adrenomedullin and Edema

Topics: Adrenomedullin; Animals; Edema; Endothelium, Vascular; Homeostasis; Humans; Ischemia; Mice; Mice, Knockout; Molecular Targeted Therapy; Neovascularization, Physiologic; Receptor Activity-Modifying Protein 2

Revealing the mechanisms underlying the functional integrity of the vascular system could make available novel therapeutic approaches. We previously showed that knocking out the widely expressed peptide adrenomedullin (AM) or receptor activity-modifying protein 2 (RAMP2), an AM-receptor accessory protein, causes vascular abnormalities and is embryonically lethal. Our aim was to investigate the function of the vascular AM-RAMP2 system directly.. We generated endothelial cell-specific RAMP2 and AM knockout mice (E-RAMP2(-/-) and E-AM(-/-)). Most E-RAMP2(-/-) mice died perinatally. In surviving adults, vasculitis occurred spontaneously. With aging, E-RAMP2(-/-) mice showed severe organ fibrosis with marked oxidative stress and accelerated vascular senescence. Later, liver cirrhosis, cardiac fibrosis, and hydronephrosis developed. We next used a line of drug-inducible E-RAMP2(-/-) mice (DI-E-RAMP2(-/-)) to induce RAMP2 deletion in adults, which enabled us to analyze the initial causes of the aforementioned vascular and organ damage. Early after the induction, pronounced edema with enhanced vascular leakage occurred. In vitro analysis revealed the vascular leakage to be caused by actin disarrangement and detachment of endothelial cells. We found that the AM-RAMP2 system regulates the Rac1-GTP/RhoA-GTP ratio and cortical actin formation and that a defect in this system causes the disruption of actin formation, leading to vascular and organ damage at the chronic stage after the gene deletion.. Our findings show that the AM-RAMP2 system is a key determinant of vascular integrity and homeostasis from prenatal stages through adulthood. Furthermore, our models demonstrate how endothelial cells regulate vascular integrity and how their dysregulation leads to organ damage.

Topics: Adrenomedullin; Age Factors; Aging; Animals; Antigens, CD; Arteriosclerosis; Cadherins; Disease Models, Animal; Edema; Endothelium, Vascular; Fibrosis; Glomerulosclerosis, Focal Segmental; Homeostasis; Kidney; Leukocytes; Mice; Mice, Knockout; Oxidative Stress; Receptor Activity-Modifying Protein 2; Vasculitis

Adrenomedullin (AM) and its receptor complexes, calcitonin receptor-like receptor (Calcrl) and receptor activity modifying protein 2/3, are highly expressed in lymphatic endothelial cells and are required for embryonic lymphatic development. To determine the role of Calcrl in adulthood, we used an inducible Cre-loxP system to temporally and ubiquitously delete Calcrl in adult mice. Following tamoxifen injection, Calcrl(fl/fl)/CAGGCre-ER™ mice rapidly developed corneal edema and inflammation that was preceded by and persistently associated with dilated corneoscleral lymphatics. Lacteals and submucosal lymphatic capillaries of the intestine were also dilated, while mesenteric collecting lymphatics failed to properly transport chyle after an acute Western Diet, culminating in chronic failure of Calcrl(fl/fl)/CAGGCre-ER™ mice to gain weight. Dermal lymphatic capillaries were also dilated and chronic edema challenge confirmed significant and prolonged dermal lymphatic insufficiency. In vivo and in vitro imaging of lymphatics with either genetic or pharmacologic inhibition of AM signaling revealed markedly disorganized lymphatic junctional proteins ZO-1 and VE-cadherin. The maintenance of AM signaling during adulthood is required for preserving normal lymphatic permeability and function. Collectively, these studies reveal a spectrum of lymphatic defects in adult Calcrl(fl/fl)/CAGGCre-ER™ mice that closely recapitulate the clinical symptoms of patients with corneal, intestinal and peripheral lymphangiectasia.

Topics: Adrenomedullin; Animals; Antigens, CD; Cadherins; Calcitonin Receptor-Like Protein; Edema; Endothelial Cells; Female; Gene Deletion; Gene Expression; Genetic Vectors; Intestinal Mucosa; Intestines; Limbus Corneae; Lymphangiectasis; Lymphatic Vessels; Male; Mice; Mice, Transgenic; Signal Transduction; Skin; Tamoxifen; Zonula Occludens-1 Protein

