RESUMEN
Alamandine is a peptide hormone belonging to the renin-angiotensin system (RAS). It acts through the Mas-related G-protein coupled receptor type D, MrgD, which is expressed in different tissues, including the brain. In the present study, we hypothesize that a lack of alamandine, through MrgD, could cause the anxiety-like behavior in transgenic rats with low brain angiotensinogen [TGR(ASrAOGEN)680]. Adult male transgenic rats exhibited a significant increase in the latency to feeding time in the novelty suppressed feeding test and a decrease in the percentage of time and entries in the open arms in the elevated plus maze. These effects were reversed by intracerebroventricular infusion of alamandine. Pretreatment with D-Pro7-Ang-(1-7), a Mas and MrgD receptor antagonist, prevented the anxiolytic effects induced by this peptide. However, its effects were not altered by the selective Mas receptor antagonist, A779. In conclusion, our data indicates that alamandine, through MrgD, attenuates anxiety-like behavior in male TGR(ASrAOGEN)680, which reinforces the importance of the counter-regulatory RAS axis as promising target for the treatment of neuropsychiatric disorders.
Asunto(s)
Angiotensinógeno , Ansiolíticos , Ansiedad , Encéfalo , Ratas Transgénicas , Receptores Acoplados a Proteínas G , Animales , Masculino , Receptores Acoplados a Proteínas G/metabolismo , Receptores Acoplados a Proteínas G/genética , Ratas , Ansiedad/tratamiento farmacológico , Ansiedad/metabolismo , Ansiolíticos/farmacología , Angiotensinógeno/metabolismo , Angiotensinógeno/genética , Encéfalo/metabolismo , Encéfalo/efectos de los fármacos , Receptores de la Hormona Gastrointestinal/metabolismo , Oligopéptidos/farmacología , Proteínas del Tejido NerviosoRESUMEN
OBJECTIVES: To evaluate the impact of genetic deletion of receptors of the counterregulatory arms of the renin-angiotensin system in depressive-like behaviours. METHODS: 8-12 weeks-old male mice wild type (WT, C57BL/6J) and mice with genetic deletion of MrgD (MrgD KO) or Mas receptors (Mas KO) were subjected to the Forced Swim Test (FST) and the Tail Suspension Test (TST). Brain-derived neurotrophic factor (BDNF) levels were measured by enzyme-linked immunosorbent assay (ELISA). Blockade of Mas was performed by acute intracerebroventricular (icv) injection of its selective antagonist, A779. RESULTS: No statistical difference in immobility time was observed between MrgD KO and WT male animals subjected to FST and TST. However, acute icv injection of A779 significantly increased the immobility time of MrgD KO male mice subjected to FST and TST, suggesting the involvement of Mas in preventing depressive-like behaviour. Indeed, Mas KO male animals showed increased immobility time in FST and TST, evidencing a depressive-like behaviour in these animals, in addition to a reduction in BDNF levels in the prefrontal cortex and hippocampus. No changes in BDNF levels were observed in MrgD KO male animals. CONCLUSION: Our data showed that Mas plays an important role in the neurobiology of depression probably by modulating BDNF expression. On the contrary, lack of MrgD did not alter depressive-like behaviour, which was supported by the lack of alterations in BDNF levels.
Asunto(s)
Factor Neurotrófico Derivado del Encéfalo , Depresión , Ratones , Masculino , Animales , Depresión/genética , Depresión/metabolismo , Factor Neurotrófico Derivado del Encéfalo/genética , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Ratones Endogámicos C57BL , Suspensión Trasera , Corteza Prefrontal/metabolismo , Hipocampo/metabolismoRESUMEN
Clinical studies have shown a correlation between thyroid disorders and cardiac diseases. High levels of triiodothyronine (T3) induce cardiac hypertrophy, a risk factor for cardiac complications and heart failure. Previous results have demonstrated that angiotensin-(1-7) is able to block T3-induced cardiac hypertrophy; however, the molecular mechanisms involved in this event have not been fully elucidated. Here, we evidenced the contribution of FOXO3 signaling to angiotensin-(1-7) effects. Angiotensin-(1-7) treatment increased nuclear FOXO3 levels and reduced p-FOXO3 levels (inactive form) in isolated cardiomyocytes. Knockdown of FOXO3 by RNA silencing abrogated the antihypertrophic effect of angiotensin-(1-7). Increased expression of antioxidant enzymes superoxide dismutase 1 (SOD1 and catalase) and lower levels of reactive oxygen species and nuclear factor-κB (NF-κB) were observed after angiotensin-(1-7) treatment in vitro. Consistent with these results, transgenic rats overexpressing angiotensin-(1-7) displayed increased nuclear FOXO3 and SOD1 levels and reduced NF-κB levels in the heart. These results provide a new molecular mechanism responsible for the antihypertrophic effect of angiotensin-(1-7), which may contribute to future therapeutic targets.
