Relaxin Does Not Improve Angiotensin II-Induced Target-Organ Damage

Relaxin is a corpus-luteum produced protein hormone with vasodilatatory, anti-fibrotic, and angiogenic properties that are opposite to angiotensin (Ang) II. We investigated whether or not relaxin ameliorates Ang II-induced target-organ damage. We used double transgenic rats harboring both human renin and angiotensinogen genes (dTGR) that develop severe hypertension, target-organ damage, and die untreated within 7–8 weeks. Recombinant relaxin at a low (26 μg/kg/d) and a high dose (240 μg/kg/d) was given to 4 week-old dTGR and age-matched Sprague-Dawley rats (SD). Systolic blood pressure increased progressively in untreated dTGRs from 162±3 mmHg at week 5 to 225±5 mmHg at week 7. Relaxin had no effect on blood pressure whereas SD rats were normotensive (106±1 mmHg). Untreated and relaxin-treated dTGR had similarly severe cardiac hypertrophy indices. Relaxin did not ameliorate albuminuria and did not prevent matrix-protein deposition in the heart and kidney in dTGR. Finally, relaxin treatment did not reduce mortality. These data suggest that pharmacological doses of relaxin do not reverse severe effects of Ang II.     

Cardiac hypertrophy is associated with altered thioredoxin and ASK-1 signaling in a mouse model of menopause

Oxidative stress is implicated in menopause-associated hypertension and cardiovascular disease. The role of antioxidants in this process is unclear. We questioned whether the downregulation of thioredoxin (TRX) is associated with oxidative stress and the development of hypertension and target-organ damage (cardiac hypertrophy) in a menopause model. TRX is an endogenous antioxidant that also interacts with signaling molecules, such as apoptosis signal-regulated kinase 1 (ASK-1), independently of its antioxidant function. Aged female wild-type (WT) and follitropin receptor knockout (FORKO) mice (20-24 wk), with hormonal imbalances, were studied. Mice were infused with ANG II (400 ng x kg(-1) x min(-1); 14 days). Systolic blood pressure was increased by ANG II in WT (166+/-8 vs. 121+/-5 mmHg) and FORKO (176+/-7 vs. 115+/-5 mmHg; P<0.0001; n=9/group) mice. In ANG II-infused FORKO mice, cardiac mass was increased by 42% (P<0.001). This was associated with increased collagen content and augmented ERK1/2 phosphorylation (2-fold). Cardiac TRX expression and activity were decreased by ANG II in FORKO but not in WT (P<0.01) mice. ASK-1 expression, cleaved caspase III content, and Bax/Bcl-2 content were increased in ANG II-infused FORKO (P<0.05). ANG II had no effect on cardiac NAD(P)H oxidase activity or on O(2)(*-) levels in WT or FORKO. Cardiac ANG II type 1 receptor expression was similar in FORKO and WT. These findings indicate that in female FORKO, ANG II-induced cardiac hypertrophy and fibrosis are associated with the TRX downregulation and upregulation of ASK-1/caspase signaling. Our data suggest that in a model of menopause, protective actions of TRX may be blunted, which could contribute to cardiac remodeling independently of oxidative stress and hypertension.     

Effects of spironolactone and eprosartan on cardiac remodeling and angiotensin-converting enzyme isoforms in rats with experimental heart failure

