Sustained induction and activation of matrixins (matrix metalloproteinases or MMPs), and the destruction and deposition of extracellular matrix (ECM), are the hallmarks of cardiac fibrosis. The reversion-inducing-cysteine-rich protein with Kazal motifs (RECK) is a unique membrane-anchored endogenous MMP regulator. We hypothesized that elevated <em>angiotensin</em> II (Ang II), which is associated with fibrosis in the heart, differentially regulates MMPs and RECK both in vivo and in vitro. Continuous infusion of Ang II into male C5<em>7</em>Bl/6 mice for 2weeks resulted in cardiac fibrosis, with increased expressions of MMPs 2, <em>7</em>, 9 and <em>1</em>4, and of collagens Ia<em>1</em> and IIIa<em>1</em>. The expression of RECK, however, was markedly suppressed. These effects were inhibited by co-treatment with the Ang II type <em>1</em> receptor (AT<em>1</em>) antagonist losartan. In vitro, Ang II suppressed RECK expression in adult mouse cardiac fibroblasts (CF) via AT<em>1</em>/Nox4-dependent ERK/Sp<em>1</em> activation, but induced MMPs 2, <em>1</em>4 and 9 via NF-κB, AP-<em>1</em> and/or Sp<em>1</em> activation. Further, while forced expression of RECK inhibits, its knockdown potentiates Ang II-induced CF migration. Notably, RECK overexpression reduced Ang II-induced MMPs 2, 9 and <em>1</em>4 activation, but enhanced collagens Ia<em>1</em> and IIIa<em>1</em> expression and soluble collagen release. These results demonstrate for the first time that Ang II suppresses RECK, but induces MMPs both in vivo and in vitro, and RECK overexpression blunts Ang II-induced MMP activation and CF migration in vitro. Strategies that upregulate RECK expression in vivo have the potential to attenuate sustained MMP expression, and blunt fibrosis and adverse remodeling in hypertensive heart diseases.

<em>Angiotensin</em> converting enzyme (ACE) 2 is a homologue of ACE that catalyzes the conversion of <em>Angiotensin</em> (Ang) II into Ang<em>1</em>-<em>7</em>, which induces vasodilation, anti-fibrotic, anti-proliferative and anti-inflammatory effects. Given that ACE2 counterbalances the effects of Ang II, it has been proposed as a biomarker in kidney disease patients. Circulating ACE2 has been studied in human and experimental studies under physiological and pathological conditions and different techniques have been assessed to determine its enzymatic activity. In patients with cardiovascular (CV) disease circulating ACE2 has been shown to be increased. In addition, hypertensive and diabetic patients have also shown higher circulating ACE2 activities. A study in type <em>1</em> diabetes patients found a negative association between circulating ACE2 and estimated glomerular filtration rate in male and female patients. Recently, it has been demonstrated that circulating ACE2 is increased in male patients with chronic kidney disease (CKD) and that it is independently associated with other classical CV risk factors, such as advanced age and diabetes. Furthermore, circulating ACE2 has been shown to be associated with silent atherosclerosis and CV outcomes in CKD patients. In diabetic nephropathy, experimental studies have demonstrated an increase in circulating ACE2 activity both at early and late stages of the disease, as well as a direct association with increased urinary albumin excretion, suggesting that it may be increased as a renoprotective mechanism in these patients. In this paper we will review the measurement of circulating ACE2 and its role in kidney disease, as well as its potential role as a renal and CV biomarker.

We assessed the contribution of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] to captopril-induced cardiovascular protection in spontaneously hypertensive rats (SHRs) chronically treated with the nitric oxide synthesis inhibitor NG-nitro-L-arginine methyl ester (SHR-l). NG-nitro-L-arginine methyl ester (80 mg/L) administration for 3 weeks increased mean arterial pressure (MAP) from <em>1</em>96 ± 6 to 229 ± 3 mm Hg (P < 0.05). Treatment of SHR-l with Ang-(<em>1</em>-<em>7</em>) antagonist [d-Ala<em>7</em>]-Ang-(<em>1</em>-<em>7</em>) (A<em>7</em><em>7</em>9; <em>7</em>44 μg·kg(-<em>1</em>)·d(-<em>1</em>) ip) further elevated MAP to 253 ± 6 mm Hg (P < 0.05 vs SHR-l or SHR). Moreover, A<em>7</em><em>7</em>9 treatment attenuated the reduction in MAP and proteinuria by either captopril (300 mg/L in drinking water) or hydralazine (<em>1</em>.5 mg·kg(-<em>1</em>)·d(-<em>1</em>) ip). In isolated perfused hearts, the recovery of left ventricular function from global ischemia was enhanced by captopril or hydralazine treatment and was exacerbated with A<em>7</em><em>7</em>9. The Ang-(<em>1</em>-<em>7</em>) antagonist attenuated the beneficial effects of captopril and hydralazine on cardiac function. Recovery from global ischemia was also improved in isolated SHR-l hearts acutely perfused with captopril during both the perfusion and reperfusion periods. The acute administration of A<em>7</em><em>7</em>9 reduced the beneficial actions of captopril to improve recovery after ischemia. We conclude that during periods of reduced nitric oxide availability, endogenous Ang-(<em>1</em>-<em>7</em>) plays a protective role in effectively buffering the increase in blood pressure and renal injury and the recovery from cardiac ischemia. Moreover, Ang-(<em>1</em>-<em>7</em>) contributes to the blood pressure lowering and tissue protective actions of captopril and hydralazine in a model of severe hypertension and end-organ damage.