Asunto(s)
Angiotensina I/farmacología , Catalasa/metabolismo , Proteína Forkhead Box O3/metabolismo , Miocitos Cardíacos/patología , FN-kappa B/metabolismo , Fragmentos de Péptidos/farmacología , Superóxido Dismutasa-1/metabolismo , Triyodotironina/efectos adversos , Regulación hacia Arriba , Animales , Antioxidantes/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Hipertrofia , Masculino , Modelos Biológicos , Miocitos Cardíacos/efectos de los fármacos , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas/metabolismo , Ratas Sprague-Dawley , Ratas Transgénicas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Regulación hacia Arriba/efectos de los fármacosRESUMEN
Activation of the angiotensin (Ang)-converting enzyme (ACE) 2/Ang-(1-7)/MAS receptor pathway of the renin-angiotensin system (RAS) induces protective mechanisms in different diseases. Herein, we describe the cardiovascular phenotype of a new transgenic rat line (TG7371) that expresses an Ang-(1-7)-producing fusion protein. The transgene-specific mRNA and the corresponding protein were shown to be present in all evaluated tissues of TG7371 with the highest expression in aorta and brain. Plasma Ang-(1-7) levels, measured by radioimmunoassay (RIA) were similar to control Sprague-Dawley (SD) rats, however high Ang-(1-7) levels were found in the hypothalamus. TG7371 showed lower baseline mean arterial pressure (MAP), assessed in conscious or anesthetized rats by telemetry or short-term recordings, associated with increased plasma atrial natriuretic peptide (ANP) and higher urinary sodium concentration. Moreover, evaluation of regional blood flow and hemodynamic parameters with fluorescent microspheres showed a significant increase in blood flow in different tissues (kidneys, mesentery, muscle, spleen, brown fat, heart and skin), with a resulting decrease in total peripheral resistance (TPR). TG7371 rats, on the other hand, also presented increased cardiac and global sympathetic tone, increased plasma vasopressin (AVP) levels and decreased free water clearance. Altogether, our data show that expression of an Ang-(1-7)-producing fusion protein induced a hypotensive phenotype due to widespread vasodilation and consequent fall in peripheral resistance. This phenotype was associated with an increase in ANP together with an increase in AVP and sympathetic drive, which did not fully compensate the lower blood pressure (BP). Here we present the hemodynamic impact of long-term increase in tissue expression of an Ang-(1-7)-fusion protein and provide a new tool to investigate this peptide in different pathophysiological conditions.
Asunto(s)
Angiotensina I/metabolismo , Sistema Cardiovascular/metabolismo , Hemodinámica , Hipertensión/prevención & control , Fragmentos de Péptidos/metabolismo , Sistema Nervioso Simpático/metabolismo , Angiotensina I/genética , Animales , Arginina Vasopresina/metabolismo , Factor Natriurético Atrial/metabolismo , Velocidad del Flujo Sanguíneo , Presión Sanguínea , Sistema Cardiovascular/fisiopatología , Modelos Animales de Enfermedad , Genotipo , Proteína Ácida Fibrilar de la Glía/genética , Proteína Ácida Fibrilar de la Glía/metabolismo , Hemodinámica/genética , Hipertensión/genética , Hipertensión/metabolismo , Hipertensión/fisiopatología , Masculino , Fragmentos de Péptidos/genética , Fenotipo , Ratas Sprague-Dawley , Ratas Transgénicas , Proteínas Recombinantes de Fusión/metabolismo , Flujo Sanguíneo Regional , Sistema Nervioso Simpático/fisiopatología , Factores de Tiempo , Resistencia VascularRESUMEN
Alamandine (Ala1-Arg2-Val3-Tyr4-Ile5-His6-Pro7), a heptapeptide hormone of the renin-angiotensin system (RAS), exerts its effects through the Mas-related G-protein coupled receptor of the type D, MrgD, which is expressed in different tissues, including the brain. In the present study, we tested the hypothesis that alamandine could attenuate the depression-like behavior observed in transgenic rats with low brain angiotensinogen, TGR (ASrAOGEN)680. Transgenic rats exhibited a significant increase in the immobility time in forced swim test, a phenotype reversed by intracerebroventricular infusion of alamandine. Pretreatment with D-Pro7-Ang-(1-7), a Mas/MrgD receptor antagonist, prevented the antidepressant-like effect induced by this peptide demonstrating, for the first time, that alamandine through MrgD receptor, can modulate depression-like behavior in TGR (ASrAOGEN)680. This result shows an action of alamandine which strengthens the importance of the counter-regulatory arms of the RAS in fight and treatment of neuropsychiatric diseases.
Asunto(s)
Angiotensinógeno/genética , Antidepresivos/farmacología , Encéfalo/efectos de los fármacos , Proteínas del Tejido Nervioso/fisiología , Oligopéptidos/farmacología , Receptores Acoplados a Proteínas G/fisiología , Angiotensina I/farmacología , Angiotensinógeno/metabolismo , Animales , Encéfalo/metabolismo , Inyecciones Intraventriculares , Masculino , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Oligopéptidos/administración & dosificación , Fragmentos de Péptidos/farmacología , Ratas , Ratas Sprague-Dawley , Ratas Transgénicas , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/metabolismoRESUMEN
Resolution failure of exacerbated inflammation triggered by Influenza A virus (IAV) prevents return of pulmonary homeostasis and survival, especially when associated with secondary pneumococcal infection. Therapeutic strategies based on pro-resolving molecules have great potential against acute inflammatory diseases. Angiotensin-(1-7) [Ang-(1-7)] is a pro-resolving mediator that acts on its Mas receptor (MasR) to promote resolution of inflammation. We investigated the effects of Ang-(1-7) and the role of MasR in the context of primary IAV infection and secondary pneumococcal infection and evaluated pulmonary inflammation, virus titers and bacteria counts, and pulmonary damage. Therapeutic treatment with Ang-(1-7) decreased neutrophil recruitment, lung injury, viral load and morbidity after a primary IAV infection. Ang-(1-7) induced apoptosis of neutrophils and efferocytosis of these cells by alveolar macrophages, but had no direct effect on IAV replication in vitro. MasR-deficient (MasR-/-) mice were highly susceptible to IAV infection, displaying uncontrolled inflammation, increased viral load and greater lethality rate, as compared to WT animals. Ang-(1-7) was not protective in MasR-/- mice. Interestingly, Ang-(1-7) given during a sublethal dose of IAV infection greatly reduced morbidity associated with a subsequent S. pneumoniae infection, as seen by decrease in the magnitude of neutrophil influx, number of bacteria in the blood leading to a lower lethality. Altogether, these results show that Ang-(1-7) is highly protective against severe primary IAV infection and protects against secondary bacterial infection of the lung. These effects are MasR-dependent. Mediators of resolution of inflammation, such as Ang-(1-7), should be considered for the treatment of pulmonary viral infections.
