angiotensin-i and Fibrosis

Pulmonary fibrosis is a chronic, fibrotic lung disease affecting 3 million people worldwide. The ACE2/Ang-(1-7)/MasR axis is of interest in pulmonary fibrosis due to evidence of its anti-fibrotic action. Current scientific evidence supports that inhibition of ACE2 causes enhanced fibrosis. ACE2 is also the primary receptor that facilitates the entry of SARS-CoV-2, the virus responsible for the current COVID-19 pandemic. COVID-19 is associated with a myriad of symptoms ranging from asymptomatic to severe pneumonia and acute respiratory distress syndrome (ARDS) leading to respiratory failure, mechanical ventilation, and often death. One of the potential complications in people who recover from COVID-19 is pulmonary fibrosis. Cigarette smoking is a risk factor for fibrotic lung diseases, including the idiopathic form of this disease (idiopathic pulmonary fibrosis), which has a prevalence of 41% to 83%. Cigarette smoke increases the expression of pulmonary ACE2 and is thought to alter susceptibility to COVID-19. Cannabis is another popular combustible product that shares some similarities with cigarette smoke, however, cannabis contains cannabinoids that may reduce inflammation and/or ACE2 levels. The role of cannabis smoke in the pathogenesis of pulmonary fibrosis remains unknown. This review aimed to characterize the ACE2-Ang-(1-7)-MasR Axis in the context of pulmonary fibrosis with an emphasis on risk factors, including the SARS-CoV-2 virus and exposure to environmental toxicants. In the context of the pandemic, there is a dire need for an understanding of pulmonary fibrotic events. More research is needed to understand the interplay between ACE2, pulmonary fibrosis, and susceptibility to coronavirus infection.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Cannabis; Cigarette Smoking; COVID-19; Fibrosis; Humans; Idiopathic Pulmonary Fibrosis; Inflammation; Lung; Pandemics; Peptide Fragments; Proto-Oncogene Mas; Respiration, Artificial; Respiratory Distress Syndrome; Respiratory Insufficiency; Risk Factors; SARS-CoV-2; Spike Glycoprotein, Coronavirus

In human pathology, SARS-CoV-2 utilizes multiple molecular pathways to determine structural and biochemical changes within the different organs and cell types. The clinical picture of patients with COVID-19 is characterized by a very large spectrum. The reason for this variability has not been clarified yet, causing the inability to make a prognosis on the evolution of the disease.. PubMed search was performed focusing on the role of ACE 2 receptors in allowing the viral entry into cells, the role of ACE 2 downregulation in triggering the tissue pathology or in accelerating previous disease states, the role of increased levels of Angiotensin II in determining endothelial dysfunction and the enhanced vascular permeability, the role of the dysregulation of the renin angiotensin system in COVID-19 and the role of cytokine storm.. The pathological changes induced by SARS-CoV-2 infection in the different organs, the correlations between the single cell types targeted by the virus in the different human organs and the clinical consequences, COVID-19 chronic pathologies in liver fibrosis, cardiac fibrosis and atrial arrhythmias, glomerulosclerosis and pulmonary fibrosis, due to the systemic fibroblast activation induced by angiotensin II are discussed.. The main pathways involved showed different pathological changes in multiple tissues and the different clinical presentations. Even if ACE2 is the main receptor of SARS-CoV-2 and the main entry point into cells for the virus, ACE2 expression does not always explain the observed marked inter-individual variability in clinical presentation and outcome, evidencing the complexity of this disorder. The proper interpretation of the growing data available might allow to better classifying COVID-19 in human pathology.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Atrial Fibrillation; Blood Coagulation; Capillary Permeability; Cardiomyopathies; COVID-19; Cytokine Release Syndrome; Cytokines; Endothelium, Vascular; Fibroblasts; Fibrosis; Humans; Liver Cirrhosis; Myocarditis; Receptors, Coronavirus; Renin-Angiotensin System; SARS-CoV-2; Systemic Inflammatory Response Syndrome; Thrombosis; Virus Internalization

This review identifies how the classical/non-classical renin-angiotensin system (RAS) and exercise influence muscle wasting. The classical RAS axis enhances muscle loss through the interaction with NADPH oxidase (NOX), ubiquitin proteasome system (UPS), protein synthesis and fibrosis pathways. The mainstream hypothesis identifies reactive oxygen species (ROS) as the key pathway in muscle, this review recognizes alternative pathways that lead to an increase in muscle wasting through the classical RAS axis. In addition, pathways in which the non-classical RAS axis and exercise inhibit the classical RAS axis are also explored. The non-classical RAS axis and exercise have a significant negative impact on ROS production and protein synthesis. The non-classical RAS axis has been identified in this review to directly affect protein synthesis pathways not by altering the pre-existing intracellular ROS level, further supporting the idea that muscle wasting caused by the classical RAS system is not entirely due to ROS production. Exercise has been identified to modify the RAS axes making it a therapeutic option.

Topics: Angiotensin I; Angiotensin II; Apoptosis; Exercise; Fibrosis; Humans; Mitochondria; Muscle, Skeletal; Muscular Atrophy; NADPH Oxidases; Peptide Fragments; Protein Biosynthesis; Reactive Oxygen Species; Renin-Angiotensin System; Ubiquitin-Protein Ligases

The renin angiotensin system (RAS) plays an important role in inflammation and fibrosis. The classical axis of the RAS, formed by angiotensin converting en-zyme (ACE), angiotensin II (Ang II) and angiotensin receptor type 1 (AT1), activates several cell functions and molecular signaling pathways related to tissue injury, inflammation and fibrosis. In sharp contrast, the RAS axis composed by angiotensin converting enzyme 2 (ACE2), angiotensin-(1-7) and Mas receptor exerts opposite effects in relation to inflammatory response and tissue fibrosis.. In this review, we have the aim to summarize recent findings on the anti-inflammatory and anti-fibrogenic role of ACE2/Ang-(1-7)/Mas axis in the context of basic research, experimental human dis-eases and clinical studies.. Several studies showed that ACE2/Angiotensin-(1-7)/Mas axis reduces cytokine release and inhibits signaling pathways of tissue fibrosis in experimental models of human diseases including atherosclerosis, cerebral ischemia, obesity, chronic kidney disease, liver diseases and asthma. On the other hand, very few data was provided by clinical studies.. Experimental studies clearly support the anti-inflammatory and anti-fibrotic effects of ACE2/ Ang-(1-7)/Mas axis. Clinical studies, especially phase III and IV trials, will be necessary to establish the therapeutic role of ACE2/Ang-(1-7)/Mas axis in controlling inflammation in different human diseases.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Animals; Anti-Inflammatory Agents; Cytokines; Fibrosis; Gene Expression Regulation; Humans; Peptide Fragments; Peptidyl-Dipeptidase A; Proto-Oncogene Mas; Proto-Oncogene Proteins; Receptors, G-Protein-Coupled; Signal Transduction

Recent advances have improved our understanding of the renin-angiotensin system (RAS). These have included the recognition that angiotensin (Ang)-(1-7) is a biologically active product of the RAS cascade. The identification of the ACE homologue ACE2, which forms Ang-(1-7) from Ang II, and the GPCR Mas as an Ang-(1-7) receptor have provided the necessary biochemical and molecular background and tools to study the biological significance of Ang-(1-7). Most available evidence supports a counter-regulatory role for Ang-(1-7) by opposing many actions of Ang II on AT₁ receptors, especially vasoconstriction and proliferation. Many studies have now shown that Ang-(1-7) by acting via Mas receptor exerts inhibitory effects on inflammation and on vascular and cellular growth mechanisms. Ang-(1-7) has also been shown to reduce key signalling pathways and molecules thought to be relevant for fibrogenesis. Here, we review recent findings related to the function of the ACE2/Ang-(1-7)/Mas axis and focus on the role of this axis in modifying processes associated with acute and chronic inflammation, including leukocyte influx, fibrogenesis and proliferation of certain cell types. More attention will be given to the involvement of the ACE2/Ang-(1-7)/Mas axis in the context of renal disease because of the known relevance of the RAS for the function of this organ and for the regulation of kidney inflammation and fibrosis. Taken together, this knowledge may help in paving the way for the development of novel treatments for chronic inflammatory and renal diseases.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Animals; Cell Proliferation; Fibrosis; Humans; Kidney; Leukocytes; Models, Biological; Nephritis; Peptide Fragments; Peptidyl-Dipeptidase A; Proto-Oncogene Mas; Proto-Oncogene Proteins; Receptors, G-Protein-Coupled; Signal Transduction

It is well known that the RAS (renin-angiotensin system) plays a key role in the modulation of many functions in the body. AngII (angiotensin II) acting on AT1R (type 1 AngII receptor) has a central role in mediating most of the actions of the RAS. However, over the past 10 years, several studies have presented evidence for the existence of a new arm of the RAS, namely the ACE (angiotensin-converting enzyme) 2/Ang-(1-7) [angiotensin-(1-7)]/Mas axis. Ang-(1-7) can be produced from AngI or AngII via endo- or carboxy-peptidases respectively. ACE2 appears to play a central role in Ang-(1-7) formation. As described for AngII, Ang-(1-7) also has a broad range of effects in different organs and tissues which goes beyond its initially described cardiovascular and renal actions. Those effects are mediated by Mas and can counter-regulate most of the deleterious effects of AngII. The interaction Ang-(1-7)/Mas regulates different signalling pathways, such as PI3K (phosphoinositide 3-kinase)/AKT and ERK (extracellularsignal-regulated kinase) pathways and involves downstream effectors such as NO, FOXO1 (forkhead box O1) and COX-2 (cyclo-oxygenase-2). Through these mechanisms, Ang-(1-7) is able to improve pathological conditions including fibrosis and inflammation in organs such as lungs, liver and kidney. In addition, this heptapeptide has positive effects on metabolism, increasing the glucose uptake and lipolysis while decreasing insulin resistance and dyslipidaemia. Ang-(1-7) is also able to improve cerebroprotection against ischaemic stroke, besides its effects on learning and memory. The reproductive system can also be affected by Ang-(1-7) treatment, with enhanced ovulation, spermatogenesis and sexual steroids synthesis. Finally, Ang-(1-7) is considered a potential anti-cancer treatment since it is able to inhibit cell proliferation and angiogenesis. Thus the ACE2/Ang-(1-7)/Mas pathway seems to be involved in many physiological and pathophysiological processes in several systems and organs especially by opposing the detrimental effects of inappropriate overactivation of the ACE/AngII/AT1R axis.

Topics: Angiogenesis Inhibitors; Angiotensin I; Animals; Anti-Inflammatory Agents; Antineoplastic Agents; Brain Ischemia; Cell Proliferation; Female; Fibrosis; Glucose; Humans; Insulin; Kidney; Lipid Metabolism; Male; Metabolic Syndrome; Peptide Fragments; Peptidyl-Dipeptidase A; Proto-Oncogene Mas; Proto-Oncogene Proteins; Receptors, G-Protein-Coupled; Renin-Angiotensin System; Reproduction; Signal Transduction

Activation of the renin-angiotensin system has a pivotal role in the pathogenesis of diabetic complications. However, recent evidence suggests that it may also contribute to the development of diabetes itself. In the endocrine pancreas, all the components of an active renin-angiotensin system are present, which modulate a range of activities including local blood flow, hormone release and prostaglandin synthesis. In both types 1 and 2 diabetes, there is an up-regulation of its expression and activity in the endocrine pancreas. Whether these changes have a direct pathogenetic role or reflect a response to local stress or tissue injury remains to be established. Angiotensin-mediated increases in oxidative stress, inflammation and free fatty acids levels potentially contribute to beta-cell dysfunction in diabetes. In addition, activation of the renin-angiotensin system appears to potentiate the action of other pathogenic pathways including glucotoxicity, lipotoxicity and advanced glycation. In experimental models of type 2 diabetes, blockade of the renin-angiotensin system with angiotensin converting enzyme inhibitors or angiotensin receptor antagonists results in the improvement of islet structure and function. Moreover, the incidence of de novo diabetes appears to be significantly reduced by blockade of the renin-angiotensin system in clinical studies. At least two large controlled trials are currently underway to study the role of renin-angiotensin system in the development of diabetes. It is hoped that these studies will demonstrate the true potential of the blockade of the renin-angiotensin system for the prevention of diabetes.

Topics: Angiotensin I; Angiotensin II; Angiotensinogen; Animals; Diabetes Mellitus; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Fibrosis; Humans; Hyperglycemia; Hypoglycemia; Islets of Langerhans; Pancreas; Peptidyl-Dipeptidase A; PPAR gamma; Receptors, Angiotensin; Renin; Renin-Angiotensin System

The heart is composed of highly differentiated cardiac myocytes, which constitute parenchyma, and stroma or connective tissue. Fibrillar collagen turnover in the heart and its valve leaflets, in particular, is dynamic and essential to tissue repair. Emerging evidence further suggests connective tissue is a metabolically active entity, where peptide hormones are generated and degraded and, in turn, these peptides regulate collagen turnover. This concept arose from quantitative in vitro autoradiography using an iodinated derivative of lisinopril (125I-351A) as ligand to localize angiotensin converting enzyme (ACE) binding density within the heart. A heterogeneous distribution was found: low-density ACE binding within atria and ventricles; high ACE binding density at sites of high collagen turnover, such as valve leaflets, adventitia, and fibrous tissue of diverse etiologic origins. ACE-producing cells at these latter sites were identified by monoclonal ACE antibody. They included valvular interstitial cells (VIC) and fibroblast-like cells each of which also contained alpha-smooth muscle actin and the transcript for type I collagen (in situ hybridization). Substrate utilization in cultured VIC was found to include angiotensin I and bradykinin. Angiotensin II and bradykinin receptor-ligand binding was observed in VIC and at fibrous tissue sites. Connective tissue ACE is independent of circulating angiotensin II. In vivo, fibrous tissue formation is attenuated by ACE inhibition or antagonism of AT1 receptor. Angiotensin II and bradykinin are stimulatory and inhibitory, respectively, to cultured adult cardiac fibroblast collagen synthesis suggesting a paradigm of reciprocal regulation to fibroblast collagen turnover. Stroma and its cellular constituents represent a dynamic metabolic entity that regulates its own peptide hormone composition and turnover of fibrillar collagen. These findings may provide insights that could be used to advantage to either promote or forestall fibrous tissue formation depending on the nature of cardiovascular disease.

Topics: Angiotensin I; Angiotensin II; Animals; Bradykinin; Connective Tissue; Fibrosis; Heart; Hormones; Humans; Models, Cardiovascular; Myocardium; Prostaglandins; Wound Healing

Renin-angiotensin system (RAS) plays an indispensable role in regulating blood pressure through its effects on fluid and electrolyte balance. As an aside, cumulative evidence from experimental to clinical studies supports the notion that dysregulation of RAS contributes to the pro-inflammatory, pro-oxidative, and pro-fibrotic processes that occur in pulmonary diseases like asthma, chronic obstructive pulmonary disease (COPD), idiopathic pulmonary fibrosis (IPF), and acute lung injury (ALI). Pharmacological intervention of the various RAS components can be a novel therapeutic strategy for the treatment of these respiratory diseases. In this chapter, we first give a recent update on the RAS, and then compile, review, and analyse recent reports on targeting RAS components as treatments for respiratory diseases. Inhibition of the pro-inflammatory renin, angiotensin-converting enzyme (ACE), angiotensin (Ang) II, and Ang II type 1 receptor (AT1R) axis, and activation of the protective ACE2, AT2R, Ang (1-7), and Mas receptor axis have demonstrated varying degrees of efficacies in experimental respiratory disease models or in human trials. The newly identified alamandine/Mas-related G-protein-coupled receptor member D pathway has shown some therapeutic promise as well. However, our understanding of the RAS ligand-and-receptor interactions is still inconclusive, and the modes of action and signaling cascade mediating the newly identified RAS receptors remain to be better characterized. Clinical data are obviously lacking behind the promising pre-clinical findings of certain well-established molecules targeting at different pathways of the RAS in respiratory diseases. Translational human studies should be the focus for RAS drug development in lung diseases in the next decade.

