Epoxyeicosatrienoic acids (EET) have antihypertensive and anti-inflammatory properties and play a role in the maintenance of renal vascular function. A novel approach to increase EET levels is to inhibit epoxide hydrolase enzymes that are responsible for conversion of biologically active EET to dihydroxyeicosatrienoic acids (DHET). We hypothesized that soluble epoxide hydrolase (SEH) inhibition would improve renal vascular function and ameliorate hypertension induced renal damage. Chronic administration of the specific SEH inhibitor <em>1</em>-cyclohexyl-3-dodecylurea (CDU, 3 mg/d) for <em>1</em>0 d lowered BP in <em>angiotensin</em> hypertensive rats. The contribution of renal vascular SEH to afferent arteriolar function in <em>angiotensin</em> hypertension was also assessed. SEH protein expression was increased in renal microvessels from hypertensive rats. Although CDU did not change afferent arteriolar responsiveness to <em>angiotensin</em> in normotensive animals, CDU treatment significantly attenuated afferent arteriolar diameter responses to <em>angiotensin</em> in hypertensive kidneys from 5<em>1</em>% +/- 8% to 28% +/- <em>7</em>%. Protection of the renal vasculature and glomerulus during chronic CDU administration was demonstrated by histology. Urinary albumin excretion, an index of renal damage, was also lower in CDU-treated hypertensive rats. These data demonstrate that SEH inhibition has antihypertensive and renal vascular protective effects in <em>angiotensin</em> hypertension and suggests that SEH inhibitors may be a useful therapeutic intervention for cardiovascular diseases.

We studied the distribution of nephrin in renal biopsies from <em>1</em><em>7</em> patients with diabetes and nephrotic syndrome (<em>7</em> type <em>1</em> and <em>1</em>0 type 2 diabetes), 6 patients with diabetes and microalbuminuria (<em>1</em> type <em>1</em> and 5 type 2 diabetes), and <em>1</em>0 normal subjects. Nephrin expression was semiquantitatively evaluated by measuring immunofluorescence intensity by digital image analysis. We found an extensive reduction of nephrin staining in both type <em>1</em> (6<em>7</em> +/- 9%; P < 0.00<em>1</em>) and type 2 (65 +/- <em>1</em>0%; P < 0.00<em>1</em>) diabetic patients with diabetes and nephrotic syndrome when compared with control subjects. The pattern of staining shifted from punctate/linear distribution to granular. In patients with microalbuminuria, the staining pattern of nephrin also showed granular distribution and reduction intensity of 69% in the patient with type <em>1</em> diabetes and of 62 +/- 4% (P < 0.00<em>1</em>) in the patients with type 2 diabetes. In vitro studies on human cultured podocytes demonstrated that glycated albumin and <em>angiotensin</em> II reduced nephrin expression. Glycated albumin inhibited nephrin synthesis through the engagement of receptor for advanced glycation end products, whereas <em>angiotensin</em> II acted on cytoskeleton redistribution, inducing the shedding of nephrin. This study indicates that the alteration in nephrin expression is an early event in proteinuric patients with diabetes and suggests that glycated albumin and <em>angiotensin</em> II contribute to nephrin downregulation.

The novel SARS coronavirus SARS-CoV-2 pandemic may be particularly deleterious to patients with underlying cardiovascular disease (CVD). The mechanism for SARS-CoV-2 infection is the requisite binding of the virus to the membrane-bound form of <em>angiotensin</em>-converting enzyme 2 (ACE2) and internalization of the complex by the host cell. Recognition that ACE2 is the co-receptor for the coronavirus has prompted new therapeutic approaches to block the enzyme or reduce its expression to prevent the cellular entry and SARS-CoV-2 infection in tissues that express ACE2 including lung, heart, kidney, brain, and gut. ACE2, however, is a key enzymatic component of the renin-<em>angiotensin</em>-aldosterone system (RAAS); ACE2 degrades Ang II, a peptide with multiple actions that promote CVD, and generates Ang-(<em>1</em>-<em>7</em>) which antagonizes the effects of Ang II. Moreover, experimental evidence suggests that RAAS blockade by ACE inhibitors, AT_<em>1</em> receptor antagonists and mineralocorticoid antagonists, as well as statins enhance ACE2 that, in part, contribute to the benefit of these regimens. In lieu of the fact that many older patients with hypertension or other CVDs are routinely treated with RAAS blockers and statins, new clinical concerns have developed regarding whether these patients are at greater risk for SARS-CoV-2 infection, whether RAAS and statin therapy should be discontinued, and the potential consequences of RAAS blockade to COVID-<em>1</em>9-related pathologies such as acute and chronic respiratory disease. The current perspective critically examines the evidence for ACE2 regulation by RAAS blockade and statins, the cardiovascular benefits of ACE2, and whether ACE2 blockade is a viable approach to attenuate COVID-<em>1</em>9.

<em>Angiotensin</em>-converting enzyme 2 (ACE2) is the first human homologue of ACE to be described. ACE2 is a type I integral membrane protein that functions as a carboxypeptidase, cleaving a single hydrophobic/basic residue from the COOH-terminus of its substrates. Because ACE2 efficiently hydrolyzes the potent vasoconstrictor <em>angiotensin</em> II to <em>angiotensin</em> (<em>1</em>-<em>7</em>), this has changed our overall perspective about the classical view of the renin <em>angiotensin</em> system in the regulation of hypertension and heart and renal function, because it represents the first example of a feedforward mechanism directed toward mitigation of the actions of <em>angiotensin</em> II. This paper reviews the new data regarding the biochemistry of <em>angiotensin</em>-(<em>1</em>-<em>7</em>)-forming enzymes and discusses key findings such as the elucidation of the regulatory mechanisms participating in the expression of ACE2 and <em>angiotensin</em>-(<em>1</em>-<em>7</em>) in the control of the circulation.

