of ACE and chymase enzyme showed that ¹²⁵I-Ang-(<em>1</em>-<em>1</em>2) was primarily converted into Ang II (65±<em>1</em>8%) by chymase while its hydrolysis into Ang II by ACE was significantly lower or undetectable. The activity of individual enzyme was calculated based on the amount of Ang II formation. These results showed very high chymase-mediated Ang II formation (28±3.<em>1</em> fmol × min⁻¹ × mg⁻¹, n = 9) from ¹²⁵I-Ang-(<em>1</em>-<em>1</em>2) and very low or undetectable Ang II formation by ACE (<em>1</em>.<em>1</em>±0.2 fmol×min⁻¹ × mg⁻¹). Paralleling these findings, these tissues showed significant content of chymase protein that by immunocytochemistry were primarily localized in atrial cardiac myocytes. In conclusion, we demonstrate for the first time in human cardiac tissue a dominant role of cardiac chymase in the formation of Ang II from Ang-(<em>1</em>-<em>1</em>2).

Our studies in the mRen2.Lewis female rat, an <em>angiotensin</em> II- and estrogen-dependent model of hypertension, revealed that chronic activation of estrogen receptor GPR30 markedly reduces blood pressure in ovariectomized females. The present studies measured acute vasodilation to the selective GPR30 agonist G-<em>1</em> and <em>1</em><em>7</em>-β-estradiol (<em>1</em>0(-9)-<em>1</em>0(-5.5) M) in isolated aortic rings and mesenteric arteries from intact mRen2.Lewis females. Maximal relaxation was greater in mesenteric vessels versus the aorta for both G-<em>1</em> (4<em>7</em>% ± 8% vs 80% ± 5% of phenylephrine preconstriction, P < 0.00<em>1</em>) and estradiol (42% ± <em>7</em>% vs 83% ± 4% of phenylephrine preconstriction, P < 0.00<em>1</em>). The GPR30 antagonist G<em>1</em>5 attenuated the response to both estradiol and G-<em>1</em>. Removal of the endothelium or pretreatment with Nitro-L-arginine methyl ester (L-NAME) partially attenuated vasorelaxation. Responses were not altered in mesenteric vessels from ovariectomized females. Immunohistochemical analysis revealed GPR30 expression in mesenteric endothelial and smooth muscle cells, and smooth muscle expression was confirmed in cultured cells. We conclude that estradiol-induced relaxation in conduit and resistance vessels from mRen2.Lewis females may be mediated by the novel estrogen receptor GPR30. The direct vasodilatory response of G-<em>1</em> in resistance vessels presents one mechanism for the reduction in blood pressure induced by chronic G-<em>1</em> administration.

We examined the influence of chronic treatment with ANG-(<em>1</em>-<em>7</em>) on development of hypertension and end-organ damage in spontaneously hypertensive rats (SHR) chronically treated with the nitric oxide synthesis inhibitor L-NAME (SHR-L-NAME). L-NAME administered orally (80 mg/l) for 4 wk significantly elevated mean arterial pressure (MAP) compared with SHR controls drinking regular water (269 +/- <em>1</em>0 vs. <em>1</em>96 +/- 6 mmHg). ANG-(<em>1</em>-<em>7</em>) (24 microg x kg(-<em>1</em>) x h(-<em>1</em>)) or captopril (300 mg/l) significantly attenuated the elevation in MAP due to L-NAME (2<em>1</em>3 +/- <em>7</em> and 228 +/- 8 mmHg, respectively), and ANG-(<em>1</em>-<em>7</em>) + captopril completely reversed the L-NAME-dependent increase in MAP (<em>1</em>93 +/- 5 mmHg). L-NAME-induced increases in urinary protein were significantly lower in ANG-(<em>1</em>-<em>7</em>)-treated animals (226 +/- 6 vs. <em>1</em>45 +/- <em>1</em>2 mg/day). Captopril was more effective (96 +/- <em>1</em>2 mg/day), and there was no additional effect of captopril + ANG-(<em>1</em>-<em>7</em>) (8<em>7</em> +/- 5 mg/day). The abnormal vascular responsiveness to endothelin-<em>1</em>, carbachol, and sodium nitroprusside in perfused mesenteric vascular bed of SHR-L-NAME was improved by ANG-(<em>1</em>-<em>7</em>) or captopril, with no additive effect of ANG-(<em>1</em>-<em>7</em>) + captopril. In isolated perfused hearts, recovery of left ventricular function from 40 min of global ischemia was significantly better in ANG-(<em>1</em>-<em>7</em>)- or captopril-treated SHR-L-NAME, with additive effects of combined treatment. The beneficial effects of ANG-(<em>1</em>-<em>7</em>) on MAP and cardiac function were inhibited when indomethacin was administered with ANG-(<em>1</em>-<em>7</em>), but indomethacin did not reverse the protective effects on proteinuria or vascular reactivity. The protective effects of the ANG-(<em>1</em>-<em>7</em>) analog AVE-099<em>1</em> were qualitatively comparable to those of ANG-(<em>1</em>-<em>7</em>) but were not improved over those of captopril alone. Thus, during reduced nitric oxide availability, ANG-(<em>1</em>-<em>7</em>) attenuates development of severe hypertension and end-organ damage; prostaglandins participate in the MAP-lowering and cardioprotective effects of ANG-(<em>1</em>-<em>7</em>); and additive effects of captopril + ANG-(<em>1</em>-<em>7</em>) on MAP, but not proteinuria or endothelial function, suggest common, as well as different, mechanisms of action for the two treatments. Together, the results provide further evidence of a role for ANG-(<em>1</em>-<em>7</em>) in protective effects of <em>angiotensin</em>-converting enzyme inhibition and suggest dissociation of factors influencing MAP and those influencing end-organ damage.

