Mas Receptor Antagonist Research Articles

Mas receptor blockade impairs exercise-induced cardiac hypertrophy

Exercise training leads to physiological cardiac hypertrophy and the protective axis of the renin-angiotensin system composed of angiotensin-converting enzyme 2, angiotensin-(1–7), and Mas receptor seems involved in this process. However, the role of the basal activity of the Mas receptor in exercise-induced physiological cardiac hypertrophy is still unclear. We evaluated the effects of the Mas receptor blockade on the left ventricular structure and function of rats submitted to running training. Rats were assigned to 4 groups: sedentary (S), sedentary + A-779 (Mas receptor antagonist, 120 µg/kg/day, i.p.; SA), trained (60-minute treadmill running sessions, five days a week, 8 weeks; T), and trained + A-779 (TA). Systolic blood pressure was higher in sedentary and trained rats treated with A-779 at the end of the experimental period. The A-779 treatment prevented the left ventricular hypertrophy evoked by physical exercise and increased collagen deposition in sedentary and trained rats. Cardiomyocytes from the SA group presented increased length and thickness of the sarcomeres, elongated mitochondria, glycogen deposits, and enlarged cisterns of the sarcoplasmic reticulum. TA group presented a reduced sarcomere thickness and cytoplasm with a degenerative aspect. These findings show that the basal activity of the Mas receptor is essential for the proper turnover of the extracellular matrix in the myocardium and the maintenance of the sarcomeric structure of cardiomyocytes.

Ang-(1-7) attenuates podocyte injury induced by high glucose in vitro.

The incidence of diabetic nephropathy (DN) is gradually increasing worldwide. Podocyte injury, such as podocyte apoptosis and loss of the slit diaphragm (SD)-specific markers are early pathogenic features of DN. The cultured mouse podocytes were separated into a high glucose-treated (HG, 30mM) group to mimic DN in vitro, a low glucose-treated (LG, 5mM) group as a control and HG+ angiotensin-(1-7)(Ang-(1-7)) and HG+Ang-(1-7) + D-Ala7-Ang-(1-7) (A779, Ang-(1-7)/Mas receptor antagonist) experimental groups. The Cell Counting Kit-8 (CCK-8) method and flow cytometry was used to detect podocyte activity and podocyte apoptosis respectively. The expression of angiotensin type 1 receptor (AT1R), Mas receptor (MasR) and podocyte-specific markers were examined by q-PCR and Western blot, respectively. The results showed that the decrease in podocyte activity; the increase in podocyte apoptosis; the decreased mRNA and protein expression of nephrin, podocin, WT-1 and MasR; and the upregulated expression of AT1R induced by HG could be reversed by Ang-(1-7). However, these effects were blocked by A779. The possible mechanisms of the Ang-(1-7)-mediated effect depended on MasR. In addition, the protective effect of Ang-(1-7) on podocyte activity was dose-dependent and most obvious at 10 µM. A779 had the greatest antagonistic action against Ang-(1-7) at a concentration of 10 μM. This study reveals that binding of Ang-(1-7) to its specific receptor MasR may counteract the effects of Ang II mediated by AT1R to significantly attenuate podocyte injury induced by high glucose. Ang-(1-7)/MasR targeting in podocytes may be a therapeutic approach to attenuate renal injury in DN.

Angiotensin-(1−7) attenuates the negative inotropic response to acetylcholine in the heart

Previous studies have suggested that the Angiotensin-(1−7) [(Ang-(1−7)] can change cardiac function by modulating the autonomic nervous system. However, it is unknown whether the Ang-(1−7) can modulate the effect of acetylcholine (ACh) in ventricular contractility. Thus, this study aimed to investigate whether Ang-(1−7) modifies the amplitude of the cardiac cholinergic effects and if these effects are intrinsic to the heart. In anesthetized Wistar rats, Ang-(1−7) attenuated the effect of ACh in decreasing the left ventricular end-systolic pressure (LVESP), dP/dtmax, and dP/dtmin, but did not modify the hypotensive effect of ACh. Similarly, Ang-(1−7) attenuated the reduction of the LVESP, dP/dtmax, and dP/dtmin evoked by ACh in isolated hearts. These effects were blocked by the Mas receptor antagonist, A-779, but not by the adenylyl cyclase inhibitor MDL-12,330 A. Ang-(1−7) also attenuated the reduction in the maximum contraction and relaxation speeds and the shortening promoted by ACh in isolated cardiomyocytes. These data show that Ang-(1−7) acting through Mas receptor counter-regulates the myocardial contractile response to ACh in an arterial pressure and heart rate-independent manner.