Angiotensin-converting enzyme (ACE)-2 is a newly described enzyme with antagonistic effects to those of the classical ACE (ACE-1). Both ANG II and aldosterone play an important role in the pathophysiology of congestive heart failure (CHF) and in the adverse cardiac remodeling during its development. In this study, we examined the effects of experimental CHF induced by an aortocaval fistula (ACF) and of its treatment with ANG II and aldosterone inhibitors on the relative levels of ACE-1 and ACE-2. We also compared the effects of spironolactone, an aldosterone antagonist, and eprosartan, an ANG II receptor antagonist, on heart hypertrophy and fibrosis in rats with ACF. Spironolactone (15 mg x kg(-1) x day(-1) ip, via minipump) or eprosartan (5 mg x kg(-1) x day(-1) ip, via minipump) was administered into rats with ACF for 14 and 28 days. Specific antibodies were used to determine the protein levels of myocardial ACE-1 and ACE-2. ACF increased the cardiac levels of ACE-1 and decreased those of ACE-2. Heart-to-body weight ratio significantly increased from 0.30 +/- 0.004% in sham-operated controls to 0.50 +/- 0.018% and 0.56 +/- 0.044% (P < 0.001) in rats with ACF, 2 and 4 wk after surgery, respectively, in association with increased plasma levels of aldosterone. The area occupied by collagen increased from 2.33 +/- 0.27% to 6.85 +/- 0.65% and 8.03 +/- 0.93% (P < 0.01), 2 and 4 wk after ACF, respectively. Both spironolactone and eprosartan decreased cardiac mass and collagen content and reversed the shift in ACE isoforms. ACF alters the ratio between ACE isoforms in a manner that increases local ANG II and aldosterone levels. Early treatment with both ANG II and aldosterone antagonists is effective in reducing this effect. Thus ACE isoform shift may represent an important component of the development of cardiac remodeling in response to hemodynamic overload, and its correction may contribute to the beneficial therapeutic effects of renin-angiotensin-aldosterone system inhibitors.     

Angiotensin II-mediated oxidative stress promotes myocardial tissue remodeling in the transgenic (mRen2) 27 Ren2 rat

Angiotensin II (ANG II) contributes to cardiac remodeling, hypertrophy, and left ventricular dysfunction. ANG II stimulation of the ANG type 1 receptor (AT(1)R) generates reactive oxygen species via NADPH oxidase, which facilitates this hypertrophy and remodeling. This investigation sought to determine whether cardiac oxidative stress and cellular remodeling could be attenuated by in vivo AT(1)R blockade (AT(1)B) (valsartan) or superoxide dismutase/catalase mimetic (tempol) treatment in a rodent model of chronically elevated tissue levels of ANG II, the transgenic (mRen2) 27 rat (Ren2). Ren2 rats overexpress the mouse renin transgene with resultant hypertension, insulin resistance, proteinuria, and cardiovascular damage. Young (6-7 wk old) male Ren2 and age-matched Sprague-Dawley rats were treated with valsartan (30 mg/kg), tempol (1 mmol/l), or placebo for 3 wk. Heart tissue NADPH oxidase (NOX) activity and immunohistochemical analysis of subunits NOX2, Rac1, and p22(phox), heart tissue malondialdehyde, and insulin-stimulated protein kinase B (Akt) activation were measured. Structural changes were assessed with cine MRI, transmission electron microscopy, and light microscopy. Increases in septal wall thickness and altered systolic function (cine MRI) were associated with perivascular fibrosis and increased mitochondria in Ren2 on light and transmission electron microscopy (P < 0.05). AT(1)B, but not tempol, reduced blood pressure (P < 0.05); significant improvements were seen with both AT(1)B and tempol on NOX activity, subunit expression, malondialdehyde, and insulin-mediated activation/phosphorylation of Akt (each P < 0.05). Collectively, these data suggest cardiac oxidative stress-induced structural and functional changes are driven, in part, by AT(1)R-mediated increases in NADPH oxidase activity.     

Potentiation of the antihypertensive action of losartan by peripheral overexpression of the ANG II type 2 receptor

