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Differential inducibility of angiotensin II AT2 receptor between SHR and WKY vascular smooth muscle cells
Abstract
Although the fetal aorta expresses a substantial amount of angiotensin II type 2 receptors, the expression level of angiotensin II type 2 receptors in the adult aorta and cultured vascular smooth muscle cells is very low or even absent. Prolonged serum depletion (6 to 8 days) with a supplement of insulin, transferrin and sodium selenite induced angiotensin II type 2 receptors and mRNA in cultured vascular smooth muscle cells from Wistar Kyoto rats. Insulin was found to be essential for the induction of the receptor. However, these receptors could not be induced in cultured vascular smooth muscle cells from spontaneously hypertensive rats. These results suggest that: (1) insulin plays an important role for the expression of the angiotensin II type 2 receptor gene; and (2) the type 2 receptor gene expression is differentially regulated between cultured vascular smooth muscle cells of Wistar Kyoto rats and spontaneously hypertensive rats.
... The values represent the average of triplicate determinations. Serum-free culture is known to increase AT 2 expression in cultured cells [34]. ...
... To evaluate the effects of Ang II and the AT 2 receptor signaling on the growth of PAN02 cells in vitro, the effect of a low concentration of Ang II (10 nM) was examined on the growth of PAN02 cells co-cultured with MSFs prepared from either wild type or AT 2 -KO mice or with AT 2 receptor over-expressing MSFs prepared from either wild type or AT 2 -KO mice. Since AT 2 receptor expression is known to be attenuated in culture [34], AT 2 receptor expression should be assured by the receptor over-expression. As shown in Figure 5, growth of PAN02 was significantly attenuated when the AT 2 receptor was over-expressed in co-cultured MSFs. ...
Pancreatic cancer is one of the most aggressive human malignancies, with a very poor prognosis. To evaluate the effect of angiotensin II (Ang II) type 2 receptor (AT2) expression in the host's body on the growth of pancreatic carcinoma, we have investigated the growth of mouse pancreatic ductal carcinoma grafts in syngeneic wild type and AT2 receptor-deficient (AT2-KO) mice.
The role of AT2 receptor-signaling in stromal cells on the growth of murine pancreatic carcinoma cells (PAN02) was studied using various in vitro and in vivo assays. In vivo cell proliferation, apoptosis, and vasculature in tumors were monitored by Ki-67 immunostaining, TUNEL assay, and von Willebrand factor immunostaining, respectively. In the co-culture study, cell proliferation was measured by MTT cell viability assay. All the data were analyzed using t-test and data were treated as significant when p < 0.05.
Our results show that the growth of subcutaneously transplanted syngeneic xenografts of PAN02 cells, mouse pancreatic ductal carcinoma cells derived from the C57/BL6 strain, was significantly faster in AT2-KO mice compared to control wild type mice. Immunohistochemical analysis of tumor tissue revealed significantly more Ki-67 positive cells in xenografts grown in AT2-KO mice than in wild type mice. The index of apoptosis is slightly higher in wild type mice than in AT2-KO mice as evaluated by TUNEL assay. Tumor vasculature number was significantly higher in AT2-KO mice than in wild type mice. In vitro co-culture studies revealed that the growth of PAN02 cells was significantly decreased when grown with AT2 receptor gene transfected wild type and AT2-KO mouse-derived fibroblasts. Faster tumor growth in AT2-KO mice may be associated with higher VEGF production in stromal cells.
These results suggest that Ang II regulates the growth of pancreatic carcinoma cells through modulating functions of host stromal cells; Moreover, Ang II AT2 receptor signaling is a negative regulator in the growth of pancreatic carcinoma cells. These findings indicate that the AT2 receptor in stromal fibroblasts is a potentially important target for chemotherapy for pancreatic cancer.
... Despite the fact that the media of rat thoracic aorta express the AT 2 receptor, smooth muscle cells derived from the aorta and cultured in vitro do not Ichiki et al., 1996). This is possibly due to down-regulation of the AT 2 receptor gene by growth factors in serum-containing culture medium since AT 2 receptor expression can be regained upon prolonged serum depletion in the presence of insulin . ...
... The cloned mouse preadipocyte cell line, Ob1771, expresses the AT 2 receptor upon differentiation in a serum-free medium and responds to Ang II by producing prostacyclin, which promotes the differentiation of the preadipocytes by a paracrine mechanism (Darimont et al., 1994). Rat VSMC in culture do not express the AT 2 receptor under regular culture conditions in the presence of fetal calf ANGIOTENSIN RECEPTORS serum (Stoll et al., 1995;Ichiki et al., 1996;Kambayashi et al., 1996), but, as outlined above, the AT 2 receptor can emerge in rat thoracic VSMC when cultured in serumdepleted medium supplemented with insulin or other insulin-like growth factors such as IGF Kambayashi et al., 1996). Thus rat VSMC grown and maintained under ordinary culture conditions do not seem to provide convenient materials for the study of the role of the AT 2 receptor. ...
