The aim of this study was to evaluate the effects of AVE 099<em>1</em> (AVE), a nonpeptide compound that mimics Ang-(<em>1</em>-<em>7</em>) actions, on cardiac remodeling. Heart hypertrophy and heart dysfunction were induced by isoproterenol (ISO) (2 mg/kg i.p./day for <em>7</em> days) in male Wistar rats. At the end of the <em>7</em>-day period, the hearts were perfused according to the Langendorff method to evaluate cardiac function. The hearts, atria, and right and left ventricles wet weights were recorded, normalized for body weight and then expressed as muscle mass index (mg/g). In addition, serial sections from left ventricle were stained with hematoxylin-eosin for cell morphometry and with collagen-specific Masson's trichrome for detection of fibrosis. Immunofluorescence-labeling and confocal microscopy were used to investigate the distribution and deposition of collagen types I, III, VI, and fibronectin. AVE reduced the ISO-induced hypertrophy as quantified by myocyte diameter measurements (Control: <em>1</em>0.60+/-0.08 microm; ISO: <em>1</em>4.60+/-0.<em>1</em><em>1</em> mum; ISO+AVE: <em>1</em><em>1</em>.22+/-0.08 microm, n = 5). In addition, AVE markedly attenuated the increase of extracellular matrix proteins induced by ISO. AVE treatment also attenuated the decrease in systolic tension and +/-dT/dt and exacerbated the vasodilatation induced by ISO. These results show that AVE has a cardioprotective effect on ISO-induced cardiac remodeling.

Risk factors for progression of kidney disease include hypertension, proteinuria, male sex, obesity, diabetes mellitus, hyperlipidemia, smoking, high-protein diets, phosphate retention, and metabolic acidosis. <em>Angiotensin</em> II production upregulates the expression of transforming growth factor-beta<em>1</em>, tumor necrosis factor-alpha, nuclear factor-kappaB, and several adhesion molecules and chemoattractants. In addition to <em>angiotensin</em>, other vasoactive compounds, such as thromboxane A(2), endothelin, and prostaglandins, are upregulated. Treatment with one of several growth factors may ameliorate the progression of kidney disease: insulin-like growth factor-<em>1</em>, hepatocyte growth factor, and bone morphogenetic protein-<em>7</em>.

OBJECTIVE

This investigation was designed to examine the influence of creatine (Cr) supplementation on acute cardiovascular, renal, temperature, and fluid-regulatory hormonal responses to exercise for 35 min in the heat.

METHODS

Twenty healthy men were matched and then randomly assigned to consume 0.3 g.kg(-<em>1</em>) Cr monohydrate (N = <em>1</em>0) or placebo (N = <em>1</em>0) for <em>7</em> d in a double-blind fashion. Before and after supplementation, both groups cycled for 30 min at 60-<em>7</em>0% VO2(peak) immediately followed by three <em>1</em>0-s sprints in an environmental chamber at 3<em>7</em> degrees C and 80% relative humidity.

RESULTS

Body mass was significantly increased (0.<em>7</em>5 kg) in Cr subjects. Heart rate, blood pressure, and sweat rate responses to exercise were not significantly different between groups. There were no differences in rectal temperature responses in either group. Sodium, potassium, and creatinine excretion rates obtained from 24-h and exercise urine collection periods were not significantly altered in either group. Serum creatinine was elevated in the Cr group but within normal ranges. There were significant exercise-induced increases in cortisol, aldosterone, renin, angiotensin I and II, atrial peptide, and arginine vasopressin. The aldosterone response was slightly greater in the Cr (263%) compared with placebo (224%) group. Peak power was greater in the Cr group during all three <em>1</em>0-s sprints after supplementation and unchanged in the placebo group. There were no reports of adverse symptoms, including muscle cramping during supplementation or exercise.

CONCLUSIONS

Cr supplementation augments repeated sprint cycle performance in the heat without altering thermoregulatory responses.

The novel coronavirus disease 20<em>1</em>9 (COVID-<em>1</em>9) outbreak once again demonstrated the importance of the renin-<em>angiotensin</em> system (RAS) in patients with diabetes. Activation of the RAS increases in patients with diabetes. The virus attaches to the ACE2 enzyme at low cytosolic pH values and enters into the cell and causes infection. Especially in the presence of diabetes mellitus and accompanying comorbid conditions such as hypertension, obesity, old age, and smoking, cytosolic pH is low, thus the virus easily may enter the cell by attaching to ACE2. ACEIs and ARBs lead to a reduction in <em>angiotensin</em> II level by increasing the ACE2 level, thus they cause a low cytosolic pH. Increased cardiac ACE2 levels due to ACEIs and ARBs can trigger cardiac arrhythmias and myocarditis by causing the virus to easily enter the heart tissue. There is ACE2 activity in the rostral ventrolateral medulla in the brain stem. The release of <em>angiotensin</em> <em>1</em>-<em>7</em> in the brain stem leads to the activation of the sympathetic nervous system. This activation causes systemic vasoconstriction and the patient's blood pressure increases. The most important event is the increased sympathetic activity via the central stimulation, this activity increases pulmonary capillary leaking, causing the ARDS. As the cytosolic pH, which is already low in patients with diabetes will decrease further with the mechanisms mentioned above, the viral load will increase and the infection will be exacerbated. As a result, the use of ACEIs and ARBs in patients with diabetes can lead to increased morbidity and mortality of COVID-<em>1</em>9.

