Mas Receptor Research Articles

Angiotensin-(1-7) ameliorates high glucose-induced vascular endothelial injury through suppressing chloride channel 3

ABSTRACT Diabetes Mellitus (DM) is a significant risk factor for cardiovascular disease (CVD), which is leading cause of deaths in DM patients. However, there are limited effective medical therapies for diabetic CVD. Vascular endothelial injury caused by DM is a critical risk factor for diabetic CVD. Previous study has indicated that Angiotensin-(1-7) (Ang-(1-7)) may prevent diabetic CVD, whereas it is not clear that Ang-(1-7) whether attenuates diabetic CVD through suppressing vascular endothelial injury. In this study, we found that Ang-(1-7) alleviated high glucose (HG)-induced endothelial injury in bEnd3 cells. Moreover, Ang-(1-7) ameliorated HG-induced endothelial injury through downregulating chloride channel 3 (CIC-3) via Mas receptor. Furthermore, HG-induced CIC-3 enhanced reactive oxygen species (ROS) and cytokine production and reduced the level of nitric oxide (NO), while Ang-(1-7) preserved the impact of HG-induced CIC-3 on productions of ROS, cytokine and NO through inhibiting CIC-3 via Mas receptor. Summarily, the present study revealed that Ang-(1-7) alleviated HG-induced vascular endothelial injury through the inhibition of CIC-3, suggested that Ang-(1-7) may preserve diabetic CVD through suppressing HG-induced vascular endothelial injury.

ACE2 pathway regulates thermogenesis and energy metabolism.

Identification of key regulators of energy homeostasis holds important therapeutic promise for metabolic disorders, such as obesity and diabetes. ACE2 cleaves angiotensin II (Ang II) to generate Ang-(1-7) which acts mainly through the Mas1 receptor. Here, we identify ACE2 pathway as a critical regulator in the maintenance of thermogenesis and energy expenditure. We found that ACE2 is highly expressed in brown adipose tissue (BAT) and that cold stimulation increases ACE2 and Ang-(1-7) levels in BAT and serum. Ace2 knockout mice (Ace2-/y) and Mas1 knockout mice (Mas1-/-) displayed impaired thermogenesis. Mice transplanted with brown adipose tissue from Mas1-/- display metabolic abnormalities consistent with those seen in the Ace2 and Mas1 knockout mice. In contrast, impaired thermogenesis of Leprdb/db obese diabetic mice and high-fat diet-induced obese mice were ameliorated by overexpression of Ace2 or continuous infusion of Ang-(1-7). Activation of ACE2 pathway was associated with improvement of metabolic parameters, including blood glucose, lipids, and energy expenditure in multiple animal models. Consistently, ACE2 pathway remarkably enhanced the browning of white adipose tissue. Mechanistically, we showed that ACE2 pathway activated Akt/FoxO1 and PKA pathway, leading to induction of UCP1 and activation of mitochondrial function. Our data propose that adaptive thermogenesis requires regulation of ACE2 pathway and highlight novel potential therapeutic targets for the treatment of metabolic disorders.

Angiotensin-(1-7)/MasR axis promotes migration of monocytes/macrophages with a regulatory phenotype to perform phagocytosis and efferocytosis.

Nonphlogistic migration of macrophages contributes to the clearance of pathogens and apoptotic cells, a critical step for the resolution of inflammation and return to homeostasis. Angiotensin-(1-7) [Ang-(1-7)] is a heptapeptide of the renin-angiotensin system that acts through Mas receptor (MasR). Ang-(1-7) has recently emerged as a novel proresolving mediator, yet Ang-(1-7) resolution mechanisms are not fully determined. Herein, Ang-(1-7) stimulated migration of human and murine monocytes/macrophages in a MasR-, CCR2-, and MEK/ERK1/2–dependent manner. Pleural injection of Ang-(1-7) promoted nonphlogistic mononuclear cell influx alongside increased levels of CCL2, IL-10, and macrophage polarization toward a regulatory phenotype. Ang-(1-7) induction of CCL2 and mononuclear cell migration was also dependent on MasR and MEK/ERK. Of note, MasR was upregulated during the resolution phase of inflammation, and its pharmacological inhibition or genetic deficiency impaired mononuclear cell recruitment during self-resolving models of LPS pleurisy and E. coli peritonitis. Inhibition/absence of MasR was associated with reduced CCL2 levels, impaired phagocytosis of bacteria, efferocytosis, and delayed resolution of inflammation. In summary, we have uncovered a potentially novel proresolving feature of Ang-(1-7), namely the recruitment of mononuclear cells favoring efferocytosis, phagocytosis, and resolution of inflammation. Mechanistically, cell migration was dependent on MasR, CCR2, and the MEK/ERK pathway.

