OBJECTIVE

To determine the independent associations of antihypertensive drugs with the risk of incident gout among people with hypertension.

METHODS

Nested case-control study.

METHODS

UK general practice database, 2000-<em>7</em>.

METHODS

All incident cases of gout (n = 24,<em>7</em>68) among adults aged 20-<em>7</em>9 and a random sample of 50,000 matched controls.

METHODS

Relative risk of incident gout associated with use of antihypertensive drugs.

RESULTS

After adjusting for age, sex, body mass index, visits to the general practitioner, alcohol intake, and pertinent drugs and comorbidities, the multivariate relative risks of incident gout associated with current use of antihypertensive drugs among those with hypertension (n = 29,138) were 0.8<em>7</em> (95% confidence interval 0.82 to 0.93) for calcium channel blockers, 0.81 (0.<em>7</em>0 to 0.94) for losartan, 2.36 (2.21 to 2.52) for diuretics, 1.48 (1.40 to 1.5<em>7</em>) for β blockers, 1.24 (1.1<em>7</em> to 1.32) for angiotensin converting enzyme inhibitors, and 1.29 (1.16 to 1.43) for non-losartan angiotensin II receptor blockers. Similar results were obtained among those without hypertension. The multivariate relative risks for the duration of use of calcium channel blockers among those with hypertension were 1.02 for less than one year, 0.88 for 1-1.9 years, and 0.<em>7</em>5 for two or more years and for use of losartan they were 0.98, 0.8<em>7</em>, and 0.<em>7</em>1, respectively (both P<0.05 for trend).

CONCLUSIONS

Compatible with their urate lowering properties, calcium channel blockers and losartan are associated with a lower risk of incident gout among people with hypertension. By contrast, diuretics, β blockers, angiotensin converting enzyme inhibitors, and non-losartan angiotensin II receptor blockers are associated with an increased risk of gout.

BACKGROUND

<em>Angiotensin</em> II (Ang II) is a vasoconstrictor but also a growth factor. However, the Ang II type <em>1</em> receptor does not have a tyrosine kinase domain that mediates the cellular signals for mitosis. We have shown that Ang II acts via "trans"-activation of the epidermal growth factor receptor (EGFR) to induce activation of tyrosine kinase and mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) in vascular smooth muscle cells (VSMCs). To examine whether EGFR is involved in the development of left ventricular hypertrophy (LVH), we inhibited EGFR with a specific antisense oligodeoxynucleotide to attenuate the Ang II-induced cardiovascular hypertrophic effects.

RESULTS

The antisense oligodeoxynucleotide to EGFR (EGFR-AS) was designed and tested on Ang II-induced ERK activation in cultured VSMCs. We also investigated the effects of EGFR-AS on LVH and blood pressure (BP) in Ang II-infused hypertensive rats. In VSMCs, EGFR-AS (2.5 micromol/L) reduced EGFR expression and inhibited the Ang II-induced phosphorylation of ERK. In rats, Ang II (<em>1</em>50 ng/h for <em>1</em>4 days) increased BP compared with controls (<em>1</em>84+/-6 mm Hg versus <em>1</em>22+/-3 mm Hg; n=7; P<0.0<em>1</em>). Continuous intravenous infusion of EGFR-AS (2 mg/kg) decreased BP (<em>1</em>69+/-8 mm Hg; n=8; P<0.05). Ang II infusion increased the left ventricular/body weight (LV/BW) ratio compared with control rats (2.75+/-0.08 versus 2.33+/-0.07; P<0.0<em>1</em>). EGFR-AS, but not EGFR-sense, normalized the LV/BW in Ang II-infused rats (2.32+/-0.06; P<0.0<em>1</em>) and attenuated Ang II-enhanced EGFR expression and ERK phosphorylation.

CONCLUSIONS

Ang II requires EGFR to mediate ERK activation in VSMCs and the heart. EGFR plays a critical role in the LVH induced by Ang II.

BACKGROUND

Although community pharmacists have historically been paid primarily for drug distribution and dispensing services, medication therapy management (MTM) services evolved in the 1990s as a means for pharmacists and other providers to assist physicians and patients in managing clinical, service, and cost outcomes of drug therapy. The Medicare Prescription Drug, Improvement, and Modernization Act of 2003 (MMA 2003) and the subsequent implementation of Medicare Part D in January 2006 for the more than 20 million Medicare beneficiaries enrolled in the Part D benefit formalized MTM services for a subset of high-cost patients. Although Medicare Part D has provided a new opportunity for defining the value of pharmacist-provided MTM services in the health care system, few publications exist which quantify changes in the provision of pharmacist-provided MTM services over time.

OBJECTIVE

To (a) describe the changes over a <em>7</em>-year period in the primary types of MTM services provided by community pharmacies that have contracted with drug plan sponsors through an MTM administrative services company, and (b) quantify potential MTM-related cost savings based on pharmacists' self-assessments of the likely effects of their interventions on health care utilization.

METHODS

Medication therapy management claims from a multistate MTM administrative services company were analyzed over the <em>7</em>-year period from January 1, 2000, through December 31, 2006. Data extracted from each MTM claim included patient demographics (e.g., age and gender), the drug and type that triggered the intervention (e.g., drug therapeutic class and therapy type as either acute, intermittent, or chronic), and specific information about the service provided (e.g., Reason, Action, Result, and Estimated Cost Avoidance [ECA]). ECA values are derived from average national health care utilization costs, which are applied to pharmacist self-assessment of the "reasonable and foreseeable" outcome of the intervention. ECA values are updated annually for medical care inflation.