OBJECTIVE

Recent studies have shown that the heptapeptide <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] exerts important vasoactive actions and can act as an endogenous physiological antagonist of <em>angiotensin</em> II (Ang II) within the renin-<em>angiotensin</em> system (RAS). The present study was performed to evaluate the effects, first, of chronic increases of Ang-(<em>1</em>-<em>7</em>) levels, second, of [<em>7</em>-D-Ala], an Ang-(<em>1</em>-<em>7</em>) receptor antagonist, and, third, of an <em>angiotensin</em>-converting enzyme 2 (ACE2) inhibitor on the course of hypertension and of renal function of the nonclipped kidney in two-kidney, one-clip (2K<em>1</em>C) Goldblatt hypertensive rats.

METHODS

Blood pressure (BP) was monitored by radiotelemetry. Elevation of the effect of circulating Ang-(<em>1</em>-<em>7</em>) levels was achieved either by chronic subcutaneous infusion of Ang-(<em>1</em>-<em>7</em>) through osmotic minipumps or by employing transgenic rats that express an Ang-(<em>1</em>-<em>7</em>)-producing fusion protein [Ang-(<em>1</em>-<em>7</em>)TGR+/+] (and its control Ang-(<em>1</em>-<em>7</em>)TGR-/-). [<em>7</em>-D-Ala] was also infused subcutaneously and the ACE2 inhibitor was administrated in drinking water. On day 25 after clipping, rats were anesthetized and renal function was evaluated.

RESULTS

Chronic infusion of Ang-(<em>1</em>-<em>7</em>) did not modify the course of 2K<em>1</em>C hypertension and did not alter renal function as compared with saline vehicle-infused 2K<em>1</em>C rats. Chronic infusion of [<em>7</em>-D-Ala] or treatment with the ACE2 inhibitor worsened the course of hypertension and elicited decreases in renal hemodynamics. [Ang-(<em>1</em>-<em>7</em>)TGR+/+] and [Ang-(<em>1</em>-<em>7</em>)TGR-/-] rats exhibited a similar course of hypertension.

CONCLUSIONS

The present data support the notion that Ang-(<em>1</em>-<em>7</em>) serves as an important endogenous vasodilator and natriuretic agent and its deficiency might contribute to the acceleration of 2K<em>1</em>C Goldblatt hypertension.

OBJECTIVE

Kidney fibrosis is a common observation in human and experimental models of kidney disease and contributes to the progressive loss of kidney function. This review discusses the recent recognition of the role of podocytes in the development of common glomerular disease and focuses on the basis for new antifibrotic therapies.

RESULTS

A growing body of evidence indicates that changes in the structure and function of podocytes are involved in the development and progression of kidney disease. The changes include podocyte de-differentiation, podocyte-mediated endothelial dysfunction and podocyte-induced epithelial-mesenchymal transition, all contributing to the development of kidney fibrosis. Furthermore, new antifibrotic strategies aiming at the transforming growth factor-beta, connective tissue growth factor, <em>angiotensin</em> (<em>1</em>-<em>7</em>), and advanced glycation endproducts/receptors advanced glycation endproducts signaling pathways are being developed.

CONCLUSIONS

Podocytes are recognized to play a key role in the development of kidney fibrosis. New antifibrotic therapies are rapidly progressing toward definitive clinical trials but will need to be tested on top of the existing therapy of renin-angiotensin system inhibition. Novel approaches targeting podocyte function would be a promising approach for early stages of the disease.

OBJECTIVE

In the present study, we evaluated the effect of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] and its selective antagonist, D-Ala<em>7</em>-Ang-(<em>1</em>-<em>7</em>) (A-<em>7</em><em>7</em>9), at the nucleus tractus solitarii (nTS), in the modulation of the bradycardic component of the baroreceptor reflex.

METHODS

Mean arterial pressure (MAP) and heart rate were continuously recorded. Reflex changes in heart rate elicited by bolus injection of graded doses of phenylephrine were evaluated before and after bilateral microinjection (glass micropipette) of Ang-(<em>1</em>-<em>7</em>) (<em>1</em>0 pmol or 25 pmol), A-<em>7</em><em>7</em>9 (50 pmol) or saline (vehicle) into the nTS of urethane anesthetized male Wistar rats or spontaneously hypertensive rats (SHR). The averaged ratio between reflex changes in heart rate and changes in MAP was used as index of baroreflex sensitivity.

RESULTS

Microinjection of Ang-(<em>1</em>-<em>7</em>) into the nTS elicited significant decreases in MAP and heart rate in both Wistar and SHR. While the decrease in MAP was similar in both strains, the changes in heart rate were smaller in SHR. A-<em>7</em><em>7</em>9 produced small changes in MAP and heart rate that were no different from those induced by saline. After microinjection of <em>1</em>0 pmol of Ang-(<em>1</em>-<em>7</em>) into the nTS of normotensive rats, there was a significant increase in baroreflex sensitivity. In SHR, only the microinjection of a higher dose (25 pmol) of Ang-(<em>1</em>-<em>7</em>) produced a significant increase in baroreflex sensitivity. A significant reduction inbaroreflex sensitivity was observed after microinjection of A-<em>7</em><em>7</em>9 (50 pmol) in both strains.

CONCLUSIONS

These results indicate that Ang-(<em>1</em>-<em>7</em>) exerts a tonic modulatory effect on the baroreflex control of heart rate at the nTS, probably through a non-AT<em>1</em> non-AT2 receptor subtype. In addition, our data showed a reduced sensitivity to Ang-(<em>1</em>-<em>7</em>) at the nTS of SHR, that could be accounting, at least in part, for the decreased baroreflex sensitivity present in this model of hypertension.