Our previous studies demonstrated that peripheral overexpression of angiotensin II (ANG II) type 2 receptors (AT(2)R) prevents hypertension-induced cardiac hypertrophy and remodeling without altering high blood pressure. This, coupled with the observations that AT(2)R play a role in the antihypertensive actions of ANG II type 1 receptor (AT(1)R) blockers (ARBs), led us to propose that peripheral overexpression of AT(2)R would improve the antihypertensive action of losartan (Los) in Sprague-Dawley (SD) rats made hypertensive via chronic infusion of ANG II. Here we utilized adenoviral vector-mediated AT(2)R gene transfer to test this hypothesis. A single intracardiac injection of adenoviral vector containing genomic AT(2)R (G-AT(2)R) DNA and enhanced green fluorescent protein (EGFP) gene controlled by cytomegalovirus (CMV) promoters (Ad-G-AT(2)R-EGFP; 5 x 10(9) infectious units) into adult SD rats produced robust AT(2)R overexpression in cardiovascular tissues (kidney, lung, heart, aorta, mesenteric artery, and renal artery) that persisted for 3-5 days postinjection. By 7 days post viral injection, the overexpressed AT(2)R are reduced toward basal values in certain tissues (lung, kidney, and heart) and are undetectable in others (kidney and blood vessels). In two separate protocols, we demonstrated that the hypotensive effect of Los (0.125, 0.5, and 1.0 mg/kg iv) was significantly greater in the AT(2)R-overexpressing animals (-40.7 +/- 4.3, -41.8 +/- 4.8, and -48.1 +/- 2.6 mmHg, respectively) compared with control vector (Ad-CMV-EGFP)-treated rats (-12.4 +/- 2.2, -20.2 +/- 3.4, and -27.3 +/- 3.4 mmHg, respectively). These results provide support for a depressor role of AT(2)R and the proposal that combined AT(2)R agonist and ARB treatment may be an improved therapeutic strategy for controlling hypertension.     

Role of cardiac overexpression of ANG II in the regulation of cardiac function and remodeling postmyocardial infarction

ANG II has a clear role in development of cardiac hypertrophy, fibrosis, and dysfunction. It has been difficult, however, to determine whether these actions are direct or consequences of its systemic hemodynamic effects in vivo. To overcome this limitation, we used transgenic mice with cardiac-specific expression of a transgene fusion protein that releases ANG II from cardiomyocytes (Tg-ANG II-cardiac) without involvement of the systemic renin-angiotensin system and tested whether increased cardiac ANG II accelerates remodeling and dysfunction postmyocardial infarction (MI), whereas those mice show no evidence of cardiac hypertrophy under the basal condition. Male 12- to 14-wk-old Tg-ANG II-cardiac mice and their wild-type littermates (WT) were subjected to sham-MI or MI by ligating the left anterior descending coronary artery for 8 wk. Cardiac ANG II levels were approximately 10-fold higher in Tg-ANG II-cardiac mice than their WT, whereas ANG II levels in plasma and other tissues did not differ between strains. Systolic blood pressure and heart rate were similar between groups with or without MI. In sham-MI, Tg-ANG II-cardiac mice had increased collagen deposition and decreased capillary density. The differences between strains became more pronounced after MI. Although cardiac function was well preserved in the Tg-ANG II-cardiac mice with sham-MI, cardiac remodeling and dysfunction post-MI were more severe than WT. Our results demonstrate that, independent of systemic hemodynamic effects, cardiac ANG II may act locally in the heart, causing interstitial fibrosis in sham-MI and accelerating deterioration of cardiac dysfunction and remodeling post-MI.     

Prevention of angiotensin II-induced cardiac remodeling by angiotensin-(1-7)

Cardiac remodeling, which typically results from chronic hypertension or following an acute myocardial infarction, is a major risk factor for the development of heart failure and, ultimately, death. The renin-angiotensin system (RAS) has previously been established to play an important role in the progression of cardiac remodeling, and inhibition of a hyperactive RAS provides protection from cardiac remodeling and subsequent heart failure. Our previous studies have demonstrated that overexpression of angiotensin-converting enzyme 2 (ACE2) prevents cardiac remodeling and hypertrophy during chronic infusion of angiotensin II (ANG II). This, coupled with the knowledge that ACE2 is a key enzyme in the formation of ANG-(1-7), led us to hypothesize that chronic infusion of ANG-(1-7) would prevent cardiac remodeling induced by chronic infusion of ANG II. Infusion of ANG II into adult Sprague-Dawley rats resulted in significantly increased blood pressure, myocyte hypertrophy, and midmyocardial interstitial fibrosis. Coinfusion of ANG-(1-7) resulted in significant attenuations of myocyte hypertrophy and interstitial fibrosis, without significant effects on blood pressure. In a subgroup of animals also administered [d-Ala(7)]-ANG-(1-7) (A779), an antagonist to the reported receptor for ANG-(1-7), there was a tendency to attenuate the antiremodeling effects of ANG-(1-7). Chronic infusion of ANG II, with or without coinfusion of ANG-(1-7), had no effect on ANG II type 1 or type 2 receptor binding in cardiac tissue. Together, these findings indicate an antiremodeling role for ANG-(1-7) in cardiac tissue, which is not mediated through modulation of blood pressure or altered cardiac angiotensin receptor populations and may be at least partially mediated through an ANG-(1-7) receptor.     

