Eliezeck, M. | Search bioRxiv

bioRxiv2024

Identification and characterization of alamandine-(1-5), a new component of the Renin-Angiotensin System with unique properties

The renin-angiotensin system (RAS) comprises a biochemical cascade that hydrolyzes angiotensinogen into several different bioactive peptides, which can activate different receptors promoting plenty of specific effects. The aim of this study was to evaluate the presence of the putative product of alamandine, the pentapeptide alamandine-(1-5) in the circulation and its biological activity. To accomplish this we have used mass spectrometry (MALDI/TOF/TOF, LC-MS/MS) and several methodologies including isolated blood vessels, isolated perfused hearts, isolated cardiomyocytes, blood pressure recording in freely-moving normotensive and hypertensive rats (SHR), high resolution echocardiography (VEVO 2100), central administration (ICV infusion and microinjection in the insular cortex), cell culture (endothelial cells and GPCR-transfected CHO cells) and wild type and Mas, MrgD or AT2R deficient mice. Our results show that alamandine-(1-5) circulates in the human and rodent blood and promotes many biological central and peripheral actions. More importantly, its plasma concentration is increased in pediatric nephropathic patients. A major role for plasma ACE activity in the formation of alamandine-(1-5) from alamandine was observed using plasma samples from Angiotensinogen-KO mice. Alamandine-(1-5) increased Baroreflex sensitivity and produced a long-lasting ([~]6 hours) anti-hypertensive effect in SHR, associated with a significant reduction in cardiac output. A particularly important effect of this pentapeptide was observed in isolated perfused heart and cardiomyocyte contractility (reduced inotropism). It was capable of stimulating NO production through all receptors from the renin-angiotensin protective arm, (MAS, MrgD and AT2R) in CHO-transfected cells. Our data shows that Alamandine-(1-5) exhibits selective actions that set it apart from traditional concepts of the vasodilatory axis of the RAS and that are possibly intricately linked to a complex interplay between Mas, MrgD and AT2 receptors. This novel finding suggests that RAS may possess a complexity that surpasses our current understanding.

physiology↗

bioRxiv2024

Evidence that the monoamine oxidase B (MAO-B) plays a central role in the inotropic dysfunction induced by genetic deletion of the Mas-related-G protein-coupled receptor D (MrgD) in mice

The renin-angiotensin system (RAS) plays a critical role in the regulation of the cardiovascular system. The Mas-related G protein receptor member D (MrgD) is the receptor of alamandine, and both are components of the RAS noncanonical arm. Alamandine/MrgD induces vasodilation, anti-inflammatory, anti-fibrotic and anti-oxidative effects. In contrast, Mrgd gene deletion leads to a remarkable dilated cardiomyopathy (DCM) in mice. Here, we aimed to investigate the molecular mechanisms of DCM triggered by the deletion of MrgD in the left ventricle and isolated ventricular cardiomyocytes from 8-12 weeks old mice using phosphoproteomics. Our findings revealed an increased oxidative stress not caused by angiotensin II/AT1 hyperactivation but instead due to the up-regulation of the monoamine oxidase B (MAO-B), leading to a higher catabolism of dopamine and epinephrine in the MrgD-KO cardiac tissues. The oxidative environment induced by MAO-B hyperactivation seems to be the cause of the observed alteration in ionic dynamics - altered Ca2+ transient and Na+/K+-ATPase activity - leading to altered resting membrane potential (RMP) and decreased contraction of MrgD-KO cardiomyocytes. In addition, cardiac Troponin-I phosphorylation, and Titin dephosphorylation seem to contribute to the contractile dysfunction observed in MrgD-KO. The treatment of cardiomyocytes from MrgD-KO mice with the MAO-B inhibitor Pargyline reverted the observed impaired contraction, corroborating the hypothesis that MAO-B hyperactivation is, at least partially, the cause of the failing heart observed in MrgD-KO mouse. The findings reported here provide important insights into the pathogenesis of heart failure and suggest a potential therapeutic target (MrgD activation) for managing failing hearts.

physiology↗