Asunto(s)
Angiotensina I/uso terapéutico , Antiinflamatorios/uso terapéutico , Fragmentos de Péptidos/uso terapéutico , Infecciones Neumocócicas/tratamiento farmacológico , Neumonía Viral/tratamiento farmacológico , Proteínas Proto-Oncogénicas/inmunología , Receptores Acoplados a Proteínas G/inmunología , Células A549 , Angiotensina I/farmacología , Animales , Antiinflamatorios/farmacología , Líquido del Lavado Bronquioalveolar/citología , Líquido del Lavado Bronquioalveolar/inmunología , Citocinas/inmunología , Perros , Humanos , Virus de la Influenza A , Pulmón/efectos de los fármacos , Pulmón/inmunología , Pulmón/patología , Células de Riñón Canino Madin Darby , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Neutrófilos/efectos de los fármacos , Neutrófilos/inmunología , Fragmentos de Péptidos/farmacología , Peroxidasa/inmunología , Fagocitosis/efectos de los fármacos , Infecciones Neumocócicas/inmunología , Infecciones Neumocócicas/patología , Neumonía Viral/inmunología , Neumonía Viral/patología , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas/genética , Receptores Acoplados a Proteínas G/genética , Streptococcus pneumoniaeRESUMEN
BACKGROUND: The renin-angiotensin system (RAS) plays an important role in erectile function. The RAS contains 2 major axes: one deleterious, composed of ACE-Ang II-AT1 receptor, and another protective, composed of ACE2-Ang-(1-7)-Mas receptor. While aging is a well-known cause for development of male sexual disorders, little is known about local regulation of the RAS in age-related erectile dysfunction (ED). AIM: The present study aimed to assess regulation of the RAS in aging-associated ED rat model and evaluate possible options for disease management through pharmacological modulation of the RAS. METHODS: Penile tissues were harvested from 3-, 12-, and 24-month-old Wistar rats. Local expression of major RAS components and ED markers was measured by RT-PCR. Protein expression of RAS components was assessed by western blot. Collagen deposition was measured by Sirius Red and immunohistochemical staining. Evaluation of collagen content was also performed in penile sections of Mas-knockout mice by Sirius Red and Masson's trichrome stainings. Finally, the effect of Ang-(1-7) pretreatment on TGF-ß-induced myofibroblast activation was studied in primary cavernosal and immortalized fibroblasts. OUTCOMES: Experimental results highlighted the essential role of the RAS in modulation of cavernosal fibrosis. RESULTS: The present study demonstrates local expression of angiotensinogen mRNA alongside with major RAS components, which suggests local autonomous functioning of the RAS within penile tissue. Gene expression analysis revealed strong positive correlation between ACE-Ang II-AT1 axis with markers for inflammation and fibrosis. While corpus cavernosum from 24-month-old rats was characterized by increased collagen deposition, protein expression of ACE, AT1, and Mas was shown to be upregulated in the penile tissue of this group. At the same time, penile sections from Mas-knockout mice (FVB/N background) were also shown to have increased collagen deposition. Finally, it was demonstrated that Ang-(1-7) treatment of primary cavernosal and immortalized fibroblasts was able to alleviate TGF-ß-induced fibroblast-to-myofibroblast transition. CLINICAL TRANSLATION: The present study suggests Ang-(1-7) treatment as a possible strategy for pharmacological management of fibrosis-associated ED in aging. STRENGTHS & LIMITATIONS: The link between the RAS and penile fibrosis, indicated by a holistic screening of different ED markers, was confirmed by in vivo and in vitro data. However, results, presented in the manuscript, need to be further reinforced by human data. Important to note, the main goal of the study was to characterize RAS regulation in aging condition rather than state any causal relationships. CONCLUSION: Present study characterizes RAS regulation in aging-associated ED and indicates its important role in cavernosal fibrosis. Bragina ME, Costa-Fraga F, Sturny M, et al. Characterization of the Renin-Angiotensin System in Aged Cavernosal Tissue and its Role in Penile Fibrosis. J Sex Med 2020;17:2129-2140.
Asunto(s)
Induración Peniana , Sistema Renina-Angiotensina , Anciano , Angiotensina I , Angiotensina II/metabolismo , Animales , Fibrosis , Humanos , Masculino , Ratones , Erección Peniana , Fragmentos de Péptidos , Peptidil-Dipeptidasa A/metabolismo , Ratas , Ratas WistarRESUMEN
Skeletal muscle atrophy, which occurs in lipopolysaccharide (LPS)-induced sepsis, causes a severe muscle function reduction. The increased autophagy contributes to sepsis-induced skeletal muscle atrophy in a model of LPS injection, increasing LC3II/LC3I ratio, autophagy flux, and autophagosomes. Angiotensin-(1-7) (Ang-(1-7)) has anti-atrophic effects via the Mas receptor in skeletal muscle. However, the impact of Ang-(1-7) on LPS-induced autophagy is unknown. In this study, we determined the effect of Ang-(1-7) on sepsis-induced muscle autophagy. C57BL6 wild-type (WT) mice and mice lacking the Mas receptor (KO Mas) were injected with LPS together with the systemic administration of Ang-(1-7) to determine autophagy in skeletal muscle. We also evaluated autophagy and p38 and c-Jun N-terminal kinase (JNK)activation. Our results show that Ang-(1-7) prevents LPS-induced autophagy in the diaphragm, tibialis anterior, and gastrocnemius of WT mice, which is demonstrated by a decrease in the LC3II/LC3I ratio and mRNA levels of lc3b and ctsl. This effect was lost in KO Mas mice, suggesting the role of the Mas receptor. The results in C2C12 cells show that Ang-(1-7) reduces several LPS-dependent effects, such as autophagy (LC3II/LC3I ratio, autophagic flux, and autophagosomes), activation of p38 and JNK, B-cell lymphoma-2 (BCL2) phosphorylation, and disassembly of the Beclin1/BCL2 complex. In conclusion, Ang-(1-7)/Mas receptor reduces LPS-induced autophagy in skeletal muscle. In vitro assays indicate that Ang-(1-7) prevents LPS-induced autophagy and modifies the MAPK signaling and the disassembly of a complex involved at the beginning of autophagy.