Sex Difference in MasR Expression and Functions in the Renal System.

Renin-angiotensin system (RAS), as a critical system for controlling body fluid and hemostasis, contains peptides and receptors, including angiotensin 1-7 (Ang 1-7) and Mas receptor (MasR). Ang 1-7 implements its function via MasR. Ang II is another peptide in RAS that performs its actions via two Ang II type 1 and 2 receptors (AT1R and AT2R). The functions of AT2R and MasR are very similar, and both have a vasodilation effect, while AT1R has a vasoconstriction role. MasR affects many mechanisms in the brain, heart, blood vessels, kidney, lung, endocrine, reproductive, skeletal muscle, and liver and probably acts like a paracrine hormone in these organs. The effect of Ang 1-7 in the kidney is complex according to the hydroelectrolyte status, the renal sympathetic nervous system, and the activity level of the RAS. The MasR expression and function seem more complex than Ang II receptors and have interacted with Ang II receptors and many other factors, including sex hormones. Also, pathological conditions including hypertension, diabetes, and ischemia-reperfusion could change MasR expression and function. In this review, we consider the role of sex differences in MasR expression and functions in the renal system under physiological and pathological conditions.

View of the Renin-Angiotensin System in Acute Kidney Injury Induced by Renal Ischemia-Reperfusion Injury.

Renal ischemia-reperfusion injury (RIRI) is a sequence of complicated events that is defined as a reduction of the blood supply followed by reperfusion. RIRI is the leading cause of acute kidney injury (AKI). Among the diverse mediators that take part in RIRI-induced AKI, the renin-angiotensin system (RAS) plays an important role via conventional (angiotensinogen, renin, angiotensin-converting enzyme (ACE), angiotensin (Ang) II, and Ang II type 1 receptor (AT1R)) and nonconventional (ACE2, Ang 1-7, Ang 1-9, AT2 receptor (AT2R), and Mas receptor (MasR)) axes. RIRI alters the balance of both axes so that RAS can affect RIRI-induced AKI. In overall, the alteration of Ang II/AT1R and AKI by RIRI is important to consider. This review has looked for the effects and interactions of RAS activities during RIRI conditions.

The Effects of Angiotensin II or Angiotensin 1-7 on Rat Pial Microcirculation during Hypoperfusion and Reperfusion Injury: Role of Redox Stress.

Renin–angiotensin systems produce angiotensin II (Ang II) and angiotensin 1-7 (Ang 1-7), which are able to induce opposite effects on circulation. This study in vivo assessed the effects induced by Ang II or Ang 1-7 on rat pial microcirculation during hypoperfusion–reperfusion, clarifying the mechanisms causing the imbalance between Ang II and Ang 1-7. The fluorescence microscopy was used to quantify the microvascular parameters. Hypoperfusion and reperfusion caused vasoconstriction, disruption of blood–brain barrier, reduction of capillary perfusion and an increase in reactive oxygen species production. Rats treated with Ang II showed exacerbated microvascular damage with stronger vasoconstriction compared to hypoperfused rats, a further increase in leakage, higher decrease in capillary perfusion and marker oxidative stress. Candesartan cilexetil (specific Ang II type 1 receptor (AT1R) antagonist) administration prior to Ang II prevented the effects induced by Ang II, blunting the hypoperfusion–reperfusion injury. Ang 1-7 or ACE2 activator administration, preserved the pial microcirculation from hypoperfusion–reperfusion damage. These effects of Ang 1-7 were blunted by a Mas (Mas oncogene-encoded protein) receptor antagonist, while Ang II type 2 receptor antagonists did not affect Ang 1-7-induced changes. In conclusion, Ang II and Ang 1-7 triggered different mechanisms through AT1R or MAS receptors able to affect cerebral microvascular injury.