Angiotensin (1‐7) exerts antioxidant effects, inhibits mTOR signaling and protects against endoplasmic reticulum stress and apoptosis in HK2 cells

Angiotensin (Ang) 1‐7, a bioactive heptapeptide of the renin‐angiotensin aldosterone system is known to exert antioxidant and nephroprotective effects. However, the cellular mechanisms involved in the beneficial effect of Ang 1‐7 is not entirely understood. Here, we studied the effect of Ang 1‐7 on H2O2‐mediated oxidative damage, endoplasmic reticulum (ER) stress and apoptosis in the human proximal tubular (HK2) cells and the specific mechanisms. HK2 cells were incubated with H2O2 (500 µM) and pre‐treated without and with Ang 1‐7 (100 nM). We measured reactive oxygen species (ROS) generation, ER stress, apoptosis and mTOR signaling in these cells. H2O2 treatment resulted in an increased oxidative stress in HK2 cells. Increased in the expression ER stress marker, binding immunoglobulin protein (BiP) and C/EBP homologous protein (CHOP), and cellular apoptosis was also observed after H2O2treatment. Moreover, H2O2 treatment resulted in the activation of mTOR complexes (mTORC1 and mTORC2) in these cells. Ang 1‐7 pre‐treatment to H2O2incubated cells significantly attenuated ROS generation, ER stress and apoptosis. These effects of Ang 1‐7 were blocked by co‐treatment with A779 (Mas receptor antagonist). Additionally, prior treatment of Ang 1‐7 inhibited the H2O2mediated mTOR activation. However, A779 treatment and transfection of HK2 cells with Mas receptor siRNA reversed the inhibitory effect of Ang 1‐7 on mTOR activities. In conclusion, our study suggests that Ang 1‐7 acting via Mas receptor could protect HK2 cells from H2O2‐induced oxidative damage through inhibition of oxidative stress, ER stress and ultimately cell apoptosis by suppressing mTOR activation.

Lateral Preoptic Area Neurons Activated by Angiotensin-(1-7) Increase Intravesical Pressure: A Novel Feature in Central Micturition Control.

Central micturition control and urine storage involve a multisynaptic neuronal circuit for the efferent control of the urinary bladder. Electrical stimulation of the lateral preoptic area (LPA) at the level of the decussation of the anterior commissure in cats evokes relaxation of the bladder, whereas ventral stimulation of LPA evokes vigorous contraction. Endogenous Angiotensin-(1–7) [(Ang-(1–7)] synthesis depends on ACE-2, and its actions on binding to Mas receptors, which were found in LPA neurons. We aimed to investigate the Ang-(1–7) actions into the LPA on intravesical pressure (IP) and cardiovascular parameters. The gene and protein expressions of Mas receptors and ACE-2 were also evaluated in the LPA. Angiotensin-(1–7) (5 nmol/μL) or A-779 (Mas receptor antagonist, 50 nmol/μL) was injected into the LPA in anesthetized female Wistar rats; and the IP, mean arterial pressure (MAP), heart rate (HR), and renal conductance (RC) were recorded for 30 min. Unilateral injection of Ang-(1–7) into the LPA increased IP (187.46 ± 37.23%) with peak response at ∼23–25-min post-injection and yielded no changes in MAP, HR, and RC. Unilateral or bilateral injections of A-779 into the LPA decreased IP (−15.88 ± 2.76 and −27.30 ± 3.40%, respectively) and elicited no changes in MAP, HR, and RC. The genes and the protein expression of Mas receptors and ACE-2 were found in the LPA. Therefore, the LPA is an important part of the circuit involved in the urinary bladder control, in which the Ang-(1–7) synthetized into the LPA activates Mas receptors for increasing the IP independent on changes in RC and cardiovascular parameters.

Angiotensin-(1-7)/Mas receptor modulates anti-inflammatory effects of exercise training in a model of chronic allergic lung inflammation