RESULTS

From a database of nearly 100,000 MTM claims, a convenience sample of 50 plan sponsors was selected. After exclusion of claims with missing or potentially duplicate data, there were <em>7</em>6,148 claims for 23,<em>7</em>98 patients from community pharmacy MTM providers in 4<em>7</em> states. Over the <em>7</em>-year period from January 1, 2000, through December 31, 2006, the mean ([SD] median) pharmacy reimbursement was $8.44 ([$5.19] $<em>7</em>.00) per MTM service, and the mean ([SD] median) ECA was $93.<em>7</em>8 ([$1,022.23] $5.00). During the <em>7</em>-year period, pharmacist provided MTM interventions changed from primarily education and monitoring for new or changed prescription therapies to prescriber consultations regarding cost-efficacy management (Pearson chi-square P<0.001). Services also shifted from claims involving acute medications (e.g. penicillin antibiotics, macrolide antibiotics, and narcotic analgesics) to services involving chronic medications (e.g., lipid lowering agents, angiotensin-converting enzyme [ACE] inhibitors, and beta-blockers; P<0.001), resulting in significant changes in the therapeutic classes associated with MTM claims and an increase in the proportion of older patients served (P<0.001). These trends resulted in higher pharmacy reimbursements and greater ECA per claim over time (P<0.001).

CONCLUSIONS

MTM interventions over a <em>7</em>-year period evolved from primarily the provision of patient education involving acute medications towards consultation-type services for chronic medications. These changes were associated with increases in reimbursement amounts and pharmacist-estimated cost savings. It is uncertain if this shift in service type is a result of clinical need, documentation requirements, or reimbursement opportunities.

<em>1</em><em>7</em>beta-Oestradiol (E2)-mediated inhibition of <em>angiotensin</em>-converting enzyme (ACE) protects the E2-replete kidney from the progression of hypertensive renal disease. <em>Angiotensin</em>-converting enzyme 2 (ACE2), a homologue of ACE, counters the actions of ACE by catalysing the conversion of <em>angiotensin</em> II (Ang II) to <em>angiotensin</em>(<em>1</em>-<em>7</em>) [Ang(<em>1</em>-<em>7</em>)]. We investigated E2 regulation of ACE2 in the renal wrap (RW) model of hypertension in rats. After 6 weeks on a high-sodium diet (4% NaCl), the activity of ACE2 was reduced in the renal cortex by 3<em>1</em>%, which was mirrored by similar decreases in ACE2 protein (30%) and mRNA expression (36%) in the ovariectomized RW rat (RW-OVX); E2 replacement prevented these effects. The RW-OVX rats exhibited greater renal injury, including <em>1</em>.<em>7</em>-fold more tubulointerstitial fibrosis and <em>1</em>.6-fold more glomerulosclerosis than E2-replete females (RW-Intact and RW-OVX+E2). <em>Angiotensin</em>(<em>1</em>-<em>7</em>) infusion prevented these exacerbating effects of ovariectomy on renal pathology; no differences in indicators of renal injury were observed between RW-OVX-Ang(<em>1</em>-<em>7</em>) and RW-Intact rats. These renal protective effects of Ang(<em>1</em>-<em>7</em>) infusion were not attributable to increased ACE2 activity or to changes in heart rate or body weight, since these parameters were unchanged by Ang(<em>1</em>-<em>7</em>) infusion. Furthermore, Ang(<em>1</em>-<em>7</em>) infusion did not attenuate renal injury by reducing mean arterial pressure (MAP), since infusion of the peptide did not lower MAP but rather caused a slight increase during a 6 week chronic treatment for Ang(<em>1</em>-<em>7</em>). These results suggest that E2-mediated upregulation of renal ACE2 and the consequent increased Ang(<em>1</em>-<em>7</em>) production contribute to E2-mediated protection from hypertensive renal disease. These findings have implications for E2-deficient women with hypertensive renal disease and suggest that therapeutics targeted towards increasing ACE2 activity and Ang(<em>1</em>-<em>7</em>) levels will be renal protective.

Acute physiological hyperinsulinemia increases skeletal muscle capillary blood volume (CBV), presumably to augment glucose and insulin delivery. We hypothesized that insulin-mediated changes in CBV are impaired in type 2 diabetes mellitus (DM) and are improved by <em>angiotensin</em>-converting enzyme inhibition (ACE-I). Zucker obese diabetic rats (ZDF, n = <em>1</em>8) and control rats (n = 9) were studied at 20 wk of age. One-half of the ZDF rats were treated with quinapril (ZDF-Q) for <em>1</em>5 wk prior to study. CBV and capillary flow in hindlimb skeletal muscle were measured by contrast-enhanced ultrasound (CEU) at baseline and at 30 and <em>1</em>20 min after initiation of a euglycemic hyperinsulinemic clamp (3 mU.min(-<em>1</em>).kg(-<em>1</em>)). At baseline, ZDF and ZDF-Q rats were hyperglycemic and hyperinsulinemic vs. controls. Glucose utilization in ZDF rats was 60-<em>7</em>0% lower (P < 0.05) than in controls after 30 and <em>1</em>20 min of hyperinsulinemia. In ZDF-Q rats, glucose utilization was impaired at 30 min but similar to controls at <em>1</em>20 min. Basal CBV was lower in ZDF and ZDF-Q rats compared with controls (<em>1</em>3 +/- 4, <em>7</em> +/- 3, and 9 +/- 2 U, respectively). With hyperinsulinemia, CBV increased by about twofold in control animals at 30 and <em>1</em>20 min, did not change in ZDF animals, and increased in ZDF-Q animals only at <em>1</em>20 min to a level similar to controls. Anatomic capillary density on immunohistology was not different between groups. We conclude that insulin-mediated capillary recruitment in skeletal muscle, which participates in glucose utilization, is impaired in animals with DM and can be partially reversed by chronic ACE-I therapy.