Renin-Angiotensin System Alterations in the Human Alzheimer's Disease Brain.

Understanding Alzheimer's disease (AD) in terms of its various pathophysiological pathways is essential to unravel the complex nature of the disease process and identify potential therapeutic targets. The renin-angiotensin system (RAS) has been implicated in several brain diseases, including traumatic brain injury, ischemic stroke, and AD. This study was designed to evaluate the protein expression levels of RAS components in postmortem cortical and hippocampal brain samples obtained from AD versus non-AD individuals. We analyzed RAS components in the cortex and hippocampus of postmortem human brain samples by western blotting and immunohistochemical techniques in comparison with age-matched non-demented controls. The expression of AT1R increased in the hippocampus, whereas AT2R expression remained almost unchanged in the cortical and hippocampal regions of AD compared to non-AD brains. The Mas receptor was downregulated in the hippocampus. We also detected slight reductions in ACE-1 protein levels in both the cortex and hippocampus of AD brains, with minor elevations in ACE-2 in the cortex. We did not find remarkable differences in the protein levels of angiotensinogen and Ang II in either the cortex or hippocampus of AD brains, whereas we observed a considerable increase in the expression of brain-derived neurotrophic factor in the hippocampus. The current findings support the significant contribution of RAS components in AD pathogenesis, further suggesting that strategies focusing on the AT1R and AT2R pathways may lead to novel therapies for the management of AD.

Pharmacodynamic assessment of orally available Mas agonists

AbstractBackgroundOxidative stress (OS) plays a crucial role in driving Alzheimer’s disease (AD) pathogenesis. Angiotensin II (A‐II), the ligand for AT1R (pathological axis of the renin angiotensin system [RAS]), promotes the development of AD in part by increasing OS. Mas receptor and its ligand angiotensin (1‐7) (A(1‐7); protective axis of RAS) inhibits A‐II's pathological actions. Two orally available small molecule Mas agonists, RASRx1902 and RASRx1911, were developed to overcome shortcomings of A(1‐7). Both RASRx1902 and RASRx1911 reduce brain OS 90 days after transaortic constriction surgery (TAC), suggesting that both drugs might reduce neurodegeneration. To understand their pharmacodynamics, RASRx1902 and RASRx1911 were tested in the TAC model to determine their optimal dose and dosing regimen, and biological activity.MethodC57BL/6J mice underwent TAC or sham surgery. After echocardiography performed, mice were randomized based on their TAC gradient. Mice received one dose of saline, RASRx1902, or RASRx1911 at a range of subcutaneous or oral doses and euthanized at 0.5, 1, 2, 4 and 6 hours post‐treatment. After dose and route of delivery optimization, mice were treated 1x/day, 2x/day, 2x/week or 3x/week for 2 weeks. Brain tissues were isolated and analyzed for pharmacokinetics, functional markers for OS, and Akt/ERK activation.ResultsOne to three weeks after TAC surgery, the levels of mitochondrial superoxide were increased in microglia, but not in neurons, oligodendrocytes, and astrocytes compared with sham surgery. Treatment with RASRx1902 and RASRx1911, administered either subcutaneously or orally, reduced microglial superoxide levels compared to saline‐treated TAC mice. RASRx1902 showed a stronger inhibitory activity on TAC‐mediated superoxide production than RASRx1911. RASRx1902 and RASRx1911 treatment reduced the TAC‐mediated decreased Akt/Erk activity in brain compared to saline treatment.ConclusionsMicroglial mitochondrial superoxide is the main contributor to promote OS, a contributor to AD neurodegeneration. Our data clearly show that our metabolically stable small molecule Mas agonists have the capacity to decrease ROS levels in microglia and might mediate neuronal survival through Akt/Erk signaling pathway. Therefore, RASRx1902 and RASRx1911 are promising novel drug for the treatment of AD.