AimsExercise training increases circulating and tissue levels of angiotensin-(1-7) [Ang-(1-7)], which was shown to attenuate inflammation and fibrosis in different diseases. Here, we evaluated whether Ang-(1-7)/Mas receptor is involved in the beneficial effects of aerobic training in a chronic model of asthma. Material and methodsBALB/c mice were subjected to a protocol of asthma induced by ovalbumin sensitization (OVA; 4 i.p. injections) and OVA challenge (3 times/week for 4 weeks). Simultaneously to the challenge period, part of the animals was continuously treated with Mas receptor antagonist (A779, 1 μg/h; for 28 days) and trained in a treadmill (TRE; 60% of the maximal capacity, 1 h/day, 5 days/week during 4 weeks). PGC1-α mRNA expression (qRT-PCR), plasma IgE and lung cytokines (ELISA), inflammatory cells infiltration (enzymatic activity assay) and airway remodeling (by histology) were evaluated. Key findingsBlocking the Mas receptor with A779 increased IgE and IL-13 levels and prevented the reduction in extracellular matrix deposition in airways in OVA-TRE mice. Mas receptor blockade prevented the reduction of myeloperoxidase activity, as well as, prevented exercise-induced IL-10 increase. These data show that activation of Ang-(1-7)/Mas receptor pathway is involved in the anti-inflammatory and anti-fibrotic effects of aerobic training in an experimental model of chronic asthma. SignificanceOur results support exercise training as a non-pharmacological tool to defeat lung remodeling induced by chronic pulmonary inflammation. Further, our result also supports development of new therapy based on Ang-(1-7) or Mas agonists as important tool for asthma treatment in those patients that cannot perform aerobic training.

Maternal and Fetal Defects of Gestational Angiotensin 1‐7 Receptor Antagonism: A Possible Preclinical Model of Preeclampsia

Preeclampsia (PE) is a complex pathological state associated with serious adverse maternal and fetal consequences. Imbalances of the pressor ACE/Ang II and depressor ACE2/Ang 1-7 arms of renin-angiotensin system are implicated in PE pathophysiology. The current experimental study investigated whether the blockade of Ang 1-7 receptors (Mas receptors) during late pregnancy would replicate clinical features of PE. Two different dosing regimens of A779 (Mas receptor antagonist, 0.750 and 1.3 mg/kg/day s.c.) were administered daily to pregnant rats for 7 consecutive days starting from gestational day 14. The developed maternal and fetal abnormalities were compared to those produced by gestational L-NAME administration (50 mg/kg/day for 7 days), a standard model of PE in rodents. Gestational administration of L-NAME or A779 reduced weight and number of placentas and pups and increased death and resorption percentages of pups. These effects were more evident with the lower dose of A779 than with the higher dose. Moreover, A779 or L-NAME provoked similar elevations in systolic blood pressure and proteinuria. Immunohistochemical studies on placental tissues revealed enhanced protein expression of NFκB in dams treated with L-NAME or the lower, but not higher, dose of A779. While placental ACE expression remained unaltered, ACE2 expression was upregulated by A779 and L-NAME. Studies in isolated perfused kidneys demonstrated diminished acetylcholine vasodilation in L-NAME, but not A779, challenged preparations. Neither drug affected renal vasoconstriction induced by Ang II or phenylephrine. The current data highlight the likelihood of exploiting pregnant dams with pharmacologically eliminated Mas receptors for PE modeling in rodents. However, more studies are needed to further validate the model fidelity and its intimate resemblance to human PE.

Cardio-protective effects of angiotensin-(1-5) via mas receptor in rats against ischemic-perfusion injury

Angiotensin-(1-5) [Ang-(1-5)], which is a metabolite of Ang-(1-7) catalyzed by angiotensin-converting enzyme, is a novel pentapeptide of the renin-angiotensin system. Ang-(1-7), Ang III and Ang IV have a cardio-protective effect via Mas receptor, Ang II type 2 receptor (AT2R) and AT4R, respectively. However, it is not clear whether Ang-(1-5) has cardio-protective effects. The aim of this study is to investigate whether Ang-(1-5) protects the heart against ischemia-reperfusion (I/R) injury. After sacrificing Sprague-Dawley rats, the hearts were perfused with Krebs-Henseleit buffer for a 20 min pre-ischemic period with and without Ang-(1-5) followed by 20 min global ischemia and 50 min reperfusion. Ang-(1-5) (1 μM) improved changes in post-ischemic left ventricular developed pressure (LVDP), ±dP/dt, and post-ischemic left ventricular end-diastolic pressure (LVEDP) induced by reperfusion compared to control hearts. Ang-(1-5) decreased myocardial infarct size and LDH activity, and increased coronary flow and the amount of atrial natriuretic peptide (ANP) in coronary effluent during reperfusion compared to control hearts. Pretreatment with Mas receptor antagonist but not with AT1R or AT2R antagonist attenuated the improvement of changes in I/R-induced ventricular hemodynamics by Ang-(1-5). Ang-(1-5) treatment decreased Bax, caspase-3 and caspase-9 protein levels, and increased Bcl-2 protein level, which were attenuated by A779 pretreatment. Ang-(1-5) treatment increased Mn-superoxide dismutase, catalase, and heme oxygenase-1 protein levels, which was attenuated by A779 pretreatment. These results suggest that the cardio-protective effects of Ang-(1-5) against I/R injury may be partly related to activating anti-oxidant and anti-apoptotic enzymes via Mas receptor.