Angiotensin-(1-7) binds to specific receptors on cardiac fibroblasts to initiate antifibrotic and antitrophic effects

ANG-(1-7) improves the function of the remodeling heart. Although this peptide is generated directly within the myocardium, the effects of ANG-(1-7) on cardiac fibroblasts that play a critical role in cardiac remodeling are largely unknown. We tested the hypothesis that specific binding of ANG-(1-7) to cardiac fibroblasts regulates cellular functions that are involved in cardiac remodeling. 125I-labeled ANG-(1-7) binding assays identified specific binding sites of ANG-(1-7) on adult rat cardiac fibroblasts (ARCFs) with an affinity of 11.3 nM and a density of 131 fmol/mg protein. At nanomolar concentrations, ANG-(1-7) interacted with specific sites that were distinct from ANG II type 1 and type 2 receptors without increasing cytosolic Ca2+ concentration. At these concentrations, ANG-(1-7) had inhibitory effects on collagen synthesis as assessed by [3H]proline incorporation and decreased mRNA expression of growth factors in ARCFs. These effects of ANG-(1-7) contrasted with effects of ANG II. Pretreatment of ARCFs with ANG-(1-7) inhibited ANG II-induced increases in collagen synthesis and in mRNA expression of growth factors, including endothelin-1 and leukemia inhibitory factor. ANG-(1-7) pretreatment also inhibited the stimulatory effects of conditioned medium from ANG II-treated ARCFs on [3H]leucine incorporation and atrial natriuretic factor mRNA expression, markers of hypertrophy, in cardiomyocytes. Thus ANG-(1-7) interacted with specific receptors on ARCFs to exert potential antifibrotic and antitrophic effects that could reverse ANG II effects. These results suggest that ANG-(1-7) may play an important role in the heart in regulating cardiac remodeling.     

Angiotensin-(1-7) attenuates angiotensin II-induced cardiac remodeling associated with upregulation of dual-specificity phosphatase 1

Chronic hypertension induces cardiac remodeling, including left ventricular hypertrophy and fibrosis, through a combination of both hemodynamic and humoral factors. In previous studies, we showed that the heptapeptide ANG-(1-7) prevented mitogen-stimulated growth of cardiac myocytes in vitro, through a reduction in the activity of the MAPKs ERK1 and ERK2. In this study, saline- or ANG II-infused rats were treated with ANG-(1-7) to determine whether the heptapeptide reduces myocyte hypertrophy in vivo and to identify the signaling pathways involved in the process. ANG II infusion into normotensive rats elevated systolic blood pressure >50 mmHg, in association with increased myocyte cross-sectional area, ventricular atrial natriuretic peptide mRNA, and ventricular brain natriuretric peptide mRNA. Although infusion with ANG-(1-7) had no effect on the ANG II-stimulated elevation in blood pressure, the heptapeptide hormone significantly reduced the ANG II-mediated increase in myocyte cross-sectional area, interstitial fibrosis, and natriuretic peptide mRNAs. ANG II increased phospho-ERK1 and phospho-ERK2, whereas cotreatment with ANG-(1-7) reduced the phosphorylation of both MAPKs. Neither ANG II nor ANG-(1-7) altered the ERK1/2 MAPK kinase MEK1/2. However, ANG-(1-7) infusion, with or without ANG II, increased the MAPK phosphatase dual-specificity phosphatase (DUSP)-1; in contrast, treatment with ANG II had no effect on DUSP-1, suggesting that ANG-(1-7) upregulates DUSP-1 to reduce ANG II-stimulated ERK activation. These results indicate that ANG-(1-7) attenuates cardiac remodeling associated with a chronic elevation in blood pressure and upregulation of a MAPK phosphatase and may be cardioprotective in patients with hypertension.     