The cardiovascular and other actions of angiotensin II (Ang II) are mediated by AT(1) and AT(2) receptors, which are seven transmembrane glycoproteins with 30% sequence similarity. Most species express a single autosomal AT(1) gene, but two related AT(1A) and AT(1B) receptor genes are expressed in rodents. AT(1) receptors are predominantly coupled to G(q/11), and signal through phospholipases A, C, D, inositol phosphates, calcium channels, and a variety of serine/threonine and tyrosine kinases. Many AT(1)-induced growth responses are mediated by transactivation of growth factor receptors. The receptor binding sites for agonist and nonpeptide antagonist ligands have been defined. The latter compounds are as effective as angiotensin converting enzyme inhibitors in cardiovascular diseases but are better tolerated. The AT(2) receptor is expressed at high density during fetal development. It is much less abundant in adult tissues and is up-regulated in pathological conditions. Its signaling pathways include serine and tyrosine phosphatases, phospholipase A(2), nitric oxide, and cyclic guanosine monophosphate. The AT(2) receptor counteracts several of the growth responses initiated by the AT(1) and growth factor receptors. The AT(4) receptor specifically binds Ang IV (Ang 3-8), and is located in brain and kidney. Its signaling mechanisms are unknown, but it influences local blood flow and is associated with cognitive processes and sensory and motor functions. Although AT(1) receptors mediate most of the known actions of Ang II, the AT(2) receptor contributes to the regulation of blood pressure and renal function. The development of specific nonpeptide receptor antagonists has led to major advances in the physiology, pharmacology, and therapy of the renin-angiotensin system.
... In cultured tubular cells, serum deprivation, TNF␣, and Ang II stimulation caused apoptosis [13,25]. Although there are no studies in cultured tubular cells, serum deprivation up-regulated AT 2 in mesangial cells and VSMC [26,27], showing that factors involved in apoptosis could also be responsible for the tubular AT 2 over-expression found in folic acid nephropathy. AT 2 may also be involved in apoptosis through the NF-B pathway. ...
... In cultured mesangial cells and VSMC, prolonged serum depletion induced AT 2 mRNA expression. This effect was only found in cells from Wistar-Kyoto rats, but not from spontaneously hypertensive rats (SHR) [26,27]. In VSMC, insulin-like growth factor upregulates AT 2 [34], and in rat neonatal myocytes, protein kinase C activators, such as Ang II down-regulated AT 2 mRNA level [35]. ...
Activation of the renin angiotensin system has been described in pathologic conditions, including kidney damage. Angiotensin II (Ang II) acts through two receptors, AT1 and AT2. Most of the known actions of Ang II, including vasoconstriction and fibrosis, are due to AT1 activation. Recent data suggest that AT2 participates in the regulation of cell growth and renal inflammatory infiltration. Therefore, we investigated the renal expression of AT2 receptors in several models of renal injury.
Investigations were done in the following experimental models of kidney damage: systemic infusion of Ang II (inflammation), folic acid nephropathy (tubular cell death), and protein overload proteinuria. AT2 expression was determined by immunohistochemistry (protein) and reverse transcription-polymerase chain reaction (RT-PCR) (gene).
In control animals, low levels of renal expression of AT2 were found. Ang II infusion resulted in an up-regulation of AT2 in tubular cells and de novo AT2 expression in glomeruli and vessels, associated with the presence of inflammatory cells. Acute tubular injury induced by folic acid was characterized by AT2 overexpression and apoptosis in tubular cells. Protein overload caused heavy proteinuria and tubular AT2 up-regulation.
AT2 is re-expressed in pathologic conditions of kidney damage, such as inflammation, apoptosis, and proteinuria, suggesting a potential role of this receptor during renal injury.
... Since the AT 2 protein expression level in this cultured cell is smaller than those reported (27,34) the AT 2 protein level may not be sufficient to exhibit the AT 2 -mediated Ang II effect in COX-2 expression. In this study, the AT 2 was over-expressed by transfecting cells with AT 2 cDNA using the AT 2 adenoviral vector. ...
... Proportion of the AT 2 receptor expression levels seems smaller than the previous measurement of the receptor expression in intestinal mucosa in vivo (27). However this result seems reasonable, since AT 2 expression is suppressed during cell culture but is inducible by removing serum from the culture medium (34). Indeed, AT 2 protein expression in RIE-1 cells was noticeably increased by serum removal (data not shown). ...