Angiotensin 1-7 and losartan worsen the cisplatin induced nephrotoxicity in female rats

Introduction: Cisplatin (CP) is an anti-cancer drug with the most common side effects of nephrotoxicity. Losartan, an angiotensin II type 1 receptor (AT1R) antagonist and angiotensin 1-7 (Ang1-7) protects the kidney against CP administration in males Moreover, the activity of the renin angiotensin system (RAS) and the incidence of CP induced nephrotoxicity are gender related. Objectives: The role of Ang1-7 and losartan against CP induced nephrotoxicity in female rats was examined. Methods: Thirty-two female Wistar rats in five experimental groups were treated with vehicle, single dose of CP (7.5 mg/kg), CP+losartan (10 ), CP+Ang1-7 (30 μg/kg/d) or CP+Ang1-7+A779 (Mas receptor antagonist, 100 μg/kg/d). The biochemical and histology measurements were conducted one week later. Results: The levels of serum creatinine (Cr) and blood urea nitrogen (BUN) in serum increased insignificantly by CP alone administration. However co-treatment of CP with losartan, Ang1-7, or Ang1-7 plus A779 showed an increase of the serum levels of BUN and Cr, and kidney tissue damage score (KTDS) (P < 0.05) when compared with control groups. Conclusion: The AT1R and Mas receptor (MasR) antagonists and Ang1-7 administration promote the CP induced damage of kidney in female rats, and special attention is needed during CP therapy in hypertensive patients who are treating with anti-hypertensive drug of losartan.

Abstract P094: Crosstalk Between Mas And ET B R Elicits Protective Effects In Endothelial Cells And Vascular Function

Mas and ET B receptors physically interact in endothelial cells (ECs) and are involved in the protective actions of angiotensin 1-7 (Ang (1-7)). We assessed whether the MAS/ET B R interaction plays a role in EC signalling and whether strategies to enhance MAS/ET B R association influence vascular responses. Human ECs were stimulated with Ang (1-7) (10 -7 M) in the presence/absence of A779 (Mas receptor antagonist, 10 -5 M) and BQ788 (ET B R antagonist, 10 -5 M). Protein expression and signalling activation were assessed by immunoblotting. NO production was evaluated by DAF-FM fluorescence and ROS production by chemiluminescence (superoxide anion) or amplex red (hydrogen peroxide (H 2 O 2 )). mRNA expression was assessed by qPCR. Endothelial function was assessed in mouse intact arteries by myography. Ang (1-7), through Mas and ET B R induced phosphorylation of eNOS (35%); followed by an increase in NO production (2.0 fold) (p<0.05 vs ctl). High throughput screening of protein:protein interaction compounds in an in-house library identified 23 potential enhancers of the MAS/ET B R interaction. Fluorescence polarization assays were used to further select the most potent enhancers and define their working concentration for testing in ECs (Enh1-4: 10 -5 M). Enh4 increased superoxide anion (55.6±26.3% vs ctl, p<0.05) and H 2 O 2 production (54.7±11.1% vs ctl, p<0.05), while Enh3 increased H 2 O 2 generation (48.1±15.4% vs ctl, p<0.05) in ECs. Moreover, Enh4 increased Nrf2 (3.0 fold), Sod1 (2.0 fold) and Nqo1 (3.1 fold) mRNA expression (p<0.05 vs ctl). Enh3 and Enh4 increased NO production (Enh3: 21.2±7.4%; Enh4: 23.6±8.2% vs veh, p<0.05) in ECs. Acetylcholine (Ach) curves were performed to assess endothelium-dependent relaxation in the absence and presence of enhancers. Enh4 increased ACh-induced relaxation (Emax%: 96.7±4.6 vs ctl: 70.4±3.3, p<0.05), while other enhancers did not improve endothelial function. Taken together, increasing MAS/ET B R interaction with specific enhancers augments protective signalling in ECs and promotes endothelial-dependent vasorelaxation, particularly with Enh4. In conclusion, enhancing interactions between MasR and ET B R may be a new vasoprotective strategy to improve vascular function in cardiovascular disease.

