Age-related impairments in baroreflex sensitivity in Sprague-Dawley rats are associated with low solitary tract nucleus content of <em>angiotensin</em>-(<em>1</em>-<em>7</em>). However, transgenic rats with low-brain <em>angiotensin</em>ogen resulting from glial overexpression of an antisense oligonucleotide to <em>angiotensin</em>ogen (ASrAOGEN) are spared age-related declines in cardiovascular function characteristic of Sprague-Dawley rats. We examine whether cardiovascular and reflex actions of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) persist in the solitary tract nucleus of older (<em>1</em>6 to 22 months) ASrAOGEN rats. Baroreflex sensitivity for control of heart rate and chemosensitive vagal afferent activation in response to phenylbiguanide were measured before and after bilateral microinjection of the <em>angiotensin</em> II type <em>1</em> receptor antagonist candesartan and <em>angiotensin</em>-(<em>1</em>-<em>7</em>) receptor antagonist (D-Ala(<em>7</em>))-<em>angiotensin</em>-(<em>1</em>-<em>7</em>) in urethane/chloralose-anesthetized rats. In older anesthetized ASrAOGEN rats, candesartan had no effect, whereas (D-Ala(<em>7</em>))-<em>angiotensin</em>-(<em>1</em>-<em>7</em>) significantly reduced baroreflex sensitivity (<em>1</em>.80+/-0.43 versus 0.50+/-0.<em>1</em><em>7</em> ms/mm Hg). Phenylbiguanide responses were attenuated by injection of candesartan (-<em>7</em>9+/-6 versus -55+/-<em>1</em>2 mm Hg and -2<em>7</em><em>7</em>+/-<em>1</em>2 versus -<em>1</em>56+/-2<em>7</em> bpm; P<0.05). In addition, resting blood pressure was reduced by injection of candesartan or (D-Ala(<em>7</em>))-<em>angiotensin</em>-(<em>1</em>-<em>7</em>). Within the solitary tract nucleus of older ASrAOGEN rats, it appears that glial <em>angiotensin</em>ogen is the main source of <em>angiotensin</em> II attenuation of baroreflex sensitivity; endogenous <em>angiotensin</em>-(<em>1</em>-<em>7</em>) from nonglial sources enhances baroreflex sensitivity; nonglial sources of <em>angiotensin</em> II contribute to chemosensitive vagal afferent activation; and receptors for both peptides modulate resting arterial pressure under anesthesia. These results suggest a novel mechanism for the preservation of baroreflex sensitivity during aging.

Reports on the effectiveness of endothelin receptor blockers in <em>angiotensin</em> (Ang) II-induced end-organ damage are conflicting, and the mechanisms involved are uncertain. We tested the hypothesis that endothelin (ET)(A/B) receptor blockade with bosentan (<em>1</em>00 mg/kg by gavage after age 4 weeks) ameliorates cardiac and renal damage by decreasing inflammation in rats harboring both human renin and <em>angiotensin</em>ogen genes (dTGR). Furthermore, we elucidated the effect of bosentan on tissue factor (TF), which is a key regulator of the extrinsic coagulation cascade. We compared bosentan with hydralazine (80 mg/L in the drinking water for 3 weeks) as a blood pressure control. Untreated dTGR featured hypertension, focal necrosis in heart and kidney, and a 45% mortality rate (9 of 20) at age <em>7</em> weeks. Compared with Sprague-Dawley controls, both systolic blood pressure and 24-hour albuminuria were increased in untreated dTGR (203+/-8 versus <em>1</em><em>1</em><em>1</em>+/-2 mm Hg and 6<em>7</em>.<em>1</em>+/-8.6 versus 0.3+/-0.06 mg/d at week <em>7</em>, respectively). Bosentan and hydralazine both reduced blood pressure and cardiac hypertrophy. Mortality rate was markedly reduced by bosentan (<em>1</em>/<em>1</em>5) and partially by hydralazine (4/<em>1</em>5). However, only bosentan decreased albuminuria and renal injury. Untreated and hydralazine-treated dTGR showed increased nuclear factor (NF)-kappaB and AP-<em>1</em> expression in the kidney and heart; the p65 NF-kappaB subunit was increased in the endothelium, vascular smooth muscles cells, infiltrating cells, glomeruli, and tubules. In the heart and kidney, ET(A/B) receptor blockade inhibited NF-kappaB and AP-<em>1</em> activation compared with hydralazine treatment. Macrophage infiltration, ICAM-<em>1</em> expression, and the integrin expression on infiltrating cells were markedly reduced. Renal vasculopathy was accompanied by increased tissue factor expression on macrophages and vessels of untreated and hydralazine-treated dTGR, which was markedly reduced by bosentan. Thus, ET(A/B) receptor blockade inhibits NF-kappaB and AP-<em>1</em> activation and the NF-kappaB- and/or AP-<em>1</em>-regulated genes ICAM-<em>1</em>, VCAM-<em>1</em>, and TF, independent of blood pressure-related effects. We conclude that Ang II-induced NF-kappaB and AP-<em>1</em> activation and subsequent inflammation and coagulation involve at least in part the ET(A/B) receptors.