Asunto(s)
Angiotensina I/farmacología , Autofagia , Músculo Esquelético/metabolismo , Fragmentos de Péptidos/farmacología , Proteínas Proto-Oncogénicas/metabolismo , Receptores Acoplados a Proteínas G/metabolismo , Animales , Catepsina L/metabolismo , Línea Celular , Lipopolisacáridos/farmacología , MAP Quinasa Quinasa 4/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Asociadas a Microtúbulos/metabolismo , Músculo Esquelético/efectos de los fármacos , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Receptores Acoplados a Proteínas G/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
We have recently described a new peptide of the renin-angiotensin system, alamandine, a derivative of angiotensin-(1-7). Mas-related G protein-coupled receptor member D (MrgD) was identified as its receptor. Although similar cardioprotective effects of alamandine to those of angiotensin-(1-7) have been described, the significance of this peptide in heart function is still elusive. We aimed to evaluate the functional role of the alamandine receptor MrgD in the heart using MrgD-deficient mice. MrgD was localized in cardiomyocytes by immunofluorescence using confocal microscopy. High-resolution echocardiography was performed in wild-type and MrgD-deficient mice (2 and 12 wk old) under isoflurane anesthesia. Standard B-mode images were obtained in the right and left parasternal long and short axes for morphological and functional assessment and evaluation of cardiac deformation. Additional heart function evaluation was performed using Langendorff isolated heart preparations and inotropic measurements of isolated cardiomyocytes. Immunofluorescence indicated that the MrgD receptor is expressed in cardiomyocytes, mainly in the membrane and perinuclear and nuclear regions. Echocardiography showed left ventricular remodeling and severe dysfunction in MrgD-deficient mice. Strikingly, MrgD-deficient mice presented a pronounced dilated cardiomyopathy with a marked decrease in systolic function. Echocardiographic changes were supported by the data obtained in isolated hearts and inotropic measurements in cardiomyocytes. Our data add new evidence for a major role for alamandine/MrgD in the heart. Furthermore, our results indicate that we have identified a new gene implicated in dilated cardiomyopathy, unveiling a new target for translational approaches aimed to treat heart diseases. NEW & NOTEWORTHY The renin-angiotensin system is a key target for cardiovascular therapy. We have recently identified a new vasodepressor/cardioprotective angiotensin, alamandine. Here, we unmasked a key role for its receptor, Mas-related G protein-coupled receptor member D (MrgD), in heart function. The severe dilated cardiomyopathy observed in MrgD-deficient mice warrants clinical and preclinical studies to unveil its potential use in cardiovascular therapy.
Asunto(s)
Cardiomiopatía Dilatada/genética , Eliminación de Gen , Receptores Acoplados a Proteínas G/genética , Animales , Cardiomiopatía Dilatada/metabolismo , Cardiomiopatía Dilatada/patología , Células Cultivadas , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Contracción Miocárdica , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Receptores Acoplados a Proteínas G/metabolismo , Remodelación VentricularRESUMEN
Physical exercise induces cell proliferation in the adult hippocampus in rodents. Serotonin (5-HT) and angiotensin (Ang) II are important mediators of the pro-mitotic effect of physical activity. Here, we examine precursor cells in the adult brain of mice lacking angiotensin-converting enzyme (ACE) 2, and explore the effect of an acute running stimulus on neurogenesis. ACE2 metabolizes Ang II to Ang-(1-7) and is essential for the intestinal uptake of tryptophan (Trp), the 5-HT precursor. In ACE2-deficient mice, we observed a decrease in brain 5-HT levels and no increase in the number of BrdU-positive cells following exercise. Targeting the Ang II/AT1 axis by blocking the receptor, or experimentally increasing Trp/5-HT levels in the brain of ACE2-deficient mice, did not rescue the running-induced effect. Furthermore, mice lacking the Ang-(1-7) receptor, Mas, presented a normal neurogenic response to exercise. Our results identify ACE2 as a novel factor required for exercise-dependent modulation of adult neurogenesis and essential for 5-HT metabolism.