Adapter molecule DOC-2 is differentially expressed in pressure and volume overload hypertrophy and inhibits collagen synthesis in cardiac fibroblasts.

DOC-2 (differentially expressed in ovarian carcinoma) is involved in Ras-, beta-integrin-, PKC-, and transforming growth factor-beta-mediated cell signaling. These pathways are implicated in the accumulation of extracellular matrix proteins during progression of hypertrophy to heart failure; however, the role of DOC-2 in cardiac pathophysiology has never been examined. This study was undertaken to 1) analyze DOC-2 expression in primary cultures of cardiac fibroblasts and cardiac myocytes and in the heart following different types of hemodynamic overloads and 2) examine its role in growth factor-mediated ERK activation and collagen production. Pressure overload and volume overload were induced for 10 wk in Sprague-Dawley rats by aortic constriction and by aortocaval shunt, respectively. ANG II (0.3 mg.kg(-1).day(-1)) was infused for 2 wk. Results showed that, compared with myocytes, DOC-2 was found abundantly expressed in cardiac fibroblasts. Treatment of cardiac fibroblasts with ANG II and TPA resulted in increased expression of DOC-2. Overexpression of DOC-2 in cardiac fibroblasts led to inhibition of hypertrophy agonist-stimulated ERK activation and collagen expression. An inverse correlation between collagen and DOC-2 was observed in in vivo models of cardiac hypertrophy; in pressure overload and after ANG II infusion, increased collagen mRNA correlated with reduced DOC-2 levels, whereas in volume overload increased DOC-2 levels were accompanied by unchanged collagen mRNA. These data for the first time describe expression of DOC-2 in the heart and demonstrate its modulation by growth-promoting agents in cultured cardiac fibroblasts and in in vivo models of heart hypertrophy. Results suggest a role of DOC-2 in cardiac remodeling involving collagen expression during chronic hemodynamic overload.     

Effects of central infusion of ANG II and losartan on the cardiac baroreflex in rabbits

The effect of chronic activation or inhibition of central ANG II receptors on cardiac baroreflex function in conscious normotensive rabbits was examined. Animals received a fourth ventricular (4V) infusion of ANG II (30 and 100 ng/h), losartan (3 and 30 microg/h), or Ringer solution (2 microl/h) for 2 wk. After 1 and 2 wk, ANG II (100 ng/h) decreased cardiac baroreflex gain by 20 and 37%, respectively (P = 0.015), whereas losartan (30 microg/h) increased baroreflex gain by 24 and 58%, respectively (P = 0.02). Within 1 wk of the end of the infusions, cardiac baroreflex gain had returned to control. Ringer solution or the lower doses of ANG II or losartan did not modify the cardiac baroreflex function. Blood pressure and heart rate were not altered by any treatment, nor was their variability affected. These data demonstrate a novel long-term modulation of cardiac baroreflexes by endogenous ANG II that is independent of blood pressure level.     

Effects of all-trans retinoic acid on angiotensin II-induced myocyte hypertrophy.