<em>Angiotensin</em> II (Ang II) is an important regulator of proximal tubule salt and water reabsorption. Recent studies indicate that rabbit proximal tubule <em>angiotensin</em> II receptors are the type-<em>1</em> (AT<em>1</em>R) subtype. We studied the effect of Ang II on proximal tubule receptor expression. Rabbits were treated with either <em>angiotensin</em> converting enzyme inhibitors or a low salt diet to modulate endogenous Ang II levels. In captopril-treated rabbits, liver and glomerular AT<em>1</em>R mRNA levels increased 242 +/- <em>1</em>25 and <em>1</em>4<em>1</em> +/- 60%, respectively (n = 6-<em>7</em>; P < 0.05), as determined by quantitative PCR. In contrast, proximal tubule AT<em>1</em>R mRNA levels decreased 40 +/- <em>1</em><em>1</em>% (n = 6; P < 0.05). Binding of <em>1</em>25I Ang II to renal cortical basolateral membranes of captopril-treated rabbits decreased from 2.9 +/- 0.55 to <em>1</em>.4 +/- 0.<em>1</em><em>7</em> fmol/mg protein (n = 6; P < 0.025). In rabbits fed a sodium chloride-deficient diet for 4 wk, AT<em>1</em>R mRNA levels decreased 52 +/- <em>1</em><em>1</em>% in liver and 43 +/- <em>7</em>% in glomeruli (n = 4-5; P < 0.05), whereas they increased <em>1</em>4<em>1</em> +/- 85% (n = 5; P < 0.05) in proximal tubule. In basolateral membranes from rabbits on the sodium chloride-deficient diet, specific binding of <em>1</em>25I Ang II increased from 2.<em>1</em> +/- 0.2 to 4.3 +/- <em>1</em>.<em>1</em> fmol/mg protein (n = <em>7</em>; P < 0.05). To determine whether Ang II directly regulates expression of proximal tubule AT<em>1</em> receptors, further studies were performed in cultured proximal tubule cells grown from microdissected S<em>1</em> segments of rabbit proximal tubules and immortalized by transfection with a replication-defective SV40 vector. Incubation of these cells with Ang II (<em>1</em>0(-<em>1</em><em>1</em>) to <em>1</em>0(-<em>7</em>) M) led to concentration-dependent increases in both AT<em>1</em>R mRNA levels and specific <em>1</em>25I Ang II binding. Pretreatment with pertussis toxin inhibited Ang II stimulation of AT<em>1</em>R mRNA. AT<em>1</em>R mRNA expression was decreased by either forskolin or a nonhydrolyzable cAMP analogue (dibutryl cAMP). Simultaneous Ang II administration overcame the inhibitory effect of forskolin but not dibutryl cAMP. These results indicate that proximal tubule AT<em>1</em>R expression is regulated by ambient Ang II levels, and Ang II increases AT<em>1</em>R mRNA at least in part by decreasing proximal tubule cAMP generation through a pertussis toxin-sensitive mechanism. Upregulation of proximal tubule AT<em>1</em>R by Ang II may be important in mediating enhanced proximal tubule sodium reabsorption in states of elevated systemic or intrarenal Ang II.

The messenger RNA (mRNA) distribution of 60 proteins was examined in the 3 fractions obtained by collagenase digestion (fat cells and the nonfat cells comprising the tissue remaining after collagenase digestion [matrix] and the stromovascular cells) of omental adipose tissue obtained from morbidly obese women undergoing bariatric surgery. Fat cells were enriched by at least 3-fold as compared with nonfat cells in the mRNAs for retinol binding protein 4, <em>angiotensin</em>ogen, adipsin, glutathione peroxidase 3, uncoupling protein 2, peroxisome proliferator-activated receptor gamma, cell death-inducing DFFA-like effector A, fat-specific protein 2<em>7</em>, <em>1</em><em>1</em>beta-hydroxysteroid dehydrogenase <em>1</em>, glycerol channel aquaporin <em>7</em>, NADPH:quinone oxidoreductase <em>1</em>, cyclic adenosine monophosphate phosphodiesterase 3B, glyceraldehyde-3-phosphate dehydrogenase, insulin receptor, and amyloid A<em>1</em>. Fat cells were also enriched by at least 26-fold in the mRNAs for proteins involved in lipolysis such as hormone-sensitive lipase, lipoprotein lipase, adipose tissue triglyceride lipase, and FAT/CD36. The relative distribution of mRNAs in cultured preadipocytes was also compared with that of in vitro differentiated adipocytes derived from human omental adipose tissue. Cultured preadipocytes had far lower levels of the mRNAs for inflammatory proteins than the nonfat cells of omental adipose tissue. The nonfat cells were enriched by at least 5-fold in the mRNAs for proteins involved in the inflammatory response such as tumor necrosis factor alpha, interleukin lbeta, cyclooxygenase 2, interleukin 24, interleukin 6, and monocyte chemoattractant protein <em>1</em> plus the mRNAs for osteopontin, vaspin, endothelin, <em>angiotensin</em> II receptor <em>1</em>, butyrylcholinesterase, lipocalin 2, and plasminogen activator inhibitor <em>1</em>. The cells in the adipose tissue matrix were enriched at least 3-fold as compared with the isolated stromovascular cells in the mRNAs for proteins related to the inflammatory response, as well as osteopontin and endothelial nitric oxide synthase. We conclude that the mRNAs for inflammatory proteins are primarily present in the nonfat cells of human omental adipose tissue.

The hypothesis that endothelium-derived relaxing factor (EDRF) modulates hypoxic pulmonary vasoconstriction (HPV) was tested in isolated, blood-perfused rat lungs ventilated with gas mixtures of 2<em>1</em>% O2-5% CO2-<em>7</em>4% N2 (normoxia) or of 3% O2-5% CO2-92% N2 (hypoxia); 30 microM NG-monomethyl-L-arginine (L-NMMA), an inhibitor of EDRF production, caused a reduction in the endothelium-dependent relaxant response to acetylcholine (ACh) from 62 +/- <em>7</em>, 88 +/- 4, and <em>1</em>00 +/- 4% to 26 +/- 8, 49 +/- <em>1</em>2, and <em>7</em>5 +/- <em>7</em>% at ACh concentrations of <em>1</em>, <em>1</em>0, and <em>1</em>00 microM, respectively (p less than 0.05 at all concentrations), indicating that L-NMMA acts via the inhibition of EDRF production. L-NMMA induced a concentration-related augmentation in HPV of 20 +/- 5, 32 +/- 8, and 34 +/- 8% at concentrations of 30, 300, and <em>1</em>,000 microM (p less than 0.05, compared with a vehicle control group at all concentrations). The pressor response to a dose of <em>angiotensin</em> II (A-II), which produced the same increase in pulmonary artery pressure as that induced by hypoxia, was also significantly augmented (2 +/- 0.6%), but to a lesser extent. The augmentation of HPV by 30 microM L-NMMA was completely reversed by <em>1</em> mM L-arginine (a precursor of EDRF), but not by D-arginine (an isomer of L-arginine). One and 6 mM L-arginine, but not 6 mM D-arginine caused a significant inhibition of HPV by 20 +/- 2 and 4<em>7</em> +/- <em>1</em>2% (p less than 0.05, compared with the vehicle control group) and a small but not significant reduction in A-II-mediated contraction.(ABSTRACT TRUNCATED AT 250 WORDS)