The generation of the Lew.Tg(mRen2) congenic hypertensive rat strain, developed through a backcross of the hypertensive (mRen2)2<em>7</em> transgenic rat with normotensive Lewis rats, provides a new model by which primary hypertension can be studied without the genetic variability found in the original strain. The purpose of this study was to characterize the Lew.Tg(mRen2) rats by dually investigating the effects of type <em>1</em> <em>angiotensin</em> II (ANG II) receptor (AT(<em>1</em>)) blockade and <em>angiotensin</em>-converting enzyme (ACE) activity inhibition on the ANG-(<em>1</em>-<em>7</em>)/ACE2 axis of the renin-<em>angiotensin</em> system in this new hypertensive model. The control of blood pressure elicited by <em>1</em>2-day administration of either lisinopril (mean difference change = 92 +/- 2, P < 0.05) or losartan (mean difference change = 69 +/- 2, P < 0.05) was associated with 54% and 33% increases in cardiac ACE2 mRNA and 54% and 43% increases in cardiac ACE mRNA, respectively. Lisinopril induced a 3.<em>1</em>-fold (P < 0.05) increase in renal cortical expression of ACE2, whereas losartan increased ACE2 mRNA 3.5-fold (P < 0.05). Both treatment regimens increased renal ACE mRNA 2.6-fold (P < 0.05). The two therapies augmented ACE2 protein activity, as well as increased cardiac and renal AT(<em>1</em>) receptor mRNAs. ACE inhibition reduced plasma ANG II levels (8<em>1</em>%, P < 0.05) and increased plasma ANG-(<em>1</em>-<em>7</em>) (265%, P < 0.05), whereas losartan had no effect on the peptides. In contrast with what had been shown in normotensive rats, ACE inhibition decreased renal ANG II excretion and transiently decreased ANG-(<em>1</em>-<em>7</em>) excretion, whereas losartan treatment was associated with a consistent decrease in ANG-(<em>1</em>-<em>7</em>) urinary excretion rates. In response to the treatments, the expression of both renal cortical renin and <em>angiotensin</em>ogen mRNAs was significantly augmented. The paradoxical effects of blockade of ANG II synthesis and activity on urinary excretion rates of the peptides and plasma <em>angiotensins</em> levels suggest that, in Lew.Tg(mRen2) congenic rats, a failure of compensatory ACE2 and ANG-(<em>1</em>-<em>7</em>)-dependent vasodepressor mechanisms may contribute both to the development and progression of hypertension driven by increased formation of endogenous ANG II.

Injections of the <em>angiotensin</em>(<em>1</em>-<em>7</em>) [Ang(<em>1</em>-<em>7</em>)] antagonist [d-Ala<em>7</em>]-Ang(<em>1</em>-<em>7</em>) into the nucleus of the solitary tract (NTS) of Sprague-Dawley rats reduce baroreceptor reflex sensitivity (BRS) for control of heart rate by approximately 40%, whereas injections of the <em>angiotensin</em> II (Ang II) type <em>1</em> receptor antagonist candesartan increase BRS by 40% when reflex bradycardia is assessed. The enzyme <em>angiotensin</em>-converting enzyme 2 (ACE2) is known to convert Ang II to Ang(<em>1</em>-<em>7</em>). We report that ACE2 activity, as well as ACE and neprilysin activities, are present in plasma membrane fractions of the dorsomedial medulla of Sprague-Dawley rats. Moreover, we show that BRS for reflex bradycardia is attenuated (<em>1</em>.<em>1</em>6 +/- 0.29 ms mmHg-<em>1</em> before versus 0.33 +/- 0.<em>1</em><em>1</em> ms mmHg-<em>1</em> after; P < 0.05; n = 8) 30-60 min following injection of the selective ACE2 inhibitor MLN4<em>7</em>60 (<em>1</em>2 pmol in <em>1</em>20 nl) into the NTS. These findings support the concept that within the NTS, local synthesis of Ang(<em>1</em>-<em>7</em>) from Ang II is required for normal sensitivity for the baroreflex control of heart rate in response to increases in arterial pressure.

Regional pulmonary glucose metabolism (MRglu; mumol h-<em>1</em> g-<em>1</em>), extravascular lung density (D(EV); g cm-3) and vascular volume (VB; ml cm-3) were measured in a single midthoracic transaxial slice (approximately 2 cm thick) using position emission tomography (PET) in seven patients with histologically proven sarcoidosis. The measurements were repeated <em>1</em>-<em>7</em> months later after steroid therapy (in two cases, no treatment) in order to assess MRglu as an index of inflammation and relate it to routine pulmonary function tests, chest radiography and serum <em>angiotensin</em> converting enzyme (SACE) levels. MRglu was computed from serial lung scans and peripheral venous blood samples for 60 min following an i.v. injection of <em>1</em>8F-2-fluoro-2-deoxy-D-glucose (<em>1</em>8FDG). Both MRglu (which was increased in six of seven patients) and elevated SACE levels returned to normal in those patients treated with high-dose steroids. Regional vascular volume was normal in six of seven cases and did not change significantly with therapy. The high tissue density measured in all patients decreased significantly in two of three patients treated with 40 mg prednisolone daily. The abnormal MRglu observed in active sarcoidosis becomes normal pari passu with SACE levels during high-dose steroid therapy. We conclude that MRglu measured with <em>1</em>8FDG and PET may reflect "disease activity" in sarcoidosis in quantitative terms (per gram lung tissue) and in respect of disease distribution.