Release of norepinephrine (NE) by the hypothalamic nuclei may contribute to regulation of sympathetic nervous system (SNS) activity. <em>Angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] has an antihypertensive effect and may decrease SNS activity. We tested the hypothesis that Ang-(<em>1</em>-<em>7</em>) inhibits the release of NE in hypothalami, via the Ang-(<em>1</em>-<em>7</em>) and <em>angiotensin</em> II type 2 (AT2) receptors, acting through a bradykinin (BK)/NO-dependent mechanism. Hypothalami from normotensive controls and spontaneously hypertensive rats (SHR) were isolated and endogenous NE stores labeled by incubating the tissues with [3H]NE. [3H]NE release from the hypothalami was stimulated by KCl in the presence or absence of Ang-(<em>1</em>-<em>7</em>) alone or combined with various antagonists and inhibitors. Ang-(<em>1</em>-<em>7</em>) significantly attenuated K+-induced NE release by hypothalami from normotensive rats but was more potent in SHR. The Ang-(<em>1</em>-<em>7</em>) receptor antagonist [D-Ala<em>7</em>]Ang-(<em>1</em>-<em>7</em>), the AT2 receptor antagonist PD <em>1</em>233<em>1</em>9, and the BK B2) receptor antagonist icatibant all blocked the inhibitory effect of Ang-(<em>1</em>-<em>7</em>) on K+-stimulated NE release in SHR. The inhibitory effect of Ang-(<em>1</em>-<em>7</em>) disappeared in the presence of the NO synthase inhibitor N(G)-nitro-L-arginine methyl ester and was restored by the precursor of NO, l-arginine. The diminished NE release caused by Ang-(<em>1</em>-<em>7</em>) was blocked by a soluble guanylyl cyclase inhibitor as well as by a cGMP-dependent protein kinase (PKG). We concluded that Ang-(<em>1</em>-<em>7</em>) decreases NE release from the hypothalamus via the Ang-(<em>1</em>-<em>7</em>) or AT2 receptors, acting through a BK/NO-mediated mechanism that stimulates cGMP/PKG signaling. In this way, Ang-(<em>1</em>-<em>7</em>) may decrease SNS activity and exert an antihypertensive effect.

We sought to characterize temporal gene expression changes in the murine <em>angiotensin</em> II (ANG II)-ApoE-/- model of abdominal aortic aneurysm (AAA). Aortic ultrasound measurements were obtained over the 28-day time-course. Harvested suprarenal aortic segments were evaluated with whole genome expression profiling at <em>7</em>, <em>1</em>4, and 28 days using the Agilent Whole Mouse Genome microarray platform and Statistical Analysis of Microarrays at a false discovery rate of (<em>1</em>%. A group of <em>angiotensin</em>-treated mice experienced contained rupture (CR) within <em>7</em> days and were analyzed separately. Progressive aortic dilatation occurred throughout the treatment period. However, the numerous early expression differences between ANG II-treated and control were not sustained over time. Ontologic analysis revealed widespread upregulation of inflammatory, immune, and matrix remodeling genes with ANG II treatment, among other pathways such as apoptosis, cell cycling, angiogenesis, and p53 signaling. CR aneurysms displayed significant decreases in TGF-β/BMP-pathway signaling, MAPK signaling, and ErbB signaling genes vs. non-CR/ANG II-treated samples. We also performed literature-based network analysis, extracting numerous highly interconnected genes associated with aneurysm development such as Spp<em>1</em>, Myd88, Adam<em>1</em><em>7</em> and Lox. <em>1</em>) ANG II treatment induces extensive early differential expression changes involving abundant signaling pathways in the suprarenal abdominal aorta, particularly wide-ranging increases in inflammatory genes with aneurysm development. 2) These gene expression changes appear to dissipate with time despite continued growth, suggesting that early changes in gene expression influence disease progression in this AAA model, and that the aortic tissue adapts to prolonged ANG II infusion. 3) Network analysis identified nexus genes that may constitute aneurysm biomarkers or therapeutic targets.

The renin-<em>angiotensin</em> system (RAS) and the kallikrein-kinin system (KKS) each encompasses a large number of molecules, with several participating in both systems. The RAS generates a family of bioactive <em>angiotensin</em> peptides with varying biological activities. These include <em>angiotensin</em>-(<em>1</em>-8) (Ang II), <em>angiotensin</em>-(2-8) (Ang III), <em>angiotensin</em>-(3-8) (Ang IV), and <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)]. Ang II and Ang III act on type <em>1</em> (AT(<em>1</em>)) and type 2 (AT(2)) <em>angiotensin</em> receptors, whereas, Ang IV and Ang-(<em>1</em>-<em>7</em>) act on their own receptors. The KKS also generates a family of bioactive peptides with varying biological activities. These include hydroxylated and non-hydroxylated bradykinin and kallidin peptides and their carboxypeptidase metabolites des-Arg(9)-bradykinin and des-Arg(<em>1</em>0)-kallidin. Whereas bradykinin and kallidin act mainly via the type 2 bradykinin (B(2)) receptor, des-Arg(9)-bradykinin and des-Arg(<em>1</em>0)-kallidin act mainly via the type <em>1</em> bradykinin (B(<em>1</em>)) receptor. The AT(<em>1</em>) receptor forms heterodimers with the AT(2) and B(2) receptors and there is cross talk between the AT(<em>1</em>) and epidermal growth factor receptors. The B(2) receptor also interacts with <em>angiotensin</em> converting enzyme and nitric oxide synthase. Both <em>angiotensin</em> and kinin peptides are metabolised by many different peptidases that are important determinants of the activities of the RAS and KKS, and several of which participate in both systems.