The mechanisms governing the pathological accumulation of collagen in the extracellular matrix following angioplasty are complex, but may involve interactions between endothelium-derived paracrine agents and vascular cellular components. We tested the hypothesis that nitric oxide (NO) directly decreases collagen levels and decreases endothelin (ET-<em>1</em>)-stimulated increases in levels of specific collagen subtypes in coronary vascular smooth muscle cells (VSMC). Cultured VSMC were incubated for 48 h with the NO donor CAS <em>7</em>54 (<em>1</em>0(-4) M), ET-<em>1</em> (<em>1</em>0(-8) M), or ET-<em>1</em> plus CAS <em>7</em>54. In some experiments, <em>angiotensin</em> II (Ang II; <em>1</em>0(-8) M) was utilized in place of ET-<em>1</em>. Soluble collagen types I and III were quantitated with an ELISA method, and cell counts were performed. CAS <em>7</em>54 significantly inhibited cell proliferation (-<em>1</em><em>7</em>+/-2% v control), basal total protein synthesis (-65+/-<em>7</em>% v control), and basal collagen type I levels (-39+/-6% v control), but not collagen type III levels. ET-<em>1</em> and Ang II both significantly stimulated cell proliferation (26+/-5% v control), total protein synthesis (<em>1</em>69+/-6% v control), and collagen type I levels (200+/-<em>1</em><em>1</em>% v control). Ang II, but not ET-<em>1</em>, significantly increased collagen type III levels. Co-incubations of ET-<em>1</em> and CAS <em>7</em>54 resulted in a significant decrease in cell proliferation, protein synthesis, and collagen levels (-23+/-2% v control, 90+/-5% v control, and 63+/-3% v control, respectively) compared to ET-<em>1</em> alone. In contrast, co-incubation of Ang II and CAS <em>7</em>54 had no significant effect on cell proliferation, protein synthesis, and collagen levels seen with Ang II alone. These results demonstrate that NO inhibits basal collagen levels and cell division. Additionally, NO alters ET-<em>1</em> stimulation of VSMC proliferation, protein synthesis, and production of extracellular matrix components. Thus, an imbalance in key endothelium-derived compounds could significantly impact upon extracellular matrix deposition following mechanical revascularization.

BACKGROUND

Omapatrilat, a new vasopeptidase inhibitor, inhibits the activity of <em>angiotensin</em>-converting enzyme (ACE) and neutral endopeptidase 24.<em>1</em><em>1</em> (NEP). Because these two enzymes participate in the degradation of the vasodilator and natriuretic peptide, <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)], we assessed whether omapatrilat treatment is associated with changes in the plasma and urinary excretion rates of the <em>angiotensins</em>.

METHODS

We investigated in spontaneously hypertensive rats (SHR) (0.24 kg body weight) the effect of omapatrilat on plasma and urinary concentrations of <em>angiotensin</em> (Ang) I, Ang II and Ang-(<em>1</em>-<em>7</em>) during <em>1</em><em>7</em> days of administration of either the drug (N = <em>1</em>5, <em>1</em>00 micromol/kg/day) or vehicle (N = <em>1</em>4) in the drinking water. Hemodynamic and renal excretory function studies were associated with histological examination of the expression of Ang-(<em>1</em>-<em>7</em>) in the kidneys of both vehicle and omapatrilat-treated SHRs.

RESULTS

Omapatrilat induced a sustained lowering of systolic blood pressure (-68 mm Hg) without changes in cardiac rate. The mild positive water balance produced by omapatrilat did not cause natriuresis or kaliuresis, although it was associated with a significant decrease in urine osmolality. Blood pressure normalization was accompanied by increases in plasma Ang I (2969%), Ang II (5<em>7</em>%), and Ang-(<em>1</em>-<em>7</em>) (<em>1</em>63%) levels, paralleling pronounced increases in urinary excretion rates of Ang I and Ang-(<em>1</em>-<em>7</em>) but not Ang II. Detection of Ang-(<em>1</em>-<em>7</em>) immunostaining in the kidneys of five other SHR exposed either to vehicle (N = 3) or omapatrilat (N = 2) ascertained the source of the Ang-(<em>1</em>-<em>7</em>) found in the urine. Intense Ang-(<em>1</em>-<em>7</em>) staining, more pronounced in omapatrilat-treated SHR, was found in renal proximal tubules throughout the outer and inner regions of the renal cortex and the thick ascending loop of Henle, whereas no Ang-(<em>1</em>-<em>7</em>)-positive immunostaining was found in glomeruli and distal tubules.

CONCLUSIONS

Omapatrilat antihypertensive effects caused significant activation of the renin-<em>angiotensin</em> system associated with increases in urinary excretion rates of Ang I and Ang-(<em>1</em>-<em>7</em>). Combined studies of Ang-(<em>1</em>-<em>7</em>) metabolism in urine and immunohistochemical studies in the kidney revealed the existence of an intrarenal source, which may account for the pronounced increase in the excretion rate of the vasodilator heptapeptide. These findings provide further evidence for a contribution of Ang-(<em>1</em>-<em>7</em>) to the regulation of renal function and blood pressure.

The goals of this study were to <em>1</em>) determine the acute effect of ANG-(<em>1</em>-<em>7</em>) on vascular tone in isolated middle cerebral arteries (MCAs) from Sprague-Dawley rats fed a normal salt (NS; 0.4% NaCl) diet, 2) evaluate the ability of chronic intravenous infusion of ANG-(<em>1</em>-<em>7</em>) (4 ng·kg(-<em>1</em>)·min(-<em>1</em>)) for 3 days to restore endothelium-dependent dilation to acetylcholine (ACh) in rats fed a high-salt (HS; 4% NaCl) diet, and 3) determine whether the amelioration of endothelial dysfunction by ANG-(<em>1</em>-<em>7</em>) infusion in rats fed a HS diet is different from the protective effect of low-dose ANG II infusion in salt-fed rats. MCAs from rats fed a NS diet dilated in response to exogenous ANG-(<em>1</em>-<em>7</em>) (<em>1</em>0(-<em>1</em>0)-<em>1</em>0(-5) M). Chronic ANG-(<em>1</em>-<em>7</em>) infusion significantly reduced vascular superoxide levels and restored the nitric oxide-dependent dilation to ACh (<em>1</em>0(-<em>1</em>0)-<em>1</em>0(-5) M) that was lost in MCAs of rats fed a HS diet. Acute vasodilation to ANG-(<em>1</em>-<em>7</em>) and the restoration of ACh-induced dilation by chronic ANG-(<em>1</em>-<em>7</em>) infusion in rats fed a HS diet were blocked by the Mas receptor antagonist [D-ALA(<em>7</em>)]-ANG-(<em>1</em>-<em>7</em>) or the ANG II type 2 receptor antagonist PD-<em>1</em>233<em>1</em>9 and unaffected by ANG II type <em>1</em> receptor blockade with losartan. The restoration of ACh-induced dilation in MCAs of HS-fed rats by chronic intravenous infusion of ANG II (5 ng·kg(-<em>1</em>)·min(-<em>1</em>)) was blocked by losartan and unaffected by d-ALA. These findings demonstrate that circulating ANG-(<em>1</em>-<em>7</em>), working via the Mas receptor, restores endothelium-dependent vasodilation in cerebral resistance arteries of animals fed a HS diet via mechanisms distinct from those activated by low-dose ANG II infusion.

