An oral formulation of angiotensin-(1-7) reverses corpus cavernosum damages induced by hypercholesterolemia.

INTRODUCTION: The renin angiotensin system plays a crucial role in erectile function. It has been shown that elevated angiotensin-II levels contribute to the development of erectile dysfunction (ED). Oppositely, angiotensin-(1-7) (Ang-[1-7]) mediates penile erection by activation of receptor Mas. Recently, we have developed a formulation based on Ang-(1-7) inclusion in cyclodextrin (CyD) [Ang-(1-7)-CyD], which allows for the oral administration of Ang-(1-7). AIM: In the present study, we evaluated the effects of chronic treatment with Ang-(1-7)-CyD on penile fibrosis, oxidative stress, and endothelial function in hypercholesterolemic mice. METHODS: Apolipoprotein(Apo)E-/- mice fed a Western-type diet for 11 weeks received Ang-(1-7)-CyD or vehicle during the final 3 weeks. Collagen content and reactive oxygen species (ROS) production within the corpus cavernosum were evaluated by Sirius red and dihydroethidium staining, respectively. Protein expression of neuronal nitric oxide synthase (nNOS) and endothelial nitric oxide synthase (eNOS), nicotinamide adenine dinucleotide phosphate (NADPH) subunits (p67-phox and p22-phox), and AT1 and Mas receptors in the penis was assessed by Western blotting. Nitric oxide (NO) production was measured by Griess assay in the mice serum. Cavernosal strips were mounted in an isometric organ bath to evaluate the endothelial function. MAIN OUTCOME MEASURES: The effect of Ang-(1-7)-CyD treatment on penile fibrosis, oxidative stress, and endothelial function in hypercholesterolemia-induced ED. RESULTS: Ang-(1-7)-CyD treatment reduced collagen content in the corpus cavernosum of ApoE-/- mice. This effect was associated with an attenuation of ROS production and a diminished expression of NADPH. Furthermore, Ang-(1-7)-CyD treatment augmented the expression of nNOS and eNOS in the penis and elevated vascular NO production. Importantly, these effects were accompanied by an improvement in cavernosal endothelial function. CONCLUSION: Long-term treatment with Ang-(1-7)-CyD reduces penile fibrosis associated with attenuation of oxidative stress. Additionally, cavernosal endothelial function in hypercholesterolemic mice was markedly improved. These results suggest that Ang-(1-7)-CyD might have significant therapeutic benefits for the treatment of erectile dysfunction.

Update on the role of cannabinoid receptors after ischemic stroke.

Cannabinoids are considered as key mediators in the pathophysiology of inflammatory diseases, including atherosclerosis. In particular, they have been shown to reduce the ischemic injury after acute cardiovascular events, such as acute myocardial infarction and ischemic stroke. These protective and anti-inflammatory properties on peripheral tissues and circulating inflammatory have been demonstrated to involve their binding with both selective cannabinoid type 1 (CB1 and type 2 (CB2) transmembrane receptors. On the other hands, the recent discoveries of novel different classes of cannabinoids and receptors have increased the complexity of this system in atherosclerosis. Although only preliminary data have been reported on the activities of novel cannabinoid receptors, several studies have already investigated the role of CB1 and CB2 receptors in ischemic stroke. While CB1 receptor activation has been shown to directly reduce atherosclerotic plaque inflammation, controversial data have been shown on neurotransmission and neuroprotection after stroke. Given its potent anti-inflammatory activities on circulating leukocytes, the CB2 activation has been proven to produce protective effects against acute poststroke inflammation. In this paper, we will update evidence on different cannabinoid-triggered avenues to reduce inflammation and neuronal injury in acute ischemic stroke.

Treatment with Angiotensin-(1-7) reduces inflammation in carotid atherosclerotic plaques.

Angiotensin (Ang)-(1-7), acting through the receptor Mas, has atheroprotective effects; however, its role on plaque vulnerability has been poorly studied. Here, we investigated the expression of the renin-angiotensin system (RAS) components in stable and unstable human carotid plaques. In addition, we evaluated the effects of the chronic treatment with an oral formulation of Ang-(1-7) in a mouse model of shear stress-determined carotid atherosclerotic plaque. Upstream and downstream regions of internal carotid plaques were obtained from a recently published cohort of patients asymptomatic or symptomatic for ischaemic stroke. Angiotensinogen and renin genes were strongly expressed in the entire cohort, indicating an intense intraplaque modulation of the RAS. Intraplaque expression of the Mas receptor mRNA was increased in the downstream portion of asymptomatic patients as compared to corresponding region in symptomatic patients. Conversely, AT1 receptor gene expression was not modified between asymptomatic and symptomatic patients. Treatment with Ang-(1-7) in ApoE-/- mice was associated with increased intraplaque collagen content in the aortic root and low shear stress-induced carotid plaques, and a decreased MMP-9 content and neutrophil and macrophage infiltration. These beneficial effects were not observed in the oscillatory shear stress-induced plaque. In vitro incubation with Ang-(1-7) did not affect ICAM-1 expression and apoptosis on cultured endothelial cells. In conclusion, Mas receptor is up regulated in the downstream portions of human stable carotid plaques as compared to unstable lesions. Treatment with the oral formulation of Ang-(1-7) enhances a more stable phenotype in atherosclerotic plaques, depending on the local pattern of shear stress forces.