The Liver and Heart: How the Two Most Beloved “Babies” in the Human Body Communicate

The Liver and Heart: How the Two Most Beloved “Babies” in the Human Body Communicate

Aerobic training-mediated DNA hypermethylation of Agtr1a and Mas1 genes ameliorate mesenteric arterial function in spontaneously hypertensive rats.

The imbalance of vasoconstrictor and vasodilator axes of the renin-angiotensin system (RAS) is observed in hypertension. Exercise regulates RAS level and improves vascular function. This study focused on the contribution of RAS axes in vascular function of mesenteric arteries and exercise-induced DNA methylation of the Agtr1a (AT1aR) and Mas1 (MasR) genes in hypertension. Spontaneously hypertensive rats (SHRs) and Wistar-Kyoto rats were randomized into exercise or sedentary group. Levels of plasma RAS components, vascular tone, and DNA methylation markers were measured. Blood pressure of SHR was markedly reduced after 12weeks of aerobic exercise. RAS peptides in plasma were all increased with an imbalanced upregulation of Ang II and Ang-(1-7) in SHR, exercise revised the level of RAS and increased Ang-(1-7)/Ang II. The vasoconstriction response induced by Ang II was mainly via type 1 receptors (AT1R), while this contraction was inhibited by Mas receptor (MasR). mRNA and protein of AT1R and MasR were both upregulated in SHR, whereas exercise significantly suppressed this imbalanced increase and increased MasR/AT1R ratio. Exercise hypermethylated Agtr1a and Mas1 genes, associating with increased DNMT1 and DNMT3b and SAM/SAH. Aerobic exercise ameliorates vascular function via hypermethylation of the Agtr1a and Mas1 genes and restores the vasoconstrictor and vasodilator axes balance.

The angiotensin converting enzyme 2 (ACE2) system in the brain: possible involvement in Neuro-Covid.

The brain has its own intrinsic renin-angiotensin system (RAS) with all its components present in the central nervous system (CNS). Recent data demonstrate that also the main components of the angiotensin concerting enzyme 2 (ACE2) system (at least ACE2 itself, as well as the biologically active angiotensin (1-7) and its cognate receptor Mas) are expressed in the brain. Aside from these members, alamadine and MrgD are discussed as further members that have neuro-active roles in the CNS. Little is known about the possible functions of MrgD within the brain. Concerning angiotensin (1-7) acting through the Mas receptor, data were accumulating that this system is involved in numerous processes contributing to neuronal plasticity and even learning and memory. Malfunctions in the brain ACE2 system are associated with disturbances in neuronal plasticity. Since SARS-CoV-2 has a high affinity towards ACE2, Neuro-Covid may directly or indirectly depend on a disturbed balance in the ACE2 derived angiotensin system in the brain. Since the ACE2 system in the brain is far from being understood, a deeper understanding of e.g. the angiotensin (1-7) / Mas system is needed, especially with regard to the roles of angiotensin (1-7) in neuronal plasticity.

A Pair of “ACEs”

The emergence of the COVID-19 viral pandemic has generated a renewed interest in pharmacologic agents that target the renin angiotensin system (RAS). Angiotensin-converting enzyme 1 (ACE1) inhibitors decrease the synthesis of angiotensin II (Ang II) from its precursor angiotensin I and inhibit the breakdown of bradykinin, while Ang II receptor blockers antagonize the action of Ang II at the receptor level downstream. The actions of both classes of drugs lead to vasodilation, a blunting of sympathetic drive and a reduction in aldosterone release, all beneficial effects in hypertension and congestive heart failure. ACE2 cleaves the vasoconstrictor Ang II to produce the anti-inflammatory cytoprotective angiotensin 1–7 (Ang 1–7) peptide, which functions through the G protein–coupled receptor MAS to counteract the pathophysiologic effects induced by Ang II via its receptors, including vasoconstriction, inflammation, hypercoagulation, and fibrosis. SARS-CoV-2 enters human cells by binding ACE2 on the cell surface, decreases ACE2 activity, competes for ACE2 receptor-binding sites, and shifts the RAS toward an overexpression of Ang II, accounting for many of the deleterious effects of the virus. Thus, there is great interest in developing recombinant ACE2 as a therapeutic for prevention or treatment of COVID-19. Notably, ACE2 is highly expressed in the oral cavity, and saliva and dorsum of the tongue are major reservoirs of SARS-CoV-2. Cost-effective methods to debulk the virus in the oral cavity may aid in the prevention of viral spread. Here we review the pharmacology of targeted small molecule inhibitors of the RAS and discuss novel approaches to employing ACE2 as a therapeutic for COVID-19.