Myogenic and <em>angiotensin</em> contractions of afferent arterioles generate reactive oxygen species. Resistance vessels express neutrophil oxidase-2 and -4. <em>Angiotensin</em> II activates p4<em>7</em>(phox)/neutrophil oxidase-2, whereas it downregulates NOX-4. Therefore, we tested the hypothesis that p4<em>7</em>(phox) enhances afferent arteriolar <em>angiotensin</em> contractions. <em>Angiotensin</em> II infusion in p4<em>7</em>(phox) +/+ but not -/- mice increased renal cortical NADPH oxidase activity (<em>7</em>±<em>1</em>-<em>1</em>2±<em>1</em> [P<0.0<em>1</em>] versus 5±<em>1</em>-<em>7</em>±<em>1</em> <em>1</em>0(3) · RLU · min(-<em>1</em>) · μg protein(-<em>1</em>) [P value not significant]), mean arterial pressure (<em>7</em><em>7</em>±2-9<em>1</em>±2 [P<0.005] versus <em>7</em>4±2-<em>7</em><em>7</em>±<em>1</em> mm Hg [P value not significant]), and renal vascular resistance (<em>7</em>.5±0.4-<em>1</em>0.<em>1</em>±0.<em>7</em> [P<0.0<em>1</em>] versus <em>7</em>.9±0.4-8.3±0.4 mm Hg/mL · min(-<em>1</em>) · gram kidney weight(-<em>1</em>) [P value not significant]). Afferent arterioles from p4<em>7</em>(phox) -/- mice had a lesser myogenic response (3.<em>1</em>±0.4 versus <em>1</em>.4±0.2 dynes · cm(-<em>1</em>) · mm Hg(-<em>1</em>); P<0.02) and a lesser (P<0.05) contraction to <em>1</em>0(-6) M <em>angiotensin</em> II (diameter change +/+: 9.3±0.2-3.4±0.6 μm versus -/-: 9.9±0.6-<em>7</em>.5±0.4 μm). <em>Angiotensin</em> and increased perfusion pressure generated significantly (P<0.05) more reactive oxygen species in p4<em>7</em>(phox) +/+ than -/- arterioles. <em>Angiotensin</em> II infusion increased the maximum responsiveness of afferent arterioles from p4<em>7</em>(phox) +/+ mice to <em>1</em>0(-6) M <em>angiotensin</em> II yet decreased the response in p4<em>7</em>(phox) -/- mice. The <em>angiotensin</em> infusion increased the sensitivity to <em>angiotensin</em> II only in p4<em>7</em>(phox) +/+ mice. We conclude that p4<em>7</em>(phox) is required to enhance renal NADPH oxidase activity and basal afferent arteriolar myogenic and <em>angiotensin</em> II contractions and to switch afferent arteriolar tachyphylaxis to sensitization to <em>angiotensin</em> during a prolonged <em>angiotensin</em> infusion. These effects likely contribute to hypertension and renal vasoconstriction during infusion of <em>angiotensin</em> II.

<em>1</em>. Plasma renin (measured in the presence of additional substrate) was significantly higher (<em>1</em>0.<em>7</em> +/- <em>1</em>.<em>1</em> S.E. of mean ng/ml.hr) in foetal lambs of <em>1</em><em>1</em><em>1</em>-<em>1</em>44 days gestation age (full term <em>1</em>4<em>7</em> days) than in their mothers (<em>1</em>.5 +/- 0.2 ng/ml.hr S.E. of mean, P < 0.00<em>1</em>) but plasma <em>angiotensin</em> II concentrations were in the same range (ewe 4<em>7</em>.3 +/- 6.6 S.E. of mean, foetus 4<em>7</em>.4 +/- <em>1</em>4.<em>1</em> S.E. of mean pg/ml.). The endogenous velocity of renin production by foetal plasma was also greater than that of maternal plasma.2. Foetal plasma [Na(+)] (<em>1</em>3<em>7</em> +/- 0.8 S.E. of mean m-equiv/l.), was lower than that in the ewe (<em>1</em>42 +/- <em>1</em>.5 m-equiv/l. S.E. of mean, P < 0.0<em>1</em>).3. Foetal plasma renin in lambs of less than <em>1</em>20 days gestation was lower (9.2 +/- 2.<em>7</em> S.E. of mean ng/ml.hr) than that in lambs of over <em>1</em>30 days gestation (<em>1</em>2.6 +/- 2.6 ng/ml.hr S.E. of mean, P < 0.0<em>1</em>). Foetal plasma [K(+)] (3.8 +/- 0.<em>1</em> S.E. of mean m-equiv/l.) was also lower in lambs of less than <em>1</em>20 days gestation than in those over <em>1</em>30 days (4.<em>1</em> +/- 0.<em>1</em> S.E. of mean m-equiv/l., P < 0.00<em>1</em>).4. When small volumes of blood (</= 3% of blood volume) were withdrawn from foetal lambs, plasma renin increased. The% increase of plasma renin in hypoxaemic foetal lambs was significantly less (P < 0.05) than in control lambs. At the end of 60 min hypoxaemia, arterial pressure and plasma [K(+)] were significantly higher in hypoxaemic than in control foetal lambs.5. During foetal hypoxaemia, plasma <em>angiotensin</em> II concentration increased concurrently with plasma renin.6. Bilateral nephrectomy was performed in two foetal lambs. Plasma renin fell to very low levels and <em>angiotensin</em> II became undetectable.<em>7</em>. Adrenaline ( approximately .0.42 mug/min.kg I.V.) infused into the foetus did not alter foetal plasma renin. When adrenaline was infused into the ewe ( approximately 0.26 mug/min.kg) maternal plasma renin increased. Maternal infusion of adrenaline raised foetal plasma renin significantly more (P < 0.05) than foetal infusion.8. It is concluded that the foetal kidney is the major source of foetal renin in the last quarter of gestation and that renin release is stimulated by very small reductions of blood volume. Hypoxaemia does not augment renin release and cannot be responsible for high levels of renin and <em>angiotensin</em> associated with vaginal delivery.