The novel corona virus disease has shaken the entire world with its deadly effects and rapid transmission rates, posing a significant challenge to the healthcare authorities to develop suitable therapeutic solution to save lives on earth. The review aims to grab the attention of the researchers all over the globe, towards the role of ACE2 in COVID-<em>1</em>9 disease. ACE2 serves as a molecular target for the SARS-CoV-2, to enter the target cell, by interacting with the viral glycoprotein spikes. However, the complexity began when numerous studies identified the protective response of ACE2 in abbreviating the harmful effects of vasoconstrictor, anti-inflammatory peptide, <em>angiotensin</em> 2, by mediating its conversion to <em>angiotensin</em>-(<em>1</em>-<em>7</em>), which exercised antagonistic actions to <em>angiotensin</em> 2. Furthermore, certain investigations revealed greater resistance among children as compared to the geriatrics, towards COVID-<em>1</em>9 infection, despite the elevated expression of ACE2 in pediatric population. Based upon such evidences, the review demonstrated possible therapeutic interventions, targeting both the protective and deleterious effects of ACE2 in COVID-<em>1</em>9 disease, primarily inhibiting ACE2-virus interactions or administering soluble ACE2. Thus, the authors aim to provide an opportunity for the researchers to consider RAAS system to be a significant element in development of suitable treatment regime for COVID-<em>1</em>9 pandemic.

<strong class="sub-title"> Keywords: </strong> ACE2; Angiotensin 2; Angiotensin-(<em>1</em>–<em>7</em>); Corona virus; Glycoprotein spikes; RAAS system.

In contrast to the relatively ubiquitous <em>angiotensin</em>-converting enzyme (ACE), expression of the mammalian ACE homologue, ACE2, was initially described in the heart, kidney and testis. ACE2 is a type I integral membrane protein with its active site domain exposed to the extracellular surface of endothelial cells and the renal tubular epithelium. Here ACE2 is poised to metabolise circulating peptides which may include <em>angiotensin</em> II, a potent vasoconstrictor and the product of <em>angiotensin</em> I cleavage by ACE. To this end, ACE2 may counterbalance the effects of ACE within the renin-<em>angiotensin</em> system (RAS). Indeed, ACE2 has been implicated in the regulation of heart and renal function where it is proposed to control the levels of <em>angiotensin</em> II relative to its hypotensive metabolite, <em>angiotensin</em>-(<em>1</em>-<em>7</em>). The recent solution of the structure of ACE2, and ACE, has provided new insight into the substrate and inhibitor profiles of these two key regulators of the RAS. As the complexity of this crucial pathway is unravelled, there is a growing interest in the therapeutic potential of agents that modulate the activity of ACE2.

Ang-(<em>1</em>-<em>7</em>) [<em>angiotensin</em>-(<em>1</em>-<em>7</em>)] is a biologically active heptapeptide component of the RAS (renin-<em>angiotensin</em> system), and is generated in the kidney at relatively high levels, via enzymatic pathways that include ACE2 (<em>angiotensin</em>-converting enzyme 2). The biological effects of Ang-(<em>1</em>-<em>7</em>) in the kidney are primarily mediated by interaction with the G-protein-coupled receptor Mas. However, other complex effects have been described that may involve receptor-receptor interactions with AT(<em>1</em>) (<em>angiotensin</em> II type <em>1</em>) or AT(2) (<em>angiotensin</em> II type 2) receptors, as well as nuclear receptor binding. In the renal vasculature, Ang-(<em>1</em>-<em>7</em>) has vasodilatory properties and it opposes growth-stimulatory signalling in tubular epithelial cells. In several kidney diseases, including hypertensive and diabetic nephropathy, glomerulonephritis, tubulointerstitial fibrosis, pre-eclampsia and acute kidney injury, a growing body of evidence supports a role for endogenous or exogenous Ang-(<em>1</em>-<em>7</em>) as an antagonist of signalling mediated by AT(<em>1</em>) receptors and thereby as a protector against nephron injury. In certain experimental conditions, Ang-(<em>1</em>-<em>7</em>) appears to paradoxically exacerbate renal injury, suggesting that dose or route of administration, state of activation of the local RAS, cell-specific signalling or non-Mas receptor-mediated pathways may contribute to the deleterious responses. Although Ang-(<em>1</em>-<em>7</em>) has promise as a potential therapeutic agent in humans with kidney disease, further studies are required to delineate its signalling mechanisms in the kidney under physiological and pathophysiological conditions.

BACKGROUND

KLF4 mediates inflammatory responses after vascular injury/disease; however, the role of KLF4 in abdominal aortic aneurysms (AAAs) remains unknown. The goals of the present study were to (<em>1</em>) determine the role of KLF4 in experimental AAA; and (2) determine the effect of KLF4 on smooth muscle (SM) cells in AAAs.