We previously demonstrated that angiotensin II (Ang II) receptor signaling is involved in azoxymethane-induced mouse colon tumorigenesis. In order to clarify the role of Ang II in COX-2 expression in the intestinal epithelium, the receptor subtype-specific effect on COX-2 expression in a rat intestinal epithelial cell line (RIE-1) has been investigated. Ang II dose- and time-dependently increased the expression of COX-2, but not COX-1 mRNA and protein. This stimulation was completely blocked by the AT(1) receptor antagonist but not the AT(2) receptor antagonist. Ang II and lipopolysaccharide (LPS) additively induced COX-2 protein in RIE-1 cells, whereas the LPS-induced COX-2 expression was significantly attenuated by low concentrations of Ang II or the AT(2) agonistic peptide CGP-42112A only in AT(2) over-expressed cells. These data indicate that Ang II bi-directionally regulates COX-2 expression via both AT(1) and AT(2) receptors. Control of COX-2 expression through Ang II signaling may have significance in cytokine-induced COX-2 induction and colon tumorigenesis.
... In contrast, Igarachi et al. have demonstrated that inhibition of AT1R restores insulin resistance induced by Ang II in VSMC derived from both normal and diabetics rat while PD123319, an AT2R antagonist had no effect (Igarashi et al., 2007). Additionally, the expression of AT2R is extremely low in adult smooth muscle cells (Ichiki et al., 1996). Therefore, it is likely that the contribution of AT2R in our system is minimal. ...
Insulin resistance is an underlying mechanism of type 2 diabetes and its vascular complications. Recent evidence suggests that crosstalk between angiotensin II (Ang II) and the insulin signaling in vascular smooth muscle cell (VSMC) may contribute to cellular insulin resistance. We hypothesized that Ang II inhibits the anti-mitogenic pathways while enhancing the mitogenic pathways stimulated by insulin via activation of Protein Tyrosine Phosphatase-1B (PTP-1B) in VSMC. We found that Ang II significantly inhibited insulin-induced phosphorylation of tyrosine 608 of IRS-1 and serine 473 of Akt, a downstream member of anti-mitogenic pathway of insulin. In contrast, Ang II increased the serine phosphorylation of IRS-1 which was not affected by the presence of insulin. Activation of p42/p44 MAPK (a mitogenic pathway) induced by insulin was further enhanced by Ang II. Transfection of VSMC with PTP-1B antisense oligonucleotide markedly reduced the effects of Ang II on insulin signaling. Furthermore, an increase in VSMC growth was attenuated by PTP-1B antisense only in the presence of both Ang II and insulin. Finally, we also showed that Ang II-induced activation of PTP-1B in VSMC was PKA/JAK2 dependent. We conclude that Ang II modulates both anti-mitogenic and mitogenic pathways of insulin via the activation of PTP-1B.
... A lack of inducible AT 2 receptor expression in cultured rat glomerular mesangial cells from stroke-prone spontaneously hypertensive rats and aortic vascular smooth muscle cells from spontaneously hypertensive rats has been described. 5,28 The AT 2 receptor not only causes vasodilation in the preglomerular afferent arteriole 7 but also exerts an antiproliferative effect 29 and induces apoptosis. 30 Recently, accelerated renal interstitial fibrosis and collagen deposition has been observed in adult AT 2 receptor null mutant mice during unilateral ureteral obstruction. ...
-This study was designed to investigate distribution and regulation of the renal AT1A and AT2 subtype receptors in rats with either systemic angiotensin II (Ang II)-induced hypertension or acute phase renal hypertension (2-kidney, 1-clip [2K1C] or 2-kidney, 1-figure-of-8-wrap [2K1W]). In normal rat kidneys, positive immunostaining for the AT1A receptor was observed in the intrarenal vasculature, glomeruli, proximal and distal tubules, and collecting ducts. The AT2 receptor was localized mainly to the glomeruli. The AT1A but not AT2 receptor protein expression was significantly reduced in rats with 10-day systemic Ang II-induced hypertension. In both 7-day 2K1C and 3-day 2K1W rats, the AT1A receptor was significantly reduced in ischemic and contralateral kidneys compared with sham-operated control rats. Reduction in AT2 receptor expression was observed only in the ischemic kidneys in 2K1C and 2K1W renal hypertensive rats. These results demonstrate that the AT1A receptor is widely distributed in the glomerulus and all other nephron segments of the rat kidney. Renal AT1A but not AT2 receptor protein is downregulated in rats with Ang II-induced hypertension. In renal hypertensive rats, the AT1A receptor is bilaterally downregulated and the AT2 receptor is downregulated only in the ischemic kidney.