Downregulation of spinal angiotensin converting enzyme 2 is involved in neuropathic pain associated with type 2 diabetes mellitus in mice

We have previously reported that the spinal angiotensin (Ang) system is involved in the modulation of streptozotocin (STZ)-induced diabetic neuropathic pain in mice. An important drawback of this model however is the fact that the neuropathic pain is independent of hyperglycemia and produced by the direct stimulation of peripheral nerves. Here, using the leptin deficient ob/ob mouse as a type 2 diabetic model, we examined whether the spinal Ang system was involved in naturally occuring diabetic neuropathic pain. Blood glucose levels were increased in ob/ob mice at 5–15 weeks of age. Following the hyperglycemia, persistent tactile and thermal hyperalgesia were observed at 11–14 and 9–15 weeks of age, respectively, which was ameliorated by insulin treatment. At 12 weeks of age, the expression of Ang-converting enzyme (ACE) 2 in the spinal plasma membrane fraction was decreased in ob/ob mice. Spinal ACE2 was expressed in neurons and microglia but the number of NeuN-positive neurons was decreased in ob/ob mice. In addition, the intrathecal administration of Ang (1-7) and SB203580, a p38 MAPK inhibitor, attenuated hyperalgesia in ob/ob mice. The phosphorylation of spinal p38 MAPK was also attenuated by Ang (1-7) in ob/ob mice. These inhibitory effects of Ang (1-7) were prevented by A779, a Mas receptor antagonist. In conclusion, we revealed that the Ang (1-7)-generating system is downregulated in ob/ob mice and is accompanied by a loss of ACE2-positive neurons. Furthermore, Ang (1-7) decreased the diabetic neuropathic pain through inhibition of p38 MAPK phosphorylation via spinal Mas receptors.

Angiotensin (1-7) Inhibits Ang II-mediated ERK1/2 Activation by Stimulating MKP-1 Activation in Vascular Smooth Muscle Cells.

The renin–angiotensin system (RAS) exerts profound physiological effects on blood pressure regulation and fluid homeostasis, mainly by modulating renal, cardiovascular, and central nervous systems. Angiotensin (Ang)-(1-7), an end-product of RAS, is recognized by its cardiovascular protective properties through stimulation of the Mas receptor, including vasodilation, anti-inflammatory, and antihypertensive actions, and consequently, counter-regulating the well-known Ang II-elicited actions. The overall hypothesis of this study is that Ang-(1-7) inhibits Ang II-induced ERK1/2 activation in vascular smooth muscle cells (VSMCs), via regulation of mitogen-activated protein phosphatase-1 (MKP-1) activity. Aortas from male Wistar rats were incubated with Ang-(1-7) or vehicle. Concentration-response curves to Ang II were performed in endothelium-denuded aortas, in the presence or absence of ERK1/2 (PD98059) inhibitor or Mas receptor (A-779) antagonist. Expression of proteins was assessed by western blot, and immunohistochemistry was conducted in VSMCs. Ang-(1-7) incubation decreased Ang II-induced contractile response in aortas, and this effect was not observed in the presence of PD98059 or A-779. Stimulation of VSMCs with Ang-(1-7) prevented Ang II-induced ERK1/2 phosphorylation, but not C-Raf-activation. Furthermore, Ang II decreased MKP-1 phosphorylation in VSMCs. Interestingly, simultaneous incubation of Ang-(1-7) with Ang II favored MKP-1 phosphorylation, negatively modulating ERK1/2 activation in VSMCs. The results suggest that Ang-(1-7) counter-regulates actions evoked by Ang II overproduction, as observed in cardiovascular diseases, mainly by modulating MKP-1 activity. This evidence suggests that the role of Ang-(1-7) in MKP-1-regulation represents a target for new therapeutic development.

Angiotensin‐(1‐7) Prevents Thyroid Hormone‐induced Cardiomyocyte Hypertrophy by FOXO3 Signaling Pathway

