To investigate the effect of the angiotensin peptides and their agonists and antagonists on cellular proliferation in proliferating infantile haemangioma (IH) in vitro explants.

Sympathoexcitation contributes to the progression of heart failure. Activation of brain angiotensin II type 1 receptors (AT1R) causes central sympathoexcitation. Thus, we assessed the hypothesis that the increase in circulating angiotensin II comparable to that reported in heart failure model affects cardiac function through the central sympathoexcitation via activating AT1R in the brain. In Sprague-Dawley rats, the subcutaneous infusion of angiotensin II for 14 days increased the circulating angiotensin II level comparable to that reported in heart failure model rats after myocardial infarction. In comparison with the control, angiotensin II infusion increased 24 hours urinary norepinephrine excretion, and systolic blood pressure. Angiotensin II infusion hypertrophied left ventricular (LV) without changing chamber dimensions while increased end-diastolic pressure. The LV pressure -: volume relationship indicated that angiotensin II did not impact on the end-systolic elastance, whereas significantly increased end-diastolic elastance. Chronic intracerebroventricular infusion of AT1R blocker, losartan, attenuated these angiotensin II-induced changes. In conclusion, circulating angiotensin II in heart failure is capable of inducing sympathoexcitation via in part AT1R in the brain, subsequently leading to LV diastolic dysfunction.

The renin-angiotensin system (RAS) controls not only systemic functions, such as blood pressure, but also local tissue-specific events. Previous studies have shown that angiotensin II receptor type 1 (AT1R) and type 2 (AT2R), two RAS components, are expressed in chondrocytes. However, the angiotensin II (ANG II) effects exerted through these receptors on chondrocyte metabolism are not fully understood. In this study, we investigated the effects of ANG II and AT1R blockade on chondrocyte proliferation and differentiation. Firstly, we observed that ANG II significantly suppressed cell proliferation and glycosaminoglycan content in rat chondrocytic RCS cells. Additionally, ANG II decreased CCN2, which is an anabolic factor for chondrocytes, via increased MMP9. In Agtr1a-deficient RCS cells generated by the CRISPR-Cas9 system, Ccn2 and Aggrecan (Acan) expression increased. Losartan, an AT1R antagonist, blocked the ANG II-induced decrease in CCN2 production and Acan expression in RCS cells. These findings suggest that AT1R blockade reduces ANG II-induced chondrocyte degeneration. Interestingly, AT1R-positive cells, which were localized on the surface of the articular cartilage of 7-month-old mice expanded throughout the articular cartilage with aging. These findings suggest that ANG II regulates age-related cartilage degeneration through the ANG II-AT1R axis.

Angiotensin II (Ang II) is a major component of the renin-angiotensin or renin-angiotensin-aldosterone system, which is the main element found to be involved in cardiopathology. Recently, long-term metabolomics studies have linked high levels of angiotensin plasma to inflammatory conditions such as coronary heart disease, obesity, and type 2 diabetes. Monocyte/macrophage cellular function and phenotype orchestrate the inflammatory response in various pathological conditions, most notably cardiometabolic disease. An activation of the Ang II system is usually associated with inflammation and cardiovascular disease; however, the direct effect on monocyte/macrophages has still not been well elucidated. Herein, we have evaluated the cellular effects of Ang II on THP-1-derived macrophages. Ang II stimulated the expression of markers involved in monocyte/macrophage cell differentiation (e.g., CD116), as well as adhesion, cell-cell interaction, chemotaxis, and phagocytosis (CD15, CD44, CD33, and CD49F). Yet, Ang II increased the expression of proinflammatory markers (HLA-DR, TNF-α, CD64, CD11c, and CD38) and decreased CD206 (mannose receptor), an M2 marker. Moreover, Ang II induced cytosolic calcium overload, increased reactive oxygen species, and arrested cells in the G1 phase. Most of these effects were induced via the angiotensin II type 1 receptor (AT1R). Collectively, our results provide new evidence in support of the effect of Ang II in inflammation associated with cardiometabolic diseases.

We evaluated the role of angiotensin II (AII) in a marrow-derived macrophage-driven model of atherosclerosis.