Topics: Angiotensin I; Angiotensin II; Angiotensins; Fibrosis; Humans; Peptide Fragments; Receptor, Angiotensin, Type 1; Renin-Angiotensin System; Respiratory Tract Diseases; Signal Transduction

To verify whether Ang-(1-7) produces an antagonistic effect on Ang II-mediated atrial remodeling. Ang II-induced HL-1 cell model and a rat model of Ang II-induced atrial remodeling were constructed and intervened with Ang II Ang-(1-7), AngII +Ang-(1-7), Ang II+ c-Src specific inhibitor (SU6656), and Ang II + Ang-(1-7) + SSG (SHP-1/2 specific inhibitor, stibogluconate), respectively. The systolic blood pressure of the rat caudal artery was detected. And trial fibrosis was detected by Picrosirius red staining and Masson's trichrome staining. Expressions of transforming growth factor-β (TGF-β), tissue inhibitor of metalloproteinases 1 (TIMP1), Matrix metalloproteinase 2 (MMP-2), connective tissue growth factor (CTGF), galectin-3, α-smooth muscle actin (α-SMA), and collagen I/III were subjected to qPCR and western blot. Furthermore, SHP-1 binding to c-Src was verified by co-immunoprecipitation (Co-IP). Results showed that the expressions of TGF-β, TIMP1, MMP-2, CTGF, α-SMA, galectin-3, and collagen I were increased markedly in the Ang II intervention group, and the expressions of p-ERK1/2, p-Akt, and p-p38MAPK were also increased dramatically. Ang-(1-7) or SU6656 addition could inhibit the action of Ang II factor, thereby minimizing the expressions of the previously described genes and proteins. Simultaneously, SSG supplement reversed the antagonistic effect of Ang-(1-7) on Ang II, and the latter elevated the blood pressure and induced atrial fibrosis in rats. Ang-(1-7) could reverse the changes related to Ang II-induced atrial fibrosis in rats. In conclusion, Ang-(1-7) antagonized Ang II-induced atrial remodeling by regulating SHP-1 and c-Src, thereby affecting the MAPKs/Akt signaling pathway.

Topics: Angiotensin I; Angiotensin II; Animals; Fibrosis; Matrix Metalloproteinase 2; p38 Mitogen-Activated Protein Kinases; Peptide Fragments; Protein Tyrosine Phosphatase, Non-Receptor Type 6; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-fyn; Rats; Rats, Sprague-Dawley; Signal Transduction; src Homology Domains; Transforming Growth Factor beta

Topics: Angiotensin I; Angiotensin II; Animals; Cardiomegaly; Disease Models, Animal; Fibrosis; Hypertension; Imidazoles; Injections, Intraperitoneal; Losartan; Male; Peptide Fragments; Proto-Oncogene Mas; Rats; Rats, Sprague-Dawley; Sulfonamides; Thiophenes; Tyrosine 3-Monooxygenase

What is the central question of this study? Eccentric contraction exercises cause damage to muscle fibres and induce inflammatory responses. The exacerbation of this process can induce deposition of fibrous connective tissue, leading to decreased muscle function. The aim of this study was to examine the role of angiotensin-(1-7) in this context. What is the main finding and its importance? Our results show that oral treatment with angiotensin-(1-7) decreases muscle damage induced by eccentric exercise, reducing inflammation and fibrosis in the gastrocnemius and soleus muscles. This study shows a potential effect of angiotensin-(1-7) for the prevention of muscle injuries induced by physical exercise.. Eccentric contraction exercises cause damage to the muscle fibres and induce an inflammatory reaction. The protective effect of angiotensin-(1-7) [Ang-(1-7)] in skeletal muscle has led us to examine the role of this peptide in modifying processes associated with inflammation and fibrogenesis induced by eccentric exercise. In this study, we sought to investigate the effects of oral administration of Ang-(1-7) formulated in hydroxypropyl β-cyclodextrin (HPβ-CD) in prevention and treatment of muscle damage after downhill running. Male Wistar rats were divided into three groups: control (untreated and not exercised; n = 10); treated/exercised HPβ-CD Ang-(1-7) (n = 40); and treated/exercised HPβ-CD (n = 40). Exercised groups were subjected to a single eccentric contraction exercise session on a treadmill inclined to -13° at a constant speed of 20 m/min, for 60 min. Oral administration of HPβ-CD Ang-(1-7) and HPβ-CD was performed 3 h before the exercise protocol and daily as a single dose, until the end of the experiment. Samples were collected 4, 12, 24, 48 and 72 h after the exercise session. The animals treated with the Ang-(1-7) showed lower levels of creatine kinase, lower levels of tumor necrosis factor-α in soleus muscle and increased levels of interleukin-10 cytokines. The inflammatory cells and deposition of fibrous connective tissue in soleus and gastrocnemius muscles were lower in the group treated with Ang-(1-7). The results of this study show that treatment with an oral formulation of Ang-(1-7) enhances the process of repair of muscle injury induced by eccentric exercise.

Topics: Administration, Oral; Angiotensin I; Animals; Fibrosis; Male; Muscle, Skeletal; Peptide Fragments; Physical Conditioning, Animal; Rats; Rats, Wistar

Tsantan Sumtang, which consists of Choerospondias axillaris (Roxb.) Burtt et Hill, Myristica fragrans Houtt and Santalum album L, is a traditional and common prescription of Tibetan medicine. Tsantan Sumtang originates from Four Tantra with properties of nourishing heart and has been used as a folk medicine for cardiovascular diseases and heart failure in Qinghai, Tibet and Inner Mongolia. Our previous studies found that Tsantan Sumtang showed beneficial effects on right ventricular structure in hypoxia rats, while the underling mechanism remains unclear.. To elucidate the underlying mechanisms of Tsantan Sumtang attenuated right ventricular (RV) remodeling and fibrosis of chronic hypoxia-induced pulmonary arterial hypertension (HPAH) rats.. Our results showed that RVHI, RV/TL and RVSP were significantly increased in HPAH rat. Furthermore, levels of collagen I, collagen III and hydroxyproline were up-regulated in RV tissue under hypoxia. We found that RV hypertrophy and fibrosis were associated with increased expression of ACE, AngII, AT1R as well as decreased expression of ACE2, Ang1-7 and Mas. RV remodeling and fibrosis were attenuated after Tsantan Sumtang administration by up-regulating ACE2 and Mas level as well as down-regulating ACE, AngII and AT1R levels in RV tissue. 35 constituents in Tsantan Sumtang were identified.. Tsantan Sumtang attenuated RV remodeling and fibrosis in rat exposed to chronic hypoxia. The pharmacological effect of Tsantan Sumtang was based on equilibrating ACE-AngII-AT1R and ACE2-Ang1-7-Mas axis of RV tissue in HPAH rat.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Disease Models, Animal; Dose-Response Relationship, Drug; Fibrosis; Hypertrophy, Right Ventricular; Hypoxia; Male; Medicine, Tibetan Traditional; Peptide Fragments; Peptidyl-Dipeptidase A; Plant Preparations; Rats; Rats, Sprague-Dawley; Receptor, Angiotensin, Type 1; Ventricular Remodeling

Sarcopenia associated with chronic liver disease (CLD) is one of the more common extrahepatic features in patients with these pathologies. Among the cellular alterations observed in the muscle tissue under CLD is the decline in the muscle strength and function, as well as the increased fatigue. Morphological changes, such as a decrease in the fiber diameter and transition in the fiber type, are also reported. At the molecular level, sarcopenia for CLD is characterized by: i) a decrease in the sarcomeric protein, such as myosin heavy chain (MHC); ii) an increase in the ubiquitin-proteasome system markers, such as atrogin-1/MAFbx1 and MuRF-1/TRIM63; iii) an increase in autophagy markers, such as LC3II/LC3I ratio. Among the regulators of muscle mass is the renin-angiotensin system (RAS). The non-classical axis of RAS includes the Angiotensin 1-7 [Ang-(1-7)] peptide and its receptor Mas, which in skeletal muscle has anti-atrophic effect in models of muscle wasting induced by immobilization, lipopolysaccharide, myostatin or angiotensin II. In this paper, we evaluated the effect of Ang-(1-7) on the sarcopenia by CLD in a murine model induced by the 5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) hepatotoxin administered through diet. Our results show that Ang-(1-7) administration prevented the decline of the function and strength of muscle and increased the fatigue detected in the DDC-fed mice. Besides, we observed that the decreased fiber diameter and MHC levels, as well as the transition of fiber types, were all abolished by Ang-(1-7) in mice fed with DDC. Finally, Ang-(1-7) can decrease the atrogin-1 and MuRF-1 expression as well as the autophagy marker in mice treated with DDC. Together, our data support the protective role of Ang-(1-7) on the sarcopenia by CLD in mice.

Topics: Angiotensin I; Animals; Autophagy; Biomarkers; Chronic Disease; Fibrosis; Liver Diseases; Male; Mice; Mice, Inbred C57BL; Muscle Fibers, Skeletal; Muscle Proteins; Muscle Strength; Muscle, Skeletal; Muscular Atrophy; Peptide Fragments; Proteasome Endopeptidase Complex; Sarcopenia; SKP Cullin F-Box Protein Ligases; Tripartite Motif Proteins; Ubiquitin; Ubiquitin-Protein Ligases

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).. 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.. 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-β-induced myofibroblast activation was studied in primary cavernosal and immortalized fibroblasts.. Experimental results highlighted the essential role of the RAS in modulation of cavernosal fibrosis.. 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-β-induced fibroblast-to-myofibroblast transition.. The present study suggests Ang-(1-7) treatment as a possible strategy for pharmacological management of fibrosis-associated ED in aging.. 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.. 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.

Topics: Aged; Angiotensin I; Angiotensin II; Animals; Fibrosis; Humans; Male; Mice; Penile Erection; Penile Induration; Peptide Fragments; Peptidyl-Dipeptidase A; Rats; Rats, Wistar; Renin-Angiotensin System

Topics: Angiotensin I; Angiotensin II; Angiotensin II Type 1 Receptor Blockers; Angiotensin-Converting Enzyme 2; Angiotensin-Converting Enzyme Inhibitors; Animals; Captopril; Cell Line; Epithelial-Mesenchymal Transition; Fibrosis; Losartan; Lung; Male; Mice, Inbred C57BL; Oligopeptides; Peptide Fragments; Peptidyl-Dipeptidase A; Renin-Angiotensin System; Silicosis

Cardiac fibrosis is associated with most of heart diseases, but its molecular mechanism remains unclear. Anoctamin-1 (ANO1), a calcium-activated chloride channels (CaCCs) protein, plays a critical role in various pathophysiological processes. In the current study, we identified ANO1 expression in myocardial infarction (MI) model of rat and verified the role of ANO1 in cardiac fibrosis using transcriptomics combined with RNAi assays. we found that ANO1 expression was increased during the first two weeks, and decreased in the third week after MI. Fluorescence double labeling showed that ANO1 was mainly expressed in cardiac fibroblasts (CFs) and displayed an increased expression in CFs with proliferation tendency. The proliferation and secretion of CFs were markedly inhibited by knockdown of ANO1. RNA-Seq showed that most of the downregulation genes were related to the proliferation of CFs and cardiac fibrosis. After ANO1 knockdown, the expressions of angiotensin II type 1 receptor (AT1R) and cell nuclear proliferation antigen were markedly reduced, and the phosphorylation levels of MEK and ERK1/2 was decreased significantly, indicating that ANO1 regulate cardiac fibrosis through ATIR-mediated MAPK signaling pathway. These findings would be useful for the development of therapeutic strategies targeting ANO1 to treat and prevent cardiac fibrosis.

Topics: Angiotensin I; Animals; Animals, Newborn; Anoctamin-1; Disease Models, Animal; Fibroblasts; Fibrosis; Male; MAP Kinase Signaling System; Myocardial Infarction; Myocardium; Rats, Sprague-Dawley; Receptor, Angiotensin, Type 1; Signal Transduction

Studies implicate that angiotensin 1-7 (Ang1-7) imparts protective effects in the kidney. However, its relevance in hypertensive kidney disease is not fully understood. The purpose of this study was to explore the role of Ang1-7 on renal damage/remodeling during hypertension and its potential underlying molecular-cellular mechanisms.. Hypertension was induced in adult Sprague-Dawley rats by infusion of aldosterone (ALDO; 0.75 μg/hour) for 4 weeks with or without co-treatment of Ang1-7 (1 mg/kg/day). Untreated rats served as controls. Systolic blood pressure was monitored by tail-cuff technique. Renal fibrosis was evaluated by picrosirius red staining and renal collagen volume fraction was quantitated using imaging analyzing system. The expression of profibrotic factors [transforming growth factor-β1 (TGF-β1), platelet-derived growth factor-D (PDGF-D), fibroblast growth factor-1 (FGF-1), vascular endothelial growth factor-D (VEGF-D), and tissue inhibitors of metalloproteinases (TIMPs)] and free radical producing enzymes (inducible nitric oxide synthase and nicotinamide adenine dinucleotide phosphate [NADPH] oxidase) in the kidney were examined by reverse transcription-polymerase chain reaction and western blot. Renal oxidative stress was assessed by malondialdehyde (MDA) measurement.. Chronic ALDO infusion caused hypertension and hypertensive renal disease represented as glomerular damage/sclerosis. Ang1-7 co-treatment did not affect blood pressure in ALDO-treated rats, but significantly attenuated the glomerular damage/fibrosis. ALDO treatment significantly elevated renal expression of profibrogenic factors, including TGF-β1, TIMP-1/TIMP-2, FGF-1, PDGF-D, and VEGF-D, whereas Ang1-7 co-treatment significantly reduced renal TGF-β1, TIMP-1/TIMP-2, and FGF-1, but not PDGF-D and VEGF-D. Furthermore, ALDO infusion elevated NADPH oxidase (gp91phox) and MDA in the kidney, which was attenuated by Ang1-7 co-treatment.. Ang1-7 plays a protective role in the hypertensive kidney disease independent of blood pressure. The beneficial effects of Ang1-7 are likely mediated via suppressing TGF-β/FGF-1 pathways and oxidative stress.

Topics: Angiotensin I; Animals; Antihypertensive Agents; Blood Pressure; Blotting, Western; Disease Models, Animal; Fibrosis; Gene Expression Regulation; Hypertension, Renal; Kidney; Lymphokines; Male; Nephritis; Oxidative Stress; Peptide Fragments; Platelet-Derived Growth Factor; Rats; Rats, Sprague-Dawley; RNA; Tissue Inhibitor of Metalloproteinases; Vascular Endothelial Growth Factor D

Cell metabolic pathways are highly conserved among species and change rapidly in response to drug stimulation. Therefore, we explore the effects of angiotensin-(1-7) in a primary cell model of cardiac fibrosis established in angiotensin II-stimulated cardiac fibroblasts via metabolomics analysis and further clarify the potential protective mechanism of angiotensin-(1-7).. After exposing cardiac fibroblasts to angiotensin II and/or angiotensin-(1-7), 172 metabolites in these cells were quantified and identified by gas chromatography-mass spectrometry. The data were subsequently analyzed by orthogonal partial least squares discriminant analysis to shortlist biochemically significant metabolites associated with the antifibrotic action of angiotensin-(1-7). Seven significant metabolites were identified: 10,13-dimethyltetradecanoic acid, arachidonic acid, aspartic acid, docosahexaenoic acid (DHA), glutathione, palmitelaidic acid, and pyroglutamic acid. By metabolic network analysis, we found that these metabolites were involved in six metabolic pathways, including arachidonic acid metabolism, leukotriene metabolism, and the γ-glutamyl cycle. Since these metabolic pathways are related to calcium balance and oxidative stress, we further verified that angiotensin-(1-7) suppressed the abnormal extracellular calcium influx and excessive accumulation of intracellular reactive oxygen species (ROS) in angiotensin II-stimulated cardiac fibroblasts. Furthermore, we found that angiotensin-(1-7) suppressed the abnormal calcium- and ROS-dependent activation of calcium/calmodulin-dependent protein kinase II delta (CaMKIIδ), the increased expression of CaMKIIδ-related proteins (NADPH oxidase 4 (Nox4), cellular communication network factor 2 (CTGF), and p-ERK1/2), and excessive collagen deposition in vitro and in vivo.. Angiotensin-(1-7) can ameliorate the angiotensin II-stimulated metabolic perturbations associated with cardiac fibroblast activation. These metabolic changes indicate that modulation of calcium- and ROS-dependent activation of CaMKIIδ mediates the activity of angiotensin-(1-7) against cardiac fibrosis. Moreover, pyroglutamic acid and arachidonic acid may be potential biomarkers for monitoring the antifibrotic action of angiotensin-(1-7).