<AbstractText>Short-term treatment for people with type 2 diabetes using a low dose of the selective endothelin A receptor antagonist atrasentan reduces albuminuria without causing significant sodium retention. We report the long-term effects of treatment with atrasentan on major renal outcomes.</AbstractText><p><div><b>METHODS</b></div>We did this double-blind, randomised, placebo-controlled trial at 689 sites in 4<em>1</em> countries. We enrolled adults aged <em>1</em>8-85 years with type 2 diabetes, estimated glomerular filtration rate (eGFR) 25-<em>7</em>5 mL/min per <em>1</em>·<em>7</em>3 m² of body surface area, and a urine albumin-to-creatinine ratio (UACR) of 300-5000 mg/g who had received maximum labelled or tolerated renin-<em>angiotensin</em> system inhibition for at least 4 weeks. Participants were given atrasentan 0·<em>7</em>5 mg orally daily during an enrichment period before random group assignment. Those with a UACR decrease of at least 30% with no substantial fluid retention during the enrichment period (responders) were included in the double-blind treatment period. Responders were randomly assigned to receive either atrasentan 0·<em>7</em>5 mg orally daily or placebo. All patients and investigators were masked to treatment assignment. The primary endpoint was a composite of doubling of serum creatinine (sustained for ≥30 days) or end-stage kidney disease (eGFR <<em>1</em>5 mL/min per <em>1</em>·<em>7</em>3 m² sustained for ≥90 days, chronic dialysis for ≥90 days, kidney transplantation, or death from kidney failure) in the intention-to-treat population of all responders. Safety was assessed in all patients who received at least one dose of their assigned study treatment. The study is registered with ClinicalTrials.gov, number NCT0<em>1</em>858532.</p><AbstractText>Between May <em>1</em><em>7</em>, 20<em>1</em>3, and July <em>1</em>3, 20<em>1</em><em>7</em>, <em>1</em><em>1</em> 08<em>7</em> patients were screened; 5<em>1</em><em>1</em><em>7</em> entered the enrichment period, and 4<em>7</em><em>1</em><em>1</em> completed the enrichment period. Of these, 2648 patients were responders and were randomly assigned to the atrasentan group (n=<em>1</em>325) or placebo group (n=<em>1</em>323). Median follow-up was 2·2 years (IQR <em>1</em>·4-2·9). <em>7</em>9 (6·0%) of <em>1</em>325 patients in the atrasentan group and <em>1</em>05 (<em>7</em>·9%) of <em>1</em>323 in the placebo group had a primary composite renal endpoint event (hazard ratio [HR] 0·65 [95% CI 0·49-0·88]; p=0·004<em>7</em>). Fluid retention and anaemia adverse events, which have been previously attributed to endothelin receptor antagonists, were more frequent in the atrasentan group than in the placebo group. Hospital admission for heart failure occurred in 4<em>7</em> (3·5%) of <em>1</em>325 patients in the atrasentan group and 34 (2·6%) of <em>1</em>323 patients in the placebo group (HR <em>1</em>·33 [95% CI 0·85-2·0<em>7</em>]; p=0·208). 58 (4·4%) patients in the atrasentan group and 52 (3·9%) in the placebo group died (HR <em>1</em>·09 [95% CI 0·<em>7</em>5-<em>1</em>·59]; p=0·65).</AbstractText><AbstractText>Atrasentan reduced the risk of renal events in patients with diabetes and chronic kidney disease who were selected to optimise efficacy and safety. These data support a potential role for selective endothelin receptor antagonists in protecting renal function in patients with type 2 diabetes at high risk of developing end-stage kidney disease.</AbstractText><AbstractText>AbbVie.</AbstractText>

BACKGROUND

Studies in animals and humans indicate a role for kinins in the actions of <em>angiotensin</em> type <em>1</em> (AT<em>1</em>) receptor blockers. However, the effect of these compounds on kinin levels in humans is unknown.

RESULTS

We measured <em>angiotensin</em> (Ang), bradykinin (BK), and kallidin peptides in subjects with essential hypertension administered placebo, losartan (50 mg OD), and eprosartan (600 mg OD) in randomized order in a double-blind, 3-period, 3-treatment, crossover trial. Peptides were measured in arterial blood using high-performance liquid chromatography-based radioimmunoassays. Losartan increased blood levels of BK-(<em>1</em>-9) and hydroxylated BK-(<em>1</em>-9) by approximately 2-fold and reduced the BK-(<em>1</em>-7)/BK-(<em>1</em>-9) ratio by 55%. There was a trend for eprosartan to produce similar changes in bradykinin levels. There were no changes in blood kallidin levels. Both losartan and eprosartan increased plasma levels of Ang I, Ang II, and Ang-(2-8), and eprosartan increased Ang-(3-8) levels. Ang-(<em>1</em>-7) and Ang-(<em>1</em>-9) levels were unchanged. There was an associated 30% to 35% reduction in Ang II/Ang I ratio and 63% to 69% reduction in Ang-(<em>1</em>-7)/Ang I ratio. Plasma ACE activity was unchanged.

CONCLUSIONS

Losartan increases bradykinin levels. The reductions in BK-(<em>1</em>-7)/BK-(<em>1</em>-9), Ang II/Ang I, and Ang-(<em>1</em>-7)/Ang I ratios suggest that the increased bradykinin levels were the result of reduced metabolism by ACE and neutral endopeptidase. Increased bradykinin levels may represent a class effect of AT<em>1</em> receptor blockers that contributes to their therapeutic actions and may also contribute to the angioedema that may accompany this therapy.

Therapeutic modulation of the renin-<em>angiotensin</em> system is not complete without taking into consideration the beneficial effects of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) in cardiovascular pathology. Various pharmacological pathways are already exploited to involve this heptapeptide in therapy as both inhibitors of <em>angiotensin</em>-converting enzyme and <em>angiotensin</em> II type <em>1</em> receptor blockers increase its levels. These drugs and administered <em>angiotensin</em>-(<em>1</em>-<em>7</em>) elicit various common effects, and some effects of the drugs are partially mediated by <em>angiotensin</em>-(<em>1</em>-<em>7</em>). The pharmacodynamic profile of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) is rather complex, and in vitro and in vivo studies demonstrated a wide palette of effects for <em>angiotensin</em>-(<em>1</em>-<em>7</em>), some of them potentially beneficial for cardiovascular disease. Using various animal models to study cardiovascular physiology and disease it was shown that <em>angiotensin</em>-(<em>1</em>-<em>7</em>) has antihypertensive, antihypertrophic, antifibrotic and antithrombotic properties, all properties that may prove beneficial in a clinical setting. We also observed a novel action of <em>angiotensin</em>-(<em>1</em>-<em>7</em>), namely its capacity to stimulate the proliferation of endothelial progenitor cells. Access of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) to the clinic, however, is restricted due to its unfavorable pharmacokinetic properties. In order to benefit of the therapeutic potential of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) it is crucial to increase its half-life, either by using more stable analogues, which are now under development, or specific delivery methods. We here review the pharmacological characteristics and therapeutic potential of <em>angiotensin</em>-(<em>1</em>-<em>7</em>), implementing the experimental strategies taken to exploit the pharmacological mechanism of this heptapeptide in a clinical setting, and present our contribution to this field of research.