We used cultured neonatal rat cardiac myocytes to test the hypothesis that all-trans retinoic acid (atRA) may act to modulate ANG II actions in inducing myocyte hypertrophy. Our observations were as follows. 1) atRA (10(-7) to approximately 10(-5) M ) inhibited ANG II-induced hyperplasia of fibroblasts in a dose-dependent manner. 2) Treatment of atRA attenuated the ANG II-induced increase in total cell protein content. 3) Treated with ANG II (10(-7) M) for 5 days, the cultured neonatal rat cardiac myocytes demonstrated an apparent accumulation of sarcomeric fiber proteins and Golgi's complex, as well as reorganization of the sarcomeric unit within individual myocytes. atRA (10(-6) M) treatment reduced the accumulation of contractile proteins and Golgi's complex without affecting the ANG II-induced reorganization of the sarcomeric unit. 4) atRA attenuated the ANG II-induced increase in intracellular Ca2+. Our results show that atRA inhibits some effects of ANG II on neonatal rat cardiac myocytes and suggest that atRA may be a therapeutic candidate for the prevention and therapy of cardiac hypertrophy and remodeling.     

Angiotensin-(1-7) attenuates hypertension in exercise-trained renal hypertensive rats

Angiotensin-(1-7) [ANG-(1-7)] plays a counterregulatory role to angiotensin II in the renin-angiotensin system. In trained spontaneous hypertensive rats, Mas expression and protein are upregulated in ventricular tissue. Therefore, we examined the role of ANG-(1-7) on cardiac hemodynamics, cardiac functions, and cardiac remodeling in trained two-kidney one-clip hypertensive (2K1C) rats. For this purpose, rats were divided into sedentary and trained groups. Each group consists of sham and 2K1C rats with and without ANG-(1-7) infusion. Swimming training was performed for 1 h/day, 5 days/wk for 4 wk following 1 wk of swimming training for acclimatization. 2K1C rats showed moderate hypertension and left ventricular hypertrophy without changing left ventricular function. Chronic infusion of ANG-(1-7) attenuated hypertension and cardiac hypertrophy only in trained 2K1C rats but not in sedentary 2K1C rats. Chronic ANG-(1-7) treatment significantly attenuated increases in myocyte diameter and cardiac fibrosis induced by hypertension in only trained 2K1C rats. The Mas receptor, ANG II type 2 receptor protein, and endothelial nitric oxide synthase phosphorylation in ventricles were upregulated in trained 2K1C rats. In conclusion, chronic infusion of ANG-(1-7) attenuates hypertension in trained 2K1C rats.     

Angiotensin II Induced Cardiac Dysfunction on a Chip

In vitro disease models offer the ability to study specific systemic features in isolation to better understand underlying mechanisms that lead to dysfunction. Here, we present a cardiac dysfunction model using angiotensin II (ANG II) to elicit pathological responses in a heart-on-a-chip platform that recapitulates native laminar cardiac tissue structure. Our platform, composed of arrays of muscular thin films (MTF), allows for functional comparisons of healthy and diseased tissues by tracking film deflections resulting from contracting tissues. To test our model, we measured gene expression profiles, morphological remodeling, calcium transients, and contractile stress generation in response to ANG II exposure and compared against previous experimental and clinical results. We found that ANG II induced pathological gene expression profiles including over-expression of natriuretic peptide B, Rho GTPase 1, and T-type calcium channels. ANG II exposure also increased proarrhythmic early after depolarization events and significantly reduced peak systolic stresses. Although ANG II has been shown to induce structural remodeling, we control tissue architecture via microcontact printing, and show pathological genetic profiles and functional impairment precede significant morphological changes. We assert that our in vitro model is a useful tool for evaluating tissue health and can serve as a platform for studying disease mechanisms and identifying novel therapeutics.     

Role of asymmetric dimethylarginine for angiotensin II-induced target organ damage in mice