It is well‐known that high levels of thyroid hormone promote cardiomyocyte hypertrophy, which in part is mediated by activation of Renin‐Angiotensin System (RAS). Recent study of our group demonstrated that the contralateral RAS axis (ACE2/Angiotensin‐(1–7)/Mas receptor) is also involved in this cardiac hypertrophy model and that transgenic rats overexpressing circulating levels of Angiotensin‐(1‐7) (Ang‐(1‐7)) present an improvement of cardiac function and attenuation of cardiac hypertrophy. However, the molecular mechanisms involved in these Ang‐(1‐7) actions have not been fully elucidated. The aim of this study was to investigate if the anti‐hypertrophic effect of Ang‐(1‐7) also occurs in isolated cardiomyocytes, as well as the involvement of the Foxo3 signaling pathway for these effects. All experiments were approved by the Ethics Committee on Animal Experimentation of ICB‐USP (n030/12/CEUA). Primary cardiomyocytes cultures obtained from neonatal rats were treated with triiodothyronine (T3) (15nM) and/or Angiotensin‐(1‐7) (Ang‐(1‐7)) (100nM). All data were analyzed by ANOVA followed by Tukey's post hoc test, and values of P<0.05 were considered statistically significant. The Ang‐(1‐7) treatment prevented the cardiomyocyte hypertrophy induced by T3, which was confirmed by reduction cell surface area, L‐[3,4,5‐3H] leucine incorporation and BNP gene expression. Combined treatment with the MAS receptor antagonist (A779, 100nM) impaired the anti‐hypertrophic effect of Ang‐(1‐7). Regarding molecular mechanisms, although Ang‐(1‐7) treatment did not impair the rapid activation of Akt promoted by T3, Ang‐(1‐7) treatment reduced the phosphorylation of Foxo3 (inactive form) and promoted the nuclear accumulation of Foxo3 in cardiomyocytes, indicating the activation of this protein. Collectively, these findings indicate an important cardioprotective effect of Ang‐(1‐7) in isolated cardiomycoytes treated with T3, occurring via MAS receptor and with possible participation of Foxo3.Support or Funding InformationFoundation for the Support of Research in the State of São Paulo (FAPESP) and National Council of Technological and Scientific Development (CNPq).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Angiotensin-(1-7) inhibits sodium transport via Mas receptor by increasing nitric oxide production in thick ascending limb.

Sodium transport in the thick ascending loop of Henle (TAL) is tightly regulated by numerous factors, especially angiotensin II (Ang II), a key end‐product of the renin‐angiotensin system (RAS). However, an alternative end‐product of the RAS, angiotensin‐(1‐7) [Ang‐(1‐7)], may counter some of the Ang II actions. Indeed, it causes vasodilation and promotes natriuresis through its effects in the proximal and distal tubule. However, its effects on the TAL are unknown. Because the TAL expresses the Mas receptor, an Ang‐(1‐7) ligand, which in turn may increase NO and inhibit Na+ transport, we hypothesized that Ang‐(1‐7) inhibits Na transport in the TAL, via a Mas receptor/NO‐dependent mechanism. We tested this by measuring transport‐dependent oxygen consumption (VO 2) in TAL suspensions. Administering Ang‐(1‐7) decreased VO 2; an effect prevented by dimethyl amiloride and furosemide, signifying that Ang‐(1‐7) inhibits transport‐dependent VO 2 in TAL. Ang‐(1‐7) also increased NO levels, known inhibitors of Na+ transport in the TAL. The effects of Ang‐(1‐7) on VO 2, as well as on NO levels, were ameliorated by the Mas receptor antagonist, D‐Ala, in effect suggesting that Ang‐(1‐7) may inhibit transport‐dependent VO 2 in TAL via Mas receptor‐dependent activation of the NO pathway. Indeed, blocking NO synthesis with L‐NAME prevented the inhibitory actions of Ang‐(1‐7) on VO 2. Our data suggest that Ang‐(1‐7) may modulate TAL Na+ transport via Mas receptor‐dependent increases in NO leading to the inhibition of transport activity.

Mas receptor antagonist (A799) alters the renal hemodynamics responses to angiotensin II administration after renal moderate ischemia/reperfusion in rats: gender related differences.

Moderate renal ischemia/reperfusion (I/R) injury is one of the major causes of kidney failure. We examined the role of Mas receptor (MasR) antagonist (A779) alone and combined with angiotensin II (Ang II) type 2 receptor (AT2R) antagonist (PD123319) on renal hemodynamic responses to Ang II after moderate I/R in male and female rats. Anaesthetized Wistar rats underwent 30 min partial ischemia by reduction of renal perfusion pressure (RPP) and subjected to block vasodepressor receptors followed by Ang II (100 and 300 ng/kg/min) infusion. Mean arterial pressure (MAP), renal blood flow (RBF), and renal vascular resistance (RVR) responses were assessed during graded Ang II infusion at controlled RPP. Thirty min post reperfusion, the Ang II infusion reduced RBF and increased RVR in a dose-related fashion (P < 0.05). However, A779 alone or A779 plus PD123319 infusion increased the RBF and RVR responses to Ang II infusion significantly (P < 0.05) in female but not in the male rats. MasR antagonist altered the RBF and RVR responses to Ang II infusion in female, and these responses were not altered statistically in dual blockade of MasR and AT2R. These findings suggest the important role of Mas receptor in renal vascular response to Ang II in female rats after moderate I/R.

Lipoxin A4 attenuates LPS-induced acute lung injury via activation of the ACE2-Ang-(1-7)-Mas axis.