RESULTS

KLF4 expression progressively increased at days 3, 7, and <em>1</em>4 after aortic elastase perfusion in C57BL/6 mice. Separately, loss of a KLF4 allele conferred AAA protection using ERTCre+ KLF4 flx/wt mice in the elastase AAA model. In a third set of experiments, SM-specific loss of <em>1</em> and 2 KLF4 alleles resulted in progressively greater protection using novel transgenic mice (MYHCre+ flx/flx, flx/wt, and wt/wt) in the elastase AAA model compared with control. Elastin degradation, MAC2, and cytokine production (MCP<em>1</em>, tumor necrosis factor-α, and interleukin-23) were significantly attenuated, whereas α-actin staining was increased in KLF4 knockout mice versus controls. Results were verified in global KLF4 and SM-specific knockout mice using an angiotensin II model of aneurysm formation. KLF4 inhibition with siRNA attenuated downregulation of SM gene expression in vitro, whereas in vivo studies demonstrated that KLF4 binds to promoters of SM genes by chromatin immunoprecipitation analysis. Finally, human aortic aneurysms demonstrated significantly higher KLF4 expression that was localized to SM cells.

CONCLUSIONS

KLF4 plays a critical role in aortic aneurysm formation via effects on SM cells. These results suggest that KLF4 regulates SM cell phenotypic switching and could be a potential therapeutic target for AAA disease.

To determine the contribution of kidney-derived renin and <em>angiotensin</em> converting enzyme to circulating and tissue levels of <em>angiotensin</em> peptides, we measured <em>angiotensin</em> (Ang)-(<em>1</em>-<em>7</em>), Ang II, Ang-(<em>1</em>-9), and Ang I in plasma, kidney, lung, heart, aorta, brown adipose tissue, adrenal, pituitary, and brain of five groups of male Sprague-Dawley rats: control rats, rats given the converting enzyme inhibitor ramipril (<em>1</em>0 mg/kg), rats nephrectomized 24 hours, rats nephrectomized 48 hours, and rats nephrectomized 48 hours and given ramipril. Plasma and tissues, apart from adrenal, showed a 63% to 98% reduction in Ang II, the ratio of Ang II to Ang I, or both after ramipril administration, indicating a major role for converting enzyme in Ang II formation. Nephrectomy caused a more than 95% decrease in plasma renin levels and a fourfold to eightfold increase in plasma <em>angiotensin</em>ogen levels. Apart from plasma and brain, tissues showed a 59% to <em>7</em>8% decrease in Ang II levels after nephrectomy, indicating a major role for kidney-derived renin in Ang II formation. The persistence of Ang II in plasma and tissues of anephric rats indicates that Ang II may be formed by a process independent of kidney-derived renin; this process may be amplified by the increased plasma <em>angiotensin</em>ogen levels that accompany nephrectomy. For lung, adrenal, and aorta, Ang II levels showed a further decrease when nephrectomized rats were given ramipril. However, for plasma and the other tissues, ramipril produced little or no decrease in Ang II levels of anephric rats, suggesting that Ang II may be formed by a pathway independent of converting enzyme. Such a pathway may involve the direct formation of Ang II from <em>angiotensin</em>ogen by a non-renin-like enzyme.

Angioedema is a rare, potentially life-threatening adverse event of renin-<em>angiotensin</em> system inhibitors. The objective of the present study was to determine the risk of angioedema from randomized clinical trials. A PubMed/CENTRAL/EMBASE search was made for randomized clinical trials from <em>1</em>980 to October 20<em>1</em><em>1</em> in patients on <em>angiotensin</em>-converting enzyme (ACE) inhibitors, <em>angiotensin</em> receptor blockers (ARBs), or direct renin inhibitor (DRI). Trials with a total number of patients ≥<em>1</em>00 and a duration of ≥8 weeks were included for analysis. Incidence of angioedema was pooled by weighing the incident rate of each trial by the inverse of the variance. Twenty-six trials with <em>7</em>4,85<em>7</em> patients in the ACE inhibitor arm with 232,523 person-years of follow-up, <em>1</em>9 trials with 35,4<em>7</em>9 patients on ARB with <em>1</em>22,293 person-years of follow-up, and 2 trials with 5,<em>1</em>4<em>1</em> patients on DRI with <em>1</em>,<em>7</em>35 person-years of follow-up met the inclusion criteria and were included in the analysis. In head-to-head comparison in <em>7</em> trials, risk of angioedema with ACE inhibitors was 2.2 times higher than with ARBs (95% confidence interval [CI] <em>1</em>.5 to 3.3). With ACE inhibitors and ARBs, incidence of angioedema was higher in heart failure trials compared to hypertension or coronary artery disease trials without heart failure (p <0.000<em>1</em>). Weighted incidence of angioedema with ACE inhibitors was 0.30% (95% CI 0.28 to 0.32) compared to 0.<em>1</em><em>1</em>% (95% CI 0.09 to 0.<em>1</em>3) with ARBs, 0.<em>1</em>3% (95% CI 0.08 to 0.<em>1</em>9) with DRIs, and 0.0<em>7</em>% with placebo (95% CI 0.05 to 0.09). In conclusion, incidence of angioedema with ARBs and DRI was (<em>1</em>/2 than that with ACE inhibitors and not significantly different from placebo. Incidence of angioedema was higher in patients with heart failure compared to those without heart failure with ACE inhibitors and ARBs.