Recent advances have improved our understanding of the renin-<em>angiotensin</em> system (RAS). These have included the recognition that <em>angiotensin</em> (Ang)-(<em>1</em>-<em>7</em>) is a biologically active product of the RAS cascade. The identification of the ACE homologue ACE2, which forms Ang-(<em>1</em>-<em>7</em>) from Ang II, and the GPCR Mas as an Ang-(<em>1</em>-<em>7</em>) receptor have provided the necessary biochemical and molecular background and tools to study the biological significance of Ang-(<em>1</em>-<em>7</em>). Most available evidence supports a counter-regulatory role for Ang-(<em>1</em>-<em>7</em>) by opposing many actions of Ang II on AT₁ receptors, especially vasoconstriction and proliferation. Many studies have now shown that Ang-(<em>1</em>-<em>7</em>) by acting via Mas receptor exerts inhibitory effects on inflammation and on vascular and cellular growth mechanisms. Ang-(<em>1</em>-<em>7</em>) has also been shown to reduce key signalling pathways and molecules thought to be relevant for fibrogenesis. Here, we review recent findings related to the function of the ACE2/Ang-(<em>1</em>-<em>7</em>)/Mas axis and focus on the role of this axis in modifying processes associated with acute and chronic inflammation, including leukocyte influx, fibrogenesis and proliferation of certain cell types. More attention will be given to the involvement of the ACE2/Ang-(<em>1</em>-<em>7</em>)/Mas axis in the context of renal disease because of the known relevance of the RAS for the function of this organ and for the regulation of kidney inflammation and fibrosis. Taken together, this knowledge may help in paving the way for the development of novel treatments for chronic inflammatory and renal diseases.

BACKGROUND

Tumor necrosis factor-alpha (TNF-alpha) and angiotensin II (Ang II) modulate heart failure in part by provoking the hypertrophic response. Signal transduction pathways of those factors are implicated in reactive oxygen intermediates (ROIs). Therefore, we hypothesized that TNF-alpha and Ang II might cause myocyte hypertrophy via the generation of ROIs.

RESULTS

To test the hypothesis, we tested whether TNF-alpha and Ang II could induce the generation of ROIs and whether antioxidants such as butylated hydroxyanisole (BHA), vitamin E, and catalase might inhibit the hypertrophy in cultured neonatal rat cardiac myocytes. ROIs were measured by the ROI-specific probe 2',7'-dichlorofluorescin diacetate in cultured cardiac myocytes. We demonstrated that TNF-alpha and Ang II induced the generation of ROIs in a dose-dependent manner. TNF-alpha (10 ng/mL) and Ang II (100 nmol/L) enlarged cardiac myocytes and increased [3H]leucine uptake, and BHA (10 micromol/L) significantly inhibited both effects. Other antioxidants, such as vitamin E (1 microg/mL) and catalase (100 U/mL), also inhibited the enlargement of cardiac myocytes induced by TNF-alpha.

CONCLUSIONS

These results indicate that TNF-alpha and Ang II cause hypertrophy in part via the generation of ROIs in cardiac myocytes.

BACKGROUND

Acute heart failure (AHF) in sub-Saharan Africa has not been well characterized. Therefore, we sought to describe the characteristics, treatment, and outcomes of patients admitted with AHF in sub-Saharan Africa.

METHODS

The Sub-Saharan Africa Survey of Heart Failure (THESUS-HF) was a prospective, multicenter, observational survey of patients with AHF admitted to <em>1</em>2 university hospitals in 9 countries. Among patients presenting with AHF, we determined the causes, treatment, and outcomes during 6 months of follow-up.

RESULTS

From July <em>1</em>, 200<em>7</em>, to June 30, 20<em>1</em>0, we enrolled <em>1</em>006 patients presenting with AHF. Mean (SD) age was 52.3 (<em>1</em>8.3) years, 5<em>1</em><em>1</em> (50.8%) were women, and the predominant race was black African (984 of 999 [98.5%]). Mean (SD) left ventricular ejection fraction was 39.5% (<em>1</em>6.5%). Heart failure was most commonly due to hypertension (n = 453 [45.4%]) and rheumatic heart disease (n = <em>1</em>43 [<em>1</em>4.3%]). Ischemic heart disease (n = <em>7</em><em>7</em> [<em>7</em>.<em>7</em>%]) was not a common cause of AHF. Concurrent renal dysfunction (estimated glomerular filtration rate, <30 mL/min/<em>1</em><em>7</em>3 m(2)), diabetes mellitus, anemia (hemoglobin level, (<em>1</em>0 g/dL), and atrial fibrillation were found in <em>7</em>3 (<em>7</em>.<em>7</em>%), <em>1</em><em>1</em>4 (<em>1</em><em>1</em>.4%), <em>1</em>4<em>7</em> (<em>1</em>5.2%), and <em>1</em>84 cases (<em>1</em>8.3%), respectively; 65 of 500 patients undergoing testing (<em>1</em>3.0%) were seropositive for the human immunodeficiency virus. The median hospital stay was <em>7</em> days (interquartile range, 5-<em>1</em>0), with an in-hospital mortality of 4.2%. Estimated <em>1</em>80-day mortality was <em>1</em><em>7</em>.8% (95% CI, <em>1</em>5.4%-20.6%). Most patients were treated with renin-<em>angiotensin</em> system blockers but not β-blockers at discharge. Hydralazine hydrochloride and nitrates were rarely used.