Topics: Angiotensin I; Angiotensin II; Animals; Arachidonic Acid; Calcium; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Cell Differentiation; Cell Proliferation; Cells, Cultured; Fibroblasts; Fibrosis; Gas Chromatography-Mass Spectrometry; Glutathione; Heart Diseases; Male; Metabolome; Oxidative Stress; Peptide Fragments; Rats, Sprague-Dawley; Reactive Oxygen Species

Cardiac pressure and humoral factors induce cardiac hypertrophy and fibrosis, which are characterized by increased stiffness, reduced contractility and altered perfusion. Angiotensin II (AngII) is well known to promote this pathology. Angiotensin-converting enzyme (ACE) 2, which cleaves AngII and forms Ang-(1-7), exerts protective anti-hypertrophy and anti-fibrosis effects. A disintegrin and metalloproteinase 17 (ADAM17), a membrane-bound enzyme reported to cleave ACE2, may participate in the pathological process of AngII perfusion-induced heart damage. However, researchers have not clearly determined whether dickkopf-3 (DKK3) regulates the ADAM17/ACE2 pathway and, if so, whether DKK3-mediated regulation is related to the glycogen synthase kinase-3β (GSK-3β)/β-catenin pathway. In this study, we explored whether DKK3 overexpression ameliorates the development of AngII-induced cardiac fibrosis and hypertrophy through the ADAM17/ACE2 and GSK-3β/β-catenin pathways.. Mice were injected with a DKK3-overexpressing adenovirus or vehicle and then infused with AngII or saline using subcutaneously implanted mini-pumps for four weeks. Hearts were stained with hematoxylin-eosin, Masson's trichrome and immunohistochemical markers for histology. Primary fibroblasts were treated with the adenovirus and AngII and then examined using western blotting, EdU (5-ethynyl-2'-deoxyuridine) assays and immunofluorescence. Additionally, siRNA silencing was performed to study the role of DKK3 and the involved pathways.. AngII-induced cardiac hypertrophy and interstitial and perivascular fibrosis were less severe in DKK3-overexpressing mice than in control mice. Moreover, the expression levels of fibrotic genes, such as collagen I and III, and the hypertrophic genes atrial natriuretic peptide (ANP) and beta-myosin heavy chain (β-MHC) were decreased. DKK3 overexpression also exerted a protective effect by inhibiting ADAM17 phosphorylation, thus increasing ACE2 expression and subsequently promoting AngII degradation. Furthermore, this process was mediated by the inhibition of GSK-3β and β-catenin and decreased translocation of β-catenin to the nucleus. On the other hand, the DKK3 knockdown by siRNA achieved opposite results.. DKK3 overexpression substantially alleviated AngII infusion-induced cardiac hypertrophy and fibrosis by regulating ADAM17/ACE2 pathway activity and inhibiting the GSK-3β/β-catenin pathway.

Topics: ADAM17 Protein; Adaptor Proteins, Signal Transducing; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Animals, Newborn; Apoptosis; beta Catenin; Cardiomegaly; Cell Proliferation; Disease Models, Animal; Fibroblasts; Fibrosis; Glycogen Synthase Kinase 3 beta; Inflammation; Intercellular Signaling Peptides and Proteins; Matrix Metalloproteinases; Mice, Inbred C57BL; Peptide Fragments; Peptidyl-Dipeptidase A; Perfusion; Phosphorylation; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta1

The development of new therapeutic strategies to control or reverse hepatic fibrosis requires thorough knowledge about its molecular and cellular basis. It is known that the heptapeptide angiotensin-(1-7) [ang-(1-7)] can reduce hepatic fibrosis and steatosis in vivo; therefore, it is important to uncover the mechanisms regulating its activity and cellular model of investigation. Ang-(1-7) is a peptide of the renin-angiotensin system (RAS), and here we investigated its modulatory effect on the expression pattern of microRNAs (miRNAs) in hepatic stellate cells (HSCs) LX-2, which transdifferentiate into fibrogenic and proliferative cells. We compared the miRNA profiles between quiesced, activated and ang-(1-7)-treated activated HSCs to identify miRNAs that may regulate their transdifferentiation. Thirteen miRNAs were pointed, and cellular and molecular analyses identified miRNA-1914-5p as a molecule that contributes to the effects of ang-(1-7) on lipid metabolism and on the pro-fibrotic environment control. In our cellular model, we also analyzed the regulators of fatty acid metabolism. Specifically, miRNA-1914-5p regulates the expression of malonyl-CoA decarboxylase (MLYCD) and phosphatidic acid phosphohydrolase (PAP or Lipin-1). Additionally, Lipin-1 was closely correlated with mRNA expression of peroxisome proliferator-activated receptors (PPAR)-α and -γ, which also contribute to lipid homeostasis and to the reduction of TGF-β1 expression. These findings provide a novel link between RAS and lipid metabolism in controlling HSCs activation.

Topics: Angiotensin I; Cell Transdifferentiation; Cells, Cultured; Fibrosis; Gene Expression Regulation; Hepatic Stellate Cells; Humans; Lipid Metabolism; MicroRNAs; Peptide Fragments; Signal Transduction; Vasodilator Agents

The renin-angiotensin system (RAS), especially the angiotensin II (Ang II)/angiotensin II type 1 receptor (AT1R) axis, plays an important role in the aging process of the kidney, through increased tissue reactive oxygen species production and progressively increased oxidative stress. In contrast, the angiotensin 1-7 (Ang 1-7)/Mas receptor (MasR) axis, which counteracts the effects of Ang II, is protective for end-organ damage. To evaluate the ability of resveratrol (RSV) to modulate the RAS in aging kidneys, eighteen-month-old male C57BL/6 mice were divided into two groups that received either normal mouse chow or chow containing resveratrol, for six months. Renal expressions of RAS components, as well as pro- and antioxidant enzymes, were measured and mouse kidneys were isolated for histopathology. Resveratrol-treated mice demonstrated better renal function and reduced albuminuria, with improved renal histologic findings. Resveratrol suppressed the Ang II/AT1R axis and enhanced the AT2R/Ang 1-7/MasR axis. Additionally, the expression of nicotinamide adenine dinucleotide phosphate oxidase 4, 8-hydroxy-2'-deoxyguanosine, 3-nitrotyrosine, collagen IV, and fibronectin was decreased, while the expression of endothelial nitric oxide synthase and superoxide dismutase 2 was increased by resveratrol treatment. These findings demonstrate that resveratrol exerts protective effects on aging kidneys by reducing oxidative stress, inflammation, and fibrosis, through Ang II suppression and MasR activation.

Topics: Albuminuria; Angiotensin I; Angiotensin II; Animals; Anti-Inflammatory Agents; Antioxidants; Collagen Type IV; Fibronectins; Fibrosis; Kidney; Male; Mice, Inbred C57BL; NADPH Oxidases; Nitric Oxide Synthase Type III; Oxidative Stress; Peptide Fragments; Plant Extracts; Proto-Oncogene Proteins; Receptors, Angiotensin; Receptors, G-Protein-Coupled; Renal Insufficiency, Chronic; Renin-Angiotensin System; Resveratrol; Superoxide Dismutase

The objectives of the present study were to investigate the effect of ANG-(1-7) on the development of cardiac hypertrophy and to identify the intracellular mechanism underlying this action of ANG-(1-7). Blood pressure and heart rate were recorded using radiotelemetry before and after chronic subcutaneous infusion of control (PBS), ANG II, ANG-(1-7), or ANG II + ANG-(1-7) for 4 wk in normotensive rats. Chronic administration of ANG-(1-7) did not affect either basal blood pressure or the ANG II-induced elevation in blood pressure. However, ANG-(1-7) significantly attenuated ANG II-induced cardiac hypertrophy and perivascular fibrosis in these rats. These effects of ANG-(1-7) were confirmed in cultured cardiomyocytes, in which ANG-(1-7) significantly attenuated ANG II-induced increases in cell size. This protective effect of ANG-(1-7) was significantly attenuated by pretreatment with A779 (a Mas receptor antagonist) or Mito-TEMPO (a mitochondria-targeting superoxide scavenger) as well as blockade of Sirt3 (a deacetylation-acting protein) by viral vector-mediated overexpression of sirtuin (Sirt)3 short hairpin (sh)RNA. Western blot analysis demonstrated that treatment with ANG-(1-7) dramatically increased Sirt3 expression. In addition, ANG-(1-7) attenuated the ANG II-induced increase in mitochondrial ROS generation, an effect that was abolished by A779 or Sirt3 shRNA. Moreover, ANG-(1-7) increased FoxO3a deacetylation and SOD2 expression, and these effects were blocked by Sirt3 shRNA. In summary, the protective effects of ANG-(1-7) on ANG II-induced cardiac hypertrophy and increased mitochondrial ROS production are mediated by elevated SOD2 expression via stimulation of Sirt3-dependent deacetylation of FoxO3a in cardiomyocytes. Thus, activation of the ANG-(1-7)/Sirt3 signaling pathway could be a novel therapeutic strategy in the management of cardiac hypertrophy and associated complications.

Topics: Angiotensin I; Angiotensin II; Angiotensin Receptor Antagonists; Animals; Cardiomegaly; Cardiotonic Agents; Cell Size; Fibrosis; Male; Myocytes, Cardiac; Peptide Fragments; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Signal Transduction; Sirtuins; Superoxide Dismutase

Myocardial remodeling serves an important role in the pathophysiology of coronary heart disease. The angiotensin-converting enzyme (ACE)2-angiotensin-(1-7) [Ang (1‑7)]‑Mas receptor (MasR) axis is a key regulator in myocardial remodeling and development of heart failure. To investigate how ACE2‑Ang‑(1‑7)‑MasR axis function on myocardial remodeling and cardiac fibrosis in post‑myocardial infarction (MI), male Sprague‑Dawley rats (weight, 200±20 g) were used to establish the model of myocardial infarction by ligating the left coronary artery. The present study suggests that telmisartan (Tel) and olmesartan (Olm) (5 mg/kg/d) can inhibit myocardial remodeling of post‑myocardial infarction through the ACE2‑Ang (1‑7)‑MasR pathway. Administration of Tel or Olm was demonstrated to significantly inhibit collagen deposition using Masson staining. In addition, telmisartan and olmesartan was indicated to antagonize angiotensin II (Ang II) and upregulate ACE2, MasR, Ang (1‑7) expression in myocardial tissue using immunoassay and ELISA test, and the effect of Olm was more marked than that of Tel at the same dosage. Simultaneously, compared with the MI or Sham group, the mRNA and protein expression of ACE2, Ang II and MasR in myocardial tissue demonstrated a remarkable increase in the Olm group, when compared with the Tel group. Taken together, our data demonstrated that ACE2‑Ang (1‑7)‑MasR axis may present a potential protective role in the development of myocardial remodeling and may provide a new target for drug development of cardiac fibrosis. In conclusion, Olm is superior to Tel in inhibiting myocardial local Ang II level reducing myocardial collagen deposition and improving myocardial remodeling by upregulating the expression of ACE2, Ang (1‑7) and MasR.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Collagen; Fibrosis; Heart Function Tests; Immunohistochemistry; Male; Myocardial Infarction; Peptide Fragments; Peptidyl-Dipeptidase A; Proto-Oncogene Mas; Proto-Oncogene Proteins; Rats, Sprague-Dawley; Receptors, G-Protein-Coupled; RNA, Messenger; Ventricular Dysfunction, Left; Ventricular Remodeling

Radiation-induced fibrosis (RIF) of musculoskeletal tissue is a common complication of radiation therapy for extremity soft-tissue sarcoma, with no standardized strategy for prevention and treatment. Angiotensin-(1-7) (Ang-[1-7]), a well-tolerated endogenous heptapeptide hormone with antitumor and antifibrotic properties, was tested as a radioprotectant for RIF and stiffening of irradiated muscles.. Male CD-1 mice were randomized to one of three treatment groups: control, simulated sarcoma radiation therapy to the gastrocnemius and soleus muscles, or radiation therapy along with continuous Ang-(1-7) delivery initiated three days before radiation therapy. The biologically equivalent dose of radiation (∼100.3 Gy) absorbed by normal musculature during the course of radiation therapy for extremity sarcoma was delivered by means of four dose fractions of 7.3 Gy over two weeks. Fibrosis (n = 5 per group) and mechanical properties (n = 4 to 6 per group) of the muscles were measured at six weeks and four months after radiation therapy, and the intramuscular concentration of the profibrotic cytokines transforming growth factor-beta (TGF-β) and connective tissue growth factor (CTGF) (n = 8 to 10 per group) were measured at six weeks.. Interstitial (p < 0.01) and perivascular (p < 0.05) fibrosis increased significantly in the muscles treated with radiation therapy alone versus the nonirradiated controls at both six weeks (interstitial, +89%; perivascular, +112%) and four months (interstitial, +154%; perivascular, +88%). The muscles treated with radiation alone also exhibited increased tension (p < 0.01) versus nonirradiated controls at both six weeks (+779%) and four months (+1761%) when placed under 5% strain, and at four months (+1390%; p < 0.001) under 10% strain. At four months, muscle stiffness had increased in the mice treated with radiation therapy alone (+90%; p = 0.002) compared with nonirradiated controls. TGF-β production was also greater in this group at six weeks (+37%; p = 0.06) versus control. Ang-(1-7) administration prevented RIF and stiffening, with no differences observed for any other outcome between those receiving radiation therapy with Ang-(1-7) and the nonirradiated controls. Likewise, Ang-(1-7) mitigated the increase in TGF-β and CTGF concentration from radiation therapy.. Ang-(1-7) attenuated RIF, stiffening, and production of profibrotic cytokines that were elevated in mouse skeletal muscles after simulated radiation therapy for extremity sarcoma.. Ang-(1-7) may serve as a potential therapy for the prevention of RIF in patients who require radiation therapy as adjuvant treatment for soft-tissue sarcoma.

Topics: Analysis of Variance; Angiotensin I; Animals; Biopsy, Needle; Disease Models, Animal; Fibrosis; Hindlimb; Immunohistochemistry; Male; Mice; Mice, Inbred Strains; Muscle Neoplasms; Muscle, Skeletal; Peptide Fragments; Random Allocation; Reference Values; Sarcoma, Experimental; Sensitivity and Specificity; Spasm

Congestive heart failure is one of the most prevalent and deadly complications of type 2 diabetes that is frequently associated with pulmonary dysfunction. Among many factors that contribute to development and progression of diabetic complications is angiotensin II (Ang2). Activation of pathological arm of renin-angiotensin system results in increased levels of Ang2 and signaling through angiotensin type 1 receptor. This pathway is well recognized for its role in induction of oxidative stress (OS), inflammation, hypertrophy and fibrosis. Angiotensin (1-7) [A(1-7)], through activation of Mas receptor, opposes the actions of Ang2 which can result in the amelioration of diabetic complications; enhancing the overall welfare of diabetic patients. In this study, 8 week-old db/db mice were administered A(1-7) daily via subcutaneous injections. After 16 weeks of treatment, echocardiographic assessment of heart function demonstrated significant improvement in cardiac output, stroke volume and shortening fraction in diabetic animals. A(1-7) also prevented cardiomyocyte hypertrophy, apoptosis, lipid accumulation, and decreased diabetes-induced fibrosis and OS in the heart tissue. Treatment with A(1-7) reduced levels of circulating proinflammatory cytokines that contribute to the low grade inflammation observed in diabetes. In addition, lung pathologies associated with type 2 diabetes, including fibrosis and congestion, were decreased with treatment. OS and macrophage infiltration were also reduced in the lungs after treatment with A(1-7). Long-term administration of A(1-7) to db/db mice is effective in improving heart and lung function in db/db mice. Treatment prevented pathological remodeling of the tissues and reduced OS, fibrosis and inflammation.