In the present paper, the modulation of the basolateral membrane (BLM) Na+ -ATPase activity of inner cortex from pig kidney by <em>angiotensin</em> II (Ang II) and <em>angiotensin</em>-(<em>1</em>-<em>7</em>) (Ang-(<em>1</em>-<em>7</em>)) was evaluated. Ang II and Ang-(<em>1</em>-<em>7</em>) inhibit the Na+ -ATPase activity in a dose-dependent manner (from <em>1</em>0(-<em>1</em><em>1</em>) to <em>1</em>0(-5) M), with maximal effect obtained at <em>1</em>0(-<em>7</em>) M for both peptides. Pharmacological evidences demonstrate that the inhibitory effects of Ang II and Ang-(<em>1</em>-<em>7</em>) are mediated by AT2 receptor: The effect of both polypeptides is completely reversed by <em>1</em>0(-8) M PD <em>1</em>233<em>1</em>9, a selective AT2 receptor antagonist, but is not affected by either (<em>1</em>0(-<em>1</em>2) - <em>1</em>0(-5) M) losartan or (<em>1</em>0(-<em>1</em>0)-<em>1</em>0(-<em>7</em>) M) A<em>7</em><em>7</em>9, selective antagonists for AT<em>1</em> and AT(<em>1</em>-<em>7</em>) receptors, respectively. The following results suggest that a PTX-insensitive, cholera toxin (CTX)-sensitive G protein/adenosine 3',5'-cyclic monophosphate (cAMP)/PKA pathway is involved in this process: (<em>1</em>) the inhibitory effect of both peptides is completely reversed by <em>1</em>0(-9) M guanosine 5'-O-(2-thiodiphosphate) (GDPbetaS; an inhibitor of the G protein activity), and mimicked by <em>1</em>0(-<em>1</em>0) M guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS; an activator of the G protein activity); (2) the effects of both peptides are mimicked by CTX but are not affected by PTX; (3) Western blot analysis reveals the presence of the Gs protein in the isolated basolateral membrane fraction; (4) (<em>1</em>0(-<em>1</em>0)-<em>1</em>0(-6) M) cAMP has a similar and non-additive effect to Ang II and Ang-(<em>1</em>-<em>7</em>); (5) PKA inhibitory peptide abolishes the effects of Ang II and Ang-(<em>1</em>-<em>7</em>); and (6) both <em>angiotensins</em> stimulate PKA activity.

<em>Angiotensin</em> (Ang)-(<em>1</em>-<em>7</em>) is a biologically active peptide of the renin-<em>angiotensin</em> system that has both vasodilatory and antiproliferative activities that are opposite the constrictive and proliferative effects of <em>angiotensin</em> II (Ang II). We studied the actions of Ang-(<em>1</em>-<em>7</em>) on the Ang II type <em>1</em> (AT(<em>1</em>)) receptor in cultured rat aortic vascular smooth muscle cells to determine whether the effects of Ang-(<em>1</em>-<em>7</em>) are due to its regulation of the AT(<em>1</em>) receptor. Ang-(<em>1</em>-<em>7</em>) competed poorly for [(<em>1</em>25)I]Ang II binding to the AT(<em>1</em>) receptor on vascular smooth muscle cells, with an IC(50) of 2.0 micromol/L compared with <em>1</em>.9 nmol/L for Ang II. The pretreatment of vascular smooth muscle cells with Ang-(<em>1</em>-<em>7</em>) followed by treatment with acidic glycine to remove surface-bound peptide resulted in a significant decrease in [(<em>1</em>25)I]Ang II binding; however, reduced Ang II binding was observed only at micromolar concentrations of Ang-(<em>1</em>-<em>7</em>). Scatchard analysis of vascular smooth muscle cells pretreated with <em>1</em> micromol/L Ang-(<em>1</em>-<em>7</em>) showed that the reduction in Ang II binding resulted from a loss of the total number of binding sites [B(max) 43<em>7</em>.<em>7</em>+/-26<em>1</em>.5 fmol/mg protein in Ang-(<em>1</em>-<em>7</em>)-pretreated cells compared with 60<em>7</em>.5+/-30<em>1</em>.2 fmol/mg protein in untreated cells, n=5, P<0.05] with no significant effect on the affinity of Ang II for the AT(<em>1</em>) receptor. Pretreatment with the AT(<em>1</em>) receptor antagonist L-<em>1</em>58,809 blocked the reduction in [(<em>1</em>25)I]Ang II binding by Ang-(<em>1</em>-<em>7</em>) or Ang II. Pretreatment of vascular smooth muscle cells with increasing concentrations of Ang-(<em>1</em>-<em>7</em>) reduced Ang II-stimulated phospholipase C activity; however, the decrease was significant (8<em>1</em>.2+/-6.4%, P<0.0<em>1</em>, n=5) only at <em>1</em> micromol/L Ang-(<em>1</em>-<em>7</em>). These results demonstrate that pharmacological concentrations of Ang-(<em>1</em>-<em>7</em>) in the micromolar range cause a modest downregulation of the AT(<em>1</em>) receptor on vascular cells and a reduction in Ang II-stimulated phospholipase C activity. Because the antiproliferative and vasodilatory effects of Ang-(<em>1</em>-<em>7</em>) are observed at nanomolar concentrations of the heptapeptide, these responses to Ang-(<em>1</em>-<em>7</em>) cannot be explained by competition of Ang-(<em>1</em>-<em>7</em>) at the AT(<em>1</em>) receptor or Ang-(<em>1</em>-<em>7</em>)-mediated downregulation of the vascular AT(<em>1</em>) receptor.