CONCLUSIONS

In African patients, AHF has a predominantly nonischemic cause, most commonly hypertension. The condition occurs in middle-aged adults, equally in men and women, and is associated with high mortality. The outcome is similar to that observed in non-African AHF registries, suggesting that AHF has a dire prognosis globally, regardless of the cause.

The incidence and prevalence of diabetes mellitus have grown significantly throughout the world, due primarily to the increase in type 2 diabetes. This overall increase in the number of people with diabetes has had a major impact on development of diabetic kidney disease (DKD), one of the most frequent complications of both types of diabetes. DKD is the leading cause of end-stage renal disease (ESRD), accounting for approximately 50% of cases in the developed world. Although incidence rates for ESRD attributable to DKD have recently stabilized, these rates continue to rise in high-risk groups such as middle-aged African Americans, Native Americans, and Hispanics. The costs of care for people with DKD are extraordinarily high. In the Medicare population alone, DKD-related expenditures among this mostly older group were nearly $25 billion in 20<em>1</em><em>1</em>. Due to the high human and societal costs, the Consensus Conference on Chronic Kidney Disease and Diabetes was convened by the American Diabetes Association in collaboration with the American Society of Nephrology and the National Kidney Foundation to appraise issues regarding patient management, highlighting current practices and new directions. Major topic areas in DKD included (<em>1</em>) identification and monitoring, (2) cardiovascular disease and management of dyslipidemia, (3) hypertension and use of renin-<em>angiotensin</em>-aldosterone system blockade and mineralocorticoid receptor blockade, (4) glycemia measurement, hypoglycemia, and drug therapies, (5) nutrition and general care in advanced-stage chronic kidney disease, (6) children and adolescents, and (<em>7</em>) multidisciplinary approaches and medical home models for health care delivery. This current state summary and research recommendations are designed to guide advances in care and the generation of new knowledge that will meaningfully improve life for people with DKD.

Blockers of the renin-<em>angiotensin</em>-aldosterone system (RAAS), that is, renin inhibitors, <em>angiotensin</em> (Ang)-converting enzyme (ACE) inhibitors, Ang II type <em>1</em> receptor antagonists, and mineralocorticoid receptor antagonists, are a cornerstone in the treatment of hypertension. How exactly they exert their effect, in particular in patients with low circulating RAAS activity, also taking into consideration the so-called Ang II/aldosterone escape that often occurs after initial blockade, is still incompletely understood. Multiple studies have tried to find parameters that predict the response to RAAS blockade, allowing a personalized treatment approach. Consequently, the question should now be answered on what basis (eg, sex, ethnicity, age, salt intake, baseline renin, ACE or aldosterone, and genetic variance) a RAAS blocker can be chosen to treat an individual patient. Are all blockers equal? Does optimal blockade imply maximum RAAS blockade, for example, by combining ≥2 RAAS blockers or by simply increasing the dose of <em>1</em> blocker? Exciting recent investigations reveal a range of unanticipated extrarenal effects of aldosterone, as well as a detailed insight in the genetic causes of primary aldosteronism, and mineralocorticoid receptor blockers have now become an important treatment option for resistant hypertension. Finally, apart from the deleterious ACE-Ang II-Ang II type <em>1</em> receptor arm, animal studies support the existence of protective aminopeptidase A-Ang III-Ang II type 2 receptor and ACE2-Ang-(<em>1</em> to <em>7</em>)-Mas receptor arms, paving the way for multiple new treatment options. This review provides an update about all these aspects, critically discussing the many controversies and allowing the reader to obtain a full understanding of what we currently know about RAAS alterations in hypertension.

Myocardial fibrosis is associated with an activated renin-<em>angiotensin</em>-aldosterone system (RAAS). In renovascular hypertension, this presents as a reactive perivascular and interstitial fibrosis in not only the pressure overloaded, hypertrophied left ventricle but also the normotensive, nonhypertrophied right ventricle. It therefore would appear that circulating hormonal and not hemodynamic factors are responsible for this adverse fibrous tissue response. To ascertain whether the RAAS effector hormones <em>angiotensin</em> II (AII) or aldosterone (ALDO) directly stimulate collagen synthesis or inhibit collagenase production we used cell culture. Adult rat cardiac fibroblasts (Fb) were cultured since these cells express mRNA for types I and III collagens, the major fibrillar collagens in the heart, and collagenase or matrix metalloproteinase <em>1</em> (MMP <em>1</em>), the key enzyme for interstitial collagen degradation. Collagen synthesis, determined by 3H-proline incorporation, and collagenase activity were measured in confluent, quiescent Fb after 24 h incubation with various concentrations of AII or ALDO (<em>1</em>0(-<em>1</em><em>1</em>)-<em>1</em>0(-6)M) in the presence or absence of either <em>1</em>0(-5)M type <em>1</em> (DuP <em>7</em>53) and type 2 (PD <em>1</em>23<em>1</em><em>7</em><em>7</em>) AII or <em>1</em>0(-9)-3 x <em>1</em>0(-6)M ALDO (spironolactone) receptor antagonists, respectively. Collagen synthesis, normalized per total protein synthesis, increased significantly (P < 0.005) after incubation with either <em>1</em>0(-9)M ALDO (5.9 +/- <em>1</em>.0%) or <em>1</em>0(-<em>7</em>)M AII (5.3 +/- <em>1</em>.2%) compared with untreated control cells (2.9 +/- 0.5%) of the same passage (p6-p<em>1</em>0). This increase in collagen synthesis could be completely abolished by either types <em>1</em> or 2 AII receptor antagonists in AII stimulated Fb or the competitive ALDO receptor antagonist, spironolactone, at equimolar concentration in ALDO stimulated Fb. AII significantly decreased collagenase activity which could be completely abolished by PD <em>1</em>23<em>1</em><em>7</em><em>7</em>, but not DuP <em>7</em>53, while ALDO had no effect on collagenase activity. The mineralocorticoid, ALDO, stimulates collagen synthesis in cultured adult rat cardiac Fb in concentrations similar to those found in plasma in renovascular hypertension and this response appears to occur via type I corticoid receptors. AII appears to stimulate collagen synthesis by both type <em>1</em> and 2 AII receptors, but only in high concentrations that could be generated locally within the myocardium. In addition, AII unlike ALDO inhibits collagenase activity that could be attenuated only by type 2 receptor blockade. These findings suggest a direct interaction between ALDO, AII and cardiac Fb in mediating myocardial fibrosis in hypertensive heart disease.