Asunto(s)
Encéfalo/metabolismo , Neurogénesis/genética , Peptidil-Dipeptidasa A/genética , Condicionamiento Físico Animal/fisiología , Carrera/fisiología , Serotonina/metabolismo , Células Madre Adultas/fisiología , Enzima Convertidora de Angiotensina 2 , Animales , Diferenciación Celular/genética , Proliferación Celular/genética , Regulación hacia Abajo/genética , Femenino , Eliminación de Gen , Redes y Vías Metabólicas/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Células-Madre Neurales/fisiología , Peptidil-Dipeptidasa A/fisiologíaRESUMEN
The renin-angiotensin system (RAS) plays a pivotal role in the pathogenesis of cardiovascular diseases. New members of this system have been characterized and shown to have biologically relevant actions. Alamandine and its receptor MrgD are recently identified components of RAS. In the cardiovascular system, alamandine actions included vasodilation, antihypertensive, and antifibrosis effects. Currently, the actions of alamandine on cardiomyocytes are unknown. Here our goal was twofold: 1) to unravel the signaling molecules activated by the alamandine/MrgD axis in cardiomyocytes; and 2) to evaluate the ability of this axis to prevent angiotensin II (ANG II)-induced hypertrophy. In cardiomyocytes from C57BL/6 mice, alamandine treatment induced an increase in nitric oxide (NO) production, which was blocked by d-Pro7-ANG-(1-7), a MrgD antagonist. This NO rise correlated with increased phosphorylation of AMPK. Alamandine-induced NO production was preserved in Mas-/- myocytes and lost in MrgD-/- cells. Binding of fluorescent-labeled alamandine was observed in wild-type cells, but it was dramatically reduced in MrgD-/- myocytes. We also assessed the consequences of prolonged alamandine exposure to cultured neonatal rat cardiomyocytes (NRCMs) treated with ANG II. Treatment of NRCMs with alamandine prevented ANG II-induced hypertrophy. Moreover, the antihypertrophic actions of alamandine were mediated via MrgD and NO, since they could be prevented by d-Pro7-ANG-(1-7) or inhibitors of NO synthase or AMPK. ß-Alanine, a MrgD agonist, recapitulated alamandine's cardioprotective effects in cardiomyocytes. Our data show that alamandine via MrgD induces AMPK/NO signaling to counterregulate ANG II-induced hypertrophy. These findings highlight the therapeutic potential of the alamandine/MrgD axis in the heart.
Asunto(s)
Proteínas Quinasas Activadas por AMP/metabolismo , Angiotensina II/toxicidad , Cardiomegalia/prevención & control , Miocitos Cardíacos/efectos de los fármacos , Proteínas del Tejido Nervioso/agonistas , Óxido Nítrico/metabolismo , Oligopéptidos/farmacología , Receptores Acoplados a Proteínas G/agonistas , Animales , Cardiomegalia/inducido químicamente , Cardiomegalia/enzimología , Cardiomegalia/patología , Células Cultivadas , Activación Enzimática , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Miocitos Cardíacos/enzimología , Miocitos Cardíacos/patología , Proteínas del Tejido Nervioso/metabolismo , Oligopéptidos/metabolismo , Fosforilación , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas/agonistas , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas/metabolismo , Ratas Wistar , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
BACKGROUND: Apelin is an endogenous peptidergic system which modulates cardiovascular function. Recent studies pointed out a fundamental contribution of apelin on atherosclerosis development; however, such reports revealed contradictory data, and to date, it is difficult to accurately define a beneficial or deleterious role. To better understand apelin function on atherosclerosis, we aimed to investigate apelin-13 treatment effects on atherosclerotic plaques composition. DESIGN: Apolipoprotein E gene-deleted mice were fed on Western-type diet for 11 weeks. Atherosclerotic plaque formation was induced in the carotid artery by a shear stress modifier device, which exposes the same vessel to distinct patterns of shear stress enabling the formation of plaques with different composition. Mice were treated with apelin-13 (2 mg kg-1 day-1 ) or vehicle for the last 3 weeks. RESULTS: Apelin-13 treatment did not alter the lipid content of low shear stress- and oscillatory shear stress-induced plaques in the carotid. However, apelin-13 greatly ameliorated plaque stability by increasing intraplaque collagen content and reducing MMP-9 expression. Furthermore, apelin-13 decreased the infiltration of inflammatory cells (neutrophil and macrophage) and intraplaque reactive oxygen species content. Interestingly, apelin-13 treatment reduced total cholesterol, LDL levels and free fatty acid serum levels, while HDL, triglycerides serum levels were not significantly changed. CONCLUSIONS: Apelin-13 treatment for 3 weeks did not alter the lesion size, but it significantly enhanced the stable phenotype of atherosclerotic plaques and improved serum lipid profile. These results indicate that activation of apelin system decreases plaque vulnerability.
Asunto(s)
Apelina/farmacología , Enfermedades de las Arterias Carótidas/fisiopatología , Placa Aterosclerótica/fisiopatología , Animales , Enfermedades de las Arterias Carótidas/metabolismo , Colágeno/metabolismo , Dieta Occidental , Metabolismo de los Lípidos/efectos de los fármacos , Lípidos/sangre , Macrófagos/efectos de los fármacos , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados para ApoE , Placa Aterosclerótica/metabolismo , Distribución Aleatoria , Especies Reactivas de Oxígeno/metabolismoRESUMEN
The Angiotensin II type 2 receptor (AT2R) promotes vasodilation by nitric oxide (NO) release from endothelial cells. However, the mechanisms underlying the AT2R-induced stimulation of endothelial NO synthase (eNOS) is still not completely understood. Therefore, we investigated whether in addition to the known AT2R-mediated phosphorylation of eNOS at Ser1177, activation of phosphatases and dephosphorylation of eNOS at Tyr657 and Thr495 are also involved. Human aortic endothelial cells (HAEC) were stimulated with the AT2R-agonist Compound 21 (C21) (1 µM) in the presence or absence of either PD123319 (10 µM; AT2R antagonist), l-NG-Nitroarginine methyl ester (l-NAME) (10 µM; eNOS inhibitor), MK-2206 (100 nM; protein kinase B (Akt) inhibitor) sodium fluoride (NaF) (1 nM; serine/threonine phosphatase inhibitor) or sodium orthovanadate (Na3VO4) (10 nM; tyrosine phosphatase inhibitor). NO release was estimated by quantifying 4-amino-5-methylamino-2',7'-difluorofluorescein diacetate (DAF-FM) fluorescence. The phosphorylation status of activating (eNOS-Ser1177) or inhibitory eNOS residues (eNOS-Tyr657, eNOS-Thr495) was determined by Western blotting. Phosphorylation of Akt at Ser473 was measured to estimate Akt activity. AT2R stimulation significantly increased NO release from HAEC, which was blocked by PD123319, l-NAME and both phosphatase inhibitors. Intracellular calcium transients were not changed by C21. AT2R stimulation resulted in phosphorylation of eNOS-Ser1177 and dephosphorylation of eNOS-Tyr657 and eNOS-Thr495 Phosphorylation at eNOS-Ser1177 was prevented by inhibition of Akt with MK-2206. From these data, we conclude that AT2R stimulation in human endothelial cells increases eNOS activity through phosphorylation of activating eNOS residues (eNOS-Ser1177) by Akt, and through dephosphorylation of inactivating eNOS residues (eNOS-Tyr657, eNOS-Thr495) by serine/threonine and tyrosine phosphatases, thus increasing NO release.