... Expression of the AT 2 receptor in the fetal aorta is substantial, while that in the adult aorta and cultured VSMCs is very low or even absent. Kambayashi and colleagues 78 reported that prolonged serum depletion (6 to 8 days) with a supplement of insulin induced expression of AT 2 receptor mRNA in cultured VSMCs from Wistar-Kyoto rats, but receptor expression could not be induced in VSMCs prepared from SHR. 79 They also reported that insulin-like growth factor upregulates AT 2 receptor expression in cultured VSMCs. Moreover, vasoactive substances with the protein kinase C-calcium pathway, such as norepinephrine and Ang II, have been reported to downregulate the AT 2 mRNA level in cultured rat neonatal myocytes. ...
Angiotensin II (Ang II) plays an important role in regulating cardiovascular hemodynamics and structure. Multiple lines of evidence have suggested the existence of Ang II receptor subtypes, and at least 2 distinct receptor subtypes have been defined on the basis of their differential pharmacological and biochemical properties and designated as type 1 (AT1) and type 2 (AT2) receptors. To date, most of the known effects of Ang II in adult tissues are attributable to the AT1 receptor. Recent cloning of the AT2 receptor contributes to reveal its physiological functions, but many functions of the AT2 receptor are still an enigma. AT1 and AT2 receptors belong to the 7-transmembrane, G protein-coupled receptor family. However, accumulating evidence demonstrates that the function and signaling mechanisms of these receptor subtypes are quite different, and these receptors may exert opposite effects in terms of cell growth and blood pressure regulation. We will review the role of the AT2 receptor in the cardiovascular system and the molecular and cellular mechanisms of AT2 receptor action.
... 10,20 It has been suggested that the lack of AT 2 receptors in vascular SMC of SHR might be partly responsible for the increased growth of vascular SMC and vascular remodeling through the AT 1 receptors in SHR. 21 Furthermore, Sabri et al 22 have also suggested that in Ang II-induced hypertension, changes in vascular SMC phenotypes are triggered primarily through the AT 1 receptors. In addition, kinin metabolism, including nitric oxide and prostaglandins and other systems, 11 might be involved in the difference in action between FK and EN on the NMHC isoform expression of SMC in SHR aortic tunica media. ...
The aim of this study was to clarify the differences between the angiotensin II type 1 (AT1) receptor antagonist and the angiotensin-converting enzyme (ACE) inhibitor on smooth muscle and nonmuscle myosin heavy chain isoforms in aortic smooth muscle cells of Wistar-Kyoto rats and spontaneously hypertensive rats. All 4 myosin heavy chain isoforms are heterogeneously expressed in the smooth muscle cells of the aortic tunica media in 20-week-old rats, and the contractile-type myosin heavy chains are highly expressed in smooth muscle cells of the aortic tunica media compared with the synthetic-type myosin heavy chains. Both the AT1 receptor antagonist and the ACE inhibitor had the same effects on hemodynamics, smooth muscle cell hypertrophy and proliferation, fibrosis, and vascular remodeling in spontaneously hypertensive rats. However, the AT1 receptor antagonist had a more potent effect on the downregulation of the synthetic-type myosin heavy chains than the ACE inhibitor in spontaneously hypertensive rat aortic tunica media. In contrast, these effects of the AT1 receptor antagonist and the ACE inhibitor on hemodynamics, morphology, fibrosis, and expression of myosin heavy chain isoforms in smooth muscle cells of the aortic tunica media were not observed in Wistar-Kyoto rats. Thus, within 6 weeks, the AT1 receptor antagonist might modulate the cellular composition of myosin heavy chain isoforms in smooth muscle cells more efficiently than the ACE inhibitor, without morphological changes in the spontaneously hypertensive rat aorta.
... Growth factors, including epidermal growth factor, nerve growth factor, and platelet-derived growth factor, also downregulate AT 2 receptor mRNA expression in PC12 cells (12) and R3T3 cells (8). On the other hand, Ichiki et al. (8,9) and Kambayashi et al. (11) reported that AT 2 receptor mRNA is upregulated by interleukin-1, insulin, and insulin-like growth factor. According to these observa- tions, it is more conceivable that AT 2 receptor is downregulated in left ventricular hypertrophy as observed in the present study, because PKC, cAMP, and growth factors are all increased in left ventricular hypertrophy. ...