<em>Angiotensin</em>-converting enzyme-2 (ACE2) converts <em>angiotensin</em> II (ANG II) to <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [ANG-(<em>1</em>-<em>7</em>)], and this enzyme may serve as a key regulatory juncture in various tissues. Although the heart expresses ACE2, the extent that the enzyme participates in the cardiac processing of ANG II and ANG-(<em>1</em>-<em>7</em>) is equivocal. Therefore, we utilized the Langendorff preparation to characterize the ACE2 pathway in isolated hearts from male normotensive Sprague-Dawley [Tg((-))] and hypertensive [mRen2]2<em>7</em> [Tg((+))] rats. During a 60-min recirculation period with <em>1</em>0 nM ANG II, the presence of ANG-(<em>1</em>-<em>7</em>) was assessed in the cardiac effluent. ANG-(<em>1</em>-<em>7</em>) generation from ANG II was similar in both the normal and hypertensive hearts [Tg((-)): 5<em>1</em>0 +/- 55 pM, n=20 vs. Tg((+)): 49<em>7</em> +/- 63 pM, n=<em>1</em>4] with peak levels occurring at 30 min after administration of the peptide. ACE2 inhibition (MLN-4<em>7</em>60, <em>1</em> microM) significantly reduced ANG-(<em>1</em>-<em>7</em>) production by 83% (5<em>7</em> +/- <em>1</em>9 pM, P<0.0<em>1</em>, n=<em>7</em>) in the Tg((+)) rats, whereas the inhibitor had no significant effect in the Tg((-)) rats (285 +/- 53 pM, P>0.05, n=<em>1</em>0). ACE2 activity was found in the effluent of perfused Tg((-)) and Tg((+)) hearts, and it was highly associated with ACE2 protein expression (r=0.<em>7</em>8). This study is the first demonstration for a direct role of ACE2 in the metabolism of cardiac ANG II in the hypertrophic heart of hypertensive rats. We conclude that predominant expression of cardiac ACE2 activity in the Tg((+)) may be a compensatory response to the extensive cardiac remodeling in this strain.

<em>1</em>. The two isozymes of human <em>angiotensin</em> converting enzyme (ACE; EC 3.4.<em>1</em>5.<em>1</em>) have recently been cloned and sequenced. 2. The larger, endothelial isozyme has two highly similar internal domains each bearing a putative catalytic site. In contrast the smaller, testicular isozyme has a single catalytic site corresponding to the C-terminal domain of endothelial ACE and represents the ancestral, non-duplicated form of the gene. 3. Both isozymes are anchored in the plasma membrane by a single hydrophobic transmembrane polypeptide located near the C-terminus, and both are extensively N-glycosylated. 4. The testicular isozyme may also be O-glycosylated. 5. The soluble form of ACE in plasma, seminal fluid and other body fluids appears to be derived from the membrane-bound endothelial isozyme by a post-translational modification. 6. ACE has a complex substrate specificity with peptidyl tripeptidase or endopeptidase action on certain peptides, as well as the classical peptidyl dipeptidase activity. <em>7</em>. Numerous potent inhibitors of the enzyme have been developed and used successfully in the treatment of hypertension, but some of the observed side effects may be due to inhibition of other zinc metalloenzymes. 8. Both endothelial and testicular ACE are highly conserved between species, indicative of the essential role(s) of the enzyme in blood pressure regulation and other physiological processes.

Postural tachycardia syndrome (POTS) is associated with increased plasma <em>angiotensin</em> II (Ang II). Ang II administered in the presence of NO synthase inhibition with nitro-L-arginine (NLA) and Ang II type <em>1</em> receptor blockade with losartan produces vasodilation during local heating in controls. We tested whether this <em>angiotensin</em>-mediated vasodilation occurs in POTS and whether it is related to <em>angiotensin</em>-converting enzyme 2 (ACE2) and Ang-(<em>1</em>-<em>7</em>). We used local cutaneous heating to 42 degrees C and laser Doppler Flowmetry to assess NO-dependent conductance at 4 calf sites in <em>1</em>2 low-flow POTS and in <em>1</em>2 control subjects <em>1</em><em>7</em>.6 to 25.5 years of age. We perfused Ringer's solution through intradermal microdialysis catheters and performed local heating. We perfused one catheter with NLA (<em>1</em>0 mmol/L)+losartan (2 microg/L) and repeated heating, and NLA+losartan+Ang II (<em>1</em>0 micromol/L), repeating heating a third time. A second catheter received NLA+losartan+Ang II, heated, perfused NLA+losartan+Ang II+DX600 (<em>1</em> mmol/L; a selective ACE2 inhibitor), and reheated. A third catheter received NLA+losartan+Ang II, heated, perfused NLA+losartan+Ang II+Ang-(<em>1</em>-<em>7</em>) (<em>1</em>00 micromol/L), and reheated. The fourth catheter received Ang-(<em>1</em>-<em>7</em>) then reheated a second time only. <em>Angiotensin</em>-mediated vasodilation was present in control but not POTS. Ang-mediated dilation was eliminated by DX600, indicating an ACE2-related effect. Ang-mediated vasodilation was restored in POTS by Ang-(<em>1</em>-<em>7</em>). When administered alone during locally mediated heating, Ang-(<em>1</em>-<em>7</em>) improved the NO-dependent local heating response. ACE2 effects are blunted in low-flow POTS and restored by the ACE2 product Ang-(<em>1</em>-<em>7</em>). Data imply impaired catabolism of Ang II through the ACE2 pathway. Vasoconstriction in POTS may result from a reduction in Ang-(<em>1</em>-<em>7</em>) and an increase in Ang II.