Asunto(s)
Óxido Nítrico Sintasa de Tipo III/metabolismo , Receptor de Angiotensina Tipo 2/metabolismo , Células Cultivadas , Activación Enzimática , Humanos , Fosforilación , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Receptor de Angiotensina Tipo 2/agonistas , Sulfonamidas , TiofenosRESUMEN
NEW FINDINGS: What is the central question of this study? Angiotensin-(1-7) decreases cerebral infarct volume and improves neurological function when delivered centrally before and during ischaemic stroke. Here, we assessed the neuroprotective effects of angiotensin-(1-7) when delivered orally post-stroke. What is the main finding and its importance? We show that oral delivery of angiotensin-(1-7) attenuates cerebral damage induced by middle cerebral artery occlusion in rats, without affecting blood pressure or cerebral blood flow. Importantly, these treatments begin post-stroke at times coincident with the treatment window for tissue plasminogen activator, providing supporting evidence for clinical translation of this new therapeutic strategy. ABSTRACT: As a target for stroke therapies, the angiotensin-converting enzyme 2-angiotensin-(1-7)-Mas [ACE2/Ang-(1-7)/Mas] axis of the renin-angiotensin system can be activated chronically to induce neuroprotective effects, in opposition to the deleterious effects of angiotensin II via its type 1 receptor. However, more clinically relevant treatment protocols with Ang-(1-7) that involve its systemic administration beginning after the onset of ischaemia have not been tested. In this study, we tested systemic post-stroke treatments using a molecule where Ang-(1-7) is included within hydroxypropyl-ß-cyclodextrin [HPßCD-Ang-(1-7)] as an orally bioavailable treatment. In three separate protocols, HPßCD-Ang-(1-7) was administered orally to Sprague-Dawley rats after induction of ischaemic stroke by endothelin-1-induced middle cerebral artery occlusion: (i) to assess its effects on cerebral damage and behavioural deficits; (ii) to determine its effects on cardiovascular parameters; and (iii) to determine whether it altered cerebral blood flow. The results indicate that post-stroke oral administration of HPßCD-Ang-(1-7) resulted in 25% reductions in cerebral infarct volumes and improvement in neurological functions (P < 0.05), without inducing any alterations in blood pressure, heart rate or cerebral blood flow. In conclusion, Ang-(1-7) treatment using an oral formulation after the onset of ischaemia induces significant neuroprotection in stroke and might represent a viable approach for taking advantage of the protective ACE2/Ang-(1-7)/Mas axis in this disease.
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Angiotensina I/farmacología , Neuroprotección/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Fragmentos de Péptidos/farmacología , Accidente Cerebrovascular/tratamiento farmacológico , 2-Hidroxipropil-beta-Ciclodextrina/farmacología , Animales , Presión Sanguínea/efectos de los fármacos , Circulación Cerebrovascular/efectos de los fármacos , Endotelina-1/metabolismo , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Infarto de la Arteria Cerebral Media/metabolismo , Masculino , Ratas , Ratas Sprague-Dawley , Sistema Renina-Angiotensina/efectos de los fármacos , Accidente Cerebrovascular/metabolismoRESUMEN
Losartan (Los), a non-peptidic orally active agent, reduces arterial pressure through specific and selective blockade of angiotensin II receptor AT1. However, this widely used AT1 antagonist presents low bioavailability and needs once or twice a day dosage. In order to improve its bioavailability, we used the host: guest strategy based on ß-cyclodextrin (ßCD). The results suggest that Los included in ßCD showed a typical pulsatile release pattern after oral administration to rats, with increasing the levels of plasma of Los. In addition, the inclusion compound presented oral efficacy for 72 h, in contrast to Los alone, which shows antagonist effect for only 6 h. In transgenic (mREN2)L27 rats, the Los/ßCD complex reduced blood pressure for about 6 d, whereas Los alone reduced blood pressure for only 2 d. More importantly, using this host: guest strategy, sustained release of Los for over a week via the oral route can be achieved without the need for encapsulation in a polymeric carrier. The proposed preformulation increased the efficacy reducing the dose or spacing between each dose intake.
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Bloqueadores del Receptor Tipo 1 de Angiotensina II/química , Bloqueadores del Receptor Tipo 1 de Angiotensina II/farmacología , Receptor de Angiotensina Tipo 1/metabolismo , Administración Oral , Animales , Antihipertensivos/química , Antihipertensivos/farmacología , Presión Sanguínea/efectos de los fármacos , Química Farmacéutica/métodos , Portadores de Fármacos/química , Losartán , Masculino , Polímeros/química , Ratas , Ratas Transgénicas , Ratas Wistar , beta-Ciclodextrinas/química , beta-Ciclodextrinas/farmacologíaRESUMEN
PURPOSE OF REVIEW: Angiotensin II is a main regulator of kidney function. Renal actions mediated by the angiotensin AT1 receptor have been well known for many years. In contrast, several details of angiotensin AT2 receptor actions in kidney physiology and pathophysiology were only described very recently. These findings are reviewed in this article. RECENT FINDINGS: Regarding the role of the angiotensin AT2 receptor in kidney physiology, a major recent finding was that the AT2 receptor-mediated inhibition of Na-H exchanger-3 and Na/K-ATPase in the renal proximal tubules is caused by internalisation of these transporters, thus reducing reabsorption and increasing natriuresis/diuresis. Regarding renal pathology, several studies demonstrated an attenuation of renal injury caused by diabetes or by obesity with or without high-salt diet through anti-inflammatory, antifibrotic, and antioxidative mechanisms. Generally, AT2 receptor expression seems increased and AT2 receptor-mediated effects stronger in female and obese animals. SUMMARY: The recent findings about the role of the angiotensin AT2 receptor in renal health and disease strongly suggest that pharmacological targeting of this receptor with selective agonists is a promising therapeutic strategy for inducing diuresis/natriuresis (also additive to established diuretics) and for the treatment of diabetic nephropathy or kidney disease of other pathogenesis.