Hypertension and kidney damage in the double transgenic rat (dTGR) harboring both human renin and human <em>angiotensin</em>ogen genes are dependent on the human components of the renin <em>angiotensin</em> system. We tested the hypothesis that monocyte infiltration and increased adhesion molecule expression are involved in the pathogenesis of kidney damage in dTGR. We also evaluated the effects of long-term <em>angiotensin</em>-converting enzyme (ACE) inhibition, AT<em>1</em> blockade, and human renin inhibition on monocyte recruitment and inflammatory response in dTGR. Systolic blood pressure and 24-hour albuminuria were markedly increased in <em>7</em>-week-old dTGR as compared with age-matched normotensive Sprague Dawley rats. We found a significant monocyte/macrophage infiltration in the renal perivascular space and increased expression of intercellular adhesion molecule-<em>1</em> (ICAM-<em>1</em>) and vascular cell adhesion molecule-<em>1</em> (VCAM-<em>1</em>) in the interstitium, intima, and adventitia of the small renal vessels. alphaLbeta2 integrin and alpha4beta<em>1</em> integrin, the corresponding ligands for ICAM-<em>1</em> and VCAM-<em>1</em>, were also found on infiltrating monocytes/macrophages. The expression of plasminogen activator inhibitor-<em>1</em> and fibronectin in the kidneys of dTGR were increased and distributed similarly to ICAM-<em>1</em>. In 4-week-old dTGR, long-term treatment with ACE inhibition (cilazapril), AT<em>1</em> receptor blockade (valsartan), and human renin inhibition (RO 65-<em>7</em>2<em>1</em>9) (each drug <em>1</em>0 mg/kg by gavage once a day for 3 weeks) completely prevented the development of albuminuria. However, only cilazapril and valsartan were able to decrease blood pressure to normotensive levels. Interestingly, the drugs were all equally effective in preventing monocyte/macrophage infiltration and the overexpression of adhesion molecules, plasminogen activator inhibitor-<em>1</em>, and fibronectin in the kidney. Our findings indicate that <em>angiotensin</em> II causes monocyte recruitment and vascular inflammatory response in the kidney by blood pressure-dependent and blood pressure-independent mechanisms. ACE inhibition, AT<em>1</em> receptor blockade, and human renin inhibition all prevent monocyte/macrophage infiltration and increased adhesion molecule expression in the kidneys of dTGR.

We compared the effects of the converting enzyme inhibitor perindopril on components of the renin-<em>angiotensin</em> system in plasma and kidney of male Sprague-Dawley rats administered perindopril in their drinking water at two doses (<em>1</em>.4 and 4.2 mg/kg) over <em>7</em> days. Eight <em>angiotensin</em> peptides were measured in plasma and kidney: <em>angiotensin</em>-(<em>1</em>-<em>7</em>), <em>angiotensin</em> II, <em>angiotensin</em>-(<em>1</em>-9), <em>angiotensin</em> I, <em>angiotensin</em>-(2-<em>7</em>), <em>angiotensin</em> III, <em>angiotensin</em>-(2-9), and <em>angiotensin</em>-(2-<em>1</em>0). In addition, <em>angiotensin</em> converting enzyme activity, renin, and <em>angiotensin</em>ogen were measured in plasma, and renin, <em>angiotensin</em>ogen, and their respective messenger RNAs were measured in kidney; <em>angiotensin</em>ogen messenger RNA was also measured in liver. In plasma, the highest dose of perindopril reduced <em>angiotensin</em> converting enzyme activity to <em>1</em><em>1</em>% of control, increased renin 200-fold, reduced <em>angiotensin</em>ogen to <em>1</em><em>1</em>% of control, increased <em>angiotensin</em>-(<em>1</em>-<em>7</em>), <em>angiotensin</em> I, <em>angiotensin</em>-(2-<em>7</em>), and <em>angiotensin</em>-(2-<em>1</em>0) levels 25-, 9-, <em>1</em>0-, and <em>1</em>3-fold, respectively; <em>angiotensin</em> II levels were not significantly different from control. By contrast, for the kidney, <em>angiotensin</em>-(<em>1</em>-<em>7</em>), <em>angiotensin</em> I, <em>angiotensin</em>-(2-<em>7</em>), and <em>angiotensin</em>-(2-<em>1</em>0) levels did not increase; <em>angiotensin</em> II levels fell to <em>1</em>4% of control, and <em>angiotensin</em>ogen fell to <em>1</em>2% of control. Kidney renin messenger RNA levels increased <em>1</em>2-fold, but renal renin content and <em>angiotensin</em>ogen messenger RNA levels in kidney and liver were not influenced by perindopril treatment. These results demonstrate a differential regulation of <em>angiotensin</em> peptides in plasma and kidney and provide direct support for the proposal that the cardiovascular effects of converting enzyme inhibitors depend on modulation of tissue <em>angiotensin</em> systems. Moreover, the failure of kidney <em>angiotensin</em> I levels to increase with perindopril treatment, taken together with the fall in kidney <em>angiotensin</em>ogen levels, suggests that <em>angiotensin</em>ogen may be a major rate-limiting determinant of <em>angiotensin</em> peptide levels in the kidney.