Topics: Angiotensin I; Animals; Anti-Inflammatory Agents; Antioxidants; Apoptosis; Cardiotonic Agents; Cytokines; Diabetes Mellitus, Type 2; Disease Models, Animal; Fibrosis; Heart; Hypoglycemic Agents; Lipid Metabolism; Lung; Male; Mice; Myocardium; Oxidative Stress; Peptide Fragments

Angiotensin-converting enzyme 2 (ACE2) has been shown to prevent atherosclerotic lesions and renal inflammation. However, little was elucidated upon the effects and mechanisms of ACE2 in atherosclerotic kidney fibrosis progression. Here, we examined regulatory roles of ACE2 in renal fibrosis in the apolipoprotein E (ApoE) knockout (KO) mice. The ApoEKO mice were randomized to daily deliver either angiotensin (Ang) II (1.5mg/kg) and/or human recombinant ACE2 (rhACE2; 2mg/kg) for 2 weeks. Downregulation of ACE2 and upregulation of phosphorylated Akt, mTOR and ERK1/2 levels were observed in ApoEKO kidneys. Ang II infusion led to increased tubulointerstitial fibrosis in the ApoEKO mice with greater activation of the mTOR/ERK1/2 signaling. The Ang II-mediated renal fibrosis and structural injury were strikingly rescued by rhACE2 supplementation, associated with reduced mRNA expression of TGF-β1 and collagen I and elevated renal Ang-(1-7) levels. In cultured mouse kidney fibroblasts, exposure with Ang II (100nmolL(-1)) resulted in obvious elevations in superoxide generation, phosphorylated levels of mTOR and ERK1/2 as well as mRNA levels of TGF-β1, collagen I and fibronectin 1, which were dramatically prevented by rhACE2 (1mgmL(-1)) or mTOR inhibitor rapamycin (10μmolL(-1)). These protective effects of rhACE2 were eradicated by the Ang-(1-7)/Mas receptor antagonist A779 (1μmolL(-1)). Our results demonstrate the importance of ACE2 in amelioration of kidney fibrosis and renal injury in the ApoE-mutant mice via modulation of the mTOR/ERK signaling and renal Ang-(1-7)/Ang II balance, thus indicating potential therapeutic strategies by enhancing ACE2 action for preventing atherosclerosis and fibrosis-associated kidney disorders.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Animals; Apolipoproteins E; Atherosclerosis; Fibrosis; Kidney; Kidney Diseases; Male; MAP Kinase Signaling System; Mice, Inbred C57BL; Mice, Knockout; Peptide Fragments; Peptidyl-Dipeptidase A; Sirolimus; TOR Serine-Threonine Kinases

The objective of this article is to investigate the renoprotecive effects of exendin-4 in a mouse model of unilateral ureteral obstruction (UUO) and explore the putative mechanisms.. Male Balbc mice underwent sham operation or UUO surgery, and then received intraperitoneal injection of vehicle or exendin-4, respectively. After 14 days, mice were sacrificed and the left kidneys were collected and analyzed by histology, immunohistochemistry, Western blot, quantitative real-time reverse transcription polymerase chain reaction, radioimmunoassay and enzyme-linked immunosorbent assay.. As compared to the sham group, mice that underwent UUO surgery developed more severe tubular injury and interstitial fibrosis, as well as higher expression of fibronectin (FN), collagen-1 (Col-1) and α-smooth muscle actin (α-SMA). Also, we observed higher expression of angiotensin-converting enzyme (ACE) while lower expression of angiotensin-converting enzyme 2 (ACE2), higher levels of intrarenal angiotensin II (Ang II) while lower levels of intrarenal angiotensin-(1-7), and higher expression of transforming growth factor β1 (TGF-β1) and phosphorylation of Smad3 (p-Smad3) in the obstructed kidneys. Impressively, these pathologic changes were significantly attenuated in the mice group of UUO treated with exendin-4.. Our present study indicates for the first time that exendin-4 exerts renoprotective effects in an experimental model of UUO, partly through regulating the balance of the intrarenal renin-angiotensin system and then inhibiting the Ang II-mediated TGF-β1/Smad3 signaling pathway.

Topics: Actins; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Collagen Type I; Exenatide; Fibronectins; Fibrosis; Kidney; Male; Mice, Inbred BALB C; Peptide Fragments; Peptides; Peptidyl-Dipeptidase A; Renin-Angiotensin System; RNA, Messenger; Signal Transduction; Smad3 Protein; Transforming Growth Factor beta1; Ureteral Obstruction; Venoms

The effect of the heptapeptide hormone Ang-(1-7) on microvascular fibrosis in rats with Ang II-induced hypertension was investigated, since vascular fibrosis/remodeling plays a prominent role in hypertension-induced end-organ damage and Ang-(1-7) inhibits vascular growth and fibrosis.. Fibrosis of cremaster microvessels was studied in male Lewis rats infused with Ang II and/or Ang-(1-7).. Ang II elevated systolic blood pressure by approximately 40 mmHg, while blood pressure was not changed by Ang-(1-7). Ang II increased perivascular fibrosis surrounding 20-50 μm arterioles as well as interstitial fibrosis; coadministration of Ang-(1-7) prevented the increases in fibrosis. The fibrotic factor CTGF and phospho-Smad 2/3, which upregulates CTGF, were increased by Ang II; this effect was prevented by coadministration of Ang-(1-7). Although TGF-β phosphorylates Smad 2/3, TGF-β was no different among treatment groups. In contrast, Ang II increased the MAP kinase phospho-ERK1/2, which also phosphorylates Smad; p-ERK was reduced by Ang-(1-7). Ang-(1-7), in the presence or absence of Ang II, upregulated the MAP kinase phosphatase DUSP1.. These results suggest that Ang-(1-7) increases DUSP1 to reduce MAP kinase/Smad/CTGF signaling and decrease fibrosis in resistance arterioles, to attenuate end-organ damage associated with chronic hypertension.

Topics: Angiotensin I; Angiotensin II; Animals; Antihypertensive Agents; Arterioles; Blood Pressure; Chronic Disease; Connective Tissue Growth Factor; Dual Specificity Phosphatase 1; Fibrosis; Hypertension; Male; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 3; Muscle, Skeletal; Peptide Fragments; Phosphorylation; Rats; Rats, Inbred Lew; Smad2 Protein; Smad3 Protein; Transforming Growth Factor beta; Vasoconstrictor Agents

We investigated the relationship between Ang-(1-7) [angiotensin-(1-7)] action, sHTN (systolic hypertension), oxidative stress, kidney injury, ACE2 (angiotensin-converting enzyme-2) and MasR [Ang-(1-7) receptor] expression in Type 1 diabetic Akita mice. Ang-(1-7) was administered daily [500 μg/kg of BW (body weight) per day, subcutaneously] to male Akita mice from 14 weeks of age with or without co-administration of an antagonist of the MasR, A779 (10 mg/kg of BW per day). The animals were killed at 20 weeks of age. Age-matched WT (wild-type) mice served as controls. Ang-(1-7) administration prevented sHTN and attenuated kidney injury (reduced urinary albumin/creatinine ratio, glomerular hyperfiltration, renal hypertrophy and fibrosis, and tubular apoptosis) without affecting blood glucose levels in Akita mice. Ang-(1-7) also attenuated renal oxidative stress and the expression of oxidative stress-inducible proteins (NADPH oxidase 4, nuclear factor erythroid 2-related factor 2, haem oxygenase 1), pro-hypertensive proteins (angiotensinogen, angiotensin-converting enzyme, sodium/hydrogen exchanger 3) and profibrotic proteins (transforming growth factor-β1 and collagen IV), and increased the expression of anti-hypertensive proteins (ACE2 and MasR) in Akita mouse kidneys. These effects were reversed by A779. Our data suggest that Ang-(1-7) plays a protective role in sHTN and RPTC (renal proximal tubular cell) injury in diabetes, at least in part, through decreasing renal oxidative stress-mediated signalling and normalizing ACE2 and MasR expression.

Topics: Analysis of Variance; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Blood Glucose; Blotting, Western; Diabetes Mellitus, Type 1; Fibrosis; Gene Expression Regulation; Histological Techniques; Hypertension; Immunohistochemistry; Injections, Subcutaneous; Kidney Diseases; Kidney Tubules, Proximal; Male; Mice; Oxidative Stress; Peptide Fragments; Peptidyl-Dipeptidase A; Proto-Oncogene Mas; Proto-Oncogene Proteins; Real-Time Polymerase Chain Reaction; Receptors, G-Protein-Coupled

In diabetic patients, left ventricular (LV) remodeling is highly prevalent; however, little is known about the impact of diabetes on right ventricular (RV) structure and function. We recently found that overexpression of angiotensin (ANG)-converting enzyme 2 (ACE2), which metabolizes ANG-II to ANG-(1-7) and ANG-I to ANG-(1-9), may improve LV remodeling in diabetic cardiomyopathy (DCM). Here, we aimed to assess whether LV remodeling and dysfunction are paralleled by RV alterations and the effects of ANG-(1-7) on RV remodeling in DCM. After 12 wk of diabetes induced by a single intraperitoneal injection of streptozotocin, rats were treated with saline, ANG-(1-7), perindopril, ANG-(1-7) plus perindopril, ANG-(1-7) plus Mas receptor antagonist A779, or ANG-(1-7) plus ANG-II type 2 receptor antagonist PD123319 for 4 wk. RV remodeling in diabetic rats was indicated by fibrosis of the RV free wall in the absence of hypertrophy and apoptosis. Treatment with ANG-(1-7) prevented diabetes-induced RV fibrosis and dysfunction. ANG-(1-7) (800 ng·kg(-1)·min(-1)) was superior to perindopril in improving RV fibrosis. The major mechanisms involved a complex interaction of ANG-II type 2 and Mas receptors for subsequent downregulation of ACE expression and activity and ANG-II type 1 receptor expression, as well as upregulation of ACE2 expression and activity and the expression of ANG-II type 2 receptor and sarco(endo)plasmic reticulum Ca(2+)-ATPase. Thus RV fibrosis and dysfunction plays a central role in DCM, and ANG-(1-7) mitigates diabetes-induced RV alterations.

Topics: Angiotensin I; Animals; Blood Glucose; Cells, Cultured; Coculture Techniques; Collagen; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Diabetic Cardiomyopathies; Fibrosis; Heart Ventricles; Lipids; Male; Oxidative Stress; Peptide Fragments; Proto-Oncogene Mas; Proto-Oncogene Proteins; Rats, Wistar; Receptor, Angiotensin, Type 2; Receptors, G-Protein-Coupled; Sarcoplasmic Reticulum Calcium-Transporting ATPases; Signal Transduction; Time Factors; Transforming Growth Factor beta1; Ventricular Dysfunction, Right; Ventricular Function, Right; Ventricular Remodeling

We recently found that overexpression of angiotensin (Ang)-converting enzyme 2, which metabolizes Ang-II to Ang-(1-7) and Ang-I to Ang-(1-9), may improve left ventricular remodeling in diabetic cardiomyopathy. Here we aimed to test whether chronic infusion of Ang-(1-7) can dose-dependently ameliorate left ventricular remodeling and function in a rat model of diabetic cardiomyopathy and whether the combination of Ang-(1-7) and Ang-converting enzyme inhibition may be superior to single therapy. Our results showed that Ang-(1-7) treatment dose-dependently ameliorated left ventricular remodeling and dysfunction in diabetic rats by attenuating myocardial fibrosis, myocardial hypertrophy and myocyte apoptosis via both the Mas receptor and angiotensin II type 2 receptor. Furthermore, combining Ang-(1-7) with perindopril provided additional cardioprotection relative to single therapy. Ang-(1-7) administration provides a novel and promising approach for treatment of diabetic cardiomyopathy.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Animals; Apoptosis; Blood Glucose; Cardiomegaly; Cell Communication; Cell Differentiation; Cell Proliferation; Collagen; Diabetic Cardiomyopathies; Disease Models, Animal; Drug Therapy, Combination; Echocardiography; Fibroblasts; Fibrosis; Heart Ventricles; Hemodynamics; Male; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Peptide Fragments; Peptidyl-Dipeptidase A; Perindopril; Phosphorylation; Rats; Receptors, Angiotensin; Transforming Growth Factor beta1; Ventricular Dysfunction, Left

Duchenne muscular dystrophy (DMD) is the most common inherited neuromuscular disease, and is characterized by the lack of dystrophin, muscle wasting, increased transforming growth factor (TGF)-β Smad-dependent signalling and fibrosis. Acting via the Mas receptor, angiotensin-1-7 [Ang-(1-7)], is part of the renin-angiotensin system, with the opposite effect to that of angiotensin II. We hypothesized that the Ang-(1-7)/Mas receptor axis might protect chronically damaged tissues as in skeletal muscle of the DMD mouse model mdx. Infusion or oral administration of Ang-(1-7) in mdx mice normalized skeletal muscle architecture, decreased local fibrosis and improved muscle function in vitro and in vivo. These positive effects were mediated by the inhibition of TGF-β Smad signalling, which in turn led to reduction of the pro-fibrotic microRNA miR-21 concomitant with a reduction in the number of TCF4 expressing fibroblasts. Mdx mice infused with Mas antagonist (A-779) and mdx deficient for the Mas receptor showed highly deteriorated muscular architecture, increased fibrosis and TGF-β signalling with diminished muscle strength. These results suggest that this novel compound Ang-(1-7) might be used to improve quality of life and delay death in individuals with DMD and this drug should be investigated in further pre-clinical trials.

Topics: Angiotensin I; Animals; Disease Models, Animal; Extracellular Matrix; Fibroblasts; Fibrosis; Humans; Male; Mice; Mice, Inbred mdx; Mice, Knockout; MicroRNAs; Muscle Strength; Muscle, Skeletal; Muscular Dystrophy, Duchenne; Peptide Fragments; Receptors, Cell Surface; Signal Transduction; Transforming Growth Factor beta

Angiotensin-converting enzyme 2 (ACE2) cleaves angiotensin (Ang) II to generate Ang1-7, which mediates cellular actions through Mas receptors (MasR). Hypertension is accompanied by high or low circulating AngII levels and cardiac/renal injury. The purpose of this study is to explore (i) whether circulating AngII affects ACE2/MasR expressions in the hypertensive heart and kidney; and (ii) whether Ang1-7 regulates cardiac repair/remodeling responses through MasR during hypertension.. In the first portion of the study, rats received either an AngII infusion (400ng/kg/min) for 4 weeks, leading to hypertension with high circulating AngII, or an aldosterone (ALDO, 0.75 μg/h) infusion for 4 weeks, leading to hypertension with low/normal circulating AngII. Cardiac and renal ACE2/MasR expressions were examined. We found that cardiac ACE2 was increased and MasR attenuated in both AngII and ALDO groups. However, renal ACE2 and MasR remained unchanged in both AngII- and ALDO-treated animals.. In the second portion, rats received AngII infusion with/without MasR antagonist (A779, 1mg/kg/day) for 4 weeks. The roles of MasR blockade in cardiac inflammation, fibrosis, apoptosis, and ventricular function were examined. Chronic AngII infusion caused scattered cardiac injuries, and A779 cotreatment exacerbated cardiac injury, resulting in aggravated inflammatory, fibrogenic, and apoptotic responses compared with the AngII group. Cardiac function, however, was unaltered in the AngII and A779 groups.. ACE2 and MasR expressions in the hypertensive heart and kidney are not regulated by circulating AngII levels. Ang1-7 is involved in multiple repair responses, suggesting that therapeutic strategies aimed at administering Ang1-7 hold potential for the management of cardiac remodeling.

Topics: Aldosterone; Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Apoptosis; Autocrine Communication; Disease Models, Animal; Fibrosis; Hypertension; Kidney; Male; Myocardium; Paracrine Communication; Peptide Fragments; Peptidyl-Dipeptidase A; Proto-Oncogene Mas; Proto-Oncogene Proteins; Rats, Sprague-Dawley; Receptors, G-Protein-Coupled; Signal Transduction; Ventricular Remodeling

Muscular dystrophies are a group of heterogeneous genetic disorders that cause progressive muscle weakness and wasting, dilated cardiomyopathy and early mortality. There are different types of muscular dystrophies with varying aetiologies but they all have a common hallmark of myofibre degeneration, atrophy and decreased mobility. Mutation in Sgcd (sarcoglycan-δ), a subunit of dystrophin glycoprotein complex, causes LGMD2F (limb girdle muscular dystrophy 2F). Previously, we have reported that Sgcd-deficient (Sgcd-/-) mice exhibit AngII (angiotensin II)-induced autonomic and skeletal muscle dysfunction at a young age, which contributes to onset of dilated cardiomyopathy and mortality at older ages. Two counter-regulatory RAS (renin-angiotensin system) pathways have been identified: deleterious actions of AngII acting on the AT1R (AngII type 1 receptor) compared with the protective actions of Ang-(1-7) [angiotensin-(1-7)] acting on the receptor Mas. We propose that the balance between the AngII/AT1R and Ang-(1-7)/Mas axes is disturbed in Sgcd-/- mice. Control C57BL/6J and Sgcd-/- mice were treated with Ang-(1-7) included in hydroxypropyl β-cyclodextrin (in drinking water) for 8-9 weeks beginning at 3 weeks of age. Ang-(1-7) treatment restored the AngII/AT1R compared with Ang-(1-7)/Mas balance, decreased oxidative stress and fibrosis in skeletal muscle, increased locomotor activity, and prevented autonomic dysfunction without lowering blood pressure in Sgcd-/- mice. Our results suggest that correcting the early autonomic dysregulation by administering Ang-(1-7) or enhancing its endogenous production may provide a novel therapeutic approach in muscular dystrophy.