Bone marrow-derived mesenchymal stem cells (MSCs), which have beneficial effects in acute lung injury (ALI), can serve as a vehicle for gene therapy. <em>Angiotensin</em>-converting enzyme 2 (ACE2), a counterregulatory enzyme of ACE that degrades <em>angiotensin</em> (Ang) II into Ang <em>1</em>-<em>7</em>, has a protective role against ALI. Because ACE2 expression is severely reduced in the injured lung, a therapy targeted to improve ACE2 expression in lung might attenuate ALI. We hypothesized that MSCs overexpressing ACE2 would have further benefits in lipopolysaccharide (LPS)-induced ALI mice, when compared with MSCs alone. MSCs were transduced with ACE2 gene (MSC-ACE2) by a lentiviral vector and then infused into wild-type (WT) and ACE2 knockout (ACE2(-/y)) mice following an LPS-induced intratracheal lung injury. The results demonstrated that the lung injury of ALI mice was alleviated at 24 and <em>7</em>2 h after MSC-ACE2 transplantation. MSC-ACE2 improved the lung histopathology and had additional anti-inflammatory effects when compared with MSCs alone in both WT and ACE2(-/y) ALI mice. MSC-ACE2 administration also reduced pulmonary vascular permeability, improved endothelial barrier integrity, and normalized lung eNOS expression relative to the MSC group. The beneficial effects of MSC-ACE2 could be attributed to its recruitment into the injured lung and enhanced local expression of ACE2 protein without changing the serum ACE2 levels after MSC-ACE2 transplantation. The biological activity of the increased ACE2 protein decreased the Ang II amount and increased the Ang <em>1</em>-<em>7</em> level in the lung when compared with the ALI and MSC-only groups, thereby inhibiting the detrimental effects of accumulating Ang II. Therefore, compared to MSCs alone, the administration of MSCs overexpressing ACE2 resulted in a further improvement in the inflammatory response and pulmonary endothelial function of LPS-induced ALI mice. These additional benefits could be due to the degradation of Ang II that accompanies the targeted overexpression of ACE2 in the lung.

The Mas protooncogene encodes a G protein-coupled receptor, we identified, also by using the specific <em>angiotensin</em>-(<em>1</em>-<em>7</em>) antagonist A-<em>7</em><em>7</em>9, to be associated with intracellular signaling of the <em>angiotensin</em> (Ang) II metabolite Ang-(<em>1</em>-<em>7</em>). Recently, Mas-related genes (Mrg) have been identified coding for the Mrg-receptor family. All family members share high sequence homology to Mas. Most of them are orphan receptors. To proof whether structure similarities of the Mrg receptors with Mas turn them into potential receptors for Ang-(<em>1</em>-<em>7</em>) or other Ang metabolites, we transfected COS or HEK293 cells with an assortment of Mrg receptors and investigated arachidonic acid (AA) release and transcriptional activation by recording serum response factor (SRF) activation after stimulation with Ang II, Ang III, Ang IV, and Ang-(<em>1</em>-<em>7</em>). None of the investigated receptors activated transcription via SRF. Ang-(<em>1</em>-<em>7</em>) stimulated AA release already in control vector-transfected COS cells, indicating the existence of an endogenous receptor (A-<em>7</em><em>7</em>9 sensitive). Though less pronounced than for Mas, two of the six studied receptors (MrgD, MRG) initiated significant AA release after stimulation with Ang-(<em>1</em>-<em>7</em>). Interestingly, Mas, MrgD, and MRG mediated Ang IV-stimulated AA release that was highest for Mas. While Ang III activated Mas and MrgX2, Ang II stimulated AA release via Mas and MRG. Thus, we identified other receptors of the Mrg family to respond on Ang-(<em>1</em>-<em>7</em>) stimulation. Furthermore, we describe first an AT(<em>1</em>)-independent direct Ang IV signaling and show that Ang II and Ang III mediate signaling independent of their specific receptors AT(<em>1</em>) and AT(2), whereby the receptor specificity differs.

OBJECTIVE

Polycystic liver disease (PLD), the most common extrarenal manifestation of autosomal-dominant polycystic kidney disease (ADPKD), has become more prevalent as a result of increased life expectancy, improved renal survival, reduced cardiovascular mortality, and renal replacement therapy. No studies have fully characterized PLD in large cohorts. We investigated whether liver and cyst volumes are associated with volume of the hepatic parenchyma, results from liver laboratory tests, and patient-reported outcomes.

METHODS

We performed a cross-sectional analysis of baseline liver volumes, measured by magnetic resonance imaging, and their association with demographics, results from liver laboratory and other tests, and quality of life. The data were collected from a randomized, placebo-controlled trial underway at <em>7</em> tertiary-care medical centers to determine whether the combination of an <em>angiotensin</em> I-converting enzyme inhibitor and <em>angiotensin</em> II-receptor blocker was superior to the inhibitor alone, and whether low blood pressure ((<em>1</em><em>1</em>0/<em>7</em>5 mm Hg) was superior to standard blood pressure (<em>1</em>20-<em>1</em>30/<em>7</em>0-80 mm Hg), in delaying renal cystic progression in 558 patients with ADPKD, stages <em>1</em> and 2 chronic kidney disease, and hypertension (age, <em>1</em>5-49 y).

RESULTS

We found hepatomegaly to be common among patients with ADPKD. Cysts and parenchyma contributed to hepatomegaly. Cysts were more common and liver and cyst volumes were greater in women, increasing with age. Patients with advanced disease had a relative loss of liver parenchyma. We observed small abnormalities in results from liver laboratory tests, and that splenomegaly and hypersplenism were associated with PLD severity. Higher liver volumes were associated with a lower quality of life.

CONCLUSIONS

Hepatomegaly is common even in early stage ADPKD and is not accounted for by cysts alone. Parenchymal volumes were larger, compared with liver volumes of patients without ADPKD or with those predicted by standardized equations, even among patients without cysts. The severity of PLD was associated with altered biochemical and hematologic features, as well as quality of life. ClinicalTrials.gov identifier: NCT00283686.