<em>Angiotensin</em>-converting enzyme (ACE) inhibition alone or in combination with the <em>angiotensin</em> type-I receptor (AT<em>1</em>) antagonist losartan augments circulating levels of the bioactive peptide <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)]. Hence, we determined whether Ang-(<em>1</em>-<em>7</em>) contributes to the hypotensive effects produced by the combined administration of lisinopril and losartan in spontaneously hypertensive rats by blocking the peptide's synthesis with either of two structurally different neprilysin inhibitors. Intravenous administration of CGS 24592 (30 mg/kg) to rats in which blood pressure was normalized by 9 days of therapy with lisinopril and losartan elicited an elevation of mean arterial pressure that was sustained throughout the infusion period and for 20 minutes thereafter. The hypertensive response was associated with a 62% reduction in circulating levels of Ang-(<em>1</em>-<em>7</em>) and no change in plasma <em>angiotensin</em> II (Ang II). Intravenous infusion of one other neprilysin inhibitor (SCH 393<em>7</em>0, 30 mg/kg) produced an increase in mean blood pressure of a magnitude similar to that found with CGS 24592. Pretreatment with the nonselective antagonist [Sar<em>1</em>,Thr8]-Ang II abolished any additional pressor effects of either neprilysin inhibitor in spontaneously hypertensive rats treated with lisinopril or losartan. However, neither the endothelin A antagonist BQ<em>1</em>23 nor the kinin B2 antagonist HOE <em>1</em>40 had an effect on basal blood pressure or altered the pressor or heart rate effects of the neprilysin inhibitors. These data suggest that inhibition of Ang-(<em>1</em>-<em>7</em>) formation in rats exposed to the combined blockade of Ang II production and activity is associated with a reversal of the antihypertensive actions produced by these therapies. Thus, endogenous Ang-(<em>1</em>-<em>7</em>) functions as a vasodilator hormone in this form of genetic hypertension.

Increasing evidence suggests elevated sympathetic outflow may be important in the genesis of hypertension. It is thought that peripheral <em>angiotensin</em> II, in addition to its pressor actions, may act centrally to increase sympathetic nerve activity (SNA). Without direct long-term recordings of SNA, testing the involvement of neural mechanisms in <em>angiotensin</em> II-induced increases in arterial pressure is difficult. Using a novel telemetry-based implantable amplifier, we made continuous recordings of renal SNA (RSNA) before, during, and after <em>1</em> week of <em>angiotensin</em> II-based hypertension in rabbits living in their home cages. <em>Angiotensin</em> II infusion (50 ng x kg(-<em>1</em>) x min(-<em>1</em>)) caused a sustained increase in arterial pressure (<em>1</em>8+/-3 mm Hg). There was a sustained decrease in RSNA from <em>1</em>8+/-2 normalized units (n.u.) before <em>angiotensin</em> II to 8+/-2 n.u. on day 2 and 9+/-2 n.u. on day <em>7</em> of the <em>angiotensin</em> II infusion (P<0.0<em>1</em>) before recovering to <em>1</em><em>7</em>+/-2 n.u. after ceasing <em>angiotensin</em> II. Analysis of the baroreflex response showed that although <em>angiotensin</em> II-induced hypertension led to resetting of the relationship between mean arterial pressure (MAP) and heart rate, there was no evidence of resetting of the MAP-RSNA relationship. We propose that the lack of resetting of the MAP-RSNA curve, with the resting point lying near the lower plateau, suggests the sustained decrease in RSNA during <em>angiotensin</em> II is baroreflex mediated. These results suggest that baroreflex control of RSNA and thus renal function is likely to play a significant role in the control of arterial pressure not only in the short term but also in the long term.

<em>Angiotensin</em> (Ang) II-induced organ damage has fascinated students of hypertension since the work of Wilson and Byrom. We are investigating a double transgenic rat (dTGR) model, in which rats transgenic for the human angiotensinogen and renin genes are crossed. These rats develop moderately severe hypertension but die of end-organ cardiac and renal damage by week <em>7</em>. The heart shows necrosis and fibrosis, whereas the kidneys resemble the hemolytic-uremic syndrome vasculopathy. Surface adhesion molecules (ICAM-<em>1</em> and VCAM-<em>1</em>) are expressed early on the endothelium, while the corresponding ligands are found on circulating leukocytes. Leukocyte infiltration in the vascular wall accompanies PAI-<em>1</em>, MCP-<em>1</em>, and VEGF expression. The expression of TGF-beta and deposition of extracellular matrix proteins follows, which is accompanied by fibrinoid vasculitis in small vessels of the heart and kidneys. <em>Angiotensin</em>-converting enzyme inhibitors and AT<em>1</em> receptor blockers each lowered blood pressure and shifted pressure natriuresis partially leftward by different mechanisms. When combined, they normalized blood pressure, pressure natriuresis, and protected from vasculopathy completely. Renin inhibition lowered blood pressure partially, but protected from vasculopathy completely. Endothelin receptor blockade had no influence on blood pressure but protected from vasculopathy and improved survival. We show evidence that Ang II stimulates oxidative stress directly or indirectly via endothelin <em>1</em> and that NFkappaB is upregulated in this model. We speculate that the transcription factors NFkappaB and AP-<em>1</em> are involved with initiating chemokine and cytokine expression, leading to the above cascade. The unique model and our pharmacological probes will enable us to test these hypotheses.

BACKGROUND

The role of the angiotensin II type 2 receptor (AT2-R) in left ventricular (LV) remodeling may depend on the underlying stimulus. We hypothesized that cardiac AT2-R overexpression in transgenic (TG) mice would attenuate remodeling after myocardial infarction (MI).

RESULTS

Ten wild-type (WT) C57BL/6 mice and 12 TG mice that overexpress the AT2-R in the heart were studied by cardiac MRI at baseline and days 1, 7, and 28 post-MI induced by 1 hour of occlusion of the LAD followed by reperfusion. Short-axis imaging from apex to base was used to determine LV mass index, end-diastolic and end-systolic volume indices (EDVI, ESVI), regional wall thickness and thickening, and ejection fraction (EF). Gadolinium-DTPA was infused 20 minutes before day 1 imaging to assess infarct size. At baseline, heart rate, blood pressure, LV mass index, and EDVI were similar between groups. Baseline ESVI was lower (0.20+/-0.07 versus 0.45+/-0.15 microL/g, P<0.001) and EF higher (82.3+/-4.9% versus 67.7+/-5.3%, P<0.001) in TG than WT. Infarct size was similar (36.6+/-7.2% in WT, 34.0+/-7.8% in TG, P=NS). When controlled for baseline differences, ESVI was significantly less and EF significantly higher at all time points in TG versus WT. At day 28, ESVI was 1.05+/-0.32 microL/g in TG and 1.63+/-0.41 microL/g in WT, P<0.03, and EF was 47.3+/-5.8% versus 34.1+/-9.2%, P<0.003, respectively. Regional wall thickness and thickening were greater in TG both at baseline and at day 28. At day 28, blood pressure and LV dP/dt were higher in TG.