OBJECTIVE

To synthesize the cost-effectiveness (CE) of interventions to prevent and control diabetes, its complications, and comorbidities.

METHODS

We conducted a systematic review of literature on the CE of diabetes interventions recommended by the American Diabetes Association (ADA) and published between January <em>1</em>985 and May 2008. We categorized the strength of evidence about the CE of an intervention as strong, supportive, or uncertain. CEs were classified as cost saving (more health benefit at a lower cost), very cost-effective (<or=$25,000 per life year gained [LYG] or quality-adjusted life year [QALY]), cost-effective ($25,00<em>1</em> to $50,000 per LYG or QALY), marginally cost-effective ($50,00<em>1</em> to $<em>1</em>00,000 per LYG or QALY), or not cost-effective >>$<em>1</em>00,000 per LYG or QALY). The CE classification of an intervention was reported separately by country setting (U.S. or other developed countries) if CE varied by where the intervention was implemented. Costs were measured in 200<em>7</em> U.S. dollars.

RESULTS

Fifty-six studies from 20 countries met the inclusion criteria. A large majority of the ADA recommended interventions are cost-effective. We found strong evidence to classify the following interventions as cost saving or very cost-effective: (I) Cost saving- <em>1</em>) ACE inhibitor (ACEI) therapy for intensive hypertension control compared with standard hypertension control; 2) ACEI or <em>angiotensin</em> receptor blocker (ARB) therapy to prevent end-stage renal disease (ESRD) compared with no ACEI or ARB treatment; 3) early irbesartan therapy (at the microalbuminuria stage) to prevent ESRD compared with later treatment (at the macroalbuminuria stage); 4) comprehensive foot care to prevent ulcers compared with usual care; 5) multi-component interventions for diabetic risk factor control and early detection of complications compared with conventional insulin therapy for persons with type <em>1</em> diabetes; and 6) multi-component interventions for diabetic risk factor control and early detection of complications compared with standard glycemic control for persons with type 2 diabetes. (II) Very cost-effective- <em>1</em>) intensive lifestyle interventions to prevent type 2 diabetes among persons with impaired glucose tolerance compared with standard lifestyle recommendations; 2) universal opportunistic screening for undiagnosed type 2 diabetes in African Americans between 45 and 54 years old; 3) intensive glycemic control as implemented in the UK Prospective Diabetes Study in persons with newly diagnosed type 2 diabetes compared with conventional glycemic control; 4) statin therapy for secondary prevention of cardiovascular disease compared with no statin therapy; 5) counseling and treatment for smoking cessation compared with no counseling and treatment; 6) annual screening for diabetic retinopathy and ensuing treatment in persons with type <em>1</em> diabetes compared with no screening; <em>7</em>) annual screening for diabetic retinopathy and ensuing treatment in persons with type 2 diabetes compared with no screening; and 8) immediate vitrectomy to treat diabetic retinopathy compared with deferred vitrectomy.

CONCLUSIONS

Many interventions intended to prevent/control diabetes are cost saving or very cost-effective and supported by strong evidence. Policy makers should consider giving these interventions a higher priority.

<em>Angiotensin</em>-converting enzyme 2 (ACE2) shares some homology with <em>angiotensin</em>-converting enzyme (ACE) but is not inhibited by ACE inhibitors. The main role of ACE2 is the degradation of Ang II resulting in the formation of <em>angiotensin</em> <em>1</em>-<em>7</em> (Ang <em>1</em>-<em>7</em>) which opposes the actions of Ang II. Increased Ang II levels are thought to upregulate ACE2 activity, and in ACE2 deficient mice Ang II levels are approximately double that of wild-type mice, whilst Ang <em>1</em>-<em>7</em> levels are almost undetectable. Thus, ACE2 plays a crucial role in the RAS because it opposes the actions of Ang II. Consequently, it has a beneficial role in many diseases such as hypertension, diabetes, and cardiovascular disease where its expression is decreased. Not surprisingly, current therapeutic strategies for ACE2 involve augmenting its expression using ACE2 adenoviruses, recombinant ACE2 or compounds in these diseases thereby affording some organ protection.