The aim of the present study was to investigate the role of the endogenous nitric oxide synthase inhibitor asymmetric dimethylarginine (ADMA) and its degrading enzyme dimethylarginine dimethylaminohydrolase (DDAH) in angiotensin II (ANG II)-induced hypertension and target organ damage in mice. Mice transgenic for the human DDAH1 gene (TG) and wild-type (WT) mice (each, n = 28) were treated with 1.0 microg kg(-1) min(-1) ANG II, 3.0 microg kg(-1) min(-1) ANG II, or phosphate-buffered saline over 4 wk via osmotic minipumps. Blood pressure, as measured by tail cuff, was elevated to the same degree in TG and WT mice. Plasma levels of ADMA were lower in TG than WT mice and were not affected after 4 wk by either dose of ANG II in both TG and WT animals. Oxidative stress within the wall of the aorta, measured by fluorescence microscopy using the dye dihydroethidium, was significantly reduced in TG mice. ANG II-induced glomerulosclerosis was similar between WT and TG mice, whereas renal interstitial fibrosis was significantly reduced in TG compared with WT animals. Renal mRNA expression of protein arginine methyltransferase (PRMT)1 and DDAH2 increased during the infusion of ANG II, whereas PRMT3 and endogenous mouse DDAH1 expression remained unaltered. Chronic infusion of ANG II in mice has no effect on the plasma levels of ADMA after 4 wk. However, an overexpression of DDAH1 alleviates ANG II-induced renal interstitial fibrosis and vascular oxidative stress, suggesting a blood pressure-independent effect of ADMA on ANG II-induced target organ damage.     

Enhanced renal expression of preproendothelin mRNA during chronic angiotensin II hypertension

The purpose of this study was to determine the role of endothelin in mediating the renal hemodynamic and arterial pressure changes observed during chronic ANG II-induced hypertension. ANG II (50 ng x kg(-1) x min(-1)) was chronically infused into the jugular vein by miniosmotic pump for 2 wk in male Sprague-Dawley rats with and without endothelin type A (ET(A))-receptor antagonist ABT-627 (5 mg x kg(-1) x day(-1)) pretreatment. Arterial pressure increased in ANG II rats compared with control rats (149 +/- 5 vs. 121 +/- 6 mmHg, P < 0.05, respectively). Renal expression of preproendothelin mRNA was increased by approximately 50% in both the medulla and cortex of ANG II rats. The hypertensive effect of ANG II was completely abolished in rats pretreated with the ET(A)-receptor antagonist (114 +/- 5 mmHg, P < 0.05). Glomerular filtration rate was decreased by 33% in ANG II rats, and this response was attenuated in rats pretreated with ET(A)-receptor antagonist. These data indicate that activation of the renal endothelin system by ANG II may play an important role in mediating chronic renal and hypertensive actions of ANG II.     

Effects of human pregnancy on fluid regulation responses to short-term exercise.

This study tested the hypothesis that human pregnancy alters fluid and electrolyte regulation responses to acute short-term exercise. Responses of 22 healthy pregnant women (PG; gestational age, 37.0 +/- 0.2 wk) and 17 nonpregnant controls (CG) were compared at rest and during cycling at 70 and 110% of the ventilatory threshold (VT). At rest, ANG II concentration was significantly (P < 0.05) higher in PG vs. CG, whereas plasma osmolality and concentrations of AVP, sodium, and potassium were significantly lower. Atrial natriuretic peptide concentration at rest was similar between groups. ANG II and AVP concentrations increased significantly from rest to 110% VT in CG only, whereas increases in atrial natriuretic peptide concentration were similar between groups. Increases in osmolality, and total protein and albumin concentrations from rest to both work rates were similar between the two groups. PG and CG exhibited similar shifts in fluid during acute short-term exercise, but the increases in ANG II and AVP were absent and attenuated, respectively, during pregnancy. This was attributed to the significantly augmented fluid volume state already present at rest in late gestation.     

Primary role of angiotensin-converting enzyme-2 in cardiac production of angiotensin-(1-7) in transgenic Ren-2 hypertensive rats