CONCLUSIONS

Cardiac AT2-R overexpression improves LV systolic function at baseline and preserves function during post-MI remodeling.

Activation of the renin-<em>angiotensin</em>-aldosterone system in arterial hypertension can lead to remodeling of the myocardial collagen network, with progressive collagen accumulation in the cardiac interstitium. This reactive myocardial fibrosis, which is not secondary to myocyte necrosis, appears to be an important determinant of diastolic dysfunction and thus of pathologic hypertrophy. To examine the effects of the aldosterone antagonist spironolactone on myocardial fibrosis, we analyzed interstitial fibrosis in <em>7</em> different models of arterial hypertension in rats: 2 kidney, <em>1</em> clip model of renovascular hypertension (RHT); continuous subcutaneous aldosterone (0.<em>7</em>5 micrograms/hr) infusion; RHT and aldosterone models treated with 20 mg/kg per day of subcutaneous spironolactone; uninephrectomized rats on a high sodium diet; and age- and sex-matched controls with or without spironolactone treatment. Systolic arterial pressure was comparably elevated in RHT and aldosterone models; it was modestly lowered but not normalized by 8 weeks of spironolactone treatment at the low doses used. Such treatment also failed to prevent left ventricular hypertrophy (LVH) in all experimental groups with hypertension. Spironolactone, however, was able to prevent myocardial fibrosis in RHT and aldosterone models of acquired arterial hypertension irrespective of the development of LVH and the presence of hypertension. These findings provide further evidence that elevated aldosterone levels play an important role in the adverse remodeling of the myocardium in arterial hypertension. The antifibrotic effects of spironolactone, if confirmed in human studies, may be a valuable strategy in treating hypertensive heart disease.

The present study was designed to investigate the effect of intracerebroventricular (icv) and intravenous (iv) infusion of <em>angiotensin</em> (ANG)-(<em>1</em>-<em>7</em>), ANG III, and ANG II on the baroreceptor control of heart rate (BHR) in conscious rats. Reflex changes in HR were elicited by bolus iv injection of either phenylephrine or sodium nitroprusside before and within <em>1</em> and 3 h of icv infusion of ANG II (n = <em>1</em>0), ANG III (n = 9), ANG-(<em>1</em>-<em>7</em>) (n = 9), or saline (n = 9) at a rate of 3 nmol.<em>7</em>.5 microliter-<em>1</em>.h-<em>1</em>. In another group of animals (n = 23), iv infusion of the same amount of ANG peptides was carried out at a rate of 0.<em>7</em> ml/h. The average ratio of changes in HR in beats per minute and changes in mean arterial pressure (MAP, mmHg) was used as an index of BHR sensitivity. ANG II and ANG III produced a significant increase in the basal levels of MAP, but only during the first hour of infusion (iv or icv). No significant changes in baseline HR were observed. ANG-(<em>1</em>-<em>7</em>) and saline infusion did not change basal levels of HR or MAP (iv or icv). ANG II (iv and icv) and ANG III (icv) caused a significant decrease in the BHR sensitivity for reflex bradycardia. In contrast, icv infusion of ANG-(<em>1</em>-<em>7</em>) induced a significant increase in BHR sensitivity for reflex bradycardia (-3.0 +/- 0.3, <em>1</em> h, and -2.8 +/- 0.<em>1</em> beats.min-<em>1</em>.mmHg-<em>1</em>, 3 h vs. -2.<em>1</em> +/- 0.2 beats.min-<em>1</em>.mmHg-<em>1</em>, before infusion).(ABSTRACT TRUNCATED AT 250 WORDS)

Understanding the physiological role of the plasma kallikrein-kinin system (KKS) has been hampered by not knowing how the proteins of this proteolytic system, when assembled in the intravascular compartment, become activated under physiological conditions. Recent studies indicate that the enzyme prolylcarboxypeptidase, an ANG II inactivating enzyme, is a prekallikrein activator. The ability of prolylcarboxypeptidase to act in the KKS and the renin-<em>angiotensin</em> system (RAS) indicates a novel interaction between these two systems. This interaction, along with the roles of <em>angiotensin</em> converting enzyme, cross talk between bradykinin and <em>angiotensin</em>-(<em>1</em>-<em>7</em>) action, and the opposite effects of activation of the ANG II receptors <em>1</em> and 2 support a hypothesis that the plasma KKS counterbalances the RAS. This review examines the interaction and cross talk between these two protein systems. This analysis suggests that there is a multilayered interaction between these two systems that are important for a wide array of physiological functions.

Recent studies have identified the presence of macrophages in the arterial wall of hypertensive animals and suggested that as is the case in atherosclerosis, macrophage products may be important mediators of the adaptive response of the arterial wall. In support of this, we have previously shown that the expression of monocyte chemoattractant protein-<em>1</em> is upregulated in the arteries of hypertensive animals. We hypothesized that macrophage recruitment is a critical step in the pathogenesis of hypertension. To obtain insights into this potential mechanism, we made use of mice deficient in the CC chemokine receptor 2 (CCR2), the receptor for monocyte chemoattractant protein-<em>1</em>. Hypertension was induced with the subcutaneous administration of <em>angiotensin</em> II (0.<em>7</em>5 mg. kg(-<em>1</em>). d(-<em>1</em>)) for <em>7</em> days. Using in situ hybridization with a probe for c-fms to identify macrophages, we found that hypertension-induced macrophage infiltration of the arterial wall was virtually eliminated in CCR2-deficient mice. In addition, vascular hypertrophy was reduced by approximately 65% compared with wild-type animals. These data demonstrate that CCR2 is essential for the recruitment of macrophages into the arterial wall in the setting of hypertension. Furthermore, the decreased hypertrophic response suggests that vascular hypertrophy occurs in part as a consequence of macrophage infiltration. In <em>angiotensin</em> II-induced hypertension, CCR2-mediated responses are critical to the process of macrophage recruitment and vascular hypertrophy and may represent one mechanism by which at least some forms of hypertension may lead to the development of atherosclerosis.