The major mechanism of agonist-induced internalization of G protein-coupled receptors (GPCRs) is beta-arrestin- and dynamin-dependent endocytosis via clathrin-coated vesicles. However, recent reports have suggested that some GPCRs, exemplified by the AT<em>1</em> <em>angiotensin</em> receptor expressed in human embryonic kidney (HEK) 293 cells, are internalized by a beta-arrestin- and dynamin-independent mechanism, and possibly via a clathrin-independent pathway. In this study, agonist-induced endocytosis of the rat AT<em>1</em>A receptor expressed in Chinese hamster ovary (CHO) cells was abolished by clathrin depletion during treatment with hyperosmotic sucrose and was unaffected by inhibition of endocytosis via caveolae with filipin. In addition, internalized fluorescein-conjugated <em>angiotensin</em> II appeared in endosomes, as demonstrated by colocalization with transferrin. Overexpression of beta-arrestin<em>1</em>(V53D) and beta-arrestin<em>1</em>(<em>1</em>-349) exerted dominant negative inhibitory effects on the endocytosis of radioiodinated <em>angiotensin</em> II in CHO cells. GTPase-deficient (K44A) mutant forms of dynamin-<em>1</em> and dynamin-2, and a pleckstrin homology domain-mutant (K535A) dynamin-2 with impaired phosphoinositide binding, also inhibited the endocytosis of AT(<em>1</em>) receptors in CHO cells. Similar results were obtained in COS-<em>7</em> and HEK 293 cells. Confocal microscopy using fluorescein-conjugated <em>angiotensin</em> II showed that overexpression of dynamin-<em>1</em>(K44A) and dynamin-2(K44A) isoforms likewise inhibited agonist-induced AT<em>1</em> receptor endocytosis in CHO cells. Studies on the <em>angiotensin</em> II concentration-dependence of AT<em>1</em> receptor endocytosis showed that at higher agonist concentrations its rate constant was reduced and the inhibitory effects of dominant negative dynamin constructs were abolished. These data demonstrate the importance of beta-arrestin- and dynamin-dependent endocytosis of the AT<em>1</em> receptor via clathrin-coated vesicles at physiological <em>angiotensin</em> II concentrations.

Human heart tissue enzymes cleave <em>angiotensin</em> (Ang) I to release Ang <em>1</em>-9, Ang II, or Ang <em>1</em>-<em>7</em>. In atrial homogenate preparations, cathepsin A (deamidase) is responsible for 65% of the liberated Ang <em>1</em>-9. Ang <em>1</em>-<em>7</em> was released (88% to <em>1</em>00%) by a metallopeptidase, as established with peptidase inhibitors. Ang II was liberated to about equal degrees by ACE and chymase-type enzymes. Cathepsin A's presence in heart tissue was also proven because it deamidated enkephalinamide substrate by immunoprecipitation of cathepsin A with antiserum to human recombinant enzyme and by immunohistochemistry. In immunohistochemistry, cathepsin A was detected in myocytes of atrial tissue. The products of Ang I cleavage, Ang <em>1</em>-9 and Ang <em>1</em>-<em>7</em>, potentiated the effect of an ACE-resistant bradykinin analog and enhanced kinin effect on the B(2) receptor in Chinese hamster ovary cells transfected to express human ACE and B(2) (CHO/AB), and in human pulmonary arterial endothelial cells. Ang <em>1</em>-9 and <em>1</em>-<em>7</em> augmented arachidonic acid and nitric oxide (NO) release by kinin. Direct assay of NO liberation by bradykinin from endothelial cells was potentiated at <em>1</em>0 nmol/L concentration, 2.4-fold (Ang <em>1</em>-9) and 2.<em>1</em>-fold (Ang <em>1</em>-<em>7</em>); in higher concentrations, Ang <em>1</em>-9 was significantly more active than Ang <em>1</em>-<em>7</em>. Both peptides had traces of activity in the absence of bradykinin. Ang <em>1</em>-9 and Ang <em>1</em>-<em>7</em> potentiated bradykinin action on the B(2) receptor by raising arachidonic acid and NO release at much lower concentrations than their 50% inhibition concentrations (IC(50)s) with ACE. They probably induce conformational changes in the ACE/B(2) receptor complex via interaction with ACE.

Bradykinin B2 receptor-deleted mice (Bdkrb2(-/-)) have delayed carotid artery thrombosis times and prolonged tail bleeding time resulting from elevated <em>angiotensin</em> II (AngII) and <em>angiotensin</em> receptor 2 (AT2R) producing increased plasma nitric oxide (NO) and prostacyclin. Bdkrb2(-/-) also have elevated plasma <em>angiotensin</em>-(<em>1</em>-<em>7</em>) and messenger RNA and protein for its receptor Mas. Blockade of Mas with its antagonist A-<em>7</em><em>7</em>9 in Bdkrb2(-/-) shortens thrombosis times (58 ± 4 minutes to 38 ± 4 minutes) and bleeding times (<em>1</em><em>7</em>0 ± <em>1</em>3 seconds to 88 ± 8 seconds) and lowers plasma nitrate (22 ± 4 μM to <em>1</em>5 ± 5 μM), and 6-keto-PGF<em>1</em>α (259 ± <em>1</em>03 pg/mL to <em>1</em>32 ± 58 pg/mL). Bdkrb2(-/-) platelets express increased NO, guanosine 3',5'-cyclic monophosphate, and cyclic adenosine monophosphate with reduced spreading on collagen, collagen peptide GFOGER, or fibrinogen. In vivo A-<em>7</em><em>7</em>9 or combined L-NAME and nimesulide treatment corrects it. Bdkrb2(-/-) platelets have reduced collagen-related peptide-induced integrin α2bβ3 activation and P-selectin expression that are partially corrected by in vivo A-<em>7</em><em>7</em>9, nimesulide, or L-NAME. Bone marrow transplantations show that the platelet phenotype and thrombosis time depends on the host rather than donor bone marrow progenitors. Transplantation of wild-type bone marrow into Bdkrb2(-/-) hosts produces platelets with a spreading defect and delayed thrombosis times. In Bdkrb2(-/-), combined AT2R and Mas overexpression produce elevated plasma prostacyclin and NO leading to acquired platelet function defects and thrombosis delay.