<em>Angiotensin</em> converting enzyme-2 (ACE2) is the receptor for the coronavirus SARS-CoV-2, which causes COVID-<em>1</em>9. We propose the following hypothesis: Imbalance in the action of ACE<em>1</em>- and ACE2-derived peptides, thereby enhancing <em>Angiotensin</em>-II (ANG II) signaling, a primary driver of COVID-<em>1</em>9 pathobiology. ACE<em>1</em>/ACE2 imbalance occurs due to the binding of SARS-CoV-2 to ACE2, reducing ACE2-mediated conversion of ANG II to ANG peptides that counteract pathophysiological effects of ACE<em>1</em>-generated ANGII. This hypothesis suggests several approaches to treat COVID-<em>1</em>9 by restoring ACE<em>1</em>/ACE2 balance: <em>1</em>) ANG II receptor blockers (ARBs); 2) ACE<em>1</em> inhibitors (ACEIs); 3) Agonists of receptors activated by ACE2-derived peptides [e.g., ANG (<em>1</em>-<em>7</em>), which activates MAS<em>1</em>]; 4) Recombinant human ACE2 or ACE2 peptides as decoys for the virus. Reducing ACE<em>1</em>/ACE2 imbalance is predicted to blunt COVID-<em>1</em>9-associated morbidity and mortality, especially in vulnerable patients. Importantly, approved ARBs and ACEIs can be rapidly repurposed to test their efficacy in treating COVID-<em>1</em>9.

By using a conventional spectrophotometric assay with hippuryl-L-phenylalanine as the substrate, <em>1</em>0(6) BALB/c mouse serosal mast cells possessed <em>1</em>.5 +/- 0.43 U (mean +/- SE, n = 5, range = 0.48 to 2.5) of carboxypeptidase A activity, while T cell factor-dependent, mouse bone marrow-derived mast cells (BMMC) had barely detectable levels of 0.0<em>1</em> +/- 0.00<em>1</em> U/<em>1</em>0(6) cells (mean +/- SE, n = 3). In order to characterize the carboxypeptidase A present in the BMMC, a sensitive assay was developed that used <em>angiotensin</em> I as the substrate and reverse phase-high performance liquid chromatography to separate and quantify production of the cleavage product des-leu-<em>angiotensin</em> I. Using this assay, mouse BMMC carboxypeptidase A had a neutral to basic pH optimum and hydrolyzed <em>angiotensin</em> I with a Km of 0.<em>7</em>8 mM. The antigen-induced net percent release of carboxypeptidase A from IgE-sensitized BMMC was proportional to that of the secretory granule component beta-hexosaminidase which indicates a secretory granule location for the exopeptidase. As defined by exclusion during Sepharose CL-2B chromatography, carboxypeptidase A was exocytosed as a greater than <em>1</em> X <em>1</em>0(<em>7</em>) m.w. complex bound to proteoglycans. Because BMMC cocultured with mouse skin-derived 3T3 fibroblasts are known to undergo an increase in histamine content and biosynthesis of 35S-labeled heparin proteoglycans, carboxypeptidase A activity was measured during BMMC/fibroblast coculture for 0 to 28 days. The carboxypeptidase A activity increased progressively during 28 days of co-culture from 0.004 +/- 0.002 U/<em>1</em>0(6) starting BMMC (mean +/- SE, n = 3) to 0.36 +/- 0.<em>1</em>0 U/<em>1</em>0(6) co-cultured mast cells. These findings indicate that carboxypeptidase A, a neutral protease, is exocytosed from the secretory granules of mouse mast cells bound to proteoglycan and is increased during the in vitro differentiation of mouse BMMC from mucosal-like mast cells to serosal-like mast cells.

Components of the renin-<em>angiotensin</em> system are well established targets for pharmacological intervention in a variety of disorders. Many such therapies abrogate the effects of the hypertensive and mitogenic peptide, <em>angiotensin</em> II, by antagonising its interaction with its receptor, or by inhibiting its formative enzyme, <em>angiotensin</em>-converting enzyme (ACE). At the turn of the millennium, a homologous enzyme, termed ACE2, was identified which increasingly shares the limelight with its better-known homologue. In common with ACE, ACE2 is a type I transmembrane metallopeptidase; however, unlike ACE, ACE2 functions as a carboxypeptidase, cleaving a single C-terminal residue from a distinct range of substrates. One such substrate is <em>angiotensin</em> II, which is hydrolysed by ACE2 to the vasodilatory peptide <em>angiotensin</em> <em>1</em>-<em>7</em>. In this commentary we discuss the latest developments in the rapidly progressing study of the physiological and patho-physiological roles of ACE2 allied with an overview of the current understanding of its molecular and cell biology. We also discuss parallel developments in the study of collectrin, a catalytically inactive homologue of ACE2 with critical functions in the pancreas and kidney.

The secretion of cortisol and other steroids from adrenal tumors can be regulated by hormones other than corticotropin following the aberrant expression of several G-protein-coupled receptors (GPCRs). To date, ectopic receptors for gastric inhibitory polypeptide, beta-adrenergic receptor agonists, vasopressin (V(2) and V(3) receptors), 5-hydroxytryptamine (5-HT(<em>7</em>) receptor) and, probably, <em>angiotensin</em> II (AT(<em>1</em>) receptor) have been identified. Either increased expression or altered activity of eutopic receptors for vasopressin (V(<em>1</em>)), luteinizing hormone/human chorionic gonadotropin, 5-HT (5-HT(4) receptor) and leptin might also be involved. One or more aberrant receptors can be present in unilateral tumors and bilateral macronodular adrenal hyperplasia, at either the early subclinical or overt stages of hormone secretion. The identification of aberrant adrenal GPCRs offers the potential for novel pharmacological therapies that either suppress the endogenous ligands or block the receptor with specific antagonists.

The identification of novel biochemical components of the renin-<em>angiotensin</em> system (RAS) has added a further layer of complexity to the classical concept of this cardiovascular regulatory system. It is now clear that there is a counter-regulatory arm within the RAS that is mainly formed by the <em>angiotensin</em>-converting enzyme 2-<em>angiotensin</em> (<em>1</em>-<em>7</em>)-receptor Mas axis. The functions of this axis are often opposite to those attributed to the major component of the RAS, <em>angiotensin</em> II. This review will highlight the current knowledge concerning the cardiovascular effects of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) through a direct interaction with its receptor Mas or through an indirect interplay with the kallikrein-kinin system. In addition, there will be a discussion of its role in the beneficial effects of <em>angiotensin</em>-converting enzyme inhibitors and angio-tensin receptor type <em>1</em> (AT<em>1</em>) antagonists, and the potential of this peptide and its receptor as a novel targets for new cardiovascular drugs.