The carboxyl-terminal cytoplasmic domain of the <em>angiotensin</em> II type <em>1</em> (AT<em>1</em>) receptor has recently been shown to interact with several classes of cytoplasmic proteins that regulate different aspects of AT<em>1</em> receptor physiology. Employing yeast two-hybrid screening of a mouse kidney cDNA library with the carboxyl-terminal cytoplasmic domain of the murine AT<em>1</em>a receptor as a bait, we have isolated a novel protein with a predicted molecular mass of <em>1</em>8 kDa, which we have named ATRAP (for AT<em>1</em> receptor-associated protein). ATRAP interacts specifically with the carboxyl-terminal domain of the AT<em>1</em>a receptor but not with those of <em>angiotensin</em> II type 2 (AT2), m3 muscarinic acetylcholine, bradykinin B2, endothelin B, and beta2-adrenergic receptors. The mRNA of ATRAP was abundantly expressed in kidney, heart, and testis but was poorly expressed in lung, liver, spleen, and brain. The ATRAP-AT<em>1</em>a receptor association was confirmed by affinity chromatography, by specific co-immunoprecipitation of the two proteins, and by fluorescence microscopy, showing co-localization of these proteins in intact cells. Overexpression of ATRAP in COS-<em>7</em> cells caused a marked inhibition of AT<em>1</em>a receptor-mediated activation of phospholipase C without affecting m3 receptor-mediated activation. In conclusion, we have isolated a novel protein that interacts specifically with the carboxyl-terminal cytoplasmic domain of the AT<em>1</em>a receptor and affects AT<em>1</em>a receptor signaling.

The prorenin/renin receptor is a recently discovered component of the renin-<em>angiotensin</em> system. The effects of aliskiren, a direct inhibitor of human renin, were compared with the handle region decoy peptide (HRP), which blocks the prorenin/renin receptor, in double-transgenic rats overexpressing the human renin and <em>angiotensin</em>ogen genes. After <em>7</em> wk, all aliskiren-treated rats were alive, whereas mortality was 40% in vehicle-treated and 58% in HRP-treated rats. Aliskiren but not the HRP reduced BP and normalized albuminuria, cystatin C, and neutrophil gelatinase-associated lipocalin, a marker of renal tubular damage, to the levels of nontransgenic controls. In vitro, human renin and prorenin induced extracellular signal-regulated kinase <em>1</em>/2 phosphorylation, independent of <em>angiotensin</em> II (AngII), in vascular smooth muscle cells. Preincubation with the HRP or aliskiren did not prevent renin- and prorenin-induced extracellular signal-regulated kinase <em>1</em>/2 phosphorylation, whereas the MAP kinase kinase (MEK<em>1</em>/2) inhibitor PD98059 prevented both. In conclusion, renin inhibition but not treatment with the HRP protects against AngII-induced renal damage in double-transgenic rats. In addition, the in vitro data do not support the use of the HRP to block AngII-independent prorenin- or renin-mediated effects.

Accumulating evidence has demonstrated that up-regulation of the <em>angiotensin</em> (Ang)-converting enzyme (ACE)/AngII/AngII type <em>1</em> receptor (AT<em>1</em>R) axis aggravates pulmonary fibrosis. The recently discovered ACE2/Ang-(<em>1</em>-<em>7</em>)/Mas axis, which counteracts the activity of the ACE/AngII/AT<em>1</em>R axis, has been shown to protect against pulmonary fibrosis. However, the mechanisms by which ACE2 and Ang-(<em>1</em>-<em>7</em>) attenuate pulmonary fibrosis remain unclear. We hypothesized that up-regulation of the ACE2/Ang-(<em>1</em>-<em>7</em>)/Mas axis protects against bleomycin (BLM)-induced pulmonary fibrosis by inhibiting the mitogen-activated protein kinase (MAPK)/NF-κB pathway. In vivo, Ang-(<em>1</em>-<em>7</em>) was continuously infused into Wistar rats that had received BLM or AngII. In vitro, human fetal lung-<em>1</em> cells were pretreated with compounds that block the activities of AT<em>1</em>R, Mas (A-<em>7</em><em>7</em>9), and MAPKs before exposure to AngII or Ang-(<em>1</em>-<em>7</em>). The human fetal lung-<em>1</em> cells were infected with lentivirus-mediated ACE2 before exposure to AngII. In vivo, Ang-(<em>1</em>-<em>7</em>) prevented BLM-induced lung fibrosis and AngII-induced lung inflammation by inhibiting the MAPK phosphorylation and NF-κB signaling cascades. However, exogenous Ang-(<em>1</em>-<em>7</em>) alone clearly promoted lung inflammation. In vitro, Ang-(<em>1</em>-<em>7</em>) and lentivirus-mediated ACE2 inhibited the AngII-induced MAPK/NF-κB pathway, thereby attenuating inflammation and α-collagen I production, which could be reversed by the Mas inhibitor, A-<em>7</em><em>7</em>9. Ang-(<em>1</em>-<em>7</em>) inhibited AngII-induced lung fibroblast apoptotic resistance via inhibition of the MAPK/NF-κB pathway and activation of the BCL-2-associated X protein/caspase-dependent mitochondrial apoptotic pathway. Ang-(<em>1</em>-<em>7</em>) alone markedly stimulated extracellular signal-regulated protein kinase <em>1</em>/2 phosphorylation and the NF-κB cascade. Up-regulation of the ACE2/Ang-(<em>1</em>-<em>7</em>)/Mas axis protected against pulmonary fibrosis by inhibiting the MAPK/NF-κB pathway. However, close attention should be paid to the proinflammatory effects of Ang-(<em>1</em>-<em>7</em>).