Podocytes play a central role in the maintenance of the glomerular filtration barrier and are cellular targets of angiotensin II (AngII). Non-hemodynamic pathways of AngII signaling regulate cellular function and mediate podocyte abnormalities that are associated with various glomerulopathies, including diabetic kidney disease. In this study we investigated the capacity of AngII to modulate glucose uptake in mouse podocytes expressing the human AT1 receptor (AT1R+) after 5 days of exposure to normal (NG, 5.6 mmol/L) or to high (HG, 30 mmol/L) glucose. Short (30 min) as well as long-term (24 h) incubations with AngII markedly enhanced glucose transport in both NG and HG cells. In podocytes cultured under NG conditions, AngII inhibited insulin-stimulated glucose uptake. Regardless of the presence or absence of AngII, no effect of insulin on glucose uptake was observed in HG cells. Stimulation of glucose transport by AngII was mediated by protein kinase C and by phosphoinositide 3-kinase. Glucose dependent surface expression of the glucose transporters GLUT1, GLUT2, and GLUT4 was modulated by AngII in a time and glucose concentration dependent manner. Furthermore, despite its inhibitory effect on insulin's action, AngII elevated the number of podocyte insulin receptors in both NG and HG cultured cells. These findings demonstrate that AngII modulates podocyte basal, as well as insulin-dependent glucose uptake by regulating glucose transporters and insulin signaling.

Angiotensin II (Ang II) type 1 (AT1) receptor is known to mediate a variety of physiological actions of Ang II including autophagy. However, the role of AT1 receptor in cardiomyocyte autophagy triggered by mechanical stress still remains elusive. The aim of this study was therefore to examine whether and how AT1 receptor participates in cardiomyocyte autophagy induced by mechanical stresses. A 48-hour mechanical stretch and a 4-week transverse aorta constriction (TAC) were imposed to cultured cardiomyocytes of neonatal rats and adult male C57B/L6 mice, respectively, to induce cardiomyocyte hypertrophy prior to the assessment of cardiomyocyte autophagy using LC3b-II. Losartan, an AT1 receptor blocker, but not PD123319, the AT2 inhibitor, was found to significantly reduce mechanical stretch-induced LC3b-II upregulation. Moreover, inhibition of p38MAP kinase attenuated not only mechanical stretch-induced cardiomyocyte hypertrophy but also autophagy. To the contrary, inhibition of ERK and JNK suppressed cardiac hypertrophy but not autophagy. Intriguingly, mechanical stretch-induced autophagy was significantly inhibited by Losartan in the absence of Ang II. Taken together, our results indicate that mechanical stress triggers cardiomyocyte autophagy through AT1 receptor-mediated activation of p38MAP kinase independently of Ang II.

Angiotensin (Ang)-converting enzyme 2 (ACE2) metabolizes Ang II to the vasodilatory peptide Ang(1-7), while neprilysin (NEP) generates Ang(1-7) from Ang I. Experiments used novel Surface Enhanced Laser Desorption Ionization-Time of Flight (SELDI-TOF) mass spectroscopic (MS) assays to study Ang processing. Mass spectroscopy was used to measure proteolytic conversion of Ang peptide substrates to their specific peptide products. We compared ACE/ACE2 activity in plasma, brain and kidney from C57BL/6 and NEP(-/-) mice. Plasma or tissue extracts were incubated with Ang I or Ang II (1296 or 1045, m/z, respectively), and generated peptides were monitored with MS. Angiotensin-converting enzyme 2 activity was detected in kidney and brain, but not in plasma. Brain ACE2 activity was highest in hypothalamus. Angiotensin-converting enzyme 2 activity was inhibited by the specific ACE2 inhibitor, DX600 (10 microm, 99% inhibition), but not by the ACE inhibitor, captopril (10 microm). Both MS and colorimetric assays showed high ACE activity in plasma and kidney with low levels in brain. To extend these findings, ACE measurements were made in ACE overexpressing mice. Angiotensin-converting enzyme four-copy mice showed higher ACE activity in kidney and plasma with low levels in hypothalamus. In hypothalamus from NEP-/- mice, generation of Ang(1-7) from Ang I was decreased, suggesting a role for NEP in Ang metabolism. With Ang II as substrate, there was no difference between NEP-/- and wild-type control mice, indicating that other enzymes may contribute to generation of Ang(1-7). The data suggest a predominant role of hypothalamic ACE2 in the processing of Ang II, in contrast to ACE, which is most active in plasma.