Advanced congestive heart failure (CHF) and chronic kidney disease (CKD) are characterized by increased <em>angiotensin</em> II (Ang II) levels and are often accompanied by significant skeletal muscle wasting that negatively impacts mortality and morbidity. Both CHF and CKD patients have respiratory muscle dysfunction, however the potential effects of Ang II on respiratory muscles are unknown. We investigated the effects of Ang II on diaphragm muscle in FVB mice. Ang II induced significant diaphragm muscle wasting (<em>1</em>8.<em>7</em>±<em>1</em>.6% decrease in weight at one week) and reduction in fiber cross-sectional area. Expression of the E3 ubiquitin ligases atrogin-<em>1</em> and muscle ring finger-<em>1</em> (MuRF-<em>1</em>) and of the pro-apoptotic factor BAX was increased after 24 h of Ang II infusion (4.4±0.3 fold, 3.<em>1</em>±0.5 fold and <em>1</em>.6±0.2 fold, respectively, compared to sham infused control) suggesting increased muscle protein degradation and apoptosis. In Ang II infused animals, there was significant regeneration of injured diaphragm muscles at <em>7</em> days as indicated by an increase in the number of myofibers with centralized nuclei and high expression of embryonic myosin heavy chain (E-MyHC, <em>1</em><em>1</em>.2±3.3 fold increase) and of the satellite cell marker M-cadherin (59.2±22.2% increase). Furthermore, there was an increase in expression of insulin-like growth factor-<em>1</em> (IGF-<em>1</em>, <em>1</em>.8±0.3 fold increase) in Ang II infused diaphragm, suggesting the involvement of IGF-<em>1</em> in diaphragm muscle regeneration. Bone-marrow transplantation experiments indicated that although there was recruitment of bone-marrow derived cells to the injured diaphragm in Ang II infused mice (26<em>7</em>.0±<em>7</em>4.6% increase), those cells did not express markers of muscle stem cells or regenerating myofibers. In conclusion, Ang II causes marked diaphragm muscle wasting, which may be important for the pathophysiology of respiratory muscle dysfunction and cachexia in conditions such as CHF and CKD.

Hemodynamic factors, circulating hormones, paracrine factors, and intracrine factors influence vascular smooth muscle growth and plasticity. The well-characterized role of <em>angiotensin</em> II in the modulation of vascular tone and cell function may be critically involved in the mechanisms by which vascular smooth muscle responds to signals associated with vascular endothelial dysfunction and increases in oxidative stress. Studies from this laboratory suggest that the trophic actions of <em>angiotensin</em> II may be intrinsically regulated by <em>angiotensin</em>-(<em>1</em>-<em>7</em>), a separate product of the <em>angiotensin</em> system derived from the common substrate, <em>angiotensin</em> I. Exposure of cultured vascular smooth muscle cells to <em>angiotensin</em>-(<em>1</em>-<em>7</em>) inhibited the trophic actions of <em>angiotensin</em> II and reduced the expression of the mitogenic effects of both normal serum and platelet-derived growth factor. The growth-inhibitory actions of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) were blocked by the selective D-alanine(<em>7</em>)-<em>angiotensin</em>-(<em>1</em>-<em>7</em>) antagonist and the nonselective <em>angiotensin</em> receptor blocker sarcosine(<em>1</em>)-threonine(8)-<em>angiotensin</em> II. In contrast, subtype-selective antagonists for the AT(<em>1</em>) and AT(2) receptors had no effect on the inhibitory actions of <em>angiotensin</em>-(<em>1</em>-<em>7</em>), a finding that is consistent with the pharmacological characterization of a high-affinity (<em>1</em>25)I-labeled <em>angiotensin</em>-(<em>1</em>-<em>7</em>) binding site in the vasculature by use of selective and nonselective <em>angiotensin</em> II receptor antagonists. The relevance of these findings to the proliferative response of vascular smooth muscle cells after endothelial injury was confirmed by assessment of the effect of a <em>1</em>2-day infusion of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) on neointimal formation. In these experiments, the proliferative response produced by injuring the carotid artery was inhibited by <em>angiotensin</em>-(<em>1</em>-<em>7</em>) through a mechanism that could not be explained by changes in arterial pressure. Because plasma <em>angiotensin</em>-(<em>1</em>-<em>7</em>) increased to levels comparable to those found in animals and human subjects given therapeutic doses of <em>angiotensin</em>-converting enzyme inhibitors, <em>angiotensin</em>-(<em>1</em>-<em>7</em>) may be one factor participating in the reversal of vascular proliferation during inhibition of <em>angiotensin</em> II formation or activity.

OBJECTIVE

To investigate the anti-inflammatory and anti-angiogenic effects of telmisartan, an <em>angiotensin</em> II type <em>1</em> receptor (AT<em>1</em>-R) antagonist, on ischemia-induced retinal neovascularization.

METHODS

C57BL/6 neonatal mice were reared in an 80% concentration of oxygen from postnatal day (P)7 to P<em>1</em>2, followed by room-air breathing until P<em>1</em>7, to induce ischemia-initiated retinal neovascularization (i.e., a murine model of ischemic retinopathy). Tissue localization of AT<em>1</em>-R was examined by immunohistochemistry for murine retinal wholemounts and human fibrovascular tissues excised at vitrectomy for proliferative diabetic retinopathy. Animals received intraperitoneal injection of telmisartan or vehicle. A concanavalin A lectin perfusion-labeling technique was used to evaluate the areas of physiological and pathologic retinal new vessels and the number of leukocytes adhering to the vasculature. Retinal mRNA and protein levels of intercellular adhesion molecule (ICAM)-<em>1</em>, vascular endothelial growth factor receptor (VEGFR)-<em>1</em>, and VEGFR-2 were examined by RT-PCR and ELISA.

RESULTS

Vessels in human fibrovascular tissues and the murine retinas were positive for AT<em>1</em>-R. Pathologic (P < 0.0<em>1</em>), but not physiologic (P>> 0.05), retinal neovascularization was significantly suppressed in telmisartan-treated mice compared with vehicle-treated animals. The number of adherent leukocytes (P < 0.0<em>1</em>) was also significantly reduced, together with retinal ICAM-<em>1</em> levels (P < 0.0<em>1</em>) in the telmisartan-treated group compared with the control group. No significant difference was detected in retinal VEGFR-2 levels between the two groups, whereas retinal VEGFR-<em>1</em> levels in the telmisartan-treated group were significantly (P < 0.05) lower than in the vehicle-treated group.

CONCLUSIONS

The present findings suggest that the AT<em>1</em>-R signaling blockade leads to the selective suppression of pathologic, but not physiological, retinal neovascularization through the inhibition of the inflammatory processes related to pathologic neovascularization.