The cellular localization of the AT(2) receptor and the regulation of its expression in hypertrophied left ventricle are not well known. We compared the expression of the cardiac AT(1) and AT(2) receptor in spontaneously hypertensive rats/Izumo strain (SHR/Izm) and Wistar Kyoto rats/Izumo strain (WKY/Izm), ages 4, 12, and 20 wk, by means of immunohistochemistry and Western blot analysis. In SHR/Izm, compared with WKY/Izm, blood pressure (161 +/- 2 vs. 120 +/- 2 mmHg at 12 wk, P </= 0.01, and 199 +/- 3 vs. 123 +/- 3 mmHg at 20 wk, P </= 0.01) and heart-to-body weight ratio (3.76 +/- 0.07 vs. 3.06 +/- 0.06 mg/g at 12 wk, P </= 0.01, and 3.90 +/- 0.08 vs. 3.01 +/- 0.12 mg/g at 20 wk, P </= 0.01) were significantly elevated. There was no difference in these values between the two strains at 4 wk of age. Histologically, 20-wk-old SHR/Izm demonstrated myocardial hypertrophy, a thickening of the smooth muscle layer of the intracardiac arteries, and perivascular fibrosis. By immunohistochemistry, the AT(2) receptor was localized to cardiomyocytes and vascular endothelial cells, but not in the vascular smooth muscle cells. No major AT(2) receptor signal was observed in perivascular fibrosis at any age in either strain of rats. No difference was detected in this localization between the two strains. By Western blotting, a single 44-kDa band for the AT(2) receptor and a single 60-kDa band for the AT(1) receptor were detected in ventricles from both strains of rats at all ages. Densitometric analysis demonstrated that the AT(2) receptor 44-kDa band was decreased by 20% at 12 wk and 32% at 20 wk (P < 0.01) in SHR/Izm compared with WKY/Izm. The intensity of the AT(1) receptor 60-kDa band was increased by 57% in 20-wk-old SHR/Izm compared with WKY/Izm (P < 0.05). There was no significant difference in the intensity of the 44- or 60-kDa bands in 4-wk-old animals of either strain. We demonstrated a decrease in the AT(2) receptor and an increase in the AT(1) receptor protein with no change in their localizations in hypertrophied left ventricular myocytes of SHR/Izm.
... However, in blood vessels of adult rodent models of hypertension, such as the SHR in vivo, the VSMC phenotype is highly differentiated, and recent studies with Ang II type 2 (AT 2 ) receptor knockout mouse strains have documented the important role of the AT 2 receptor subtype in the mediation of vasculogenesis and, more recently, in the mediation of VSMC differentiation. 23 Because this receptor subtype has recently been shown to mediate VSMC apoptosis in vitro 24 and has been previously shown to have different levels of expression in VSMCs from models of hypertension compared with respective controls, 25 it is also possible that the differences in growth and apoptosis in VSMCs isolated from the SHRSP compared with the WKY, as reported in the present study, may be related to altered AT 1 /AT 2 receptor expression levels or function. Such cellular mechanisms are presently hypothetical because they were not assessed in this study. ...
The present study was designed to assess vascular smooth muscle cell (VSMC) proliferation and apoptosis in primary cultured VSMCs prepared from the aortic tunica media of adult (4 to 5 months old) age- and gender-matched groups of stroke-prone spontaneously hypertensive rats (SHRSP) and the normotensive reference strain, Wistar-Kyoto (WKY) rats. In the present study, VSMC proliferation was assessed with measurement of DNA synthesis in response to stimulation of G(0)/G(1) arrested VSMCs with 10% serum, whereas apoptosis was measured in response to serum deprivation. Apoptosis in aortic VSMCs was assessed in vitro with the technique of Annexin V binding in combination with propidium iodide exclusion with bivariate flow cytometric analysis. The percentage of necrotic VSMCs in the cell populations was assessed simultaneously. The light-scattering properties of the cells were assessed to provide further information on cell shrinkage and chromatin condensation. Results of the present study have shown enhanced DNA synthesis in VSMCs from SHRSP (n=10; 5.2+/-0.9 cpmx10(3)/mg protein) compared with WKY (n=12; 2.4+/-0.7 cpmx10(3) /mg protein; P<0.05, 95% CI, -5271 to -296). In addition, the results of the present study have demonstrated the role of serum in the survival of VSMCs in vitro, because SHRSP VSMCs underwent significantly more apoptosis in response to insult by serum deprivation (n=13; 10.21+/-1.8%) than WKY VSMCs (n=7; 3.44+/-1.4%; P<0.01, 95% CI, -11.5 to -2.0). Thus, it appears that both proliferation and apoptosis are enhanced in synthetic phenotype aortic medial VSMCs from the SHRSP in vitro.
... In support of this speculation the present study clearly indicated that induction of adenomatous transformation was associated with significantly increased expression of the Ang II receptor protein (Fig. 5). Consistent with the present results we have previously demonstrated that AT 2 expression is inducible in vitro (Ichiki et al. 1996) and in vivo (Tamura et al. 2000). Although it is difficult to speculate a specific function of AT 2 in carcinogen-induced tumorigenesis, AT 2 likely play a role in lung carcinogenesis. ...