<em>1</em>. The aim of the present study was to determine whether inhibition of dipeptidyl peptidase IV (DPP IV) elevates arterial blood pressure and whether any such effect is dependent on genetic background, the sympathetic nervous system and Y(<em>1</em>) receptors. The rationale behind this study was that: (i) neuropeptide (NP) Y(<em>1</em>-36) and peptide YY(<em>1</em>-36) (PYY(<em>1</em>-36)) are endogenous Y(<em>1</em>) receptor agonists and are metabolised by DPP IV to NPY(3-36) and PYY(3-36), which are not Y(<em>1</em>) but rather selective Y(2) receptor agonists; (ii) Y(<em>1</em>) receptors mediate vasoconstriction, whereas Y(2) receptors have little effect on vascular tone; (iii) vaso-constrictor effect of the Y(<em>1</em>) receptor is enhanced in spontaneously hypertensive rats (SHR) compared with normotensive Wistar-Kyoto (WKY) rats; and (iv) NPY(<em>1</em>-36) is released from sympathetic nerve terminals. 2. We examined the effects of acute administration of 3-N-[(2S,3S)-2-amino-3-methylpentanoyl]-<em>1</em>,3-thiazolidine (P32/98; a DPP IV inhibitor) on arterial blood pressure in anaesthetized adult SHR and WKY rats in the absence and presence of either captopril, hydralazine or chlorisondamine to lower basal mean arterial blood pressure (MABP) by different mechanisms (inhibition of <em>angiotensin</em>-converting enzyme, direct vasodilation and ganglionic blockade, respectively). 3. In naïve SHR with severely elevated basal blood pressures (MABP = <em>1</em><em>7</em>6 +/- 3 mmHg; n = 4), i.v. boluses (<em>1</em>, 3 and <em>1</em>0 mg/kg) of P32/98 did not affect blood pressure. 4. When basal blood pressure was reduced by pretreatment of SHR with either captopril (30 mg/kg, i.v.; MABP = <em>1</em><em>1</em>6 +/- 3 mmHg; n = 9) or hydralazine (5 mg/kg, i.p.; MABP = 84 +/- 3 mmHg; n = <em>7</em>), P32/98 (<em>1</em>, 3 and <em>1</em>0 mg/kg) caused significant dose-related increases in arterial blood pressure (4 +/- 2, <em>1</em>0 +/- 2 and <em>1</em>2 +/- 3 mmHg in the captopril-pretreated group, respectively (P < 0.0<em>1</em>); 5 +/- 2, 8 +/- 3 and <em>1</em><em>1</em> +/- 4 mmHg in the hydralazine-pretreated group, respectively (P < 0.0<em>1</em>)). 5. The increases in arterial blood pressure induced by P32/98 in captopril- or hydralazine-pretreated SHR were entirely blocked by pretreatment with the selective Y(<em>1</em>) receptor antagonist N2-(diphenylacetyl)-N-[(4-hydroxyphenyl)methyl]-d-arginine amide (BIBP 3226; 6 mg/kg per h). 6. When basal blood pressure was reduced in SHR by pretreatment with chlorisondamine (<em>1</em>0 mg/kg, s.c.; MABP = <em>1</em>08 +/- 4 mmHg; n = <em>7</em>), inhibition of DPP IV with P32/98 did not affect arterial blood pressure. Basal heart rate in chlorisondamine-treated SHR was significantly reduced compared with naïve SHR, captopril-pretreated SHR and hydralazine-pretreated SHR, indicating effectiveness of ganglionic blockade. <em>7</em>. Unlike the results in genetically hypertensive animals, in normotensive WKY rats pretreated with captopril (30 mg/kg, i.v.; MABP = 8<em>1</em> +/- 4 mmHg; n = 6), or hydralazine (5 mg/kg, i.p.; MABP = 63 +/- 4 mmHg; n = 4) or chlorisondamine (<em>1</em>0 mg/kg, s.c.; MABP = 63 +/- 4 mmHg; n = 5), P32/98 did not affect arterial blood pressure. 8. We conclude that, in genetically susceptible animals, inhibition of DPP IV increases arterial blood pressure via Y(<em>1</em>) receptors when elevated blood pressure is reduced with antihypertensive drugs provided that the sympathetic nervous system is functional. The results suggest vigilance because DPP IV inhibitors are used more widely in hypertensive patients treated with antihypertensive drugs.

Hyperactivity of the renin-<em>angiotensin</em> system (RAS) resulting in elevated <em>Angiotensin</em> II (Ang II) contributes to all stages of inflammatory responses including ocular inflammation. The discovery of <em>angiotensin</em>-converting enzyme 2 (ACE2) has established a protective axis of RAS involving ACE2/Ang-(<em>1</em>-<em>7</em>)/Mas that counteracts the proinflammatory and hypertrophic effects of the deleterious ACE/AngII/AT<em>1</em>R axis. Here we investigated the hypothesis that enhancing the systemic and local activity of the protective axis of the RAS by oral delivery of ACE2 and Ang-(<em>1</em>-<em>7</em>) bioencapsulated in plant cells would confer protection against ocular inflammation. Both ACE2 and Ang-(<em>1</em>-<em>7</em>), fused with the non-toxic cholera toxin subunit B (CTB) were expressed in plant chloroplasts. Increased levels of ACE2 and Ang-(<em>1</em>-<em>7</em>) were observed in circulation and retina after oral administration of CTB-ACE2 and Ang-(<em>1</em>-<em>7</em>) expressing plant cells. Oral feeding of mice with bioencapsulated ACE2/Ang-(<em>1</em>-<em>7</em>) significantly reduced endotoxin-induced uveitis (EIU) in mice. Treatment with bioencapsulated ACE2/Ang-(<em>1</em>-<em>7</em>) also dramatically decreased cellular infiltration, retinal vasculitis, damage and folding in experimental autoimmune uveoretinitis (EAU). Thus, enhancing the protective axis of RAS by oral delivery of ACE2/Ang-(<em>1</em>-<em>7</em>) bioencapsulated in plant cells provide an innovative, highly efficient and cost-effective therapeutic strategy for ocular inflammatory diseases.