Angiotensin-converting enzyme-2 (ACE2) converts angiotensin II (ANG II) to angiotensin-(1-7) [ANG-(1-7)], and this enzyme may serve as a key regulatory juncture in various tissues. Although the heart expresses ACE2, the extent that the enzyme participates in the cardiac processing of ANG II and ANG-(1-7) is equivocal. Therefore, we utilized the Langendorff preparation to characterize the ACE2 pathway in isolated hearts from male normotensive Sprague-Dawley [Tg((-))] and hypertensive [mRen2]27 [Tg((+))] rats. During a 60-min recirculation period with 10 nM ANG II, the presence of ANG-(1-7) was assessed in the cardiac effluent. ANG-(1-7) generation from ANG II was similar in both the normal and hypertensive hearts [Tg((-)): 510 +/- 55 pM, n=20 vs. Tg((+)): 497 +/- 63 pM, n=14] with peak levels occurring at 30 min after administration of the peptide. ACE2 inhibition (MLN-4760, 1 microM) significantly reduced ANG-(1-7) production by 83% (57 +/- 19 pM, P<0.01, n=7) in the Tg((+)) rats, whereas the inhibitor had no significant effect in the Tg((-)) rats (285 +/- 53 pM, P>0.05, n=10). ACE2 activity was found in the effluent of perfused Tg((-)) and Tg((+)) hearts, and it was highly associated with ACE2 protein expression (r=0.78). This study is the first demonstration for a direct role of ACE2 in the metabolism of cardiac ANG II in the hypertrophic heart of hypertensive rats. We conclude that predominant expression of cardiac ACE2 activity in the Tg((+)) may be a compensatory response to the extensive cardiac remodeling in this strain.     

Sar1Ile7]angiotensin III, a new selective antagonist of the pressor effect of angiotensin III in conscious rats

Two analogues of angiotensin III were compared as antagonists of the pressor response to angiotensin II (ANG II) and angiotensin III (ANG III) in conscious, unrestrained rats. Dose-mean arterial pressure (MAP) response curves were obtained for ANG II and ANG III in the absence or presence of [Ile7]ANG III (1.3 x 10(-7) mol/kg) or [Sar1 Ile7]ANG III (1.2 x 10(-7) mol/kg). In the presence of [Ile7]ANG III, the dose-MAP response curves for ANG II and ANG III were significantly displaced to the right. [Ile7]ANG III behaved as a partial agonist on ANG II but not ANG III receptors. In the presence of [Sar1 Ile7]ANG III, the dose-MAP response curve for ANG III but not ANG II was significantly displaced to the right. This suggests that [Sar1 Ile7]ANG III is a selective antagonist of ANG III in the vasculature. [Ile7]ANG III, on the other hand, antagonizes both ANG II and ANG III receptors. Our results support the hypothesis of the existence of a sub-class of angiotensin receptors activated by ANG III in the vascular smooth muscle.     

ANG II-induced cardiac molecular and cellular events: role of aldosterone

Chronic elevation of circulating ANG II is associated with cardiac remodeling in patients with hypertension and heart failure. The underlying mechanisms, however, are not completely defined. Herein, we studied ANG II-induced molecular and cellular events in the rat heart as well as their links to the redox state. We also addressed the potential contribution of aldosterone (ALDO) on ANG II-induced cardiac remodeling. In ANG II-treated rats, and compared with controls, we found: 1) the expression of proinflammatory/profibrogenic mediators was significantly increased in the perivascular space and at the sites of microscopic injury in both ventricles; 2) macrophages and myofibroblasts were primary repairing cells at these sites, together with increased fibrillar collagen volume; 3) apoptotic macrophages and myofibroblasts were evident at the same sites; 4) NADPH oxidase (gp91phox) was significantly enhanced at these regions and primarily expressed by macrophages, whereas superoxide dismutase and catalase levels remained unchanged; 5) plasma 8-isoprostane levels were significantly increased; and 6) blood pressure was significantly elevated. Losartan treatment completely prevented cardiac oxidative stress as well as molecular/cellular responses and normalized blood pressure. Spironolactone treatment partially suppressed the cardiac inflammatory/fibrogenic responses and redox state. Thus chronic elevation of circulating ANG II is accompanied by a proinflammatory/profibrogenic phenotype involving vascular and myocardial remodeling in both ventricles. Enhanced reactive oxygen species production at these sites and increased plasma 8-isoprostane indicate the involvement of oxidative stress in ANG II-induced cardiac injury. ALDO contributes, in part, to ANG II-induced cardiac molecular and cellular responses.