To gain insight into the mechanism by which angiotensin II type 2 receptor (AT(2)) regulates carcinogen-induced lung tumorigenesis, we have newly developed anti-AT(2) single chain variable fragment (ScFv) antibodies using a rodent phage-displayed recombinant antibody library with various peptide fragments of the receptor protein, and investigated the expression of the AT(2) receptor protein. The specificity of the antibodies was verified using AT(2) over-expressing COS-7 cells and AT(2) naturally expressing PC12W cells. In control wild type mouse lung, a stronger immunoreactivity was observed in bronchial epithelial cells. A moderate immunoreactivity was detected in pulmonary vascular walls and vascular endothelial cells. In the lungs possessing tobacco-specific nitrosamine (NNK)-induced tumors, significantly increased AT(2) and AT(1 )immunostaining was observed in adenomatous lesions. These data suggest that the increase in both receptors' expression in the alveolar epithelial cells may be accompanied with the onset of NNK-induced tumorigenesis and hence play important roles in lung tumorigenesis.
It has been well-accepted that an up-regulation of not only the systemic but also local cardiac renin angiotensin system (RAS) leads to end-stage damage in the heart under pathological conditions. So, a well-controlled RAS has an important impact on the prevention of morbidity and mortality of patients with cardiovascular disorders. Local cardiac RAS is composed of renin, angiotensin-converting enzyme (ACE), angiotensin I (Ang I), and it products Ang(1–9), angiotensin II (Ang II), and its product Ang(1–7), while the main effector peptide of the RAS is Ang II. Ang II plays prominent roles in cardiovascular pathology through its fundamental roles in the modulation of cellular signaling mechanisms, such as participation in immunity, lipid peroxidation, and insulin resistance. An overall insight to the literature data, activation of cardiac RAS may be pivotal in the pathogenesis of cardiac dysfunction in diabetes, metabolic syndrome, obesity, and/or other types of pathological conditions. Herein, the aim of this review article is particularly focused to discuss the evidence on the role of crosstalk between Ang II cell signaling and abnormal electrical activity and in the hyperglycemic mammalian heart. In the content of the article, it will be discussed characteristics and mechanisms of Ang II-related cardiac remodeling such as effects of Ang II receptor activation on electrical and mechanical activities of the heart under pathological stimuli, in both cellular and organelle level alterations. It will be also documented the literature data related to the roles of Ang II receptors on structural and electrophysiological remodeling in the hyperglycemic and insulin-resistant heart.KeywordsDiabetesInsulin resistanceMetabolic syndromeAgingHeart functionRenin angiotensin system
Angiotensin (Ang) II operates both as a systemic hormone modulating blood pressure and as a paracrine factor that is synthesized in response to stress. Regardless of its origin, the response of cardiovascular tissues to Ang II is mediated by specific cell surface receptors. In vascular tissues, two angiotensin receptors have been characterized according to their sensitivity to the specific antagonists losartan and PD123319 [1]. The AT1 receptor (losartan-sensitive) accounts for the majority of Ang II binding activity in adult tissue. It is significant, however, that upon injury and during fetal development, the proportion of AT2 receptors (PD123319-sensitive) increases [2]. Until recently the AT1 receptor has been regarded as the principal mediator of the responses evoked by Ang II. As a consequence the AT2 receptor has been studied less intensively and fewer details of its biological functions have been defined. The developmental regulation of AT2 receptor expression, however, suggests that it may be important in ontogeny. Furthermore, the generation of an AT2 receptor knockout mouse has revealed that this receptor, although its absence is not lethal, governs both the drinking response and motility [3]. In our laboratory, which has made use of porcine tissue for its studies, the application of antagonists specific to either the AT1 or the AT2 receptor has revealed that both receptors are independently required for VSMC growth. This review will, therefore, summarize recent advances in our understanding of Ang II-mediated SMC growth with respect to the role of individual angiotensin receptor subtypes and their associated signaling systems. Since both receptors have also been found essential for the vascular response to injury, an emphasis has been placed on defining the relationship between SMC growth and Ang II as it applies to the development of coronary artery disease and restenosis.