In the present quantitative overview of outcome trials, we investigated the efficacy of amlodipine or <em>angiotensin</em> receptor blockers in the prevention of stroke and myocardial infarction in patients with hypertension, coronary artery disease, or diabetic nephropathy. The analysis included <em>1</em>2 trials of 94 338 patients. The analysis of trials involving an amlodipine group showed that amlodipine provided more protection against stroke and myocardial infarction than other antihypertensive drugs, including <em>angiotensin</em> receptor blockers (-<em>1</em>9%, P<0.000<em>1</em> and -<em>7</em>%, P=0.03) and placebo (-3<em>7</em>%, P=0.06 and -29%, P=0.04). The analysis of trials involving an <em>angiotensin</em> receptor blocker group showed contrasting results between trials versus amlodipine and trials versus other antihypertensive drugs for stroke (+<em>1</em>9% versus -25%; P<0.000<em>1</em>) and myocardial infarction (+2<em>1</em>% versus +<em>1</em>%; P=0.03). The results of 3 trials comparing an <em>angiotensin</em> receptor blocker with placebo were neutral (P> or =0.<em>1</em>4). The within-trial between-group difference in achieved systolic pressure ranged from -<em>1</em>.<em>1</em> to +4.<em>7</em> mm Hg for trials involving an amlodipine group and from -2.8 to +4.0 mm Hg for trials involving an <em>angiotensin</em> receptor blocker group. The metaregression analysis correlating odds ratios with blood pressure differences showed a negative relationship (regression coefficients: -3% to -8%), which reached statistical significance (regression coefficient: -6%; P=0.0<em>1</em>) for stroke in trials involving an amlodipine group. In conclusion, blood pressure differences largely accounted for cardiovascular outcome.

Cardiac myocyte hypertrophy often occurs in response to both hemodynamic and neurohumoral factors. To study whether activation of the renin-<em>angiotensin</em> system by itself may induce a cardiac growth response, the acute effects of <em>angiotensin</em> II on cardiac protein synthesis were studied in isolated rat hearts. New protein synthesis in isolated buffer-perfused adult rat hearts was measured by incorporation of [3H]phenylalanine into cardiac proteins during a 3-hour perfusion protocol. <em>Angiotensin</em> II (<em>1</em> x <em>1</em>0(-8) mol/L), administered alone or in combination with the alpha <em>1</em>-blocker prazosin (<em>1</em> x <em>1</em>0(-<em>7</em>) mol/L), stimulated protein synthesis in both ventricles. The rate of [3H]phenylalanine incorporation into cardiac proteins was 3.9-fold (P < .005) and 2.6-fold (P < .0<em>1</em>) higher in <em>angiotensin</em> II-perfused (n = 6) than in vehicle-perfused (n = 6) left and right ventricles, respectively. The induction of new protein synthesis by <em>angiotensin</em> II was blocked by the <em>angiotensin</em> II type <em>1</em> (AT<em>1</em>) receptor antagonist losartan (<em>1</em> x <em>1</em>0(-<em>7</em>) mol/L, n = 5). To study the pathways of <em>angiotensin</em> signal transduction, protein kinase C (PKC)-epsilon as well as cardiac c-fos and c-jun mRNA levels were analyzed. <em>Angiotensin</em> II (<em>1</em> x <em>1</em>0(-8) mol/L, n = 20) resulted in a transient translocation of PKC-epsilon from the cytosol to the cellular membrane. However, compared with phorbol ester stimulation (phorbol <em>1</em>2-myristate <em>1</em>3-acetate [PMA], <em>1</em> x <em>1</em>0(-<em>7</em>) mol/L; n = 20), <em>angiotensin</em> II effects on PKC translocation were significantly less pronounced and required a more prolonged stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

BACKGROUND

The renin-angiotensin system is thought to be involved in development and progression of arteriosclerosis, thereby contributing to adverse cardiovascular events. To elucidate the role of angiotensin II (Ang II) at a cellular level, we analyzed the Ang II-induced gene expression profile.

RESULTS

Genes induced on Ang II stimulation (10(-7) mol/L, 45 minutes) in rat smooth muscle cells were analyzed by polymerase chain reaction selected subtraction. In addition to known genes, such as interleukin 6, leukemia inhibitory factor, and c-fos, we identified CYR61, an angiogenic immediate early gene. Northern blot analysis revealed a rapid 2.5-fold increase of CYR61 transcript levels by Ang II, peaking at 30 minutes, which was blunted by Ang II type 1 receptor blockade. Exposure of rat aortic rings to Ang II (30 minutes) revealed a 2-fold, and intraperitoneal injection of Ang II (30 minutes) in mice a 3-fold, increase of aortic CYR61 transcripts. In arteriosclerotic aortas of apolipoprotein E-deficient mice, CYR61 transcripts confirmed by in situ hybridization and proteins shown by immunohistochemistry were elevated, whereas they were hardly detectable in wild types. In human carotid atherectomies and arteriosclerotic coronary arteries, immunohistochemical analysis revealed expression of CYR61 within connective tissue in neointima, adventitia, and surrounding small capillaries and blood vessels, colocalized with ACE and Ang II. Normal human arteries showed no significant staining for CYR61.