Renal oxidative stress and nitric oxide (NO) deficiency are key events in hypertension. Stimulation of a nitrate-nitrite-NO pathway with dietary nitrate reduces blood pressure, but the mechanisms or target organ are not clear. We investigated the hypothesis that inorganic nitrate and nitrite attenuate reactivity of renal microcirculation and blood pressure responses to <em>angiotensin</em> II (ANG II) by modulating nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activity and NO bioavailability. Nitrite in the physiological range (<em>1</em>0(-<em>7</em>)-<em>1</em>0(-5) mol/L) dilated isolated perfused renal afferent arterioles, which were associated with increased NO. Contractions to ANG II (34%) and simultaneous NO synthase inhibition (56%) were attenuated by nitrite (<em>1</em>8% and 26%). In a model of oxidative stress (superoxide dismutase-<em>1</em> knockouts), abnormal ANG II-mediated arteriolar contractions (90%) were normalized by nitrite (44%). Mechanistically, effects of nitrite were abolished by NO scavenger and xanthine oxidase inhibitor, but only partially attenuated by inhibiting soluble guanylyl cyclase. Inhibition of NADPH oxidase with apocynin attenuated ANG II-induced contractility (35%) similar to that of nitrite. In the presence of nitrite, no further effect of apocynin was observed, suggesting NADPH oxidase as a possible target. In preglomerular vascular smooth muscle cells and kidney cortex, nitrite reduced both basal and ANG II-induced NADPH oxidase activity. These effects of nitrite were also abolished by xanthine oxidase inhibition. Moreover, supplementation with dietary nitrate (<em>1</em>0(-2) mol/L) reduced renal NADPH oxidase activity and attenuated ANG II-mediated arteriolar contractions and hypertension (99±2-<em>1</em>46±2 mm Hg) compared with placebo (<em>1</em>00±3-<em>1</em>68±3 mm Hg). In conclusion, these novel findings position NADPH oxidase in the renal microvasculature as a prime target for blood pressure-lowering effects of inorganic nitrate and nitrite.

Gene-targeting studies in mice demonstrate that the renin-<em>angiotensin</em> system is required for the proper development of the renal medulla. In the absence of <em>angiotensin</em> II (ANG II) or the ANG II type <em>1</em> (AT<em>1</em>) receptor, mice exhibit poor papillary development and a severe urinary-concentrating defect. These findings imply that the ureteric bud (UB) and its branches are targets for ANG II actions during renal development. However, direct evidence linking ANG II with UB-branching morphogenesis does not exist. Using immunohistochemistry, we demonstrated that UB-derived epithelia express <em>angiotensin</em>ogen (Ao) and the AT<em>1</em> receptor during murine metanephrogenesis. Ao and AT<em>1</em> receptors are expressed in the UB branches and to a lesser extent in the stromal mesenchyme. AT<em>1</em> receptor expression in UB-derived epithelia increased from embryo day <em>1</em>2 to day <em>1</em>6 and was observed on both luminal and basolateral membranes. In accord with these findings, cultured murine UB cells express AT<em>1</em> receptor protein and mRNA. Treatment of UB cells cultured in three-dimensional type I collagen gels with ANG II (<em>1</em>0-<em>7</em> to <em>1</em>0-5 M) elicits a dose-related increase in the number of cells that have primary and secondary branches. These effects of ANG II on UB branching are abrogated by pretreatment with the AT<em>1</em> receptor antagonist candesartan. These data demonstrate a direct and independent role for ANG II acting via AT<em>1</em> receptors on UB cell branching in vitro. The presence of Ao in the stroma and AT<em>1</em> on UB cells supports the notion that cross talk between stroma and epithelial cells is crucial to epithelial branching morphogenesis in the developing kidney.

BACKGROUND

Hypercortisolism as a sign of hypothamamus-pituitary-adrenocortical (HPA) axis overactivity and sleep EEG changes are frequently observed in depression. Closely related to the HPA axis is the renin-<em>angiotensin</em>-aldosterone system (RAAS) as <em>1</em>. adrenocorticotropic hormone (ACTH) is a common stimulus for cortisol and aldosterone, 2. cortisol release is suppressed by mineralocorticoid receptor (MR) agonists 3. <em>angiotensin</em> II (ATII) releases CRH and vasopressin from the hypothalamus. Furthermore renin and aldosterone secretion are synchronized to the rapid eyed movement (REM)-nonREM cycle.

METHODS

Here we focus on the difference of sleep related activity of the RAAS between depressed patients and healthy controls. We studied the nocturnal plasma concentration of ACTH, cortisol, renin and aldosterone, and sleep EEG in 7 medication free patients with depression (<em>1</em> male, 6 females, age: (mean +/-SD) 53.3 +/- <em>1</em>4.4 yr.) and 7 age matched controls (2 males, 5 females, age: 54.7 +/- <em>1</em>9.5 yr.). After one night of accommodation a polysomnography was performed between 23.00 h and 7.00 h. During examination nights blood samples were taken every 20 min between 23.00 h and 7.00 h. Area under the curve (AUC) for the hormones separated for the halves of the night (23.00 h to 3.00 h and 3.00 h to 7.00 h) were used for statistical analysis, with analysis of co variance being performed with age as a covariate.

RESULTS

No differences in ACTH and renin concentrations were found. For cortisol, a trend to an increase was found in the first half of the night in patients compared to controls (p < 0.06). Aldosterone was largely increased in the first (p < 0.05) and second (p < 0.0<em>1</em>) half of the night. Cross correlations between hormone concentrations revealed that in contrast to earlier findings, which included only male subjects, in our primarily female sample, renin and aldosterone secretion were not coupled and no difference between patients and controls could be found, suggesting a gender difference in RAAS regulation. No difference in conventional sleep EEG parameters were found in our sample.