This paper addresses clinical controversies and uncertainties regarding thoracic aortic aneurysm and its treatment. <em>1</em>) Estimating true aortic size is confounded by obliquity, asymmetry, and noncorresponding sites: both echocardiography and computed tomography/magnetic resonance imaging are necessary for complete assessment. 2) Epidemiology of thoracic aortic aneurysm. There has been a bona fide increase in incidence of aortic aneurysm making aneurysm disease the <em>1</em>8th most common cause of death. 3) Aortic growth rate. Although a virulent disease, thoracic aortic aneurysm is an indolent process. The thoracic aorta grows slowly-0.<em>1</em> cm/year. 4) Evidence-based intervention criteria. It is imperative to extirpate the thoracic aorta before rupture or dissection occurs; surgery at 5.0- to 5.5-cm diameter will prevent most adverse natural events. Symptomatic (painful) aneurysms must be resected regardless of size. 5) Development of nonsize criteria. Mechanical properties of the aorta deteriorate at the same 6 cm at which dissection occurs; elastic properties of the aorta may soon become useful intervention criteria. 6) Medical treatment of aortic aneurysm. Medical treatment is of unproven value, even beta-blockers and <em>angiotensin</em>-receptor blockers. <em>7</em>) A genetic disease. Even non-Marfan aneurysms have a strong genetic basis. 8) Need for biomarkers. Virulent but silent, TAA cries out for a biomarker that can predict the onset of adverse events. Pathophysiologic understanding has led to identification of promising biomarkers, especially metalloproteinases. 9) Endovascular therapy for aneurysms. Endovascular therapy has burgeoned, despite the fact that the EVAR-2, DREAM, and INSTEAD trials showed no benefit at mid-term over medical or conventional surgical therapy. We must avoid "irrational exuberance." <em>1</em>0) Inciting events for acute aortic dissection. Recent evidence shows that dissections are preceded by a specific severe exertional or emotional event. <em>1</em><em>1</em>) "Silver lining" of aortic disease. Proximal aortic root disease seems to protect against arteriosclerosis.

BACKGROUND

Data from studies of <em>angiotensin</em>-converting enzyme inhibitors provide evidence that the renin-<em>angiotensin</em>-aldosterone system plays a role as a mediator of atrial remodeling in atrial fibrillation. The present study has evaluated the effect of treatment with the <em>angiotensin</em> I type <em>1</em> receptor blocker irbesartan on maintaining sinus rhythm after conversion from persistent atrial fibrillation.

RESULTS

To be included in the present study, patients must have had an episode of persistent atrial fibrillation for >7 days. The patients were then randomized and scheduled for electrical cardioversion. Two groups of patients were compared: Group I was treated with amiodarone, and group II was treated with amiodarone plus irbesartan. The primary end point was the length of time to a first recurrence of atrial fibrillation. From a total of <em>1</em>86 patients assessed in the study, <em>1</em>54 were analyzed with the use of intention-to-treat analysis. Seventy-five patients were randomly allocated to group I and 79 to group II. After 2 months of follow-up in the intention-to-treat analysis, the group treated with irbesartan had fewer patients with recurrent atrial fibrillation (Kaplan-Meier analysis, 84.79% versus 63.<em>1</em>6%, P=0.008). The Kaplan-Meier analysis of time to first recurrence during the follow-up period (median time, 254 days [range, 60 to 7<em>1</em>0]) also showed that patients treated with irbesartan had a greater probability of remaining free of atrial fibrillation (79.52% versus 55.9<em>1</em>%, P=0.007).

CONCLUSIONS

Patients treated with amiodarone plus irbesartan had a lower rate of recurrence of atrial fibrillation than did patients treated with amiodarone alone.

BACKGROUND

We previously identified the G-protein-coupled receptor Mas, encoded by the Mas proto-oncogene, as an endogenous receptor for the heptapeptide <em>angiotensin</em>-(<em>1</em>-<em>7</em>); however, the receptor is also suggested to be involved in actions of <em>angiotensin</em> II. We therefore tested whether this could be mediated indirectly through an interaction with the <em>angiotensin</em> II type <em>1</em> receptor, AT<em>1</em>.

RESULTS

In transfected mammalian cells, Mas was not activated by <em>angiotensin</em> II; however, AT<em>1</em> receptor-mediated, <em>angiotensin</em> II-induced production of inositol phosphates and mobilization of intracellular Ca2+ was diminished by 50% after coexpression of Mas, despite a concomitant increase in <em>angiotensin</em> II binding capacity. Mas and the AT<em>1</em> receptor formed a constitutive hetero-oligomeric complex that was unaffected by the presence of agonists or antagonists of the 2 receptors. In vivo, Mas acts as an antagonist of the AT<em>1</em> receptor; mice lacking the Mas gene show enhanced <em>angiotensin</em> II-mediated vasoconstriction in mesenteric microvessels.

CONCLUSIONS

These results demonstrate that Mas can hetero-oligomerize with the AT<em>1</em> receptor and by so doing inhibit the actions of <em>angiotensin</em> II. This is a novel demonstration that a G-protein-coupled receptor acts as a physiological antagonist of a previously characterized receptor. Consequently, the AT<em>1</em>-Mas complex could be of great importance as a target for pharmacological intervention in cardiovascular diseases.