It is postulated that central effects of angiotensin (Ang) II may be indirect due to rapid conversion to Ang III by aminopeptidase A (APA). Previously, we showed that Ang II and Ang III induced mitogen-activated protein (MAP) kinases ERK1/2 and stress-activated protein kinase/Jun-terminal kinases (SAPK/JNK) phosphorylation in cultured rat astrocytes. Most importantly, both peptides were equipotent in causing phosphorylation of these MAP kinases. In these studies, we used brainstem and cerebellum astrocytes to determine whether Ang II's phosphorylation of these MAP kinases is due to the conversion of the peptide to Ang III. We pretreated astrocytes with 10  μ M amastatin A or 100  μ M glutamate phosphonate, selective APA inhibitors, prior to stimulating with either Ang II or Ang III. Both peptides were equipotent in stimulating ERK1/2 and SAPK/JNK phosphorylation. The APA inhibitors failed to prevent Ang II- and Ang III-mediated phosphorylation of the MAP kinases. Further, pretreatment of astrocytes with the APA inhibitors did not affect Ang II- or Ang III-induced astrocyte growth. These findings suggest that both peptides directly induce phosphorylation of these MAP kinases as well as induce astrocyte growth. These studies establish both peptides as biologically active with similar intracellular and physiological effects.

The current study was aimed at examining the effects of moderate‑intensity endurance exercise on the expression of angiotensin II (AngII) and AngII type 1 receptor (AT1R) in the rat heart. Male Sprague‑Dawley rats were randomly divided into the control group (n=20) and moderate‑intensity endurance exercise group (n=20). Cardiac hypertrophy was induced by treadmill endurance training for 8 weeks. The mRNA expression of AngII and AT1R were assessed by reverse transcription‑quantitative polymerase chain reaction. The immune response positive area and optical density of AngII and AT1R was measured by immunohistochemistry. AngII was primarily expressed in the cytoplasm and membrane, however infrequently in coronary vascular wall smooth muscle cells. AT1R was primarily expressed in the coronary vessel wall smooth muscle, rarely in cardiac cells. The mRNA expression of cardiac AngII was significantly increased after the 8‑week exercise period, while AT1R was significantly decreased. Immunohistochemistry indicated a significant increase in the AngII immune‑positive area and optical density after the 8‑week exercise. The AT1R immune‑positive area and optical density was significantly reduced following the 8‑week exercise. In conclusion, subsequent to 8‑weeks endurance training, the AngII expression was increased and the AT1R expression was decreased. AT1R may expand the coronary artery, thereby increasing coronary blood flow and ensuring the energy supply of heart during exercise. The expression change in AngII does not reflect the character of cardiac hypertrophy. The exercise‑induced change in the expression of AngII and AT1R may be a protective mechanism to avoid cardiac pathological hypertrophy.

Angiotensin-(1-9) [Ang-(1-9)], generated from Ang I by Ang II converting enzyme 2, has been reported to have protective effects on cardiac and vascular remodeling. However, there is no report about the effect of Ang-(1-9) on pulmonary hypertension. The aim of the present study is to investigate whether Ang-(1-9) improves pulmonary vascular remodeling in monocrotaline (MCT)-induced pulmonary hypertensive rats. Sprague-Dawley rats received Ang-(1-9) (576 µg/kg/day) or saline via osmotic mini-pumps for 3 weeks. Three days after implantation of osmotic mini-pumps, 50 mg/kg MCT or vehicle were subcutaneously injected. MCT caused increases in right ventricular weight and systolic pressure, which were reduced by co-administration of Ang-(1-9). Ang-(1-9) also attenuated endothelial damage and medial hypertrophy of pulmonary arterioles as well as pulmonary fibrosis induced by MCT. The protective effects of Ang-(1-9) against pulmonary hypertension were inhibited by Ang type 2 receptor (AT2R) blocker, but not by Mas receptor blocker. Additionally, the levels of LDH and inflammatory cytokines, such as TNF-α, MCP-1, IL-1β, and IL-6, in plasma were lower in Ang-(1-9) co-treated MCT group than in vehicle-treated MCT group. Changes in expressions of apoptosis-related proteins such as Bax, Bcl-2, Caspase-3 and -9 in the lung tissue of MCT rats were attenuated by the treatment with Ang-(1-9). These results indicate that Ang-(1-9) improves MCT-induced pulmonary hypertension by decreasing apoptosis and inflammatory reaction via AT2R.