CONCLUSIONS

Hyperaldosteronism could be a sensitive marker for depression. Further our findings point to an altered renal mineralocorticoid sensitivity in patients with depression.

OBJECTIVE

The transactivation of the epidermal growth factor (EGF) receptor appears to be an important central transduction mechanism in mediating diabetes-induced vascular dysfunction. <em>Angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] via its Mas receptor can prevent the development of hyperglycaemia-induced cardiovascular complications. Here, we investigated whether Ang-(<em>1</em>-<em>7</em>) can inhibit hyperglycaemia-induced EGF receptor transactivation and its classical signalling via ERK<em>1</em>/2 and p38 MAPK in vivo and in vitro.

METHODS

Streptozotocin-induced diabetic rats were chronically treated with Ang-(<em>1</em>-<em>7</em>) or AG<em>1</em>4<em>7</em>8, a selective EGF receptor inhibitor, for 4 weeks and mechanistic studies performed in the isolated mesenteric vasculature bed as well as in primary cultures of vascular smooth muscle cells (VSMCs).

RESULTS

Diabetes significantly enhanced phosphorylation of EGF receptor at tyrosine residues Y992, Y<em>1</em>068, Y<em>1</em>086, Y<em>1</em><em>1</em>48, as well as ERK<em>1</em>/2 and p38 MAPK in the mesenteric vasculature bed whereas these changes were significantly attenuated upon Ang-(<em>1</em>-<em>7</em>) or AG<em>1</em>4<em>7</em>8 treatment. In VSMCs grown in conditions of high glucose (25 mM), an Src-dependent elevation in EGF receptor phosphorylation was observed. Ang-(<em>1</em>-<em>7</em>) inhibited both Ang II- and glucose-induced transactivation of EGF receptor. The inhibition of high glucose-mediated Src-dependant transactivation of EGF receptor by Ang-(<em>1</em>-<em>7</em>) could be prevented by a selective Mas receptor antagonist, D-Pro<em>7</em>-Ang-(<em>1</em>-<em>7</em>).

CONCLUSIONS

These results show for the first time that Ang-(<em>1</em>-<em>7</em>) inhibits EGF receptor transactivation via a Mas receptor/Src-dependent pathway and might represent a novel general mechanism by which Ang-(<em>1</em>-<em>7</em>) exerts its beneficial effects in many disease states including diabetes-induced vascular dysfunction.

Endothelial-to-mesenchymal transition (EndMT) emerges as an important source of fibroblasts. MicroRNA let-<em>7</em> exhibits anti-EndMT effects and fibroblast growth factor (FGF) receptor has been shown to be an important in microRNA let-<em>7</em> expression. The endogenous antifibrotic peptide N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP) is a substrate of <em>angiotensin</em>-converting enzyme (ACE). Here, we found that AcSDKP inhibited the EndMT and exhibited fibrotic effects that were associated with FGF receptor-mediated anti-fibrotic program. Conventional ACE inhibitor plus AcSDKP ameliorated kidney fibrosis and inhibited EndMT compared to therapy with the ACE inhibitor alone in diabetic CD-<em>1</em> mice. The endogenous AcSDKP levels were suppressed in diabetic animals. Cytokines induced cultured endothelial cells into EndMT; coincubation with AcSDKP inhibited EndMT. Expression of microRNA let-<em>7</em> family was suppressed in the diabetic kidney; antifibrotic and anti-EndMT effects of AcSDKP were associated with the restoration of microRNA let-<em>7</em> levels. AcSDKP restored diabetes- or cytokines-suppressed FGF receptor expression/phosphorylation into normal levels both in vivo and in vitro. These results suggest that AcSDKP is an endogenous antifibrotic molecule that has the potential to cure diabetic kidney fibrosis via an inhibition of the EndMT associated with the restoration of FGF receptor and microRNA let-<em>7</em>.

BACKGROUND

Excess dietary sodium has been linked to the development of hypertension and other cardiovascular diseases. In humans, the effects of sodium consumption on endothelial function have not been separated from the effects on blood pressure. The present study was designed to determine if dietary sodium intake affected endothelium-dependent dilation (EDD) independently of changes in blood pressure.

METHODS

Fourteen healthy salt-resistant adults were studied (9M, 5F; age 33 ± 2.4 years) in a controlled feeding study. After a baseline run-in diet, participants were randomized to a <em>7</em>-day high-sodium (300-350 mmol/day) and <em>7</em>-day low-sodium (20 mmol/day) diet. Salt resistance, defined as a 5 mmHg or less change in a 24-h mean arterial pressure, was individually assessed while on the low-sodium and high-sodium diets and confirmed in the participants undergoing study (low-sodium: 85 ± <em>1</em> mmHg; high-sodium: 85 ± 2 mmHg). EDD was determined in each participant via brachial artery flow-mediated dilation on the last day of each diet.

RESULTS

Sodium excretion increased during the high-sodium diet (P < 0.0<em>1</em>). EDD was reduced on the high-sodium diet (low: <em>1</em>0.3 ± 0.9%, high: <em>7</em>.3 ± 0.<em>7</em>%; P < 0.05). The high-sodium diet significantly suppressed plasma renin activity (PRA), plasma angiotensin II, and aldosterone (P < 0.05).

CONCLUSIONS

These data demonstrate that excess salt intake in humans impairs endothelium-dependent dilation independently of changes in blood pressure.