Previous studies have suggested that the expression of angiotensin type 2 receptor was inversely related to cell proliferation. We examined the effects of insulin-like growth factor (IGF-1), basic fibroblast growth factor (bFGF), transforming growth factor β1 (TGFβ1) and fetal calf serum (FCS) on cell proliferation and AT2 binding sites and mRNA level in PC12W (rat pheochromocytoma cell line) and R3T3 (mouse fibroblast cell line) which express abundant AT2 receptors. In both cell lines, serum deprivation markedly increased both AT2 receptor number and mRNA. However, in the absence of serum cell proliferation continued in PC12W and R3T3 at late passages (R3T3 LP) but not at early passages (R3T3 EP). In PC12W, none of the three growth factors studied stimulated cell proliferation, but TGFβ1 and more particularly bFGF markedly reduced AT2 expression. In R3T3 LP, IGF-1 and bFGF, but not TGFβ1, slightly stimulated cell proliferation, but the three factors, specially bFGF, reduced AT2 expression. In contrast, in R3T3 EP, the three growth factors significantly increased cell proliferation, but whereas TGFβ1 and bFGF markedly reduced AT2 binding sites and mRNA, IGF-1 caused the opposite effects. These results indicate that regulation of AT2 expression is not correlated with cell proliferation and appears to be more complex than initially suspected. In addition, they show that the same factor can have an opposite effect on AT2 expression in the same cell line depending upon the cell passage.
This phase II study investigated efficacy and safety of weekly alternating Bevacizumab (BEV)/Irinotecan (CPT-11) or Oxaliplatin (OHP) associated to weekly 5-Fluorouracil (5-FU) in first line treatment of metastatic colorectal carcinoma (MCRC).
Simon two-step design: delta 20% (p0 50%, p1 70%), power 80%, α 5%, β 20%. Projected objective responses (ORR): I step, 8/15 patients (pts); II step 26/43 pts. Schedule: weekly 12 h-timed-flat-infusion/5-FU 900 mg/m2, days 1-2, 8-9, 15-16, 22-23; CPT-11 160 mg/m2 plus BEV 5 mg/kg, days 1,15; OHP at three dose-levels, 60-70-80 mg/m2, days 8, 22; every 4 weeks.
Fifty consecutive, unselected pts < 75 years were enrolled: median age 63; young-elderly (yE) 24 (48%); liver metastases (LM) 33 pts, 66% Achieved OHP recommended dose, 80 mg/m2. ORR 82% intent-to-treat and 84% as-treated analysis. Median progression-free survival 12 months. Equivalent efficacy was obtained in yE pts. Liver metastasectomies were performed in 26% of all pts and in 39% of pts with LM. After a median follow-up of 21 months, median overall survival was 28 months. Cumulative G3-4 toxicities per patient: diarrhea 28%, mucositis 6%, neutropenia 10%, hypertension 2%. They were equivalent in yE pts. Limiting toxicity syndromes (LTS), consisting of the dose-limiting toxicity, associated or not to G2 or limiting toxicities: 44% overall, 46% in yE. Multiple versus single site LTS, respectively: overall, 24% versus 20%; yE pts, 37.5% versus 8%.
Poker combination shows high activity and efficacy in first line treatment of MCRC. It increases liver metastasectomies rate and can be safely administered.
Osservatorio Nazionale sulla Sperimentazione Clinica dei Medicinali (OsSC) Agenzia Italiana del Farmaco (AIFA) Numero EudraCT 2007-004946-34.
Angiotensin II acts on at least two receptor subtypes, AT1 and AT2. Although the physiological role of the AT2 receptor is still poorly defined, it may be implicated in inhibition of cell growth, vasorelaxation, and apoptosis. In the present study, to investigate the role of the AT2 receptor in the kidney and its implication in hypertensive states, we examined its expression using cultured mesangial cells (MC) from normotensive Wistar-Kyoto rats (WKY) and from stroke-prone spontaneously hypertensive rats (SHRSP). Receptor binding assays were performed using a nonselective ligand, [Sar1,Ile8]angiotensin II, or AT2-selective CGP42112A. Binding assays revealed that MC from WKY exhibited both AT1 and AT2 receptors, the ratio of which was confluence-dependent. In contrast, MC from SHRSP, whose proliferation activity was much higher than those from WKY, showed only the AT1 subtype. In receptor binding and Northern blot analyses, expression of the AT2 receptor of WKY-MC was low in the growing state but significantly induced upon confluence to become abundant in the post-confluent state, whereas that of SHRSP-MC was undetectable in either state. Gene expressions of AT1A and AT1B receptors were not significantly altered in either strain during the time in culture. These results indicate that the mesangial AT2-receptor expression is growth-dependent and suggest a role in the inhibition of MC growth in WKY. Much lower expression of the AT2 receptor in MC from SHRSP may suggest involvement in their higher proliferation activity and possibly in consequent renal disorders.