Neurofilament light: a possible prognostic biomarker for treatment of vascular contributions to cognitive impairment and dementia

BackgroundDecreased cerebral blood flow and systemic inflammation during heart failure (HF) increase the risk for vascular contributions to cognitive impairment and dementia (VCID) and Alzheimer disease-related dementias (ADRD). We previously demonstrated that PNA5, a novel glycosylated angiotensin 1–7 (Ang-(1–7)) Mas receptor (MasR) agonist peptide, is an effective therapy to rescue cognitive impairment in our preclinical model of VCID. Neurofilament light (NfL) protein concentration is correlated with cognitive impairment and elevated in neurodegenerative diseases, hypoxic brain injury, and cardiac disease. The goal of the present study was to determine (1) if treatment with Ang-(1–7)/MasR agonists can rescue cognitive impairment and decrease VCID-induced increases in NfL levels as compared to HF-saline treated mice and, (2) if NfL levels correlate with measures of cognitive function and brain cytokines in our VCID model.MethodsVCID was induced in C57BL/6 male mice via myocardial infarction (MI). At 5 weeks post-MI, mice were treated with daily subcutaneous injections for 24 days, 5 weeks after MI, with PNA5 or angiotensin 1–7 (500 microg/kg/day or 50 microg/kg/day) or saline (n = 15/group). Following the 24-day treatment protocol, cognitive function was assessed using the Novel Object Recognition (NOR) test. Cardiac function was measured by echocardiography and plasma concentrations of NfL were quantified using a Quanterix Simoa assay. Brain and circulating cytokine levels were determined with a MILLIPLEX MAP Mouse High Sensitivity Multiplex Immunoassay. Treatment groups were compared via ANOVA, significance was set at p < 0.05.ResultsTreatment with Ang-(1–7)/MasR agonists reversed VCID-induced cognitive impairment and significantly decreased NfL levels in our mouse model of VCID as compared to HF-saline treated mice. Further, NfL levels were significantly negatively correlated with cognitive scores and the concentrations of multiple pleiotropic cytokines in the brain.ConclusionsThese data show that treatment with Ang-(1–7)/MasR agonists rescues cognitive impairment and decreases plasma NfL relative to HF-saline-treated animals in our VCID mouse model. Further, levels of NfL are significantly negatively correlated with cognitive function and with several brain cytokine concentrations. Based on these preclinical findings, we propose that circulating NfL might be a candidate for a prognostic biomarker for VCID and may also serve as a pharmacodynamic/response biomarker for therapeutic target engagement.

Shedding Light on the Pharmacological Interactions between μ-Opioid Analgesics and Angiotensin Receptor Modulators: A New Option for Treating Chronic Pain.

The current protocols for neuropathic pain management include µ-opioid receptor (MOR) analgesics alongside other drugs; however, there is debate on the effectiveness of opioids. Nevertheless, dose escalation is required to maintain their analgesia, which, in turn, contributes to a further increase in opioid side effects. Finding novel approaches to effectively control chronic pain, particularly neuropathic pain, is a great challenge clinically. Literature data related to pain transmission reveal that angiotensin and its receptors (the AT1R, AT2R, and MAS receptors) could affect the nociception both in the periphery and CNS. The MOR and angiotensin receptors or drugs interacting with these receptors have been independently investigated in relation to analgesia. However, the interaction between the MOR and angiotensin receptors has not been excessively studied in chronic pain, particularly neuropathy. This review aims to shed light on existing literature information in relation to the analgesic action of AT1R and AT2R or MASR ligands in neuropathic pain conditions. Finally, based on literature data, we can hypothesize that combining MOR agonists with AT1R or AT2R antagonists might improve analgesia.