Topics: Administration, Oral; Angiotensin I; Animals; Dystrophin; Fibrosis; Mice; Mice, Inbred C57BL; Mice, Knockout; Motor Activity; Muscle, Skeletal; Muscular Dystrophies; Peptide Fragments; Phenotype; Sarcoglycans

The renin-angiotensin system, especially angiotensin II (ANG II), plays a key role in the development and progression of diabetic nephropathy. ANG 1-7 has counteracting effects on ANG II and is known to exert beneficial effects on diabetic nephropathy. We studied the mechanism of ANG 1-7-induced beneficial effects on diabetic nephropathy in db/db mice. We administered ANG 1-7 (0.5 mg·kg(-1)·day(-1)) or saline to 5-mo-old db/db mice for 28 days via implanted micro-osmotic pumps. ANG 1-7 treatment reduced kidney weight and ameliorated mesangial expansion and increased urinary albumin excretion, characteristic features of diabetic nephropathy, in db/db mice. ANG 1-7 decreased renal fibrosis in db/db mice, which correlated with dephosphorylation of the signal transducer and activator of transcription 3 (STAT3) pathway. ANG 1-7 treatment also suppressed the production of reactive oxygen species via attenuation of NADPH oxidase activity and reduced inflammation in perirenal adipose tissue. Furthermore, ANG 1-7 treatment decreased lipid accumulation in db/db kidneys, accompanied by increased expressions of renal adipose triglyceride lipase (ATGL). Alterations in ATGL expression correlated with increased SIRT1 expression and deacetylation of FOXO1. The upregulation of angiotensin-converting enzyme 2 levels in diabetic nephropathy was normalized by ANG 1-7. ANG 1-7 treatment exerts renoprotective effects on diabetic nephropathy, associated with reduction of oxidative stress, inflammation, fibrosis, and lipotoxicity. ANG 1-7 can represent a promising therapy for diabetic nephropathy.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Animals; Diabetic Nephropathies; Fibrosis; Forkhead Box Protein O1; Forkhead Transcription Factors; Kidney; Lipase; Male; Mice; Mice, Inbred C57BL; Oxidative Stress; Peptide Fragments; Peptidyl-Dipeptidase A; Reactive Oxygen Species; Sirtuin 1; STAT3 Transcription Factor; Triglycerides

Duchenne muscular dystrophy (DMD) is the most common inherited neuromuscular disease and is characterized by absence of the cytoskeletal protein dystrophin, muscle wasting, and fibrosis. We previously demonstrated that systemic infusion or oral administration of angiotensin-(1-7) (Ang-(1-7)), a peptide with opposing effects to angiotensin II, normalized skeletal muscle architecture, decreased local fibrosis, and improved muscle function in mdx mice, a dystrophic model for DMD. In this study, we investigated the presence, activity, and localization of ACE2, the enzyme responsible for Ang-(1-7) production, in wild type (wt) and mdx skeletal muscle and in a model of induced chronic damage in wt mice. All dystrophic muscles studied showed higher ACE2 activity than wt muscle. Immunolocalization studies indicated that ACE2 was localized mainly at the sarcolemma and, to a lesser extent, associated with interstitial cells. Similar results were observed in the model of chronic damage in the tibialis anterior (TA) muscle. Furthermore, we evaluated the effect of ACE2 overexpression in mdx TA muscle using an adenovirus containing human ACE2 sequence and showed that expression of ACE2 reduced the fibrosis associated with TA dystrophic muscles. Moreover, we observed fewer inflammatory cells infiltrating the mdx muscle. Finally, mdx gastrocnemius muscles from mice infused with Ang-(1-7), which decreases fibrosis, contain less ACE2 associated with the muscle. This is the first evidence supporting ACE2 as an important therapeutic target to improve the dystrophic skeletal muscle phenotype.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Animals; Cytoskeletal Proteins; Fibrosis; Humans; Inflammation; Mice; Mice, Inbred C57BL; Mice, Inbred mdx; Muscle Fibers, Skeletal; Muscular Dystrophy, Animal; Muscular Dystrophy, Duchenne; Peptide Fragments; Peptidyl-Dipeptidase A

: Interstitial fibrosis is a common pathological change in various heart diseases, especially cardiac hypertrophy. Arginine vasopressin (AVP), one of the hallmarks of heart failure, exhibits a profibrotic effect by promoting the proliferation and differentiation of cardiac fibroblasts (CFs). In contrast, angiotensin-(1-7) [Ang-(1-7)] was reported to be beneficial for cardiac remodeling by its antifibrotic effect. To evaluate the effect of Ang-(1-7) on AVP-stimulated CFs and the subsequent signaling molecules involved, CFs isolated from neonatal rat hearts were incubated with AVP and treated with or without Ang-(1-7). Cell proliferation, cell cycle, collagen production, and related cellular signaling molecules were then assessed. The results showed that Ang-(1-7) dose-dependently inhibited cell proliferation and collagen production in AVP-stimulated CFs. In addition, Ang-(1-7) also significantly suppressed calcineurin activity in a dose-dependent manner in AVP-stimulated CFs, which was associated with reduced collagen production. Accordingly, the nuclear translocation and transcriptional activity of nuclear factor-kappa B (NF-κB), downstream signal of calcineurin, were also notably restrained by Ang-(1-7) in AVP-stimulated CFs. Furthermore, the inhibitory effect of Ang-(1-7) on AVP-activated calcineurin-NF-κB signaling was completely reversed by the Mas receptor antagonist A-799. These findings suggest that Ang-(1-7) exerts an antifibrotic effect by inhibiting AVP-stimulated CF proliferation and collagen synthesis by inactivating Mas receptor-calcineurin-NF-κB signaling pathway.

Topics: Angiotensin I; Animals; Animals, Newborn; Arginine Vasopressin; Calcineurin; Cell Differentiation; Cell Proliferation; Collagen; Dose-Response Relationship, Drug; Fibroblasts; Fibrosis; NF-kappa B; Peptide Fragments; Proto-Oncogene Mas; Proto-Oncogene Proteins; Rats; Rats, Sprague-Dawley; Receptors, G-Protein-Coupled; Signal Transduction

We investigated the relationship among oxidative stress, hypertension, renal injury, and angiotensin-converting enzyme-2 (ACE2) expression in type 1 diabetic Akita mice. Blood glucose, blood pressure, and albuminuria were monitored for up to 5 mo in adult male Akita and Akita catalase (Cat) transgenic (Tg) mice specifically overexpressing Cat, a key antioxidant enzyme in their renal proximal tubular cells (RPTCs). Same-age non-Akita littermates and Cat-Tg mice served as controls. In separate studies, adult male Akita mice (14 wk) were treated with ANG 1-7 (500 μg·kg⁻¹·day⁻¹ sc) ± A-779, an antagonist of the Mas receptor (10 mg·kg⁻¹·day⁻¹ sc), and euthanized at the age of 18 wk. The left kidneys were processed for histology and apoptosis studies. Renal proximal tubules were isolated from the right kidneys to assess protein and gene expression. Urinary angiotensinogen (AGT), angiotensin II (ANG II), and ANG 1-7 were quantified by specific ELISAs. Overexpression of Cat attenuated renal oxidative stress; prevented hypertension; normalized RPTC ACE2 expression and urinary ANG 1-7 levels (both were low in Akita mice); ameliorated glomerular filtration rate, albuminuria, kidney hypertrophy, tubulointerstitial fibrosis, and tubular apoptosis; and suppressed profibrotic and proapoptotic gene expression in RPTCs of Akita Cat-Tg mice compared with Akita mice. Furthermore, daily administration of ANG 1-7 normalized systemic hypertension in Akita mice, which was reversed by A-779. These data demonstrate that Cat overexpression prevents hypertension and progression of nephropathy and highlight the importance of intrarenal oxidative stress and ACE2 expression contributing to hypertension and renal injury in diabetes.

Topics: Albuminuria; Angiotensin I; Angiotensin-Converting Enzyme 2; Angiotensinogen; Animals; Apoptosis; Catalase; Diabetes Mellitus, Type 1; Diabetic Nephropathies; Fibrosis; Hypertension; Kidney; Kidney Tubules; Kidney Tubules, Proximal; Male; Mice; Mice, Transgenic; Oxidative Stress; Peptide Fragments; Peptidyl-Dipeptidase A

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).. 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.. 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.. The effect of Ang-(1-7)-CyD treatment on penile fibrosis, oxidative stress, and endothelial function in hypercholesterolemia-induced ED.. 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.. 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.

Topics: Administration, Oral; Angiotensin I; Animals; Apolipoproteins E; Collagen; Cyclodextrins; Disease Models, Animal; Endothelium, Vascular; Fibrosis; Hypercholesterolemia; Impotence, Vasculogenic; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type III; Oxidative Stress; Penile Erection; Penis; Peptide Fragments; Phosphoproteins; Proto-Oncogene Mas; Proto-Oncogene Proteins; Reactive Oxygen Species; Receptor, Angiotensin, Type 1; Receptors, G-Protein-Coupled; Vasodilation; Vasodilator Agents

Angiotensin-converting enzyme 2 (ACE2), a monocarboxypeptidase which metabolizes angiotensin II (Ang II) to generate Ang-(1-7), has been shown to prevent cardiac hypertrophy and injury but the mechanism remains elusive. Irbesartan has the dual actions of angiotensin receptor blockade and peroxisome proliferator-activated receptor-γ (PPARγ) activation. We hypothesized that irbesartan would exert its protective effects on ACE2 deficiency-mediated myocardial fibrosis and cardiac injury via the PPARγ signaling.. 10-week-old ACE2 knockout (ACE2KO; Ace2(-/y)) mice received daily with irbesartan (50 mg/kg) or saline for 2 weeks. The wild-type mice (Ace2(+/y)) were used to the normal controls. We examined changes in myocardial ultrastructure, fibrosis-related genes and pathological signaling by real-time PCR gene array, Western blotting, Masson trichrome staining and transmission electron microscope analyses, respectively.. Compared with the Ace2(+/y) mice, cardiac expression of PPARα and PPARγ were reduced in Ace2(-/y) mice and the myocardial collagen volume fraction (CVF) and expression of fibrosis-related genes were increased, including transforming growth factor-β1 (TGFβ1), connective tissue growth factor (CTGF), collagen I and collagen III. Moreover, ACE2 deficiency triggered cardiac hypertrophy, increased myocardial fibrosis and adverse ultrastructure injury in ACE2KO hearts with higher levels of atrial natriuretic factor (ANF) and phosphorylated extracellular signal-regulated kinase 1/2 (ERK1/2), without affecting cardiac systolic function. Intriguingly, treatment with irbesartan significantly reversed ACE2 deficiency-mediated pathological hypertrophy and myocardial fibrosis in Ace2(-/y) mice linked with enhancement of plasma Ang-(1-7) level and downregulation of AT1 receptor in heart. Consistent with attenuation of myocardial fibrosis and ultrastructure injury, the myocardial CVF and levels of ANF, TGFβ1, CTGF, collagen I, collagen III and phosphorylated ERK1/2 were lower, and expression of PPARγ was higher in ACE2KO mice in response to irbesartan treatment, without affecting cardiac expression of PPARα, PPARδ, β-myosin heavy chain, TGFβ2 and fibronectin.. We conclude that irbesartan prevents ACE2 deficiency-mediated pathological hypertrophy and myocardial fibrosis in ACE2 mutant mice via activation of the PPARγ signaling and suppression of the TGFβ-CTGF-ERK signaling, resulting in attenuation of myocardial injury. Drugs targeting ACE2 and PPARγ represent potential candidates to prevent and treat myocardial injury and related cardiac disorders.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Biphenyl Compounds; Cardiomegaly; Cardiotonic Agents; Collagen; Connective Tissue Growth Factor; Extracellular Signal-Regulated MAP Kinases; Fibrosis; Gene Expression Regulation; Irbesartan; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocardium; Peptide Fragments; Peptidyl-Dipeptidase A; Phosphorylation; PPAR alpha; PPAR delta; PPAR gamma; Receptor, Angiotensin, Type 1; RNA, Messenger; Signal Transduction; Tetrazoles; Transforming Growth Factor beta

Chronic hypertension induces cardiac remodeling, including left ventricular hypertrophy and fibrosis, through a combination of both hemodynamic and humoral factors. In previous studies, we showed that the heptapeptide ANG-(1-7) prevented mitogen-stimulated growth of cardiac myocytes in vitro, through a reduction in the activity of the MAPKs ERK1 and ERK2. In this study, saline- or ANG II-infused rats were treated with ANG-(1-7) to determine whether the heptapeptide reduces myocyte hypertrophy in vivo and to identify the signaling pathways involved in the process. ANG II infusion into normotensive rats elevated systolic blood pressure >50 mmHg, in association with increased myocyte cross-sectional area, ventricular atrial natriuretic peptide mRNA, and ventricular brain natriuretric peptide mRNA. Although infusion with ANG-(1-7) had no effect on the ANG II-stimulated elevation in blood pressure, the heptapeptide hormone significantly reduced the ANG II-mediated increase in myocyte cross-sectional area, interstitial fibrosis, and natriuretic peptide mRNAs. ANG II increased phospho-ERK1 and phospho-ERK2, whereas cotreatment with ANG-(1-7) reduced the phosphorylation of both MAPKs. Neither ANG II nor ANG-(1-7) altered the ERK1/2 MAPK kinase MEK1/2. However, ANG-(1-7) infusion, with or without ANG II, increased the MAPK phosphatase dual-specificity phosphatase (DUSP)-1; in contrast, treatment with ANG II had no effect on DUSP-1, suggesting that ANG-(1-7) upregulates DUSP-1 to reduce ANG II-stimulated ERK activation. These results indicate that ANG-(1-7) attenuates cardiac remodeling associated with a chronic elevation in blood pressure and upregulation of a MAPK phosphatase and may be cardioprotective in patients with hypertension.

Topics: Angiotensin I; Angiotensin II; Animals; Antihypertensive Agents; Cardiomegaly; Drug Interactions; Dual Specificity Phosphatase 1; Fibrosis; Hypertension; Male; MAP Kinase Signaling System; Myocardium; Peptide Fragments; Rats; Rats, Sprague-Dawley; Up-Regulation; Vasoconstrictor Agents; Ventricular Remodeling

The renin-angiotensin system regulates cardiovascular physiology via angiotensin II engaging the angiotensin type 1 or type 2 receptors. Classic actions are type 1 receptor mediated, whereas the type 2 receptor may counteract type 1 receptor activity. Angiotensin-converting enzyme 2 metabolizes angiotensin II to angiotensin-(1-7) and angiotensin I to angiotensin-(1-9). Angiotensin-(1-7) antagonizes angiotensin II actions via the receptor Mas. Angiotensin-(1-9) was shown recently to block cardiomyocyte hypertrophy via the angiotensin type 2 receptor. Here, we investigated in vivo effects of angiotensin-(1-9) via the angiotensin type 2 receptor. Angiotensin-(1-9) (100 ng/kg per minute) with or without the angiotensin type 2 receptor antagonist PD123 319 (100 ng/kg per minute) or PD123 319 alone was infused via osmotic minipump for 4 weeks into stroke-prone spontaneously hypertensive rats. We measured blood pressure by radiotelemetry and cardiac structure and function by echocardiography. Angiotensin-(1-9) did not affect blood pressure or left ventricular mass index but reduced cardiac fibrosis by 50% (P<0.01) through modulating collagen I expression, reversed by PD123 319 coinfusion. In addition, angiotensin-(1-9) inhibited fibroblast proliferation in vitro in a PD123 319-sensitive manner. Aortic myography revealed that angiotensin-(1-9) significantly increased contraction to phenylephrine compared with controls after N-nitro-l-arginine methyl ester treatment, an effect abolished by PD123 319 coinfusion (area under the curve: angiotensin-(1-9) N-nitro-l-arginine methyl ester=98.9±11.8%; control+N-nitro-l-arginine methyl ester=74.0±10.4%; P<0.01), suggesting that angiotensin-(1-9) improved basal NO bioavailability in an angiotensin type 2 receptor-sensitive manner. In summary, angiotensin-(1-9) reduced cardiac fibrosis and altered aortic contraction via the angiotensin type 2 receptor supporting a direct role for angiotensin-(1-9) in the renin-angiotensin system.