Central infusion of <em>angiotensin</em> IV or its more stable analogues facilitates memory retention and retrieval in normal animals and reverses amnesia induced by scopolamine or by bilateral perforant pathway lesions. These peptides bind with high affinity and specificity to a novel binding site designated the <em>angiotensin</em> AT(4) receptor. Until now, the AT(4) receptor has eluded molecular characterization. Here we identify the AT(4) receptor, by protein purification and peptide sequencing, to be insulin-regulated aminopeptidase (IRAP). HEK 293T cells transfected with IRAP exhibit typical AT(4) receptor binding characteristics; the AT(4) receptor ligands, <em>angiotensin</em> IV and LVV-hemorphin <em>7</em>, compete for the binding of [(<em>1</em>25)I]Nle(<em>1</em>)-<em>angiotensin</em> IV with IC(50) values of 32 and <em>1</em>40 nm, respectively. The distribution of IRAP and its mRNA in the brain, determined by immunohistochemistry and hybridization histochemistry, parallels that of the AT(4) receptor determined by radioligand binding. We also show that AT(4) receptor ligands dose-dependently inhibit the catalytic activity of IRAP. We have therefore demonstrated that the AT(4) receptor is IRAP and propose that AT(4) receptor ligands may exert their effects by inhibiting the catalytic activity of IRAP thereby extending the half-life of its neuropeptide substrates.

OBJECTIVE

<em>Angiotensin</em>-converting enzyme 2 (ACE2) converts <em>angiotensin</em> II (Ang<em>1</em>-8) to <em>angiotensin</em> <em>1</em>-<em>7</em> (Ang<em>1</em>-<em>7</em>), a functional antagonist of Ang<em>1</em>-8, with vasodilatory, antiproliferative, antiangiogenic, and anti-inflammatory properties. In conditions with an unbalanced renin-<em>angiotensin</em>-aldosterone system with elevated Ang<em>1</em>-8, administration of ACE2 has shown promising effects in a variety of animal models. Enhancing ACE2 activity by exogenous administration of ACE2 might also be beneficial in human diseases with pathologically elevated Ang<em>1</em>-8. As a first step we performed a first-in-man study to determine pharmacokinetics, pharmacodynamics, safety, and tolerability of recombinant ACE2 in healthy volunteers.

METHODS

Recombinant human ACE2 (rhACE2) was administered intravenously to healthy human subjects in a randomized, double-blind, placebo-controlled, single-dose, dose-escalation study followed by an open-label multiple-dose study. ACE2 concentrations were determined by quantifying ACE2 activity and ACE2 content in plasma samples. Concentrations of the <em>angiotensin</em> system effector peptides Ang<em>1</em>-8, Ang<em>1</em>-<em>7</em>, and Ang<em>1</em>-5 were determined using a liquid chromatography-tandem mass spectrometry method.

RESULTS

Single rhACE2 doses of <em>1</em>00-<em>1</em>,200 μg/kg caused a dose-dependent increase of systemic exposure with biphasic elimination and a dose-independent terminal half-life of <em>1</em>0 h. In all single-dose cohorts, Ang<em>1</em>-8 decreased within 30 min postinfusion, <em>angiotensin</em> <em>1</em>-<em>7</em> (Ang<em>1</em>-<em>7</em>) either increased (<em>1</em>00 and 200 μg/kg doses), decreased, or remained unchanged (400-<em>1</em>,200 μg/kg doses), whereas <em>angiotensin</em> <em>1</em>-5 (Ang<em>1</em>-5) transiently increased for all doses investigated. With the exception of the lowest rhACE2 dose, the decrease in Ang<em>1</em>-8 levels lasted for at least 24 h. Repeated dosing (400 μg/kg for 3 or 6 days) caused only minimal accumulation of ACE2, and Ang<em>1</em>-8 levels were suppressed over the whole application period.

CONCLUSIONS

Administration of rhACE2 was well tolerated by healthy human subjects. Exposure was dose dependent with a dose-independent terminal elimination half-life in the range of <em>1</em>0 h. Despite marked changes in <em>angiotensin</em> system peptide concentrations, cardiovascular effects were absent, suggesting the presence of effective compensatory mechanisms in healthy volunteers.

BACKGROUND

Raised blood pressure is common in acute stroke, and is associated with an increased risk of poor outcomes. We aimed to examine whether careful blood-pressure lowering treatment with the angiotensin-receptor blocker candesartan is beneficial in patients with acute stroke and raised blood pressure.

METHODS

Participants in this randomised, placebo-controlled, double-blind trial were recruited from 146 centres in nine north European countries. Patients older than 18 years with acute stroke (ischaemic or haemorrhagic) and systolic blood pressure of 140 mm Hg or higher were included within 30 h of symptom onset. Patients were randomly allocated to candesartan or placebo (1:1) for 7 days, with doses increasing from 4 mg on day 1 to 16 mg on days 3 to 7. Randomisation was stratified by centre, with blocks of six packs of candesartan or placebo. Patients and investigators were masked to treatment allocation. There were two co-primary effect variables: the composite endpoint of vascular death, myocardial infarction, or stroke during the first 6 months; and functional outcome at 6 months, as measured by the modified Rankin Scale. Analyses were by intention to treat. The study is registered, number NCT00120003 (ClinicalTrials.gov), and ISRCTN13643354.

RESULTS

2029 patients were randomly allocated to treatment groups (1017 candesartan, 1012 placebo), and data for status at 6 months were available for 2004 patients (99%; 1000 candesartan, 1004 placebo). During the 7-day treatment period, blood pressures were significantly lower in patients allocated candesartan than in those on placebo (mean 147/82 mm Hg [SD 23/14] in the candesartan group on day 7 vs 152/84 mm Hg [22/14] in the placebo group; p<0·0001). During 6 months' follow-up, the risk of the composite vascular endpoint did not differ between treatment groups (candesartan, 120 events, vs placebo, 111 events; adjusted hazard ratio 1·09, 95% CI 0·84-1·41; p=0·52). Analysis of functional outcome suggested a higher risk of poor outcome in the candesartan group (adjusted common odds ratio 1·17, 95% CI 1·00-1·38; p=0·048 [not significant at p≤0·025 level]). The observed effects were similar for all prespecified secondary endpoints (including death from any cause, vascular death, ischaemic stroke, haemorrhagic stroke, myocardial infarction, stroke progression, symptomatic hypotension, and renal failure) and outcomes (Scandinavian Stroke Scale score at 7 days and Barthel index at 6 months), and there was no evidence of a differential effect in any of the prespecified subgroups. During follow-up, nine (1%) patients on candesartan and five (<1%) on placebo had symptomatic hypotension, and renal failure was reported for 18 (2%) patients taking candesartan and 13 (1%) allocated placebo.