Treatment with cyclosporine A (CysA), a potent immunosuppressive agent, is associated with systemic and renal vasoconstriction, leading to hypertension. The present study was conducted to elucidate the contribution of <em>angiotensin</em> II (Ang II) to CysA-induced hypertension and reactive oxygen species (ROS) generation. CysA (30 mg/kg per day SC), given for 3 weeks in rats, increased systolic blood pressure (SBP) from <em>1</em><em>1</em>9+/-2 to <em>1</em>45+/-3 mm Hg (n=<em>7</em>). Plasma and kidney Ang II levels were significantly higher in CysA-treated rats (<em>1</em>36+/-<em>1</em>0 fmol/mL and 5<em>1</em>6+/-<em>7</em>0 fmol/g) than in vehicle-treated (<em>1</em> mL olive oil) rats (<em>7</em>6+/-<em>1</em>0 fmol/mL and 222+/-2<em>1</em> fmol/g, n=<em>7</em>). CysA treatment increased AT<em>1</em> receptor protein expression in the aorta (by 25<em>1</em>+/-35%), whereas it was reduced in the kidney (by -32+/-4%). Superoxide anion production in aortic segments and kidney thiobarbituric acid-reactive substance (TBARS) contents were higher in CysA-treated rats (26+/-2 counts/min per milligram and 3<em>7</em>+/-3 nmol/g) than in vehicle-treated rats (<em>1</em><em>7</em>+/-<em>1</em> counts/min per milligram and 24+/-3 nmol/g). Concurrent administration of an AT<em>1</em> receptor antagonist, valsartan (30 mg/kg per day, in drinking water), to CysA-treated rats (n=<em>7</em>) significantly decreased SBP (<em>1</em><em>1</em>3+/-4 mm Hg) and prevented increases in vascular superoxide (<em>1</em>6+/-2 counts/min per milligram) and kidney TBARS contents (2<em>1</em>+/-3 nmol/g). Similarly, treatment with a superoxide dismutase mimetic, 4-hydroxy-2,2,6,6,-tetramethylpiperidine-N-oxyl (Tempol; 3 mmol/L in drinking water, n=<em>7</em>), prevented CysA-induced increases in SBP (<em>1</em><em>1</em>5+/-3 mm Hg), vascular superoxide (<em>1</em>6+/-<em>1</em> counts/min per milligram), and kidney TBARS contents (<em>1</em>9+/-2 nmol/g). These data suggest that ROS generation induced by augmented Ang II levels contributes to the development of CysA-induced hypertension.

<AbstractText>The morbidity and mortality of patients with functional mitral regurgitation (MR) remain high, but no pharmacological therapy has been proven effective. The hypothesis of this study was that sacubitril/valsartan would be superior to valsartan alone in improving functional MR via dual inhibition of the renin-<em>angiotensin</em> system and neprilysin.</AbstractText><AbstractText>In this double-blind trial, we randomly assigned <em>1</em><em>1</em>8 patients with heart failure with chronic functional MR secondary to left ventricular (LV) dysfunction to receive either sacubitril/valsartan or valsartan, in addition to standard medical therapy for heart failure. The primary end point was the change in effective regurgitant orifice area of functional MR from baseline to the <em>1</em>2-month follow-up. Secondary end points included changes in regurgitant volume, LV end-systolic volume, LV end-diastolic volume, and incomplete mitral leaflet closure area.</AbstractText><p><div><b>RESULTS</b></div>The decrease in effective regurgitant orifice area was significantly greater in the sacubitril/valsartan group than in the valsartan group (-0.058±0.095 versus -0.0<em>1</em>8±0.<em>1</em>05 cm²; P=0.032) in an intention-to-treat analysis including <em>1</em><em>1</em><em>7</em> (99%) patients. Regurgitant volume was also significantly decreased in the sacubitril/valsartan group in comparison with the valsartan group (mean difference, -<em>7</em>.3 mL; 95% CI, -<em>1</em>2.6 to -<em>1</em>.9; P=0.009). There were no significant between-group differences regarding the changes in incomplete mitral leaflet closure area and LV volumes, with the exception of LV end-diastolic volume index ( P=0.044). We noted no significant difference in the change of blood pressure between the treatment groups, and <em>7</em> patients (<em>1</em>2%) in the sacubitril/valsartan group and 9 (<em>1</em>6%) in the valsartan group had ≥<em>1</em> serious adverse events ( P=0.54).</p><AbstractText>Among patients with secondary functional MR, sacubitril/valsartan reduced MR to a greater extent than did valsartan. Our findings suggest that an <em>angiotensin</em> receptor-neprilysin inhibitor might be considered for optimal medical therapy of patients with heart failure and functional MR.</AbstractText><AbstractText>URL: https://www.clinicaltrials.gov . Unique identifier: NCT0268<em>7</em>932.</AbstractText>

<em>Angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] has beneficial effects against hypertension-induced damage in heart and kidney, but its effects in brain are not clear as yet. The present study aimed to investigate the protective effects of Ang-(<em>1</em>-<em>7</em>) on the physiopathologic changes caused by hypertension in brain of spontaneously hypertensive rats (SHRs). Wistar-Kyoto rats received intracerebroventricular (I.C.V.) infusion of artificial cerebrospinal fluid (aCSF) while SHRs received I.C.V. infusion of Ang-(<em>1</em>-<em>7</em>), Mas receptor antagonist A-<em>7</em><em>7</em>9 and aCSF for 4 weeks. Brain tissues were collected and analyzed by western blot, enzyme immunoassay, spectrophotometric assays and terminal deoxynucleotidyl transferase-mediated dUTP end-labeling (TUNEL) staining. Our study showed that infusion of Ang-(<em>1</em>-<em>7</em>) for 4 weeks significantly reduced the expression of <em>Angiotensin</em> II and <em>Angiotensin</em> II type <em>1</em> receptors in SHR brain. Additionally, it decreased the levels of malondialdehyde and elevated total superoxide dismutase activity, which was accompanied by reductions of NADPH oxidase subunit gp9<em>1</em>(phox) and inducible nitric oxide synthase in the brain of SHR. The increases of the percentage of TUNEL-positive neurons and Bax to Bcl-2 ratio in SHR brain were also attenuated by Ang-(<em>1</em>-<em>7</em>). The anti-oxidative and anti-apoptosis effects of Ang-(<em>1</em>-<em>7</em>) are independent of blood pressure reduction and can be partially abolished by A-<em>7</em><em>7</em>9. These findings suggest that chronic treatment with Ang-(<em>1</em>-<em>7</em>) is beneficial to attenuate hypertension-induced physiopathologic changes in brain and may be helpful to prevent hypertension-related cerebrovascular diseases.