Prospective, placebo-controlled clinical trials suggest that estrogen may have adverse effects on the vascular system in women. The goal of this study was to determine if 17beta-estradiol (E2) would have adverse effects on the renovasculature in a rat model of renal injury characterized by low nitric oxide (NO) and high angiotensin II (AngII). We studied female Wistar rats that were sham-operated (sham), ovariectomized (OVX), or ovariectomized and replaced with E2 (OVX/E2). All rats were maintained on a high salt diet and renovascular injury was caused by treating rats with an inhibitor of NO synthase, N(omega)-nitro-L-arginine-methyl-ester (L-NAME), for 14 days, plus AngII on days 11 through 14. L-NAME/AngII treatment, as compared to placebo, caused proteinuria, glomerular injury, and fibrinoid necrosis of renal arterioles in sham-operated rats. Ovariectomy reduced L-NAME/AngII-induced renal damage, whereas E2 treatment increased L-NAME/AngII-induced damage in OVX rats. In rats treated with L-NAME/AngII, levels of AngII type 1 receptor (AT(1)R) protein were higher in the renal cortex of sham and OVX/E2 rats than in OVX rats. AT(1)R protein correlated with renal injury. E2 treatment also increased expression of AT(1)R mRNA. Thus, under conditions of low NO and high AngII, E2 exacerbated renal injury. E2-mediated increases in renal cortical AT(1)R expression may represent a novel mechanism for the adverse renovascular effects of estrogen.

The angiotensin II (Ang II) is the principal effector peptide of the RAS system. It has a pleiotropic effect and, beside its physiological role, it has the property to stimulate angiogenesis and activate multiple signalling pathways related to cell proliferation. The purpose of the study was to determinate the Ang II expression and localization in Sardinian pterygium and normal conjunctiva by immunohistochemistry, and its possible involvement in the development and progression of the disease. Twenty-three pterygiums and eleven normal conjunctiva specimens obtained from Sardinian patients, were processed for paraffin embedding and assessed for the immunohistochemical revelation of Ang II. Significant Ang II expression was identified in pterygium and conjuntica. Particularly, thirteen pterygium specimens (n=13) displayed exclusively moderate to strong nuclear staining; some specimens (n=5) showed exclusively a moderate cytoplasmatic immunoreactivity, and few specimens (n=2) displayed moderate to strong immunoreactivity in both cytoplasm and nucleus. Statistical significance difference in respect of nuclear and cytoplasmatic localization was observed between normal conjunctiva and pterygium (P=0.038).The results showed a predominant intranuclear localization of Ang II in pterygium epithelial cells, in spite of conjunctiva that mainly showed cytoplasmatic localization. In view of these results, we hypothesized a possible gene expression modulator role played by Ang II in pterygium.