CONCLUSIONS

There was no indication that careful blood-pressure lowering treatment with the angiotensin-receptor blocker candesartan is beneficial in patients with acute stroke and raised blood pressure. If anything, the evidence suggested a harmful effect.

BACKGROUND

South-Eastern Norway Regional Health Authority; Oslo University Hospital Ullevål; AstraZeneca; Takeda.

<em>Angiotensin</em> (Ang)-converting enzyme (ACE) 2 cleaves Ang-II into the vasodilator peptide Ang-(<em>1</em>-<em>7</em>), thus acting as a pivotal element in balancing the local effects of these peptides. ACE2 has been identified in various tissues and is supposed to be a modulator of cardiovascular function. Decreases in ACE2 expression and activity have been reported in models of hypertension, heart failure, atherosclerosis, diabetic nephropathy and others. In addition, the expression level and/or activity are affected by other renin-<em>angiotensin</em> system components (e.g., ACE and AT<em>1</em> receptors). Local inhibition or global deletion of brain ACE2 induces a reduction in baroreflex sensitivity. Moreover, ACE2-null mice have been shown to exhibit either blood pressure or cardiac dysfunction phenotypes. On the other hand, over-expression of ACE2 exerts protective effects in local tissues, including the brain. In this review, we will first summarize the major findings linking ACE2 to cardiovascular function in the periphery then focus on recent discoveries related to ACE2 in the CNS. Finally, we will unveil new tools designed to address the importance of central ACE2 in various diseases, and discuss the potential for this carboxypeptidase as a new target in the treatment of hypertension and other cardiovascular diseases.

BACKGROUND

Pioglitazone and rosiglitazone, thiazolidinedione peroxisome proliferator-activated receptor-gamma (PPARgamma) activators, reduce blood pressure (BP) in some hypertensive models by unclear mechanisms. We tested the hypothesis that pioglitazone or rosiglitazone would prevent BP elevation and vascular dysfunction in angiotensin (Ang) II-infused rats by direct vascular effects.

RESULTS

Sprague-Dawley rats received Ang II (120 ng x kg(-1) x min(-1) SC) with or without pioglitazone (10 mg x kg(-1) x d(-1)) or rosiglitazone (5 mg x kg(-1) x d(-1)) for 7 days. Systolic BP, elevated in Ang II-infused rats (176+/-5 mm Hg) versus controls (109+/-2 mm Hg, P<0.01), was reduced by pioglitazone (134+/-2 mm Hg) or rosiglitazone (123+/-2 mm Hg). 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 the thiazolidinediones. In Ang II-infused rats, vascular DNA synthesis (by 3H-thymidine incorporation); expression of cell cycle proteins cyclin D1 and cdk4, angiotensin II type 1 receptors, vascular cell adhesion molecule-1, and platelet and endothelial cell adhesion molecule; and nuclear factor-kappaB activity were increased. These changes were abrogated by pioglitazone or rosiglitazone.

CONCLUSIONS

Thiazolidinedione PPAR-gamma activators attenuated the development of hypertension, corrected structural abnormalities, normalized cell growth, and improved endothelial dysfunction induced by Ang II and prevented upregulation of angiotensin II type 1 receptors, cell cycle proteins, and proinflammatory mediators. Thiazolidinediones may be useful in the prevention and/or treatment of hypertension, particularly when it is associated with insulin resistance or diabetes mellitus.

<em>1</em>. When applied directly to the brain, <em>angiotensin</em> II amide, as either the valine(5) octapeptide, causes rats in normal fluid balance to drink water.2. The drinking response to <em>angiotensin</em> injections is copious, rapid, repeatable within the same test session, and stable over months of testing in the same animal.3. The response is motivationally potent and specific. After injection the animals move directly to the source of water and drink. There is typically no preliminary hyperactivity or subsequent depression. The animals do not eat, gnaw or exhibit other behaviours that are not normally seen during spontaneous drinking. The injections rouse sleeping animals to drink and interrupt eating in animals deprived of food for two days.4. The region of the brain that is most sensitive to <em>angiotensin</em> includes the anterior hypothalamus, the preoptic region, and the septum including the nucleus accumbens.5. Intracranial renin elicited drinking. Bradykinin and vasopressin did not, nor did adrenaline, noradrenaline or aldosterone. In the most sensitive region, sites positive for <em>angiotensin</em> also yielded drinking to carbachol.6. Responses were obtained with 5 ng (ca. 5 p-mole) and occurred reliably with 50 ng <em>angiotensin</em> or more. The dose-response curve for amount drunk rose from 5 to <em>1</em>00 ng and levelled off thereafter. <em>Angiotensin</em> is therefore the most potent dipsogen known and is effective at doses that are reasonably within the concentration range for circulating endogenous <em>angiotensin</em>.<em>7</em>. Injections into the sensitive region of doses of <em>angiotensin</em> that were effective for drinking did not produce peripheral haemodynamic changes in lightly anaesthetized rats.8. This work strengthens the suggestion that <em>angiotensin</em> is a natural hormone of drinking behaviour that participates in extracellular thirst by its release from the kidney and subsequent direct action on a specific chemoreceptive region in the anterior diencephalon and limbic lobe.