Two of the primary sites of actions for <em>angiotensin</em> (ANG)-(<em>1</em>---<em>7</em>) are the vasculature and the kidney. Because little information exists concerning the metabolism of ANG-(<em>1</em>---<em>7</em>) in these tissues, we investigated the hydrolysis of the peptide in rat lung and renal brush-border membrane (BBM) preparations. Radiolabeled ANG-(<em>1</em>---<em>7</em>) was hydrolyzed primarily to ANG-(<em>1</em>---5) by pulmonary membranes. The ANG-converting enzyme (ACE) inhibitor lisinopril abolished the generation of ANG-(<em>1</em>---5), as well as that of smaller metabolites. Kinetic studies of the hydrolysis of ANG-(<em>1</em>---<em>7</em>) to ANG-(<em>1</em>---5) by somatic (pulmonary) and germinal (testes) forms of rat ACE yielded similar values, suggesting that the COOH-domain is responsible for the hydrolysis of ANG-(<em>1</em>---<em>7</em>). Pulmonary metabolism of ANG-(<em>1</em>---5) yielded ANG-(3---5) and was independent of ACE but may involve peptidyl or dipeptidyl aminopeptidases. In renal cortex BBM, ANG-(<em>1</em>---<em>7</em>) was rapidly hydrolyzed to mono- and dipeptide fragments and ANG-(<em>1</em>---4). Aminopeptidase (AP) inhibition attenuated the hydrolysis of ANG-(<em>1</em>---<em>7</em>) and increased ANG-(<em>1</em>---4) formation. Combined treatment with AP and neprilysin (Nep) inhibitors abolished ANG-(<em>1</em>---4) formation and preserved ANG-(<em>1</em>---<em>7</em>). ACE inhibition had no effect on the rate of hydrolysis or the metabolites formed in the BBM. In conclusion, ACE was the major enzymatic activity responsible for the metabolism of ANG-(<em>1</em>---<em>7</em>) in the lung, which is consistent with the ability of ACE inhibitors to increase the half-life of circulating ANG-(<em>1</em>---<em>7</em>) and raise endogenous levels of the peptide. An alternate pathway of metabolism was revealed in the renal cortex, where increased AP and Nep activities, relative to ACE activity, promote conversion of ANG-(<em>1</em>---<em>7</em>) to ANG-(<em>1</em>---4) and smaller fragments.

BACKGROUND

Recent studies have shown that both steroids and angiotensin-converting enzyme (ACE) inhibitors improve kidney survival and decrease proteinuria in patients with immunoglobulin A nephropathy. In this study, we aim to investigate whether the addition of steroids to ACE-inhibitor therapy produces a more potent antiproteinuric effect and better protection of kidney function than an ACE inhibitor alone.

METHODS

Randomized controlled trial.

METHODS

Patients with biopsy-proven immunoglobulin A nephropathy with proteinuria of 1 to 5 g/d of protein.

METHODS

63 patients were randomly assigned to either cilazapril alone (ACE-inhibitor group; n = 30) or steroid plus cilazapril (combination group; n = 33).

METHODS

The primary end point was kidney survival, defined as a 50% increase in baseline serum creatinine level.

RESULTS

After follow-up for up to 48 months, 7 patients in the ACE-inhibitor group (24.1%) reached the primary end point compared with 1 patient (3%) in the combination group. Kaplan-Meier kidney survival was significantly better in the combination group than the ACE-inhibitor group after 24 and 36 months (96.6% versus 75.7%, 96.6% versus 66.2%; P = 0.001). Urine protein excretion significantly decreased in patients in the combination group compared with the ACE-inhibitor group (time-average proteinuria, 1.04 +/- 0.54 versus 1.57 +/- 0.86 g/d of protein; P = 0.01). Multivariate analysis showed that combination treatment (hazard ratio, 0.1; 95% confidence interval, 0.014 to 0.946) and time-average proteinuria (hazard ratio, 14.3; 95% confidence interval, 2.86 to 71.92) were independent predictors of kidney survival.

CONCLUSIONS

Small sample size, a single center, and slight imbalances at baseline.

CONCLUSIONS

Our results suggest that the addition of steroid to ACE-inhibitor therapy provided additional benefit compared with an ACE inhibitor alone. However, this was a pilot study with a small number of participants achieving the end points, and thus further validation is necessary.