Microglial cells are important contributors to the neuroinflammation and blood vessel damage that occurs in ischemic retinopathies. We hypothesized that key effectors of the renin-angiotensin aldosterone system, angiotensin II (Ang II) and aldosterone, increase the density of microglia in the retina and stimulate their production of reactive oxygen species (ROS) as well as pro-angiogenic and pro-inflammatory factors. Two animal models were studied that featured up-regulation of Ang II or aldosterone and included transgenic Ren-2 rats which overexpress renin and Ang II in tissues including the retina, and Sprague Dawley rats with ischemic retinopathy and infused with aldosterone. Complementary studies were performed in primary cultures of retinal microglia from neonatal Sprague Dawley rats exposed to hypoxia (0.5% O2) and inhibitors of the angiotensin type 1 receptor (valsartan), the mineralocorticoid receptor (spironolactone) or aldosterone synthase (FAD286). In both in vivo models, the density of ionized calcium-binding adaptor protein-1 labelled microglia/macrophages was increased in retina compared to genetic or vehicle controls. In primary cultures of retinal microglia, hypoxia increased ROS (superoxide) levels as well as the expression of the NADPH oxidase (NOX) isoforms, NOX1, NOX2 and NOX4. The elevated levels of ROS as well as NOX2 and NOX4 were reduced by all of the treatments, and valsartan and FAD286 also reduced NOX1 mRNA levels. A protein cytokine array of retinal microglia revealed that valsartan, spironolactone and FAD286 reduced the hypoxia-induced increase in the potent pro-angiogenic and pro-inflammatory agent, vascular endothelial growth factor as well as the inflammatory factors, CCL5 and interferon γ. Valsartan also reduced the hypoxia-induced increase in IL-6 and TIMP-1 as well as the chemoattractants, CXCL2, CXCL3, CXCL5 and CXCL10. Spironolactone and FAD286 reduced the levels of CXCL2 and CXCL10, respectively. In conclusion, our findings that both Ang II and aldosterone influence the activation of retinal microglia implicates the renin-angiotensin aldosterone system in the pathogenesis of ischemic retinopathies.

Background Angiotensin II (Ang II) can cause hypertension and tissue impairment via AGTR1 (Ang II receptor type 1), particularly in renal proximal tubule cells, and can cause DNA damage in renal cells via nicotinamide adenine dinucleotide phosphate oxidase. BubR1 (budding uninhibited by benzimidazole-related 1) is a multifaceted kinase that functions as a mitotic checkpoint. BubR1 expression can be induced by Ang II in smooth muscle cells in vitro, but the relationship between systemic BubR1 expression and the Ang II response is unclear. Methods and Results Twenty 24-week-old male BubR1 low-expression mice (BubR1L/L mice) and age-matched BubR1+/+ mice were used in this study. We investigated how Ang II stimulation affects BubR1L/L mice. The elevated systolic blood pressure caused by Ang II stimulation in BubR1+/+ mice was significantly attenuated in BubR1L/L mice. Additionally, an attenuated level of Ang II-induced perivascular fibrosis was observed in the kidneys of BubR1L/L mice. Immunohistochemistry revealed that the overexpression of AGTR1 induced by Ang II stimulation was repressed in BubR1L/L mice. We evaluated AGTR1 and Nox-4 (nicotinamide adenine dinucleotide phosphate oxidase-4) levels to determine the role of BubR1 in the Ang II response. Results from in vitro assays of renal proximal tubule cells suggest that treatment with small interfering RNA targeting BubR1 suppressed Ang II-induced overexpression of AGTR1. Similarly, the upregulation in Nox4 and Jun N-terminal kinase induced by Ang II administration was repressed by treatment with small interfering RNA targeting BubR1. Conclusions Ang II-induced hypertension is caused by AGTR1 overexpression in the kidneys via the upregulation of BubR1 and Nox4.

Cardiac hypertrophy is an early hallmark during the clinical course of heart failure and regulated by various signalling pathways. Recently, we observed that mouse embryonic fibroblasts from CD38 knockout mice were significantly resistant to oxidative stress such as H2 O2 -induced injury and hypoxia/reoxygenation-induced injury. In addition, we also found that CD38 knockout mice protected heart from ischaemia reperfusion injury through activating SIRT1/FOXOs-mediated antioxidative stress pathway. However, the role of CD38 in cardiac hypertrophy is not explored. Here, we investigated the roles and mechanisms of CD38 in angiotensin II (Ang-II)-induced cardiac hypertrophy. Following 14 days of Ang-II infusion with osmotic mini-pumps, a comparable hypertension was generated in both of CD38 knockout and wild-type mice. However, the cardiac hypertrophy and fibrosis were much more severe in wild-type mice compared with CD38 knockout mice. Consistently, RNAi-induced knockdown of CD38 decreased the gene expressions of atrial natriuretic factor (ANF) and brain natriuretic peptide (BNP) and reactive oxygen species generation in Ang-II-stimulated H9c2 cells. In addition, the expression of SIRT3 was elevated in CD38 knockdown H9c2 cells, in which SIRT3 may further activate the FOXO3 antioxidant pathway. The intracellular Ca2+ release induced by Ang-II markedly decreased in CD38 knockdown H9c2 cells, which might be associated with the decrease of nuclear translocation of NFATc4 and inhibition of ERK/AKT phosphorylation. We concluded that CD38 plays an essential role in cardiac hypertrophy probably via inhibition of SIRT3 expression and activation of Ca2+ -NFAT signalling pathway. Thus, CD38 may be a novel target for treating cardiac hypertrophy.