<em>Angiotensin</em>-converting enzyme 2 (ACE2) is a carboxypeptidase that cleaves <em>angiotensin</em> II to <em>angiotensin</em> <em>1</em>-<em>7</em>. Recently, it was reported that mice lacking ACE2 (ACE2(-/y) mice) exhibited reduced cardiac contractility. Because mechanical pressure overload activates the cardiac renin-<em>angiotensin</em> system, we used ACE2(-/y) mice to analyze the role of ACE2 in the response to pressure overload. Twelve-week-old ACE2(-/y) mice and wild-type (WT) mice received transverse aortic constriction (TAC) or sham operation. Sham-operated ACE2(-/y) mice exhibited normal cardiac function and had morphologically normal hearts. In response to TAC, ACE2(-/y) mice developed cardiac hypertrophy and dilatation. Furthermore, their hearts displayed decreased cardiac contractility and increased fetal cardiac gene induction, compared with WT mice. In response to chronic pressure overload, ACE2(-/y) mice developed pulmonary congestion and increased incidence of cardiac death compared with WT mice. On a biochemical level, cardiac <em>angiotensin</em> II concentration and activity of mitogen-activated protein (MAP) kinases were markedly increased in ACE2(-/y) mice in response to TAC. Administration of candesartan, an AT<em>1</em> subtype <em>angiotensin</em> receptor blocker, attenuated the hypertrophic response and suppressed the activation of MAP kinases in ACE2(-/y) mice. Activation of MAP kinases in response to <em>angiotensin</em> II was greater in cardiomyocytes isolated from ACE2(-/y) mice than in those isolated from WT mice. ACE2 plays an important role in dampening the hypertrophic response to pressure overload mediated by <em>angiotensin</em> II. Disruption of this regulatory function may accelerate cardiac hypertrophy and shorten the transition period from compensated hypertrophy to cardiac failure.

Peptide hormones such as ANG II and endothelin contribute to cardiac remodeling after myocardial infarction by stimulating myocyte hypertrophy and myofibroblast proliferation. In contrast, <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [ANG-(<em>1</em>-<em>7</em>)] infusion after myocardial infarction reduced myocyte size and attenuated ventricular dysfunction and remodeling. We measured the effect of ANG-(<em>1</em>-<em>7</em>) on protein and DNA synthesis in cultured neonatal rat myocytes to assess the role of the heptapeptide in cell growth. ANG-(<em>1</em>-<em>7</em>) significantly attenuated either fetal bovine serum- or endothelin-<em>1</em>-stimulated [(3)H]leucine incorporation into myocytes with no effect on [(3)H]thymidine incorporation. [d-Ala(<em>7</em>)]-ANG-(<em>1</em>-<em>7</em>), the selective ANG type <em>1</em>-<em>7</em> (AT(<em>1</em>-<em>7</em>)) receptor antagonist, blocked the ANG-(<em>1</em>-<em>7</em>)-mediated reduction in protein synthesis in cardiac myocytes, whereas the AT(<em>1</em>) and AT(2) <em>angiotensin</em> peptide receptors were ineffective. Serum-stimulated ERK<em>1</em>/ERK2 mitogen-activated protein kinase activity was significantly decreased by ANG-(<em>1</em>-<em>7</em>) in myocytes, a response that was also blocked by [d-Ala(<em>7</em>)]-ANG-(<em>1</em>-<em>7</em>). Both rat heart and cardiac myocytes express the mRNA for the mas receptor, and a 59-kDa immunoreactive protein was identified in both extracts of rat heart and cultured myocytes by Western blot hybridization with the use of an antibody to mas, an ANG-(<em>1</em>-<em>7</em>) receptor. Transfection of cultured myocytes with an antisense oligonucleotide to the mas receptor blocked the ANG-(<em>1</em>-<em>7</em>)-mediated inhibition of serum-stimulated MAPK activation, whereas a sense oligonucleotide was ineffective. These results suggest that ANG-(<em>1</em>-<em>7</em>) reduces the growth of cardiomyocytes through activation of the mas receptor. Because ANG-(<em>1</em>-<em>7</em>) is elevated after treatment with <em>angiotensin</em>-converting enzyme inhibitors or AT(<em>1</em>) receptor blockers, ANG-(<em>1</em>-<em>7</em>) may contribute to their beneficial effects on cardiac dysfunction and ventricular remodeling after myocardial infarction.

The protease renin is the key enzyme of the renin-<em>angiotensin</em>-aldosterone cascade, which is relevant under both physiological and pathophysiological settings. The kidney is the only organ capable of releasing enzymatically active renin. Although the characteristic juxtaglomerular position is the best known site of renin generation, renin-producing cells in the kidney can vary in number and localization. (Pro)renin gene transcription in these cells is controlled by a number of transcription factors, among which CREB is the best characterized. Pro-renin is stored in vesicles, activated to renin, and then released upon demand. The release of renin is under the control of the cAMP (stimulatory) and Ca(2+) (inhibitory) signaling pathways. Meanwhile, a great number of intrarenally generated or systemically acting factors have been identified that control the renin secretion directly at the level of renin-producing cells, by activating either of the signaling pathways mentioned above. The broad spectrum of biological actions of (pro)renin is mediated by receptors for (pro)renin, <em>angiotensin</em> II and <em>angiotensin</em>-(<em>1</em>-<em>7</em>).