Topics: Angiotensin I; Angiotensin II Type 2 Receptor Blockers; Animals; Blood Pressure; Cell Proliferation; Cells, Cultured; Disease Models, Animal; Echocardiography; Endothelium, Vascular; Fibroblasts; Fibrosis; Heart; Hypertension; Imidazoles; In Vitro Techniques; Male; Myocardium; Peptide Fragments; Pyridines; Rats; Receptor, Angiotensin, Type 2; Renin-Angiotensin System; Stroke

Previous studies showed that angiotensin-(1-7) [Ang-(1-7)] attenuates cardiac remodeling by reducing both interstitial and perivascular fibrosis. Although a high affinity binding site for Ang-(1-7) was identified on cardiac fibroblasts, the molecular mechanisms activated by the heptapeptide hormone were not identified. We isolated cardiac fibroblasts from neonatal rat hearts to investigate signaling pathways activated by Ang-(1-7) that participate in fibroblast proliferation. Ang-(1-7) reduced (3)H-thymidine, -leucine and -proline incorporation into cardiac fibroblasts stimulated with serum or the mitogen endothelin-1 (ET-1), demonstrating that the heptapeptide hormone decreases DNA, protein and collagen synthesis. The reduction in DNA synthesis by Ang-(1-7) was blocked by the AT((1-7)) receptor antagonist [d-Ala(7)]-Ang-(1-7), showing specificity of the response. Treatment of cardiac fibroblasts with Ang-(1-7) reduced the Ang II- or ET-1-stimulated increase in phospho-ERK1 and -ERK2. In contrast, Ang-(1-7) increased dual-specificity phosphatase DUSP1 immunoreactivity and mRNA, suggesting that the heptapeptide hormone increases DUSP1 to reduce MAP kinase phosphorylation and activity. Incubation of cardiac fibroblasts with ET-1 increased cyclooxygenase 2 (COX-2) and prostaglandin synthase (PGES) mRNAs, while Ang-(1-7) blocked the increase in both enzymes, suggesting that the heptapeptide hormone alters the concentration and the balance between the proliferative and anti-proliferative prostaglandins. Collectively, these results indicate that Ang-(1-7) participates in maintaining cardiac homeostasis by reducing proliferation and collagen production by cardiac fibroblasts in association with up-regulation of DUSP1 to reduce MAP kinase activities and attenuation of the synthesis of mitogenic prostaglandins. Increased Ang-(1-7) or agents that enhance production of the heptapeptide hormone may prevent abnormal fibrosis that occurs during cardiac pathologies.

Topics: Angiotensin I; Angiotensin II; Animals; Animals, Newborn; Cell Proliferation; Cells, Cultured; Collagen; Cyclooxygenase 2; DNA; Dual Specificity Phosphatase 1; Endothelin-1; Fibroblasts; Fibrosis; Gene Expression Regulation; Heart Diseases; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Myocardium; Peptide Fragments; Phosphorylation; Prostaglandin-Endoperoxide Synthases; Rats; Signal Transduction

Angiotensin (ANG)-(1-7) is known to attenuate diabetic nephropathy; however, its role in the modulation of renal inflammation and oxidative stress in type 2 diabetes is poorly understood. Thus in the present study we evaluated the renal effects of a chronic ANG-(1-7) treatment in Zucker diabetic fatty rats (ZDF), an animal model of type 2 diabetes and nephropathy. Sixteen-week-old male ZDF and their respective controls [lean Zucker rats (LZR)] were used for this study. The protocol involved three groups: 1) LZR + saline, 2) ZDF + saline, and 3) ZDF + ANG-(1-7). For 2 wk, animals were implanted with subcutaneous osmotic pumps that delivered either saline or ANG-(1-7) (100 ng·kg(-1)·min(-1)) (n = 4). Renal fibrosis and tissue parameters of oxidative stress were determined. Also, renal levels of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), ED-1, hypoxia-inducible factor-1α (HIF-1α), and neutrophil gelatinase-associated lipocalin (NGAL) were determined by immunohistochemistry and immunoblotting. ANG-(1-7) induced a reduction in triglyceridemia, proteinuria, and systolic blood pressure (SBP) together with a restoration of creatinine clearance in ZDF. Additionally, ANG-(1-7) reduced renal fibrosis, decreased thiobarbituric acid-reactive substances, and restored the activity of both renal superoxide dismutase and catalase in ZDF. This attenuation of renal oxidative stress proceeded with decreased renal immunostaining of IL-6, TNF-α, ED-1, HIF-1α, and NGAL to values similar to those displayed by LZR. Angiotensin-converting enzyme type 2 (ACE2) and ANG II levels remained unchanged after treatment with ANG-(1-7). Chronic ANG-(1-7) treatment exerts a renoprotective effect in ZDF associated with a reduction of SBP, oxidative stress, and inflammatory markers. Thus ANG-(1-7) emerges as a novel target for treatment of diabetic nephropathy.

Topics: Acute-Phase Proteins; Angiotensin I; Animals; Blood Pressure; Diabetic Nephropathies; Fibrosis; Hypertriglyceridemia; Hypoxia-Inducible Factor 1, alpha Subunit; Interleukin-6; Kidney; Lipocalin-2; Lipocalins; Male; Oxidative Stress; Peptide Fragments; Proteinuria; Proto-Oncogene Proteins; Rats; Rats, Zucker; Tumor Necrosis Factor-alpha

Angiotensin converting enzyme-2 (ACE-2) is a monocarboxypeptidase that metabolises angiotensin (ANG)-II into angiotensin 1-7 (ANG 1-7), thereby functioning as a negative regulator of the renin-angiotensin system. We investigated whether treatment with ANG-II type 1 receptor blocker, olmesartan medoxomil is associated with the attenuation of cardiac myosin-induced dilated cardiomyopathy (DCM) through recently established new axis of ACE-2/ANG 1-7 mas receptor. DCM was elicited in Lewis rats by immunisation with cardiac myosin, and 28 days after immunisation, the surviving Lewis rats were divided into two groups and treated with either olmesartan medoxomil (10 mg/kg/day) or vehicle. Myocardial protein and mRNA levels of ACE-2, ANG 1-7 mas receptor were upregulated in the olmesartan-treated group compared with those of vehicle-treated DCM rats. In contrast, Olmesartan treatment effectively suppressed the myocardial protein and mRNA expressions of inflammatory markers in comparison to the vehicle-treated DCM rats. Olmesartan treatment significantly reduced fibrosis, hypertrophy and their marker molecules (OPN, CTGF, ANP and GATA-4, respectively), as well as matrix metalloproteinases compared with those of vehicle-treated DCM rats. Enhanced myocardial protein levels of phospho-p38 MAPK, phospho-JNK and phospho MAPKAPK-2 in the vehicle-treated DCM rats were prevented by olmesartan treatment. In addition, olmesartan treatment significantly lowered the protein expressions (Nitrotyrosine, p47phox and p67phox) and superoxide radical production compared with those of vehicle-treated DCM rats. Our present study might serve as a new therapeutic target of DCM in cardiovascular diseases and cardiac myosin-induced DCM via the modulation of ACE-2/ANG 1-7 mas receptor axis in rats with DCM after myosin-immunisation.

Topics: Angiotensin I; Angiotensin II Type 1 Receptor Blockers; Angiotensin-Converting Enzyme 2; Animals; Biomarkers; Cardiac Myosins; Cardiomyopathy, Dilated; Endopeptidases; Fibrosis; Gene Expression Regulation; Heart; Hypertrophy, Left Ventricular; Imidazoles; Mitogen-Activated Protein Kinases; Olmesartan Medoxomil; Peptide Fragments; Peptidyl-Dipeptidase A; Proto-Oncogene Mas; Proto-Oncogene Proteins; Rats; Receptors, G-Protein-Coupled; RNA, Messenger; Tetrazoles

Our previous studies have indicated that chronic treatment with 1-[(2-dimethylamino) ethylamino]-4-(hydroxymethyl)-7-[(4-methylphenyl) sulfonyl oxy]-9H-xanthene-9-one (XNT), an angiotensin-converting enzyme 2 (ACE2) activator, reverses hypertension-induced cardiac and renal fibrosis in spontaneously hypertensive rats (SHRs). Furthermore, XNT prevented pulmonary vascular remodelling and right ventricular hypertrophy and fibrosis in a rat model of monocrotaline-induced pulmonary hypertension. The aim of this study was to determine the mechanisms underlying the protective effects of XNT against cardiac fibrosis. Hydroxyproline assay was used to measure cardiac collagen content in control and XNT-treated (200 ng kg(-1) min(-1) for 28 days) SHRs. Cardiac ACE2 activity and protein levels were determined using the fluorogenic peptide assay and Western blot analysis, respectively. Extracellular signal-regulated kinases (ERKs; p44 and p42) and angiotensin II type 1 (AT(1)) receptor levels were quantified by Western blotting. Cardiac ACE2 protein levels were ∼15% lower in SHRs compared with Wistar-Kyoto control animals (ACE2/glyceraldehyde 3-phosphate dehydrogenase ratio: Wistar-Kyoto, 1.00 ± 0.02 versus SHR, 0.87 ± 0.01). However, treatment of SHRs with XNT completely restored the decreased cardiac ACE2 levels. Also, chronic infusion of XNT significantly increased cardiac ACE2 activity in SHRs. This increase in ACE2 activity was associated with decreased cardiac collagen content. Furthermore, the antifibrotic effect of XNT correlated with increased cardiac angiotensin-(1-7) immunostaining, though no change in cardiac AT(1) protein levels was observed. The beneficial effects of XNT were also accompanied by a reduction in ERK phosphorylation (phospho-ERK/total ERK ratio: Wistar-Kyoto, 1.00 ± 0.04; control SHR, 1.46 ± 0.25; treated SHR, 0.86 ± 0.02). Our observations demonstrate that XNT activates cardiac ACE2 and inhibits fibrosis. These effects are associated with increases in angiotensin-(1-7) and inhibition of cardiac ERK signalling.

Topics: Angiotensin I; Angiotensin-Converting Enzyme 2; Animals; Cell Culture Techniques; Collagen; Enzyme Activation; Extracellular Signal-Regulated MAP Kinases; Fibroblasts; Fibrosis; Heart; Hypertension; Male; Myocardium; Peptide Fragments; Peptidyl-Dipeptidase A; Phosphorylation; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Receptor, Angiotensin, Type 1; Signal Transduction; Xanthones

This study was designed to investigate the effect of angiotensin (1-7), a Mas receptor agonist, and A-779, a Mas receptor antagonist, in rats with diabetic cardiomyopathy (DC).. Rats treated with a single injection of streptozotocin (50 mg/kg, intraperitoneal), developed DC after 8 weeks. The extent of DC was assessed by measuring the left ventricular weight/body weight (LVW/BW) ratio, absolute LVW, left ventricular developed pressure (LVDP), maximum change in left ventricular pressure over time (dp/dtmax), minimum change in left ventricular pressure over time (dp/dtmin), left ventricular (LV) protein content, LV collagen content, lipid profile, and serum nitrite/nitrate concentration. Test drug treatment was given from week 4 to week 8.. Angiotensin (1-7) treatment attenuated DC by significantly increasing LVDP, dp/dtmax, dp/dtmin, serum nitrite/nitrate concentration and significantly decreasing the LVW/BW ratio and LV collagen content. For the first time, this study has documented that endogenous angiotensin (1-7) regulates lipid profile in rats, and that treatment with angiotensin (1-7) significantly attenuates diabetes-induced changes in lipid profile. However, LV protein content and absolute LVW remain unaffected after treatment.. Angiotensin (1-7) significantly attenuates DC in rats because of vasodilatory, antiproliferative and anifibrotic properties but also because of a significant decrease in dyslipidemia, the major culprit for cardiac dysfunctions in diabetes.

Topics: Angiotensin I; Angiotensin II; Animals; Blood Glucose; Collagen; Diabetes Mellitus, Experimental; Diabetic Cardiomyopathies; Dyslipidemias; Fibrosis; Heart Ventricles; Hypertrophy, Left Ventricular; Lipids; Nitrates; Nitrites; Peptide Fragments; Proto-Oncogene Mas; Proto-Oncogene Proteins; Rats; Rats, Wistar; Receptors, G-Protein-Coupled; Time Factors; Ventricular Function, Left; Ventricular Pressure

Recent identification of a counterregulatory axis of the renin-angiotensin system, called angiotensin-converting enzyme 2-angiotensin-(1-7) [ANG-(1-7)]-Mas receptor, may offer new targets for the treatment of renal fibrosis. We hypothesized that therapy with ANG-(1-7) would improve glomerulosclerosis through counteracting ANG II in experimental glomerulonephritis. Disease was induced in rats with the monoclonal anti-Thy-1 antibody, OX-7. Based on a three-dose pilot study, 576 microg x kg(-1) x day(-1) ANG-(1-7) was continuously infused from day 1 using osmotic pumps. Measures of glomerulosclerosis include semiquantitative scoring of matrix proteins stained for periodic acid Schiff, collagen I, and fibronectin EDA+ (FN). ANG-(1-7) treatment reduced disease-induced increases in proteinuria by 75%, glomerular periodic acid Schiff staining by 48%, collagen I by 24%, and FN by 25%. The dramatic increases in transforming growth factor-beta1, plasminogen activator inhibitor-1, FN, and collagen I mRNAs seen in disease control animals compared with normal rats were all significantly reduced by ANG-(1-7) administration (P < 0.05). These observations support our hypothesis that ANG-(1-7) has therapeutic potential for reversing glomerulosclerosis. Several results suggest ANG-(1-7) acts by counteracting ANG II effects: 1) renin expression in ANG-(1-7)-treated rats was dramatically increased as it is with ANG II blockade therapy; and 2) in vitro data indicate that ANG II-induced increases in mesangial cell proliferation and plasminogen activator inhibitor-1 overexpression are inhibited by ANG-(1-7) via its binding to a specific receptor known as Mas.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Animals; Cell Proliferation; Cells, Cultured; Collagen Type I; Disease Models, Animal; Dose-Response Relationship, Drug; Fibronectins; Fibrosis; Gene Expression Regulation; Glomerulonephritis; Infusion Pumps, Implantable; Infusions, Subcutaneous; Isoantibodies; Kidney; Male; Mesangial Cells; Peptide Fragments; Peptidyl-Dipeptidase A; Plasminogen Activator Inhibitor 1; Proteinuria; Rats; Rats, Sprague-Dawley; Renin; RNA, Messenger; Time Factors; Transforming Growth Factor beta1

It has been shown that Ang-(1-7) has cardioprotective actions. To directly investigate the effects of Ang-(1-7) specifically in the heart, we generated and characterized transgenic (TG) rats which express an Ang-(1-7)-producing fusion protein driven by the alpha-MHC promoter.. After microinjection of the transgene into fertilized rat zygotes, we obtained four different transgenic lines. Homozygous animals were analyzed with regard to the expression profile of the transgene by ribonuclease protection assay. Transgene expression was detected mainly in the heart with weak or no expression in other organs. Heterozygous TG(hA-1-7)L7301 rats presented a significant increase in cardiac Ang-(1-7) concentration compared with control rats (17.1+/-2.1 versus 3.9+/-1.4 pg/mg protein in SD rats). Radiotelemetry analysis revealed that TG rats presented no significant changes in blood pressure and heart rate compared with normal rats. Overexpression of Ang-(1-7) in the heart produced slight improvement in resting cardiac function (+ dT/dt: 81530+/-1305.0 versus 77470+/-345.5 g/s bpm in SD rats, p < 0.05), which was in keeping with the enhanced [Ca(2+)] handling observed in cardiomyocytes of TG rats. TG(hA-1-7)L7301 rats also showed a greater capacity to withstand stress since TG rats showed a less pronounced deposition of collagen type III and fibronectin induced by isoproterenol treatment in the subendocardial area than in corresponding controls. In addition, hearts from TG rats showed reduced incidence and duration of reperfusion arrhythmias in comparison with SD rats.. These results indicate that Ang-(1-7) has blood pressure-independent, antifibrotic effects, acting directly in the heart.