Since the discovery of nonpeptidic ligands, the receptors for angiotensin (Ang) II have been classified into 2 subtypes (Ang II type 1 receptor [AT1-R] and Ang II type 2 receptor [AT2-R]). AT1-R mediates most of the cardiovascular actions of Ang II. AT2-R is expressed at very high levels in the developing fetus. Its expression is very low in the cardiovascular system of the adult. The expression of AT2-R can be modulated by pathological states associated with tissue remodeling or inflammation. In failing hearts or neointima formation after vascular injury, AT2-R is reexpressed in cells proliferating in interstitial regions or neointima and exerts an inhibitory effect on Ang II-induced mitogen signals or synthesis of extracellular matrix proteins, resulting in attenuation of the tissue remodeling. An extreme form of cell growth inhibition ends in programmed cell death, and this process, which is initiated by the withdrawal of growth factors, is also enhanced by AT2-R. Cardiac myocyte- or vascular smooth muscle-specific mice that overexpress AT2-R display an inhibition of Ang II-induced chronotropic or pressor actions, suggesting the role of AT2-R on the activity of cardiac pacemaker cells and the maintenance of vascular resistance. AT2-R also activates the kinin/nitric oxide/cGMP system in the cardiovascular and renal systems, resulting in AT2-R-mediated cardioprotection, vasodilation, and pressure natriuresis. These effects, transmitted by AT2-R, are mainly exerted by stimulation of protein tyrosine or serine/threonine phosphatases in a Gi protein-dependent manner. The expression level of AT2-R is much higher in human hearts than in rodent hearts, and the AT2-R-mediated actions are likely enhanced, especially by clinical application of AT1-R antagonists. Thus, in this review, the regulation of AT2-R expression, its cellular localization, its pathological role in cardiovascular and kidney diseases, and pharmacotherapeutic effects of AT2-R stimulation are discussed.
The expression of angiotensin II type 2 (AT(2)) receptor is closely associated with cell growth, differentiation, and/or injury. We examined the effect of interferon (IFN)-gamma on AT(2) receptor expression in mouse fibroblast R3T3 cells and demonstrated that IFN-gamma treatment increased the expression of AT(2) receptor mRNA as well as its binding. Interferon regulatory factor (IRF)-1 was induced in mouse fibroblast R3T3 cells after IFN-gamma stimulation, and electrophoretic mobility shift assay showed an increase in IRF-1 binding with the IRF-specific binding sequence in the AT(2) receptor gene promoter region after IFN-gamma stimulation. The IRF-1 gene promoter contains an IFN-gamma-activated sequence (GAS) motif for possible binding of signal transducer(s) and activator(s) of transcription (STAT). Indeed, in R3T3 cells, IFN-gamma treatment resulted in rapid activation of Janus kinase (Jak) 1, Jak2, and STAT1 via tyrosine phosphorylation. Electrophoretic mobility shift assay with the GAS probe revealed increased STAT1 binding to the IRF-1 gene promoter in response to IFN-gamma stimulation. Transfection of GAS-binding oligonucleotides inhibited the effect of IFN-gamma on IRF-1 production, resulting in the AT(2) receptor trans-activation. Taken together, our data show that IFN-gamma upregulates AT(2) receptor expression in R3T3 cells via the activation of the intracellular Jak/STAT pathway and production of IRF-1.
Abstract—Angiotensin II exerts a potent role in the control of hemodynamic,and renal homeostasis. Angiotensin II is also a local and biologically active mediator involved in both endothelial and smooth,muscle,cell function acting on 2 receptor subtypes: type 1 (AT1R) and type 2 (AT2R). Whereas the key role of AT2R in the development,of the embryo has been extensively studied, the role of AT2R in the adult remains more questionable, especially in humans. In vitro studies in cultured cells and in isolated segments of aorta have shown,that AT 2R stimulation could lead to the production of vasoactive substances, among which NO is certainly the most cited, suggesting that acute AT2R stimulation will produce vasodilation. However, in different organs or in small arteries isolated from different type of tissues, other vasoactive substances may also mediate AT2R-dependent dilation. Sometimes, such as in large renal arteries, AT 2R stimulation may lead to vasoconstriction, although it is not always seen. In isolated arteries submitted to physiological
The aim of this study was to evaluate the effect of angiotensin II on models of acute inflammation. This study shows that angiotensin II potentiates the carrageenan- and dextran-induced paw edema. The administration of angiotensin II does not change the myeloperoxidase activity, neither the tissue content of interleukin-1 beta and tumor necrosis alpha nor the neutrophil migration to the peritoneal cavity, but induces significant enhancement of mast cell degranulation. The anti-histamine, mepyramine, and the anti-serotonin, metisergyde, reduce the angiotensin II-facilitated dextran-induced edema. Our results suggest that angiotensin II increases the vascular permeability through induction of mast cell degranulation and that this effect is mediated by the angiotensin AT2 receptor, since the angiotensin AT1 receptor antagonist and the angiotensin AT2 receptor agonist potentiated the paw edema.
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