Ischemia-reperfusion (I/R) is a model of acute kidney injury (AKI) that is characterized by vasoconstriction, oxidative stress, apoptosis and inflammation. Previous studies have shown that activation of the renin-<em>angiotensin</em> system (RAS) may contribute to these processes. <em>Angiotensin</em> converting enzyme 2 (ACE2) metabolizes <em>angiotensin</em> II (Ang II) to <em>angiotensin</em>-(<em>1</em>-<em>7</em>), and recent studies support a beneficial role for ACE2 in models of chronic kidney disease. However, the role of ACE2 in models of AKI has not been fully elucidated. In order to test the hypothesis that ACE2 plays a protective role in AKI we assessed I/R injury in wild-type (WT) mice and ACE2 knock-out (ACE2 KO) mice. ACE2 KO and WT mice exhibited similar histologic injury scores and measures of kidney function at 48 hours after reperfusion. Loss of ACE2 was associated with increased neutrophil, macrophage, and T cell infiltration in the kidney. mRNA levels for pro-inflammatory cytokines, interleukin-<em>1</em>β, interleukin-6 and tumour necrosis factor-α, as well as chemokines macrophage inflammatory protein 2 and monocyte chemoattractant protein-<em>1</em>, were increased in ACE2 KO mice compared to WT mice. Changes in inflammatory cell infiltrates and cytokine expression were also associated with greater apoptosis and oxidative stress in ACE2 KO mice compared to WT mice. These data demonstrate a protective effect of ACE2 in I/R AKI.

Elevated circulating fatty acid-binding protein 4 (FABP4/A-FABP/aP2), an adipokine, is associated with obesity, insulin resistance, hypertension and cardiovascular events. However, how circulating FABP4 level is modified by pharmacological agents remains unclear. We here examined the effects of <em>angiotensin</em> II receptor blockers (ARBs) on serum FABP4 level. First, essential hypertensives were treated with ARBs: candesartan (8 mg day(-<em>1</em>); n=<em>7</em>) for 2 weeks, olmesartan (20 mg day(-<em>1</em>); n=9) for <em>1</em>2 weeks, and valsartan (80 mg day(-<em>1</em>); n=94) or telmisartan (40 mg day(-<em>1</em>); n=9<em>1</em>) for 8 weeks added to amlodipine (5 mg day(-<em>1</em>)). Treatment with ARBs significantly decreased blood pressure and serum FABP4 concentrations by 8-20% without significant changes in adiposity or lipid variables, though the M value determined by hyperinsulinemic-euglycemic glucose clamp, a sensitive index of insulin sensitivity, was significantly increased by candesartan. Next, alterations in FABP4 secretion from 3T3-L<em>1</em> adipocytes were examined under several agents. Lipolytic stimulation of the β-adrenoceptor in 3T3-L<em>1</em> adipocytes by isoproterenol increased FABP4 secretion, and conversely, insulin suppressed FABP4 secretion. However, treatment of 3T3-L<em>1</em> adipocytes with <em>angiotensin</em> II or ARBs for 2 h had no effect on gene expression or secretion of FABP4 regardless of β-adrenoceptor stimulation. In conclusion, treatment with structurally different ARBs similarly decreases circulating FABP4 concentrations in hypertensive patients as a class effect of ARBs, which is not attributable to blockade of the <em>angiotensin</em> II receptor in adipocytes. Reduction of FABP4 levels by ARBs might be involved in suppression of cardiovascular events.

The implication of the renin-<em>angiotensin</em> system (RAS) in the regulation of the cardiovascular system has been well known for many years. Accordingly, many pharmaceutical inhibitors have been developed to treat several pathologies, like hypertension and heart failure, and <em>angiotensin</em> converting enzyme (ACE) became one of the major target in the treatment of these cardiovascular diseases. In the last decade however, it has become apparent that the classical view of the RAS was not quite accurate. For instance, ACE has been shown to work not only by generating <em>angiotensin</em>-II but also by interacting with receptors outside the renin-<em>angiotensin</em> system. Moreover, it has been shown that many local RAS are present in different tissues, such as the heart, brain, kidney and vasculature. However, in the past, it was impossible to determine the role of these local systems as they were pharmacologically indistinguishable from the systemic RAS. Hence, in recent years, the development of transgenic animals has allowed us to determine that these local systems are implicated in the roles that had been originally attributed exclusively to the systemic action of the RAS. However, with almost 30% of the medicated hypertensive patients harboring an uncontrolled blood pressure, a need for new drugs and new targets appears necessary. With the new century came the discovery of a new homolog of ACE, called ACE2, and early studies suggest that it may play a pivotal role in the RAS by controlling the balance between the vasoconstrictor effects of <em>angiotensin</em>-II and the vasodilatory properties of the <em>angiotensin</em>(<em>1</em>-<em>7</em>) peptide. Like ACE, ACE2 appears to hydrolyze peptides not related with the RAS and the enzyme has also been identified as a receptor for the severe acute respiratory syndrome (SARS) coronavirus. Although the tissue localization of ACE2 was originally though to be very restricted, new studies have emerged showing a more widespread distribution. Therefore, the whole dynamics of the RAS has to be re-evaluated in light of this new information. In this review, we will compare the structures, distributions and properties of ACE and its new homologue in the context of cardiovascular function, focusing on the autocrine/paracrine cardiac and brain renin-<em>angiotensin</em> systems and we will present recent data from the literature and our laboratory offering a new perspective on this potential target for the treatment of cardiovascular diseases.