Cardiac dysfunction in rats prone to audiogenic epileptic seizures.

PURPOSE: Cardiac dysfunction is one of the possible causes of sudden unexpected death in epilepsy (SUDEP). Therefore, the objective of this study was to evaluate cardiac and electrocardiographic parameters in rats with audiogenic epileptic seizures (WAR--Wistar audiogenic rats). METHODS: In vivo arterial pressure, heart rate (HR), autonomic tone and electrocardiography (ECG) were measured in awake animals in order to examine cardiac function and rhythm. Ex vivo, the Langendorff technique was used to analyze the cardiac function and the severity of reperfusion arrhythmias. In vitro, confocal microscopy was used to evaluate calcium transient parameters of isolated ventricular cardiomyocytes. RESULTS: In vivo autonomic tone evaluation revealed enhanced sympathetic activity, changes in cardiac function with increased systolic arterial pressure and higher basal HR in WAR. In addition, ECG analysis demonstrated electrical alterations with prolongation of the QT interval and QRS complex in these animals. Ex vivo, we observed a decrease in systolic tone and HR and an increase in the duration of ischemia/reperfusion arrhythmias in WAR. Moreover, intracellular Ca2+ handling analysis revealed an increase in the peak of calcium and calcium transient decay in audiogenic rats. Treatment with atenolol (ß1-adrenergic antagonist) normalized the systolic tone, reduced cardiac hypertrophy and the associated increase in the susceptibility to reperfusion arrhythmias observed in WAR. CONCLUSION: We present evidence that chronic disturbances in sympathetic tone in WAR cause increases the risk to life-threatening arrhythmias. Our results support a relationship between seizures, cardiac dysfunction and cardiac arrhythmias, which may contribute to the occurrence of SUDEP.

The novel Mas agonist, CGEN-856S, attenuates isoproterenol-induced cardiac remodeling and myocardial infarction injury in rats.

CGEN-856S is a novel Mas agonist. Herein, we examined the effects of this peptide on isoproterenol (ISO)-induced cardiac remodeling and myocardial infarction (MI) injury. We also sought to determine whether CGEN-856S activates the underlying mechanisms related to Mas receptor activation. Heart hypertrophy and fibrosis were induced by ISO (2 mg·kg(-1)·day(-1)) in Wistar rats. After a 7-day treatment period with CGEN-856S (90 µg·kg(-1)·day(-1)) or vehicle, the cardiomyocyte diameter was evaluated in left ventricular sections stained with hematoxylin and eosin, and immunofluorescence labeling and quantitative confocal microscopy were used to quantify the deposition of type I and III collagen and fibronectin in the left ventricles. MI was induced by coronary artery ligation, and CGEN-856S (90 µg·kg(-1)·day(-1)) or saline was administered for 14 days. The Langendorff technique was used to evaluate cardiac function, and left ventricular sections were stained with Masson's trichrome dye to quantify the infarct area. Using Chinese hamster ovary cells stably transfected with Mas cDNA, we evaluated whether CGEN-856S alters AKT and endothelial nitric oxide synthase (eNOS) phosphorylation. CGEN-856S reduced the degree of ISO-induced hypertrophy (13.91±0.17 µm vs. 12.41±0.16 µm in the ISO+CGEN-856S group). In addition, the Mas agonist attenuated the ISO-induced increase in collagen I, collagen III, and fibronectin deposition. CGEN-856S markedly attenuated the MI-induced decrease in systolic tension, as well as in +dT/dt and -dT/dt. Furthermore, CGEN-856S administration significantly decreased the infarct area (23.68±2.78% vs. 13.95±4.37% in the MI+CGEN-856S group). These effects likely involved the participation of AKT and NO, as CGEN-856S administration increased the levels of p-AKT and p-eNOS. Thus, our results indicate that CGEN-856S exerts cardioprotective effects on ISO-induced cardiac remodeling and MI-mediated heart failure in rats through a mechanism likely involving the eNOS/AKT pathway.

Beneficial effects of long-term administration of an oral formulation of Angiotensin-(1-7) in infarcted rats.

In this study was evaluated the chronic cardiac effects of a formulation developed by including angiotensin(Ang)-(1-7) in hydroxypropyl ß-cyclodextrin (HPßCD), in infarcted rats. Myocardial infarction (MI) was induced by left coronary artery occlusion. HPßCD/Ang-(1-7) was administered for 60 days (76 µg/Kg/once a day/gavage) starting immediately before infarction. Echocardiography was utilized to evaluate usual cardiac parameters, and radial strain method was used to analyze the velocity and displacement of myocardial fibers at initial time and 15, 30, and 50 days after surgery. Real-time PCR was utilized to evaluate the fibrotic signaling involved in the remodeling process. Once-a-day oral HPßCD/Ang-(1-7) administration improved the cardiac function and reduced the deleterious effects induced by MI on TGF-ß and collagen type I expression, as well as on the velocity and displacement of myocardial fibers. These findings confirm cardioprotective effects of Ang-(1-7) and indicate HPßCD/Ang-(1-7) as a feasible formulation for long-term oral administration of this heptapeptide.