Adrenomedullin (AM) is a peptide involved both in the pathogenesis of cardiovascular diseases and in circulatory homeostasis. The high-affinity AM receptor is composed of receptor activity-modifying protein 2 or 3 (RAMP2 or -3) and the GPCR calcitonin receptor-like receptor. Testing our hypothesis that RAMP2 is a key determinant of the effects of AM on the vasculature, we generated and analyzed mice lacking RAMP2. Similar to AM-/- embryos, RAMP2-/- embryos died in utero at midgestation due to vascular fragility that led to severe edema and hemorrhage. Vascular ECs in RAMP2-/- embryos were severely deformed and detached from the basement membrane. In addition, the abnormally thin arterial walls of these mice had a severe disruption of their typically multilayer structure. Expression of tight junction, adherence junction, and basement membrane molecules by ECs was diminished in RAMP2-/- embryos, leading to paracellular leakage and likely contributing to the severe edema observed. In adult RAMP2+/- mice, reduced RAMP2 expression led to vascular hyperpermeability and impaired neovascularization. Conversely, ECs overexpressing RAMP2 had enhanced capillary formation, firmer tight junctions, and reduced vascular permeability. Our findings in human cells and in mice demonstrate that RAMP2 is a key determinant of the effects of AM on the vasculature and is essential for angiogenesis and vascular integrity.

Topics: Adrenomedullin; Animals; Arteries; Calcitonin Receptor-Like Protein; Capillary Permeability; Cardiovascular Diseases; Cells, Cultured; Edema; Embryo Loss; Embryonic Stem Cells; Endothelium, Vascular; Female; Homeostasis; Humans; Intracellular Signaling Peptides and Proteins; Male; Membrane Proteins; Mice; Mice, Knockout; Neovascularization, Physiologic; Pregnancy; Receptor Activity-Modifying Protein 2; Receptor Activity-Modifying Proteins; Receptors, Adrenomedullin; Receptors, Calcitonin; Receptors, Peptide; Tight Junctions

Growth of blood and lymphatic vessels is essential in the developing embryo and in the pathogenesis of human diseases such as cancer, but the signaling pathways that regulate vessel growth and function are not yet well characterized. In this issue of the JCI, studies by Fritz-Six et al. and Ichikawa-Shindo et al. demonstrate that loss of signaling by the adrenomedullin peptide results in embryonic edema and death (see the related articles beginning on pages 40 and 29, respectively). Remarkably, this phenotype is attributed to defects in lymphatic vessels by one group and to defects in blood vessels by the other. In addition to defining what I believe to be a novel angiogenic signaling pathway, these studies demonstrate the intricate molecular, genetic, and physiologic relationships between blood and lymphatic vessels that must be considered by investigators of vascular biology.

Topics: Adrenomedullin; Animals; Arteries; Calcitonin Receptor-Like Protein; Capillary Permeability; Cardiovascular Diseases; Cells, Cultured; Edema; Embryo Loss; Embryonic Stem Cells; Endothelium, Vascular; Female; Homeostasis; Humans; Intracellular Signaling Peptides and Proteins; Male; Membrane Proteins; Mice; Mice, Knockout; Neovascularization, Physiologic; Pregnancy; Receptor Activity-Modifying Proteins; Receptors, Adrenomedullin; Receptors, Calcitonin; Receptors, Peptide; Tight Junctions

Our aim was to assess whether adrenomedullin (AM), a potent vasodilator peptide with a variety of cardioprotective effects, has a therapeutic potential for the treatment of acute myocarditis in a rat model.. One week after myosin injection, rats received a continuous infusion of AM or vehicle for 2 weeks, and pathological and physiological investigations were performed.. AM treatment significantly reduced the infiltration of inflammatory cells in myocarditic hearts, and decreased the expressions of macrophage chemoattractant protein-1, matrix metalloproteinase-2 and transforming growth factor-beta. Myocardial edema indicated by increased heart weight to body weight ratio and wall thickness was attenuated by AM infusion (5.7+/-0.5 vs. 6.5+/-0.4 g/kg, and 1.9+/-0.3 vs. 2.8+/-0.5 mm, respectively). Infusion of AM significantly improved left ventricular maximum dP/dt and fractional shortening of myocarditic hearts (4203+/-640 vs. 3450+/-607 mm Hg/s, and 21.3+/-4.1 vs. 14.7+/-5.1%, respectively).. Infusion of AM improved cardiac function and pathological findings in a rat model of acute myocarditis. Thus, infusion of AM may be a potent therapeutic strategy for acute myocarditis.