Previous studies have reported that lipoxin A4 (LXA4) and the angiotensin I-converting enzyme 2 (ACE2), angiotensin-(1-7) [Ang-(1-7)], and its receptor Mas [ACE2-Ang-(1-7)-Mas] axis play important protective roles in acute lung injury (ALI). However, there is still no direct evidence of LXA4-mediated protection via the ACE2-Ang-(1-7)-Mas axis during ALI. This work was performed using an LPS-induced ALI mouse model and the data indicated the following. First, the animal model was established successfully and LXA4 ameliorated LPS-induced ALI. Second, LXA4 could increase the concentration and activity of ACE2 and the levels of Ang-(1-7) and Mas markedly. Third, LXA4 decreased the levels of TNF-α, IL-1β, and reactive oxygen species while increasing IL-10 levels. Fourth, LXA4 inhibited the activation of the NF-κB signal pathway and repressed the degradation of inhibitor of NF-κB, the phosphorylation of NF-κB, and the translocation of NF-κB. Finally, and more importantly, BOC-2 (LXA4 receptor inhibitor), MLN-4760 (ACE2 inhibitor), and A779 (Mas receptor antagonist) were found to reverse all of the effects of LXA4. Our data provide evidence that LXA4 protects the lung from ALI through regulation of the ACE2-Ang-(1-7)-Mas axis.

Abstract 713: Angiotensin 1-7 Suppresses Experimental Abdominal Aortic Aneurysms

Objectives: Abdominal aortic aneurysm (AAA) is a chronic inflammatory disease, the pathogenesis of which remains incompletely defined. We and others have demonstrated the critical role of angiotensin (Ang) II and its receptor AT1 in experimental AAAs. Although Ang 1-7 and its receptor Mas counteract many AT1 functions, the role of the Ang 1-7/Mas pathway has not been studied in AAAs. This study evaluated the influences of Ang 1-7 and Mas on experimental AAAs. Methods: AAAs were created in male C57BL/6J mice via porcine pancreatic elastase (PPE) infusion. Mice were treated with Ang 1-7 (0.5 mg/kg/day via s.c) or vehicle starting immediately prior to, or 4 days following, PPE infusion. Additional mice were co-treated with Ang 1-7 and the Mas receptor antagonist A779 (0.5 mg/kg/day) beginning immediately prior to PPE infusion. Influences on AAAs were evaluated by serial ultrasound imaging and histology. Results: Figure summarizes major results. In ultrasound imaging, at 14 days Ang 1-7 treatment initiated immediately following aneurysm creation substantially suppressed aortic enlargement as compared to vehicle treatment. Histologically, Ang 1-7 treatment was associated with attenuation of aortic medial elastin and smooth muscle cell depletion, mural macrophage, CD4 + , CD8 + T and B cell density and neoangiogenesis. Co-treatment with Ang 1-7 receptor Mas antagonist A779 “rescued” the aneurysm phenotype. Additionally, Ang 1-7 treatment initiated even 4 days following AAA creation was still effective in limiting further aneurysmal enlargement and pathologic evolution. Conclusions: Ang 1-7 treatment, initiated either at, or up to 4 days following, AAA initiation limits further progression of experimental AAAs. This inhibitory effect seems likely related to activation of the Mas receptor. These results suggest that either Ang 1-7 or alternative Mas receptor agonists may have clinical utility in suppressing progression of early AAA disease.

Angiotensin 1–7 Action in the Lateral Preoptic Area is Involved in Urinary Bladder Regulation in Female Wistar Rats.