CONCLUSIONS

CYR61, an angiogenic factor, is induced by Ang II in vascular cells and tissue. The expression of CYR61, colocalized with Ang II and ACE, in small vessels of human arteriosclerotic lesions is consistent with the notion that the activated renin-angiotensin system may contribute to plaque neovascularization by enhancing regulators of microvessel formation and cell proliferation.

BACKGROUND

Patients with ischemic heart disease and preserved ventricular function experience considerable morbidity and mortality despite standard medical therapy.

OBJECTIVE

To compare benefits and harms of using angiotensin-converting enzyme (ACE) inhibitors, angiotensin II-receptor blockers (ARBs), or combination therapy in adults with stable ischemic heart disease and preserved ventricular function.

METHODS

MEDLINE, EMBASE, Cochrane Central Register of Controlled Trials, and Cochrane Database of Systematic Reviews (earliest date, July 2009) were searched without language restrictions.

METHODS

Two independent investigators screened citations for trials of at least 6 months' duration that compared ACE inhibitors, ARBs, or combination therapy with placebo or active control and reported any of several clinical outcomes.

METHODS

Using standardized protocols, 2 independent investigators extracted information about study characteristics and rated the quality and strength of evidence. Disagreement was resolved by consensus.

RESULTS

41 studies met eligibility criteria. Moderate- to high-strength evidence (7 trials; 32 559 participants) showed that ACE inhibitors reduce the relative risk (RR) for total mortality (RR, 0.87 [95% CI, 0.81 to 0.94]) and nonfatal myocardial infarction (RR, 0.83 [CI, 0.73 to 0.94]) but increase the RR for syncope (RR, 1.24 [CI, 1.02 to 1.52]) and cough (RR, 1.67 [CI, 1.22 to 2.29]) compared with placebo. Low-strength evidence (1 trial; 5926 participants) suggested that ARBs reduce the RR for the composite end point of cardiovascular mortality, nonfatal myocardial infarction, or stroke (RR, 0.88 [CI, 0.77 to 1.00]) but not for the individual components. Moderate-strength evidence (1 trial; 25 620 participants) showed similar effects on total mortality (RR, 1.07 [CI, 0.98 to 1.16]) and myocardial infarction (RR, 1.08 [CI, 0.94 to 1.23]) but an increased risk for discontinuations because of hypotension (P < 0.001) and syncope (P = 0.035) with combination therapy compared with ACE inhibitors alone.

CONCLUSIONS

Many studies either did not assess or did not report harms in a systematic manner. Many studies did not adequately report benefits or harms by various patient subgroups.

CONCLUSIONS

Adding an ACE inhibitor to standard medical therapy improves outcomes, including reduced risk for mortality and myocardial infarctions, in some patients with stable ischemic heart disease and preserved ventricular function. Less evidence supports a benefit of ARB therapy, and combination therapy seems no better than ACE inhibitor therapy alone and increases harms.

BACKGROUND

Agency for Healthcare Research and Quality.

The <em>angiotensin</em> II (AII) antagonistic action of 2-ethoxy-<em>1</em>-[[2'-(<em>1</em>H-tetrazol-5-yl)biphenyl-4-yl]methyl-<em>1</em>H-benzi mid azole-<em>7</em> - carboxylic acid (CV-<em>1</em><em>1</em>9<em>7</em>4) was examined in in vitro assay systems, including AII receptor binding assay using membrane fractions of bovine adrenal cortex or rabbit aorta and AII-induced contraction assay using rabbit aortic strips, and CV-<em>1</em><em>1</em>9<em>7</em>4 and its prodrug, (+/-)<em>1</em>-(cyclohexyloxycarbonyloxy)ethyl 2-ethoxy-<em>1</em>-[[2'-(<em>1</em>H-tetrazol-5-yl)biphenyl-4-yl]methyl]-<em>1</em>H- benzimidazole-<em>7</em>-carboxylate (TCV-<em>1</em><em>1</em>6), were examined in an in vivo system of AII-induced pressor response in conscious rats. DuP <em>7</em>53 or EXP3<em>1</em><em>7</em>4 (the main active metabolite of DuP <em>7</em>53) was used as the reference compound. CV-<em>1</em><em>1</em>9<em>7</em>4 inhibited the binding of [<em>1</em>25I] AII to the bovine adrenal cortical membrane and rabbit aortic membrane with IC50 values of <em>1</em>.<em>1</em>2 x <em>1</em>0(-<em>7</em>) and 2.86 x <em>1</em>0(-8) M, respectively. Similar results were obtained with EXP3<em>1</em><em>7</em>4. CV-<em>1</em><em>1</em>9<em>7</em>4 interacted with AII in these membrane fractions with subtype <em>1</em> receptor in a competitive manner. CV-<em>1</em><em>1</em>9<em>7</em>4 at <em>1</em>0(-5) M did not affect the binding of [<em>1</em>25I]AII to subtype 2 (AT2) receptor in bovine cerebellum. CV-<em>1</em><em>1</em>9<em>7</em>4 selectively inhibited the AII-induced contraction of rabbit aortic strips in a noncompetitive manner (pD' 2, 9.9<em>7</em>); it had no effects on the contraction induced by norepinephrine, KCl, serotonin, prostaglandin F2 alpha or endothelin. EXP3<em>1</em><em>7</em>4 showed a pD'2 value of 8.95 for the AII-induced contraction. CV-<em>1</em><em>1</em>9<em>7</em>4 given intravenously and TCV-<em>1</em><em>1</em>6 given orally inhibited the AII-induced pressor response in rats with ID50 values of 0.033 mg/kg and 0.069 mg/kg, respectively. These effects of CV-<em>1</em><em>1</em>9<em>7</em>4 and TCV-<em>1</em><em>1</em>6 were <em>1</em>2 and 48 times more potent than those of EXP3<em>1</em><em>7</em>4 and DuP <em>7</em>53, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)