Topics: Acute Disease; Adrenomedullin; Animals; Biomarkers; Blotting, Western; Chemokine CCL2; Echocardiography; Edema; Heart Rate; Immunohistochemistry; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Myocarditis; Myocardium; Myosins; Neovascularization, Physiologic; Rats; Rats, Inbred Lew; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta

Peptidylglycine alpha-amidating monooxygenase (PAM) catalyzes the COOH-terminal amidation of peptide hormones. We previously had found high expression of PAM in several regions of the developing rodent. To determine the function of PAM during mouse embryogenesis, we produced a null mutant of the PAM gene. Homozygous mutants die in utero between e14.5 and e15.5 with severe edema that is likely due to cardiovascular deficits. These defects include thinning of the aorta and carotid arteries and are very similar to those of the recently characterized adrenomedullin (AM) gene KO despite the presence of elevated immunoreactive AM in PAM KO embryos. No peptide amidation activity was detected in PAM mutant embryos, and there was no moderation of the AM-like phenotype that could be expected if any alternative peptide amidation mechanism exists in the mouse. Despite the proposed contribution of amidated peptides to neuronal cell proliferation, no alteration in neuroblast proliferation was observed in homozygous mutant embryos prior to lethality. Mice heterozygous for the mutant PAM allele develop normally and express wildtype levels of several amidated peptides despite having one half the wildtype levels of PAM activity and PAM protein. Nonetheless, both an increase in adiposity and a mild glucose intolerance developed in aged (>10 months) heterozygous mice compared to littermate controls. Ablation of PAM thus demonstrates an essential function for this gene during mouse development, while alterations in PAM activity in the adult may underlie more subtle physiologic effects.

Topics: Adrenomedullin; Animals; Blood Vessels; Brain; Edema; Embryo, Mammalian; Female; Glucose Tolerance Test; Heart Ventricles; Lung; Male; Mice; Mice, Knockout; Mixed Function Oxygenases; Multienzyme Complexes; Mutation; Peptides; Yolk Sac

Calcitonin gene-related peptide (CGRP), a neuropeptide, is a potent vasodilator. Adrenomedullin (ADM) is suggested to be produced by vascular cells in inflamed tissue. ADM shares some structural homology with CGRP. We have compared the ability of CGRP and ADM to modulate microvascular and thermal hyperalgesic responses in rat skin. Vasodilator activity was assessed by laser Doppler flowmetry, inflammatory oedema by the extravascular accumulation of intravenously-injected labelled albumin, and neutrophil accumulation by tissue myeloperoxidase, in dorsal skin. Hyperalgesia was assessed by a thermal hyperalgesimeter in paw skin. ADM (10-300 pmol) was 3 fold less potent than CGRP (3-100 pmol) as a direct vasodilator. CGRP (30 pmol) potentiated oedema formation induced by mediators of increased microvascular permeability, as expected (P<0.01). However, ADM (30-100 pmol) was without a potentiating effect, although ADM (300 pmol) was effective (P<0.01). By comparison ADM (100 pmol) potentiated neutrophil accumulation induced by interleukin-1beta (P<0.05), whereas CGRP (30 pmol) did not. No thermal hyperalgesia was observed to either CGRP or ADM, when given as single or repeated treatments. Thus despite a dilator activity neither CGRP nor ADM appears to mediate hyperalgesic activity in the periphery. However ADM, like CGRP, has the ability to potentiate inflammatory oedema formation and, in addition, ADM can potentiate neutrophil accumulation. ADM may, as suggested for CGRP, act as a modulator of the vascular phases of inflammation. The property of the two compounds of evoking differential microvascular responses and neutrophil accumulation may be due to differing mechanisms of action.

Topics: Adrenomedullin; Animals; Calcitonin Gene-Related Peptide; Edema; Hyperalgesia; Male; Microcirculation; Neutrophils; Peptide Fragments; Rats; Rats, Wistar; Regional Blood Flow; Skin; Vasodilator Agents

The antiinflammatory effect of ADM was studied in different models of inflammation and compared to the one of CGRP. Peptides were active against acetic acid-induced peritonitis in the rats. ADM and CGRP exerted the antiinflammatory effect at different doses, 400 and 20 ng/kg respectively, but with different efficacy (ADM >CGRP). This effect was blocked by pretreatment with CGRP (8-37) fragment or with L-NAME. No antiinflammatory activity was evidenced against serotonin- or carrageenin-induced rat paw edema. Our data suggest that ADM exerts antiinflammatory activity in the model characterized by a vascular component. This effect involves CGRP receptors and appears to be mediated by nitric oxide system.

Topics: Acetic Acid; Adrenomedullin; Animals; Anti-Inflammatory Agents, Non-Steroidal; Calcitonin Gene-Related Peptide; Disease Models, Animal; Dose-Response Relationship, Drug; Edema; Enzyme Inhibitors; Humans; Mice; NG-Nitroarginine Methyl Ester; Peptides; Peritonitis; Rats; Rats, Sprague-Dawley