BackgroundChemical lesions of the lateral preoptic area (LPA) enhance water intake induced by subcutaneous injection of hypertonic saline in rats. On the other hand, the LPA section abolishes the increase in intravesical pressure (IP) induced by olfactory tubercle stimulation in dogs. Immunohistochemical labeling has demonstrated the presence of Mas receptors for Angiotensin‐(1–7) in LPA. It is not known whether neurons with Mas receptors in LPA are involved in urinary bladder regulation.AimWe investigated the effects of activation or blockade of Mas receptors for angiotensin‐(1–7) in LPA on intravesical pressure in anesthetized female rats.Materials and methodsFemale Wistar rats (~240 g, N=6) underwent to a stereotaxic surgery for implantation of stainless steel guide cannulas into the LPA under ketamine and xylazin anesthesia. After 7 days, the animals were anesthetized and maintained with 2% isoflurane in 100% O2 and submitted to cannulation of the femoral artery and vein, placement of a miniaturized Doppler flow probe around the left renal artery to measure the renal blood flow and cannulation of the urinary bladder for IP measurements. After baseline IP and cardiovascular parameters recordings for 15 min, angiotensin‐(1–7) (5 nmol/ uL) or saline (1 uL) or A‐779 (Mas receptor antagonist, 50 nmol/uL) was injected into the LPA and the variables were measured for additional 60 min.ResultsUnilateral injection of angiotensin‐(1–7) into LPA evoked a significant increase in IP (187.5±37.2%) compared to saline (−2.1±1.9%). Unilateral injection of A‐779 into LPA decreased the IP (−15.9±2.8%) in comparison to saline (2.0±1.1%). Interestingly, the decrease in IP was enhanced after bilateral injection of A‐779 into LPA (−27.3±3.4%) compared to the unilateral injection of this drug and also in comparison to saline (−2.9±1.6%) bilaterally into LPA. No significant changes in cardiovascular parameters after angiotensin‐(1–7) or saline or A‐779 in APL were observed.ConclusionThe data suggest that LPA is an important part of the circuit that regulates the urinary bladder activity mediated by angiotensin‐(1–7).Support or Funding InformationFinancial support: FAPESP and NEPAS‐FMABCThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Angiotensin 1-7 ameliorates 6-hydroxydopamine lesions in hemiparkinsonian rats through activation of MAS receptor/PI3K/Akt/BDNF pathway and inhibition of angiotensin II type-1 receptor/NF-κB axis

MAS receptor (MASR), expressed in several brain areas, conferred neuroprotection against neurodegenerative disorders when activated by angiotensin (Ang) 1-7; however, its role in Parkinson’s disease (PD) remains elusive. Intra-striatal post-administration of Ang1-7, using a 6-hydroxydopamine (OHDA) PD model, improved motor performance and muscle coordination. On the molecular level, Ang1-7 upregulated the striatal expression of MASR and caused upsurge in its downstream targets (p-PI3K/p-Akt/p-CREB/BDNF) to phosphorylate TrKB, which in a positive feedback upregulates MASR. Moreover, Ang1-7 increased substantia nigral tyrosine hydroxylase (TH) expression and striatal dopamine (DA) content to indicate the preservation of the dopaminergic neuronal signal. This effect extended to inhibit the striatal expression of Ang II type-1 receptor (AT-1R) to hold the neurodegenerative effect and to boost Ang1-7 anti-inflammatory/antioxidant effects by abating NADPH oxidase, along with lipid peroxidation. Indeed, Ang1-7 was able to decrease p-MAPK p38/NF-κB p65 to level the inflammatory and oxidative stress events off. The Ang1-7-mediated activation of MASR cue and the suppression of the AT-1R cascade were partially reversed by the intrastartial injection of A-779, a MASR antagonist. The current data suggests a novel therapeutic potential for the Ang1-7 against neurotoxicity associated motor impairment related to PD. The anti-parkinsonian effect of Ang1-7, is in part, mediated by its binding to MASR and the initiation of PI3K/Akt/CREB/BDNF/TrKB cue to increase DA synthesis, besides the downregulation/inhibition of AT-1R/MAPK p38/NF-κB p65/NADPH oxidase pathway.

The Role of Vasodilator Receptors of Renin-angiotensin System on Nitric Oxide Formation and Kidney Circulation after Angiotensin II Infusion in Renal Ischemia/Reperfusion Rats.

Background:Nitric oxide (NO) as a vasodilator factor has renoprotective effect against renal ischemia. The balance between angiotensin II (Ang II) and NO can affect kidney homeostasis. The aim of this study was to determine NO alteration in response to renin–Ang system vasodilator receptors antagonists (PD123319; Ang II type 2 receptor antagonist and A779; Mas receptor antagonist) in renal ischemia/reperfusion injury (IRI) in rats.Materials and Methods:Sixty-three Wistar male and female rats were used. Animals from each gender were divided into four groups received saline, Ang II, PD123319 + Ang II, and A779 + Ang II after renal IRI. Renal IRI induced with an adjustable hook. Blood pressure and renal blood flow (RBF) measured continuously. The nitrite levels were measured in serum, kidney, and urine samples.Results:In female rats, the serum and kidney nitrite levels increased significantly by Ang II (P < 0.05) and decreased significantly (P < 0.05) when PD123319 was accompanied with Ang II. Such observation was not seen in male. Ang II decreased RBF significantly in all groups (P < 0.05), while PD + Ang II group showed significant decrease in RBF in comparison with the other groups in female rats (P < 0.05).Conclusion:Males show more sensibility to Ang II infusion; in fact, it is suggested that there is gender dimorphism in the Ang II and NO production associated with vasodilator receptors.