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Nefropatías Diabéticas/metabolismo , Túbulos Renales Proximales/metabolismo , Obesidad/metabolismo , Receptor de Angiotensina Tipo 2/metabolismo , Animales , Nefropatías Diabéticas/tratamiento farmacológico , Diuresis , Humanos , Natriuresis , Receptor de Angiotensina Tipo 2/agonistas , Factores Sexuales , Intercambiador 3 de Sodio-Hidrógeno/antagonistas & inhibidores , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidoresRESUMEN
STUDY QUESTION: Do angiotensin (Ang)-(1-7) levels in human ovarian follicular fluid (FF) correlate with the number and proportion of mature oocytes obtained for IVF? SUMMARY ANSWER: The present study shows for the first time that Ang-(1-7) levels in human FF correlate with the proportion of mature oocytes collected upon ovarian stimulation for IVF. WHAT IS KNOWN ALREADY: Ang-(1-7) is an active peptide of the renin-angiotensin system that stimulates oocyte maturation in isolated rabbit and rat ovaries. However, its role in human ovulation remains unexplored. STUDY DESIGN, SIZE, DURATION: This was a prospective cohort study including 64 participants from a single IVF center. Sample size was calculated to achieve a statistical power of 80% in detecting 20% differences in the proportion of mature oocytes between groups. The participants were enrolled in the study during six consecutive months. PARTICIPANTS/MATERIALS, SETTING, METHODS: Plasma samples were obtained from all subjects at Day 21 of the last menstrual cycle before starting pituitary blockade and controlled ovarian stimulation (COS). Plasma and FF samples were quickly mixed with a protease inhibitor cocktail and stored at -80°C. Ang-(1-7) was quantified in plasma and FF samples by a highly sensitive and specific radioimmunoassay, which was preceded by solid phase extraction, speed vacuum concentration and sample reconstitution in assay buffer. FF Ang-(1-7) levels were stratified into tertiles and the patients of each tertile were compared for COS/IVF outcomes using Kruskal-Wallis ANOVA. Multiple regression analysis was used to adjust correlations for potential confounders. The mRNA encoding for Mas, a receptor for Ang-(1-7), was investigated by real-time PCR in luteinized granulosa cells purified from the FF. MAIN RESULTS AND THE ROLE OF CHANCE: There was a four-fold increase in plasma Ang-(1-7) after ovulation induction (median 160.9 vs 41.4 pg/ml, P < 0.0001). FF Ang-(1-7) levels were similar to (169.9 pg/ml) but did not correlate with plasma Ang-(1-7) levels (r = -0.05, P = 0.665). Patients at the highest FF Ang-(1-7) tertile had a higher proportion of mature oocytes compared to patients at the lower FF Ang-(1-7) tertile (median 100% vs 70%, P < 0.01). There was a linear correlation between FF Ang-(1-7) and the proportion of mature oocytes (r = 0.380, P < 0.01), which remained significant after adjustment for age and duration of infertility (r = 0.447, P < 0.001). The luteinized granulosa cells expressed Mas receptor mRNA, which was positively correlated to the number of mature oocytes in women with more than three mature oocytes retrieved (r = 0.42, P < 0.01). LIMITATIONS, REASONS FOR CAUTION: This is an observational study, therefore, no causal relationship can be established between Ang-(1-7) and human oocyte maturation. Mas protein expression was not quantified due to limited availability of granulosa cells. WIDER IMPLICATIONS OF THE FINDINGS: Since this peptide promotes oocyte maturation in other species, it deserves further investigation as a potential maturation factor to human oocytes. STUDY FUNDING AND COMPETING INTEREST(S): Research supported by Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG). The authors have nothing to disclose.
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Angiotensina I/agonistas , Fármacos para la Fertilidad Femenina/uso terapéutico , Líquido Folicular/efectos de los fármacos , Infertilidad Femenina/terapia , Oogénesis/efectos de los fármacos , Inducción de la Ovulación , Fragmentos de Péptidos/agonistas , Regulación hacia Arriba/efectos de los fármacos , Adulto , Angiotensina I/sangre , Angiotensina I/metabolismo , Blastocisto/citología , Blastocisto/patología , Estudios de Cohortes , Composición Familiar , Femenino , Fertilización In Vitro , Líquido Folicular/metabolismo , Humanos , Infertilidad Femenina/sangre , Infertilidad Femenina/metabolismo , Infertilidad Femenina/patología , Infertilidad Masculina , Masculino , Recuperación del Oocito , Fragmentos de Péptidos/sangre , Fragmentos de Péptidos/metabolismo , Estudios Prospectivos , Radioinmunoensayo , Extracción en Fase SólidaRESUMEN
BACKGROUND: Neutrophil-mediated inflammation was recently identified as an active contributor to athero-progression. Therapeutic strategies inhibiting neutrophil degranulation or recruitment were hypothesized to positively impact on plaque vulnerability. In this study, we investigated whether treatment with the recently discovered agonist of the Mas-related G-coupled receptor type D (MrgD) alamandine would impact on neutrophil degranulation in vivo and in vitro. MATERIALS AND METHODS: Fifteen-week-old ApoE-/- mice were fed with a Western-type diet for an additional 11 weeks. After the first 2 weeks of diet, mice were surgically implanted with a carotid 'cast' device that alters the blood shear stress and induces different carotid plaque phenotypes. During the last 4 weeks before euthanasia, mice were randomly assigned to subcutaneously receive vehicle (NaCl 0·15 M) or alamandine (24 µg/kg/h) by micropump. For in vitro experiments, neutrophils were obtained after thioglycollate intraperitoneal injection in ApoE-/- mice. RESULTS: Treatment with alamandine was well-tolerated, but failed to affect lipid, macrophage, neutrophil or collagen content within carotid and aortic root plaques. Also, treatment with alamandine did not affect Th-cell polarization in lymphoid organs. Conversely, alamandine administration was associated with a reduction in serum levels of neutrophil granule enzymes, such as MMP-9 and MPO as well as MMP-9 content within aortic root plaques. In vitro, preincubation with alamandine dose-dependently abrogated PMA-induced neutrophil degranulation of MMP-9 and MPO. CONCLUSION: These results suggest that treatment with the MrgD agonist alamandine led to a reduced release of neutrophil granule products, potentially interfering with pro-atherosclerotic neutrophil activation.