Renin-Angiotensin System Induced Secondary Hypertension: The Alteration of Kidney Function and Structure

Long-term hypertension is known as a major risk factor for cardiovascular and chronic kidney disease (CKD). The Renin-angiotensin system (RAS) plays a key role in hypertension pathogenesis. Angiotensin II (Ang II) enhancement in Ang II-dependent hypertension leads to progressive CKD and kidney fibrosis. In the two-kidney one-clip model (2K1C), more renin is synthesized in the principal cells of the collecting duct than juxtaglomerular cells (JGCs). An increase of renal Ang I and Ang II levels and a decrease of renal cortical and medullary Ang 1–7 occur in both kidneys of the 2K1C hypertensive rat model. In addition, the activity of the angiotensin-converting enzyme (ACE) increases, while ACE2's activity decreases in the medullary region of both kidneys in the 2K1C hypertensive model. Also, the renal prolyl carboxypeptidase (PrCP) expression and its activity reduce in the clipped kidneys. The imbalance in the production of renal ACE, ACE2, and PrCP expression causes the progression of renal injury. Intrarenal angiotensinogen (AGT) expression and urine AGT (uAGT) excretion rates in the unclipped kidney are greater than the clipped kidney in the 2K1C hypertensive rat model. The enhancement of Ang II in the clipped kidney is related to renin secretion, while the elevation of intrarenal Ang II in the unclipped kidney is related to stimulation of AGT mRNA and protein in proximal tubule cells by a direct effect of systemic Ang II level. Ang II-dependent hypertension enhances macrophages and T-cell infiltration into the kidney which increases cytokines, and AGT synthesis in proximal tubules is stimulated via cytokines. Accumulation of inflammatory cells in the kidney aggravates hypertension and renal damage. Moreover, Ang II-dependent hypertension alters renal Ang II type 1 & 2 receptors (AT1R & AT2R) and Mas receptor (MasR) expression, and the renal interstitial fluid bradykinin, nitric oxide, and cGMP response to AT1R, AT2R, or BK B2-receptor antagonists. Based on a variety of sources including PubMed, Google Scholar, Scopus, and Science-Direct, in the current review, we will discuss the role of RAS-induced secondary hypertension on the alteration of renal function.

NHERF4 hijacks Mas-mediated PLC/AKT signaling to suppress the invasive potential of clear cell renal cell carcinoma cells.

The Mas receptor has been reported to promote migration and invasion of clear cell renal cell carcinoma (ccRCC) cells via Ang-(1-7)-dependent AKT signaling. However, the mechanism underlying the regulation of Mas function remains unknown. Here, eight PDZ domain-containing proteins were identified as Mas interactors using surface plasmon resonance (SPR) coupled to mass spectrometry (MS). NHERF4 was the only downregulated gene across multiple independent ccRCC datasets. GST pull-down and co-immunoprecipitation assays confirmed physical interaction between NHERF4 and Mas. Using NHERF4 overexpression and knockdown assays, we found that NHERF4 inhibited Mas-induced migration, invasion and in vivo metastasis of ccRCC cells. Mechanistically, NHERF4 suppressed Mas-stimulated AKT phosphorylation and the PLC/Ca2+ response. We further demonstrated that NHERF4 compromised Mas-mediated migration and invasion of ccRCC cells via regulation of the PLC/AKT signaling axis. Analysis of the ccRCC dataset revealed that low levels of NHERF4 expression were correlated with higher TNM stage, and independently predicted poor prognosis of ccRCC patients. Overall, our study identified NHERF4 as a novel regulator of ccRCC invasiveness, and a prognostic biomarker, which may be beneficial for determining optimal therapeutic strategies for ccRCC patients.

Activation of angiotensin-converting enzyme 2/angiotensin (1-7)/mas receptor axis triggers autophagy and suppresses microglia proinflammatory polarization via forkhead box class O1 signaling.