Topics: Angiotensin I; Animals; Arrhythmias, Cardiac; Blood Pressure; Calcium; Disease Models, Animal; Fibrosis; Gene Expression Regulation; Heart Rate; Heart Ventricles; Isoproterenol; Male; Myocardial Reperfusion Injury; Myocytes, Cardiac; Myosin Heavy Chains; Peptide Fragments; Promoter Regions, Genetic; Rats; Rats, Sprague-Dawley; Rats, Transgenic; Telemetry

Renin-angiotensin system (RAS) is activated in the fibrillating atria. Angiotensin-(1-7) [Ang-(1-7)] counterbalances the actions of angiotensin II (Ang II). To investigate the effects of Ang-(1-7) on the long-term atrial tachycardia-induced atrial fibrosis and atrial fibrillation (AF) vulnerability, eighteen dogs were assigned to sham group, paced group, or paced+Ang-(1-7) group, 6 dogs in each group. Rapid atrial pacing at 500 bpm was maintained for 14 days, but dogs in the sham group were instrumented without pacing. During the pacing, Ang-(1-7) (6 microg x kg(-1) x h(-1)) was given intravenously. After pacing, atrial mRNA expression of ERK1/ERK2 and atrial fibrosis were assessed, the inducibility and duration of AF were measured. Compared with sham, ERK1/ERK2 mRNA expression was increased in the paced group (P<0.05). Atrial tissue from the paced dogs showed a large amount of interstitial fibrosis, and the inducible rate of AF was increased at various BCLs in paced dogs (P<0.01). Compared with the paced group, Ang-(1-7) prevented the increase of ERK1/ERK2 mRNA expression (P<0.01 and P<0.05, respectively), and attenuated the interstitial fibrosis (P<0.01). Inducibility and duration of AF were reduced by Ang-(1-7) at various BCLs. In conclusion, Ang-(1-7) reduced AF vulnerability in chronic paced atria, and antifibrotic actions contributed to its preventive effects on AF.

Topics: Angiotensin I; Animals; Atrial Fibrillation; Base Sequence; DNA Primers; Dogs; Fibrosis; Heart Atria; Peptide Fragments; Reverse Transcriptase Polymerase Chain Reaction; Tachycardia

Emerging evidence suggests that cardiac angiotensin-converting enzyme 2 (ACE2) may contribute to the regulation of heart function and hypertension-induced cardiac remodeling. We tested the hypothesis that inhibition of ACE2 in the hearts of (mRen2)27 hypertensive rats may accelerate progression of cardiac hypertrophy and fibrosis by preventing conversion of angiotensin II (Ang II) into the antifibrotic peptide, angiotensin-(1-7) (Ang-(1-7)).. Fourteen male (mRen2)27 transgenic hypertensive rats (12 weeks old, 401 + or - 7 g) were administered either vehicle (0.9% saline) or the ACE2 inhibitor, MLN-4760 (30 mg/kg/day), subcutaneously via mini-osmotic pumps for 28 days.. Although ACE2 inhibition had no effect on average 24-h blood pressures, left ventricular (LV) Ang II content increased 24% in rats chronically treated with the ACE2 inhibitor (P < 0.05). Chronic ACE2 inhibition had no effect on plasma Ang II or Ang-(1-7) levels. Increased cardiac Ang II levels were associated with significant increases in both LV anterior, posterior, and relative wall thicknesses, as well as interstitial collagen fraction area and cardiomyocyte hypertrophy in the transgenic animals chronically treated with the ACE2 inhibitor. Cardiac remodeling was not accompanied by any further alterations in LV function.. These studies demonstrate that chronic inhibition of ACE2 causes an accumulation of cardiac Ang II, which exacerbates cardiac hypertrophy and fibrosis without having any further impact on blood pressure or cardiac function.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Angiotensin-Converting Enzyme Inhibitors; Animals; Blood Pressure; Cardiomegaly; Fibrosis; Heart Ventricles; Imidazoles; Leucine; Male; Peptide Fragments; Peptidyl-Dipeptidase A; Rats; Rats, Transgenic

Analogs of vitamin D attenuate renal injury in several models of kidney disease, but the mechanism underlying this renoprotective effect is unknown. To address the role of the vitamin D receptor (VDR) in renal fibrogenesis, we subjected VDR-null mice to unilateral ureteral obstruction for 7 days. Compared with wild-type mice, VDR-null mice developed more severe renal damage in the obstructed kidney, with marked tubular atrophy and interstitial fibrosis. Significant induction of extracellular matrix proteins (fibronectin and collagen I), profibrogenic and proinflammatory factors (TGF-beta, connective tissue growth factor, and monocyte chemoattractant protein 1), and epithelial-to-mesenchymal transition accompanied this histologic damage. Because VDR ablation activates the renin-angiotensin system and leads to accumulation of angiotensin II (AngII) in the kidney, we assessed whether elevated AngII in the VDR-null kidney promotes injury. Treatment with the angiotensin type 1 antagonist losartan eliminated the difference in obstruction-induced interstitial fibrosis between wild-type and VDR-null mice, suggesting that AngII contributes to the enhanced renal fibrosis observed in obstructed VDR-null kidneys. Taken together, these results suggest that the VDR attenuates obstructive renal injury at least in part by suppressing the renin-angiotensin system.

Topics: Angiotensin I; Animals; Cells, Cultured; Chemokine CCL2; Collagen Type I; Connective Tissue Growth Factor; Disease Models, Animal; Fibronectins; Fibrosis; Kidney; Kidney Diseases; Losartan; Mice; Mice, Inbred C57BL; Mice, Knockout; Receptors, Calcitriol; Renin-Angiotensin System; Transforming Growth Factor beta

Angiotensin-(1-7) displays antihypertensive and antiproliferative properties although its effect on cardiac remodeling and hypertrophy in hypertension has not been fully elucidated. The present study was designed to examine the effect of chronic angiotensin-(1-7) treatment on myocardial remodeling, cardiac hypertrophy and underlying mechanisms in spontaneous hypertension. Adult male spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats were treated with or without angiotensin-(1-7) or the angiotensin-(1-7) antagonist A-779 for 24 weeks. Mean arterial pressure, left ventricular geometry, expression of the hypertrophic markers ANP and β-MHC, collagen contents (type I and III), collagenase (MMP-1), matrix metalloproteinase-2 (MMP-2) and tissue inhibitor of MMPs-1 (TIMP-1) were evaluated in WKY and SHR rats with or without treatment. Our data revealed that chronic angiotensin-(1-7) treatment significantly suppressed hypertension, left ventricular hypertrophy, expression of ANP and β-MHC as well as myocardial fibrosis in SHR rats, the effects of which were nullified by the angiotensin-(1-7) receptor antagonist A-779. In addition, angiotensin-(1-7) treatment significantly counteracted hypertension-induced changes in the mRNA expression of MMP-2 and TIMP-1 and collagenase activity, the effects of which were blunted by A-779. In vitro study revealed that angiotensin-(1-7) directly increased the activity of MMP-2 and MMP-9 while decreasing the content of TIMP-1 and TIMP-2. Taken together, our results revealed a protective effect of angiotensin-(1-7) against cardiac hypertrophy and collagen deposition, which may be related to concerted changes in MMPs and TIMPs levels. These data indicated the therapeutic potential of angiotensin-(1-7) in spontaneous hypertension-induced cardiac remodeling.

Topics: Angiotensin I; Animals; Antihypertensive Agents; Atrial Natriuretic Factor; Blood Pressure; Cardiomegaly; Collagen; Collagenases; Fibrosis; Hypertension; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Myocardium; Myosin Heavy Chains; Peptide Fragments; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Tissue Inhibitor of Metalloproteinases

Angiotensin-(1-7) [Ang-(1-7)] is an endogenous 7-amino acid peptide hormone of the renin-angiotensin system that has antiproliferative properties. In this study, Ang-(1-7) inhibited the growth of cancer-associated fibroblasts (CAF) and reduced fibrosis in the tumor microenvironment. A marked decrease in tumor volume and weight was observed in orthotopic human breast tumors positive for the estrogen receptor (BT-474 or ZR-75-1) and HER2 (BT-474) following Ang-(1-7) administration to athymic mice. Ang-(1-7) concomitantly reduced interstitial fibrosis in association with a significant decrease in collagen I deposition, along with a similar reduction in perivascular fibrosis. In CAFs isolated from orthotopic breast tumors, the heptapeptide markedly attenuated in vitro growth as well as reduced fibronectin, transforming growth factor-β (TGF-β), and extracellular signal-regulated kinase 1/2 kinase activity. An associated increase in the mitogen-activated protein kinase (MAPK) phosphatase DUSP1 following treatment with Ang-(1-7) suggested a potential mechanism by which the heptapeptide reduced MAPK signaling. Consistent with these in vitro observations, immunohistochemical analysis of Ang-(1-7)-treated orthotopic breast tumors revealed reduced TGF-β and increased DUSP1. Together, our findings indicate that Ang-(1-7) targets the tumor microenvironment to inhibit CAF growth and tumor fibrosis.

Topics: Angiotensin I; Animals; Antihypertensive Agents; Blotting, Western; Breast Neoplasms; Carcinoma, Ductal, Breast; Dual Specificity Phosphatase 1; Female; Fibronectins; Fibrosis; Fluorescent Antibody Technique; Humans; Immunoenzyme Techniques; Lung Diseases, Interstitial; Mice; Mice, Nude; Mitogen-Activated Protein Kinase 3; Peptide Fragments; Phosphorylation; Transforming Growth Factor beta; Tumor Cells, Cultured

To explore the effects of Angiotensin (ANG)-(1-7) on postangioplasty fibrotic remodeling and the involvement of TGF-beta/Smad signaling pathway in this process.. Thirty two healthy New Zealand white rabbits were randomly divided into 4 groups: sham group, control group, ANG-(1-7) group and ANG-(1-7) + A-779 group. Rabbits underwent angioplasty in the abdominal aorta or sham surgery. Subsequently, an osmotic minipump was implanted for saline, ANG-(1-7) (576 microg x kg(-1) x d(-1)) or ANG-(1-7) + A-779 (576 microg x kg(-1) x d(-1)) delivery. Before and after 4 weeks treatment, the levels of ANG II in plasma were measured by ELISA. At week 4, angiography and histomorphometric analysis were performed, mRNA levels of collagen I and III were assayed by RT-PCR and protein levels of TGF-beta1 and Smad2 in local vessel were assayed by Western blot.. Following 4 weeks treatment, ANG-(1-7) and ANG-(1-7) + A-779 group displayed a significant elevation in lumen diameter [(4.11 +/- 0.10) mm and (3.34 +/- 0.11) mm vs. (2.88 +/- 0.08) mm, P < 0.05, respectively] and reduction in neointimal thickness [(208 +/- 17) microm and (407 +/- 25) microm vs. (448 +/- 15) microm, P < 0.05, respectively], neointimal area [(0.27 +/- 0.09) mm2 and (0.38 +/- 0.01) mm2 vs. (0.41 +/- 0.02) mm2, P < 0.05, respectively] and restenosis rate [(28.1 +/- 2.7)% and (36.8 +/- 2.2)% vs. (40.1 +/- 2.7)%, P < 0.05, respectively] compared with control group. Collagen I, III mRNA and TGF-beta1, Smad2 protein levels were significantly elevated in control group, ANG-(1-7) group and ANG-(1-7) +A-779 group compared to sham group (P < 0.01 or P < 0.05) and reduced in ANG-(1-7) group compared to control group (all P < 0.05). Co-treatment with A-779 reversed the inhibitory action of ANG-(1-7). Plasma levels of ANG II postangioplasty were similar in control and ANG-(1-7) group and both were significantly higher than preoperation levels.. ANG-(1-7) attenuates postangioplasty collagen synthesis in rabbits possibly through down-regulating the expression of TGF-beta1 and inhibiting the activation of Smad2 pathway.

Topics: Angiotensin I; Animals; Aorta, Abdominal; Collagen Type I; Collagen Type III; Fibrosis; Muscle, Smooth, Vascular; Peptide Fragments; Rabbits; Signal Transduction; Smad2 Protein; Transforming Growth Factor beta1

Angiotensin-converting enzyme 2 (ACE2), a homologue of angiotensin-converting enzyme (ACE), converts angiotensin (Ang) I to Ang(1-9) and Ang II to Ang(1-7), but does not directly process Ang I to Ang II. Cardiac function is compromised in ACE2 null mice; however, the importance of ACE2 in the processing of angiotensin peptides within the murine heart is not known. We determined the metabolism of angiotensins in wild-type (WT), ACE (ACE(-/-)) and ACE2 null mice (ACE2(-/-)). Angiotensin II was converted almost exclusively to Ang(1-7) in the cardiac membranes of WT and ACE(-/-) strains, although generation of Ang(1-7) was greater in the ACE(-/-) mice (27.4 +/- 4.1 versus 17.5 +/- 3.2 nmol(-1) mg h(-1) for WT). The ACE2 inhibitor MLN4760 significantly attenuated Ang II metabolism and the subsequent formation of Ang(1-7) in both strains. In the ACE2(-/-) hearts, Ang II metabolism and the generation of Ang(1-7) were significantly attenuated; however, the ACE2 inhibitor reduced the residual Ang(1-7)-forming activity in this strain. Angiotensin I was primarily converted to Ang(1-9) (WT, 28.9 +/- 3.1 nmol(-1) mg h(-1); ACE(-/-), 49.8 +/- 5.3 nmol(-1) mg h(-1); and ACE2(-/-), 35.9 +/- 5.4 nmol(-1) mg h(-1)) and to smaller quantities of Ang(1-7) and Ang II. Although the ACE2 inhibitor had no effect on Ang(1-9) formation, the carboxypeptidase A inhibitor benzylsuccinate essentially abolished the formation of Ang(1-9) and increased the levels of Ang I in cardiac membranes. In conclusion, our studies in the murine heart suggest that ACE2 is the primary pathway for the metabolism of Ang II and the subsequent formation of Ang(1-7), a peptide that, in contrast to Ang II, exhibits both antifibrotic and antiproliferative actions.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme 2; Angiotensin-Converting Enzyme Inhibitors; Angiotensins; Animals; Carboxypeptidases A; Cell Proliferation; Fibrosis; Heart; Imidazoles; Immunohistochemistry; Leucine; Male; Membranes; Mice; Mice, Knockout; Myocardium; Peptide Fragments; Peptidyl-Dipeptidase A; Protease Inhibitors; Succinates

The brain renin-angiotensin system contributes significantly to progressive left ventricular (LV) dysfunction in rats after myocardial infarction (MI). The present study evaluated the effects of central versus peripheral plus central angiotensin-converting enzyme (ACE) blockade on sympathetic activity, and LV anatomy and function after MI.. Wistar rats were treated for 4 weeks after MI with the lipophilic ACE inhibitor trandolapril at 5 mg/kg/day or the hydrophilic blocker lisinopril at 50 mg/kg/day by once daily subcutaneous injection, or with a central infusion of lisinopril at 0.1 mg/kg/day.. At 24 hours after the last dose, subcutaneous trandolapril caused 70% to 80% ACE inhibition in both brain and kidneys; lisinopril caused 10% to 20% less. Central infusion of lisinopril caused 70% inhibition of brain ACE and minimal (6%) inhibition in the kidneys. All three treatments similarly improved sympathetic reactivity and arterial baroreflex function. All three treatments lowered cardiac Ang I and II, and similarly attenuated the increases in LV end diastolic pressure, circumference, and fibrosis. Both subcutaneous treatments further decreased LV peak systolic pressure and dP/dt max, whereas icv lisinopril caused no change.. Despite marked differences in the extent of peripheral blockade, all three treatments similarly affected sympathetic activity and decreased cardiac Ang II, preload and remodeling after MI. One may speculate that central and peripheral ACE-mediated mechanisms are sequential and therefore only minor additional effects of peripheral ACE blockade are noted.