The renin-<em>angiotensin</em> system not only plays a critical role in blood pressure control but is also involved in learning and memory mechanisms. In addition to <em>angiotensin</em> (Ang) II, Ang-(<em>1</em>-<em>7</em>) may also have important biological activities in the brain. Here, we show for the first time that Ang-(<em>1</em>-<em>7</em>) enhances long-term potentiation (LTP) in the CA<em>1</em> region of the hippocampus. Our studies with AT<em>1</em> receptor antagonists and selective Ang-(<em>1</em>-<em>7</em>) receptor antagonists demonstrate the existence of a distinct Ang-(<em>1</em>-<em>7</em>) receptor in the brain, the G-protein-coupled receptor Mas, encoded by the Mas protooncogene. We also show that the genetic deletion of this receptor abolishes the Ang-(<em>1</em>-<em>7</em>)-induced enhancement of LTP. Thus, we firstly demonstrate that Ang-(<em>1</em>-<em>7</em>) influences the induction of LTP in limbic structures implicating its distinct function in learning and memory mechanisms; secondly, we have identified Mas as a functional receptor for Ang-(<em>1</em>-<em>7</em>) in the brain.

In the present study, we investigated the potentiating effect of <em>angiotensin</em>-(<em>1</em>-<em>7</em>) [Ang-(<em>1</em>-<em>7</em>)] on bradykinin (BK)-induced vasodilation in the mesenteric vascular bed of anesthetized spontaneously hypertensive rats using intravital microscopy. Topical application of BK and Ang-(<em>1</em>-<em>7</em>) induced vasodilation in mesenteric arterioles. The BK-induced effect, but not acetylcholine, sodium nitroprusside, or histamine responses, was potentiated in the presence of Ang-(<em>1</em>-<em>7</em>). This interaction was abolished by BK-B(2) and Ang-(<em>1</em>-<em>7</em>) antagonists (HOE <em>1</em>40 and A-<em>7</em><em>7</em>9, respectively), a K(+) channel blocker (tetraethylammonium), and cyclooxygenase inhibitors (indomethacin and diclofenac); however, nitric oxide synthase inhibition (Nomega-nitro-L-arginine methyl ester) did not modify the Ang-(<em>1</em>-<em>7</em>)-potentiating activity. Long-term <em>angiotensin</em>-converting enzyme (ACE) inhibition increased BK and Ang-(<em>1</em>-<em>7</em>)-induced vasodilation. The BK potentiation by Ang-(<em>1</em>-<em>7</em>) was preserved after ACE inhibition, Ang II type <em>1</em> receptor blockade, or the combination of both treatments. The most striking finding of this study was the unexpected observation that the potentiation of BK vasodilation in spontaneously hypertensive rats treated short- or long-term with ACE inhibitors was reverted by the Ang-(<em>1</em>-<em>7</em>) antagonist A-<em>7</em><em>7</em>9. Our results unmasked a key role for an Ang-(<em>1</em>-<em>7</em>)-related mechanism in mediating BK potentiation by ACE inhibitors.

Sleep apnea is characterized by increased sympathetic activity and is associated with systemic hypertension. <em>Angiotensin</em> (Ang) peptides have previously been shown to participate in the regulation of sympathetic tone and arterial pressure in the hypothalamic paraventricular nucleus (PVN) neurons. We investigated the role of endogenous Ang peptides within the PVN to control blood pressure in a rat model of sleep apnea-induced hypertension. Male Sprague-Dawley rats (250 g), instrumented with bilateral guide cannulae targeting the PVN, received chronic infusion of Ang antagonists (A-<em>7</em><em>7</em>9, Ang-(<em>1</em>-<em>7</em>) antagonist; losartan and ZD<em>7</em><em>1</em>55, AT(<em>1</em>) antagonists; PD<em>1</em>233<em>1</em>9, AT(2) receptor antagonist, or saline vehicle). A separate group received an infusion of the GABA(A) receptor agonist (muscimol) to inhibit PVN neuronal activity independent of <em>angiotensin</em> receptors. After cannula placement, rats were exposed during their sleep period to eucapnic intermittent hypoxia (IH; nadir 5% O(2); 5% CO(2) to peak 2<em>1</em>% O(2); 0% CO(2)) 20 cycles/h, <em>7</em> h/day, for <em>1</em>4 days while mean arterial pressure (MAP) was measured by telemetry. In rats receiving saline, IH exposure significantly increased MAP (+<em>1</em>2±2 mm Hg vs. Sham -2±<em>1</em> mm Hg P<0.0<em>1</em>). Inhibition of PVN neurons with muscimol reversed the increase in MAP in IH rats (MUS: -9±4 mm Hg vs. vehicle +<em>1</em>2±2 mm Hg; P<0.0<em>1</em>). Infusion of any of the Ang antagonists also prevented the rise in MAP induced by IH (A-<em>7</em><em>7</em>9: -5±<em>1</em> mm Hg, losartan: -9±4 mm Hg, ZD<em>7</em><em>1</em>55: -<em>1</em><em>1</em>±4 mm Hg and PD<em>1</em>233<em>1</em>9: -4±3 mm Hg; P<0.0<em>1</em>). Our results suggest that endogenous Ang peptides acting in the PVN contribute to IH-induced increases in MAP observed in this rat model of sleep apnea-induced hypertension.

We investigated the effects of dual renin-<em>angiotensin</em> system (RAS) blockade on <em>angiotensin</em>-converting enzyme-2 (Ace2) expression, hypertension, and renal proximal tubular cell (RPTC) apoptosis in type <em>1</em> diabetic Akita <em>angiotensin</em>ogen (Agt)-transgenic (Tg) mice that specifically overexpress Agt in their RPTCs. Adult (<em>1</em><em>1</em> wk old) male Akita and Akita Agt-Tg mice were treated with two RAS blockers (ANG II receptor type <em>1</em> blocker losartan, 30 mg·kg(-<em>1</em>)·day(-<em>1</em>)) and <em>angiotensin</em>-converting enzyme (ACE) inhibitor perindopril (4 mg·kg(-<em>1</em>)·day(-<em>1</em>)) in drinking water. Same-age non-Akita littermates and Agt-Tg mice served as controls. Blood pressure, blood glucose, and albuminuria were monitored weekly. The animals were euthanized at age <em>1</em>6 wk. The left kidneys were processed for immunohistochemistry and apoptosis studies. Renal proximal tubules were isolated from the right kidneys to assess gene and protein expression. Urinary ANG II and ANG <em>1</em>-<em>7</em> were quantified by ELISA. RAS blockade normalized renal Ace2 expression and urinary ANG <em>1</em>-<em>7</em> levels (both of which were low in untreated Akita and Akita Agt-Tg), prevented hypertension, albuminuria, tubulointerstitial fibrosis and tubular apoptosis, and inhibited profibrotic and proapoptotic gene expression in RPTCs of Akita and Akita Agt-Tg mice compared with non-Akita controls. Our results demonstrate the effectiveness of RAS blockade in preventing intrarenal RAS activation, hypertension, and nephropathy progression in diabetes and support the important role of intrarenal Ace2 expression in modulating hypertension and renal injury in diabetes.