Angiotensin II type 1 receptor (AT1R)-associated protein (ATRAP) promotes AT1R internalization along with suppression of pathological activation of tissue AT1R signaling. However, the functional significance of ATRAP in renal sodium handling and blood pressure regulation under pathological stimuli is not fully resolved. Here we show the blood pressure of mice with a gene-targeted disruption of ATRAP was comparable to that of wild-type mice at baseline. However, in ATRAP-knockout mice, angiotensin II-induced hypertension was exacerbated and the extent of positive sodium balance was increased by angiotensin II. Renal expression of the sodium-proton antiporter 3, a major sodium transporter in the proximal tubules, urinary pH, renal angiotensinogen production, and angiotensin II content was unaffected. Stimulation of the renal expression and activity of the epithelial sodium channel (ENaC), a major sodium transporter in the distal tubules, was significantly enhanced by chronic angiotensin II infusion. The circulating and urinary aldosterone levels were comparable. The blood pressure response and renal ENaC expression by aldosterone were not affected. Thus, ATRAP deficiency exacerbated angiotensin II-mediated hypertension by pathological activation of renal tubular AT1R by angiotensin II. This directly stimulates ENaC in the distal tubules and enhances sodium retention in an aldosterone-independent manner.

Angiotensin II (Ang II) has been shown to activate multiple downstream pathways resulting in endothelial dysfunction and oxidative stress. Baicalin, a natural flavone, exerts anti-oxidant and anti-apoptotic effects in cardiovascular diseases. In the present study, we hypothesized that baicalin has beneficial effects in Ang II-induced endothelial cells injury. Here, we shown that baicalin improved endothelial fuction impaired by Ang II through promoting endothelial-dependent vasodilation and suppressing the apoptosis of HUVECs in which baicalin decreased the expression of bax and cleaved caspase-3, and increased bcl-2 expression. Additionally, baicalin significantly conversed Ang II to angiotensin-1-7 [Ang-(1-7)] by activating angiotensin-converting enzyme 2 (ACE2) and Mas receptor mRNA expression and protein expression. Moreover, treatment with baicalin significantly reduced cell oxidative damage induced by Ang II through MDA/ROS decrease and NO/T-AOC increase. This antioxidant capacity was related to the increases of PI3K, phosphor-AKT (Ser-473) and phosphor-eNOS (Ser-1177). In conclusion, our results implicate that baicalin could protect endothelial cells from Ang II-induced endothelial dysfunction and oxidative stress via modulating the expression of bax, bcl-2 and cleaved caspase-3, activating ACE2/Ang-(1-7)/Mas axis and up-regulating PI3K/AKT/eNOS pathway.

The aims of this study were to assess the renal expression of angiotensin II type 1 receptor (AT1R), angiotensin II type 2 receptor (AT2R), and MAS receptor in human type 2 diabetic nephropathy (DN).

Angiotensin II (Ang II), whose generation largely depends on angiotensin-converting enzyme (ACE) activity, mediates most of the renin-angiotensin-system (RAS) effects. Elastase-2 (ELA-2), a chymotrypsin-serine protease elastase family member 2A, alternatively generates Ang II in rat arteries. Myocardial infarction (MI) leads to intense RAS activation, but mechanisms involved in Ang II-generation in resistance arteries are unknown. We hypothesized that ELA-2 contributes to vascular Ang II generation and cardiac damage in mice subjected to MI.