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Aterosclerosis/fisiopatología , Degranulación de la Célula/efectos de los fármacos , Neutrófilos/efectos de los fármacos , Oligopéptidos/farmacología , Placa Aterosclerótica/fisiopatología , Animales , Aorta Torácica/efectos de los fármacos , Aterosclerosis/tratamiento farmacológico , Arterias Carótidas/efectos de los fármacos , Progresión de la Enfermedad , Técnicas In Vitro , Metabolismo de los Lípidos/efectos de los fármacos , Metaloproteinasa 9 de la Matriz/metabolismo , Ratones Noqueados , Neutrófilos/fisiología , Peroxidasa/metabolismo , Placa Aterosclerótica/tratamiento farmacológico , Distribución Aleatoria , Receptores Acoplados a Proteínas G/agonistasRESUMEN
Nonsteroidal antiinflammatory drugs, including ibuprofen, are among the most commonly used medications and produce their antiinflammatory effects by blocking cyclooxygenase (COX)-2. Their use is associated with increased risk of heart attacks caused by blocking COX-2 in the vasculature and/or kidney, with our recent work implicating the endogenous NOS inhibitor asymmetric dimethylarginine (ADMA), a cardiotoxic hormone whose effects can be prevented by l-arginine. The ibuprofen salt ibuprofen arginate (Spididol) was created to increase solubility but we suggest that it could also augment the NO pathway through codelivery of arginine. Here we investigated the idea that ibuprofen arginate can act to simultaneously inhibit COX-2 and preserve the NO pathway. Ibuprofen arginate functioned similarly to ibuprofen sodium for inhibition of mouse/human COX-2, but only ibuprofen arginate served as a substrate for NOS. Ibuprofen arginate but not ibuprofen sodium also reversed the inhibitory effects of ADMA and NG-nitro-l-arginine methyl ester on inducible NOS (macrophages) and endothelial NOS in vitro (aorta) and in vivo (blood pressure). These observations show that ibuprofen arginate provides, in one preparation, a COX-2 inhibitor and NOS substrate that could act to negate the harmful cardiovascular consequences mediated by blocking renal COX-2 and increased ADMA. While remarkably simple, our findings are potentially game-changing in the nonsteroidal antiinflammatory drug arena.-Kirkby, N. S., Tesfai, A., Ahmetaj-Shala, B., Gashaw, H. H., Sampaio, W., Etelvino, G., Leão, N. M., Santos, R. A., Mitchell, J. A. Ibuprofen arginate retains eNOS substrate activity and reverses endothelial dysfunction: implications for the COX-2/ADMA axis.
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Arginina/análogos & derivados , Ciclooxigenasa 2/metabolismo , Ibuprofeno/farmacología , Óxido Nítrico Sintasa de Tipo III/metabolismo , Animales , Arginina/metabolismo , Arginina/farmacología , Combinación de Medicamentos , Endotelio/efectos de los fármacos , Endotelio/metabolismo , Inhibidores Enzimáticos/farmacología , Humanos , Riñón/metabolismo , Masculino , Ratones Endogámicos C57BL , Óxido Nítrico/metabolismo , Especificidad por SustratoRESUMEN
Angiotensin-(1-7) counterbalances angiotensin II cardiovascular effects. However, it has yet to be determined how cardiovascular autonomic modulation may be affected by chronic and acute elevation of Ang-(1-7). Hemodynamics and cardiovascular autonomic profile were evaluated in male Sprague-Dawley (SD) rats and transgenic rats (TGR) overexpressing Ang-(1-7) [TGR(A1-7)3292]. Blood pressure (BP) was directly measured while cardiovascular autonomic modulation was evaluated by spectral analysis. TGR received A-779 or vehicle and SD rats received Ang-(1-7) or vehicle and were monitored for 5 h after i.v. administration. In another set of experiments with TGR, A-779 was infused for 7 days using osmotic mini pumps. Although at baseline no differences were observed, acute administration of A-779 in TGR produced a marked long-lasting increase in BP accompanied by increased BP variability (BPV) and sympathetic modulation to the vessels. Likewise, chronic administration of A-779 with osmotic mini pumps in TGR increased heart rate, sympathovagal balance, BPV, and sympathetic modulation to the vessels. Administration of Ang-(1-7) to SD rats increased heart rate variability values in 88% accompanied by 8% of vagal modulation increase and 18% of mean BP reduction. These results show that both acute and chronic alteration in the Ang-(1-7)-Mas receptor axis may lead to important changes in the autonomic control of circulation, impacting either sympathetic and (or) parasympathetic systems.