Brain renin‐angiotensin (Ang) system (RAS) is implicated in neuroinflammation, a major characteristic of aging process. Angiotensin (Ang) II, produced by angiotensin‐converting enzyme (ACE), activates immune system via angiotensin type 1 receptor (AT1), whereas Ang(1–7), generated by ACE2, binds with Mas receptor (MasR) to restrain excessive inflammatory response. Therefore, the present study aims to explore the relationship between RAS and neuroinflammation. We found that repeated lipopolysaccharide (LPS) treatment shifted the balance between ACE/Ang II/AT1 and ACE2/Ang(1–7)/MasR axis to the deleterious side and treatment with either MasR agonist, AVE0991 (AVE) or ACE2 activator, diminazene aceturate, exhibited strong neuroprotective actions. Mechanically, activation of ACE2/Ang(1–7)/MasR axis triggered the Forkhead box class O1 (FOXO1)‐autophagy pathway and induced superoxide dismutase (SOD) and catalase (CAT), the FOXO1‐targeted antioxidant enzymes. Meanwhile, knockdown of MasR or FOXO1 in BV2 cells, or using the selective FOXO1 inhibitor, AS1842856, in animals, suppressed FOXO1 translocation and compromised the autophagic process induced by MasR activation. We further used chloroquine (CQ) to block autophagy and showed that suppressing either FOXO1 or autophagy abrogated the anti‐inflammatory action of AVE. Likewise, Ang(1–7) also induced FOXO1 signaling and autophagic flux following LPS treatment in BV2 cells. Cotreatment with AS1842856 or CQ all led to autophagic inhibition and thereby abolished Ang(1–7)‐induced remission on NLRP3 inflammasome activation caused by LPS exposure, shifting the microglial polarization from M1 to M2 phenotype. Collectively, these results firstly illustrated the mechanism of ACE2/Ang(1–7)/MasR axis in neuroinflammation, strongly indicating the involvement of FOXO1‐mediated autophagy in the neuroimmune‐modulating effects triggered by MasR activation.

Local renin angiotensin system and sperm DNA fragmentation.

The renin angiotensin system (RAS) appears to influence male fertility at multiple levels. In this work, we analyzed the relationship between the RAS and DNA integrity. Fifty male volunteers were divided into two groups (25 each): control (DNA fragmentation ≤20%) and pathological (DNA fragmentation >20%) cases. Activities of five peptidases controlling RAS were measured fluorometrically: prolyl endopeptidase (which converts angiotensin [A] I and A II to A 1–7), neutral endopeptidase (NEP/CD10: A I to A 1–7), aminopeptidase N (APN/CD13: A III to A IV), aminopeptidase A (A II to A III) and aminopeptidase B (A III to A IV). Angiotensin-converting enzyme (A I to A II), APN/CD13 and NEP/CD10 were also assessed by semiquantitative cytometry and quantitative flow cytometry assays, as were the receptors of all RAS components: A II receptor type 1 (AT1R), A II receptor type 2 (AT2R), A IV receptor (AT4R or insulin-regulated aminopeptidase [IRAP]), (pro)renin receptor (PRR) and A 1–7 receptor or Mas receptor (MasR) None of the enzymes that regulate levels of RAS components, except for APN/CD13 (decrease in fragmented cells), showed significant differences between both groups. Micrographs of RAS receptors revealed no significant differences in immunolabeling patterns between normozoospermic and fragmented cells. Labeling of AT1R (94.3% normozoospermic vs 84.1% fragmented), AT4R (96.2% vs 95.3%) and MasR (97.4% vs 87.2%) was similar between the groups. AT2R (87.4% normozoospermic vs 63.1% fragmented) and PRR (96.4% vs 48.2%) were higher in non-fragmented spermatozoa. These findings suggest that fragmented DNA spermatozoa have a lower capacity to respond to bioactive RAS peptides.