Topics: Angiotensin I; Angiotensin II; Angiotensin-Converting Enzyme Inhibitors; Animals; Baroreflex; Brain; Fibrosis; Hemodynamics; Indoles; Infusions, Intravenous; Injections, Subcutaneous; Kidney; Lisinopril; Male; Myocardial Infarction; Myocardium; Rats; Rats, Wistar; Renin-Angiotensin System; Sympathetic Nervous System; Ventricular Dysfunction, Left

Cardiac remodeling, which typically results from chronic hypertension or following an acute myocardial infarction, is a major risk factor for the development of heart failure and, ultimately, death. The renin-angiotensin system (RAS) has previously been established to play an important role in the progression of cardiac remodeling, and inhibition of a hyperactive RAS provides protection from cardiac remodeling and subsequent heart failure. Our previous studies have demonstrated that overexpression of angiotensin-converting enzyme 2 (ACE2) prevents cardiac remodeling and hypertrophy during chronic infusion of angiotensin II (ANG II). This, coupled with the knowledge that ACE2 is a key enzyme in the formation of ANG-(1-7), led us to hypothesize that chronic infusion of ANG-(1-7) would prevent cardiac remodeling induced by chronic infusion of ANG II. Infusion of ANG II into adult Sprague-Dawley rats resulted in significantly increased blood pressure, myocyte hypertrophy, and midmyocardial interstitial fibrosis. Coinfusion of ANG-(1-7) resulted in significant attenuations of myocyte hypertrophy and interstitial fibrosis, without significant effects on blood pressure. In a subgroup of animals also administered [d-Ala(7)]-ANG-(1-7) (A779), an antagonist to the reported receptor for ANG-(1-7), there was a tendency to attenuate the antiremodeling effects of ANG-(1-7). Chronic infusion of ANG II, with or without coinfusion of ANG-(1-7), had no effect on ANG II type 1 or type 2 receptor binding in cardiac tissue. Together, these findings indicate an antiremodeling role for ANG-(1-7) in cardiac tissue, which is not mediated through modulation of blood pressure or altered cardiac angiotensin receptor populations and may be at least partially mediated through an ANG-(1-7) receptor.

Topics: Analysis of Variance; Angiotensin I; Angiotensin II; Animals; Blood Pressure; Cardiomegaly; Disease Models, Animal; Fibrosis; Heart; Hypertension; Male; Myocardium; Peptide Fragments; Proto-Oncogene Mas; Proto-Oncogene Proteins; Rats; Rats, Sprague-Dawley; Receptor, Angiotensin, Type 1; Receptor, Angiotensin, Type 2; Receptors, G-Protein-Coupled; Time Factors; Transforming Growth Factor beta; Ventricular Remodeling

Studies in experimental animals and younger women suggest a protective role for estrogen; however, clinical trials may not substantiate this effect in older females. Therefore, the present study assessed the outcome of ovariectomy in older mRen2. Lewis rats subjected to a high-salt diet for 4 wk. Intact or ovariectomized (OVX, 15 wk of age) mRen2. Lewis rats were aged to 60 wk and then placed on a high-salt (HS, 8% sodium chloride) diet for 4 wk. Systolic blood pressures were similar between groups [OVX 169 +/- 6 vs. Intact 182 +/- 7 mmHg; P = 0.22] after the 4-wk diet; however, proteinuria [OVX 0.8 +/- 0.2 vs. Intact 11.5 +/- 2.6 mg/mg creatinine; P < 0.002, n = 6], renal interstitial fibrosis, glomerular sclerosis, and tubular casts were lower in OVX vs. Intact rats. Kidney injury molecule-1 mRNA, a marker of tubular damage, was 53% lower in the OVX HS group. Independent from blood pressure, OVX HS rats exhibited significantly lower cardiac (24%) and renal (32%) hypertrophy as well as lower C-reactive protein (28%). Circulating insulin-like growth factor-I (IGF-I) levels were not different between the Intact and OVX groups; however, renal cortical IGF-I mRNA and protein were attenuated in OVX rats [P < 0.05, n = 6]. We conclude that ovariectomy in the older female mRen2. Lewis rat conveys protection against salt-dependent increase in renal injury.

Topics: Aging; Angiotensin I; Angiotensin II; Animals; Animals, Congenic; Blood Pressure; C-Reactive Protein; Cell Adhesion Molecules; Disease Models, Animal; Female; Fibrosis; Hypertension; Hypertrophy; Insulin-Like Growth Factor I; Intracellular Signaling Peptides and Proteins; Kidney; Kidney Diseases; Membrane Proteins; Ovariectomy; Peptide Fragments; Proteinuria; Rats; Rats, Inbred Lew; Renin; Renin-Angiotensin System; RNA, Messenger; Sodium Chloride, Dietary

Pancreatic stellate cells (PSCs) are involved in pancreatic inflammation and fibrosis. Recent studies have shown that blocking the renin-angiotensin system (RAS) attenuates pancreatic inflammation and fibrosis. However, there are few data about the direct effects of high glucose on extracellular matrix (ECM) protein synthesis and angiotensin II (Ang II) induction in PSCs. PSCs were isolated from male Sprague-Dawley rats and cultured in medium containing 5.5 mM (LG group) or 27 mM D-glucose (HG group). Levels of Ang II and transforming growth factor-beta (TGF-beta) in culture media were measured and Ang II-positive cells were counted. We used real-time polymerase chain reaction (PCR) to detect Ang II receptor expression and Western blot analysis for the expression of ECM proteins such as connective-tissue growth factor (CTGF) and collagen type IV. Cells were also treated with an Ang II-receptor antagonist (candesartan, 10 microM) or angiotensin-converting enzyme (ACE) inhibitor (ramiprilat, 100 nM). Thymidine uptake by PSCs increased fourfold with high glucose treatment. Ang II levels and the proportion of Ang II-positive PSCs were significantly increased after 6 h under high-glucose conditions. TGF-beta concentrations also increased significantly with high glucose. After 72 h, the expression of CTGF and collagen type IV proteins in high-glucose cultures increased significantly and this increase was effectively attenuated by the candesartan or the ramiprilat. All together, high glucose induced PSCs proliferation and ECM protein synthesis, and these effects were attenuated by an Ang II-receptor antagonist. The data suggest that pancreatic inflammation and fibrosis aggravated by hyperglycemia, and Ang II play an important role in this pathogenesis.

Topics: Angiotensin I; Angiotensin II; Animals; Benzimidazoles; Biphenyl Compounds; Collagen; Connective Tissue Growth Factor; Dose-Response Relationship, Drug; Extracellular Matrix; Extracellular Matrix Proteins; Fibronectins; Fibrosis; Glucose; Immediate-Early Proteins; Intercellular Signaling Peptides and Proteins; Male; Pancreas; Pancreatitis; Ramipril; Rats; Rats, Sprague-Dawley; Receptor, Angiotensin, Type 1; Renin-Angiotensin System; Tetrazoles; Transforming Growth Factor beta

Cardiac remodeling is a hallmark hypertension-induced pathophysiology. In the current study, the role of the angiotensin-(1-7) fragment in modulating cardiac remodeling was examined. Sprague-Dawley rats underwent uninephrectomy surgery and were implanted with a deoxycorticosterone acetate (DOCA) pellet. DOCA animals had their drinking water replaced with 0.9% saline solution. A subgroup of DOCA-salt animals was implanted with osmotic minipumps, which delivered angiotensin-(1-7) chronically (100 ng.kg(-1).min(-1)). Control animals underwent sham surgery and were maintained on normal drinking water. Blood pressure was measured weekly with the use of the tail-cuff method, and after 4 wk of treatment, blood pressure responses to graded doses of angiotensin II were determined by direct carotid artery cannulation. Ventricle size was measured, and cross sections of the heart ventricles were paraffin embedded and stained using Masson's Trichrome to measure interstitial and perivascular collagen deposition and myocyte diameter. DOCA-salt treatment caused significant increases in blood pressure, cardiac hypertrophy, and myocardial and perivascular fibrosis. Angiotensin-(1-7) infusion prevented the collagen deposition effects without any effect on blood pressure or cardiac hypertrophy. These results indicate that angiotensin-(1-7) selectively prevents cardiac fibrosis independent of blood pressure or cardiac hypertrophy in the DOCA-salt model of hypertension.

Topics: Angiotensin I; Animals; Blood Pressure; Cardiomegaly; Collagen; Coronary Vessels; Desoxycorticosterone; Dose-Response Relationship, Drug; Fibrosis; Heart Diseases; Hypertension; Male; Peptide Fragments; Rats; Rats, Sprague-Dawley; Ventricular Remodeling

To test the hypothesis that chronic administration of angiotensin-(1-7) [Ang-(1-7)] attenuates cardiac hypertrophy in rats in vivo.. Coarctation of the suprarenal abdominal aorta was performed in 41 8-week-old male Sprague Dawley rats. Twenty-four hours after the operation, osmotic minipumps were surgically implanted subcutaneously in the rats, which were randomly divided into 3 groups, including a sham-operation group (n=15) receiving infusion with normal saline, a suprarenal aortic coarctation group (n=12), and a suprarenal aortic coarctation group (n=14) with Ang-(1-7) treatment at the dose of 25 mug x kg(-1) x h(-1). Four weeks later, the systolic and diastolic blood pressures were measured and the left ventricular mass index (LVMI, mg/g) was calculated from the ratio of left ventricular weight to body weight. The concentrations of Ang II in the plasma and myocardium were measured by radioimmunoassay, and myocardial interstitial collagen volume fraction (ICVF) was determined by quantitative morphometry of the sections with Picrosirius red staining using an automated image analyzer.. Suprarenal abdominal aortic coarctation induced a significant increase in carotid artery systolic and diastolic blood pressure, heart weight, LVMI, ICVF, and the concentration of Ang II in the myocardium (P<0.01). Chronic administration of Ang-(1-7) attenuated the increase in the heart weight, LVMI, ICVF and left ventricular diastolic end pressure (LVEDP) caused by suprarenal abdominal aortic coarctation (P<0.05). Ang-(1-7) also increased the formerly decreased maximum left ventricular pressure reduction rate (-dP/dt(max)) (P<0.05), but had no effect on blood pressure and the concentration of Ang II in the myocardium. No difference was noted in plasma concentration of Ang II between the 3 groups.. Ang-(1-7) attenuates cardiac hypertrophy and fibrosis and preserved the impaired left ventricular function induced by left ventricular pressure-overload in rats. These effects are not associated with the changes in the concentrations of Ang II in the left ventricular myocardium and plasma.

Topics: Angiotensin I; Angiotensin II; Animals; Antihypertensive Agents; Aortic Coarctation; Blood Pressure; Fibrosis; Hypertrophy, Left Ventricular; Male; Myocardium; Peptide Fragments; Random Allocation; Rats; Rats, Sprague-Dawley; Ventricular Function, Left

ANG-(1-7) improves the function of the remodeling heart. Although this peptide is generated directly within the myocardium, the effects of ANG-(1-7) on cardiac fibroblasts that play a critical role in cardiac remodeling are largely unknown. We tested the hypothesis that specific binding of ANG-(1-7) to cardiac fibroblasts regulates cellular functions that are involved in cardiac remodeling. 125I-labeled ANG-(1-7) binding assays identified specific binding sites of ANG-(1-7) on adult rat cardiac fibroblasts (ARCFs) with an affinity of 11.3 nM and a density of 131 fmol/mg protein. At nanomolar concentrations, ANG-(1-7) interacted with specific sites that were distinct from ANG II type 1 and type 2 receptors without increasing cytosolic Ca2+ concentration. At these concentrations, ANG-(1-7) had inhibitory effects on collagen synthesis as assessed by [3H]proline incorporation and decreased mRNA expression of growth factors in ARCFs. These effects of ANG-(1-7) contrasted with effects of ANG II. Pretreatment of ARCFs with ANG-(1-7) inhibited ANG II-induced increases in collagen synthesis and in mRNA expression of growth factors, including endothelin-1 and leukemia inhibitory factor. ANG-(1-7) pretreatment also inhibited the stimulatory effects of conditioned medium from ANG II-treated ARCFs on [3H]leucine incorporation and atrial natriuretic factor mRNA expression, markers of hypertrophy, in cardiomyocytes. Thus ANG-(1-7) interacted with specific receptors on ARCFs to exert potential antifibrotic and antitrophic effects that could reverse ANG II effects. These results suggest that ANG-(1-7) may play an important role in the heart in regulating cardiac remodeling.

Topics: Angiotensin I; Animals; Binding Sites; Calcium; Cells, Cultured; Collagen; Dose-Response Relationship, Drug; Fibroblasts; Fibrosis; Growth Substances; Heart Ventricles; Male; Peptide Fragments; Protein Binding; Rats; Rats, Sprague-Dawley

The efficacy of ACE inhibitors (ACEIs) in the treatment of chronic heart failures is well documented. However, ACEIs may provide incomplete blockade of the renin-angiotensin system (RAS) because of the alternative pathways for angiotensin II (All) production. We hypothesized that more complete blockade of RAS by adding an AT1 receptor blocker (ARB) may have greater potential to decrease mortality associated with heart failure and improve cardiac function than monotherapy with ACEIs. The objective of this study was to evaluate the effect of combined therapy on cardiac functions and survival in cardiomyopathic hamsters. Male cardiomyopathic hamsters (BIO TO2) were administered either placebo (group C), enalapril (30 mg/kg/day) (group E), or enalapril (30 mg/kg/day) + valsartan (500 mg/ kg/day) (group EV), starting at the age of 6 weeks. Kaplan-Meier analysis was performed to assess the differences in survival. Cardiac functions were evaluated by echocardiogram and cardiac catheterization. Group EV showed significant increases in fractional shortening, LV dP/dTmax, and deceleration time, and showed significant decreases in left ventricular diastolic dimension, LV dP/dTmin, and early diastolic mitral velocity/atrial systolic velocity. Treatment with enalapril resulted in longer survival compared with placebo. Moreover, life expectancy (median probability of survival: 433 days) increased significantly in group EV compared with group E (P<0.05) as well as group C (P<0.001). It is concluded that combined therapy improved cardiac function and survival compared to placebo or enalapril monotherapy.

Topics: Angiotensin I; Angiotensin Receptor Antagonists; Angiotensin-Converting Enzyme Inhibitors; Animals; Cardiomyopathy, Dilated; Cell Count; Chronic Disease; Cricetinae; Drug Therapy, Combination; Echocardiography, Doppler; Enalapril; Fibrosis; Hemodynamics; Male; Myocytes, Cardiac; Random Allocation; Receptors, Angiotensin; Renin-Angiotensin System

Following pericardiotomy in rats, subsequent fibrosis of the visceral pericardium becomes a site of high-density angiotensin-converting enzyme (ACE) binding. This study was undertaken to determine whether this exteriorized site of ACE activity is associated with angiotensin II (AngII) production. Four weeks after pericardiotomy, hearts were isolated and maintained by Krebs-Henseleit perfusion: coronary venous and Thebesian drainage were removed by cannulae. Following a 30-min period of stabilization, a balloon containing superfusate was placed around the heart. Superfusate composition was controlled and included either lisinopril (10(-7) mol/l), angiotensin I (AngI, 10(-7) mol/l), or angiotensinogen (10(-6) mol/l). Sixty min later, superfusate AngII concentration was determined (high-performance liquid chromatography followed by radioimmunoassay). Pericardial fibrosis was confirmed by picrosirius red staining and its high-density ACE binding by quantitative in vitro autoradiography (125I-351A). ACE activity was measured by hippuryl-histidyl-leucine degradation. In coronary effluent, AngII concentration and ACE activity were not different between controls and hearts with pericardial fibrosis. Compared to unoperated, age/sex-matched control hearts, however, we found those with pericardial fibrosis to have: (a) significantly (P < 0.05) greater tissue ACE activity (118.42 +/- 6.66 v 89.45 +/- 7.70 nmol/min/g): (b) significantly (P < 0.01) greater superfusate AngII concentration (4.98 +/- 0.94 v 1.43 +/- 0.28 pg/ml); (c) lisinopril markedly attenuated superfusate AngII concentration to that seen in controls; (d) exogenous AngI markedly increased AngII production (13.76 +/- 1.65 v 4.98 +/- 0.94 pg/ml); and (e) exogenous angiotensinogen did not alter superfusate AngII. Thus, high-density ACE binding and ACE activity of fibrosed pericardium is responsible for AngII production in this in vitro model. Cells involved in generating AngI at this site are uncertain and may involve fibroblast-like cells that express ACE and have ACE activity. The role of local AngII production is unknown, but its autocrine/paracrine properties may regulate collagen turnover of these cells.

Topics: Angiotensin I; Angiotensin II; Animals; Fibrosis; In Vitro Techniques; Male; Myocardium; Peptidyl-Dipeptidase A; Pericardium; Rats; Rats, Sprague-Dawley; Reference Values