Abstract P394: Angiotensin-(1-7) Attenuates Diastolic Dysfunction And Cardiac Remodeling In Murine Model Of Heart Failure With Preserved Ejection Fraction

Background: Heart failure with preserved ejection fraction (HFpEF) is a global public health epidemic that accounts for half of the heart failure cases. Various therapeutic approaches have been tested to block the activation of the Renin-Angiotensin System (RAS), including AT1R blockers (ARBs), Angiotensin-Converting Enzyme (ACE) inhibitors (ACEi), and direct renin inhibitors (DRIs) with modest to negligible benefits. The discovery of ACE2, a novel homolog of ACE, has advanced our understanding of the RAS. ACE2 is a monocarboxypeptidase that degrades Ang II into Ang-(1-7), which works via the activation of the Mas receptor. It has been well understood that the actions of Ang-(1-7) attenuate cardiac remodeling, production of ROS, and cardiac fibrosis. Objective: To determine the therapeutic role of Ang-(1-7) in HFpEF and identify the molecular mechanism related to its action. Methods and Results: To generate a murine model of HFpEF, male WT mice (n=24) were subjected to HFD in addition to eNOS inhibition with L-NAME (0.5 g l-1 in drinking water), as previously described. The control group (n=12) received chow diet and normal tap water. The murine model of HFpEF was validated using the non-invasive transthoracic echocardiography and invasive pressure-volume (PV) loop analyses, which exhibited diastolic dysfunction as well as cardiac hypertrophy. To evaluate the effects of Ang-(1-7) on HFpEF, animals were administered with either saline (n=12) or Ang-(1-7) (n=12) (24 μg/kg/day) for four weeks. Ang-(1-7) treatment improved diastolic function by reducing LVEDP (Ctrl: 8.267±1.254; HFD+L-NAME: 17.64±1.925; Ang-(1-7): 9.100±1.578) and Tau value (Ctrl: 7.365±0.5752; HFD+L-NAME: 9.224±0.3569; Ang-(1-7): 7.381±0.3041). Furthermore, Ang-(1-7) reduced cardiac hypertrophy by reducing the phosphorylation level of MAPK ERK 1/2 (Ctrl: 0.9074±0.1088; HFD+L-NAME: 1.212±0.1369; Ang-(1-7): 0.5615±0.1502) and increasing the phosphorylation level of AMPK (Ctrl: 0.1502±0.1502; HFD+L-NAME: 0.6127±0.06414; Ang-(1-7): 0.7852±0.1006). Ang-(1-7) treatment also reduces cardiomyocytes’ size and decreases interstitial fibrosis, as indicated by WGA and PSR staining. Conclusion: Ang-(1-7) treatment attenuated the development of HFD+L-NAME-induced HFpEF, reduced cardiac hypertrophy, and improved metabolic function.

Abstract MP67: Differential Effects Of Selective Renal Denervation On Intrarenal Neurohormone System Involved In Blood Pressure Regulation

Aims: To investigate alterations of intrarenal neurohormonal system involved in BP regulation after renal nerve stimulation (RNS)-guided renal denervation (RDN), and compare the significance of ablation on different BP-elevated magnitude sites. Methods and Results: Twenty-one dogs that met inclusion criteria were randomized to receive RDN at strong (SRA group, n=7) or weak (WRA group, n=7) BP-elevation response sites guided by RNS, or underwent RNS only (RNS-control, RSC, n=7), and fully recovered from the acute experiments. After 4 weeks, renal sympathetic and renin angiotensin system activity parameters, and major transporter expression were assessed by immunology and histochemistry. Compared with RSC group, RDN therapy significantly attenuated renal norepinephrine and tyrosine hydroxylase levels, alleviated renin release, and inhibited onsite generation of angiotensinogen. Meanwhile, the expression of exciting axis components, including angiotensin-converting enzyme (ACE), angiotensin II and angiotensin II type-1 receptor, were down-regulated, while the protective axis components such as ACE2 and Mas receptors, were up-regulated in both WRA and SRA group. Moreover, RDN reduced the abundance of aquaporin-1 and aquaporin-2 in kidneys. Although RDN had a minimal effect on overall NKCC2 expression, its activation (p-NKCC2) and directional enrichment in the apical membrane (mNKCC2) were dramatically blunted. Further analysis showed that all these changes were more significant in SRA group than WRA group. Conclusions: RDN effectively reduced systemic BP by affecting renal neurohormone-receptor axis, as well as expression and/or activation of major transporters. Selective RDN at strong BP-response sites presented a more superior beneficial effect, reconfirmed the feasibility of RNS-guided RDN.