A monoclonal antibody to mammalian angiotensin II AT1 receptor recognizes one of the angiotensin II receptor isoforms expressed by the eel (Anguilla anguilla)

1996 ◽  
Vol 16 (1) ◽  
pp. 45-56 ◽  
Author(s):  
S Marsigliante ◽  
A Muscella ◽  
S Vilella ◽  
G Nicolardi ◽  
L Ingrosso ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Angiotensin Ii ◽  
Ligand Binding ◽  
Molecular Mass ◽  
Chloride Cells ◽  
Ang Ii ◽  
Angiotensin Ii Receptor ◽  
Receptor Complexes ◽  
Intestinal Membrane ◽  
Liver Membranes

ABSTRACT Using labelled ligand-binding methods, previous studies have identified specific angiotensin II receptors (Ang II-Rs) in eel liver, kidney and intestine membranes. Isoelectric focusing on polyacrylamide gels also showed that there are two Ang II-R isoforms in eel liver, focusing at isoelectric points (pI) 6·5 and 6·7. These may have different functions. In contrast, eel enterocyte plasma membrane and renal brush border membranes contain only the pI 6·5 form. To characterize the eel receptors more fully, a newly developed monoclonal antibody (6313/G2) which selectively recognizes the AT1 subtype of mammalian Ang II-R was used. In ligand-binding experiments, the preincubation of eel liver membranes with 6313/G2 antibody eliminated the specific [3,5-3H]Tyr4-Ile5-Ang II binding. Moreover, Ang II—receptor complexes from solubilized liver membranes, which were immunoprecipitated by 6313/G2-coated beads, had a pI of 6·5. In immunoblotting experiments, the antibody recognized the isoform focusing at pI 6·5 in eel intestine and liver preparations, but not the liver pI 6·7 isoform. Immunoblotting of SDS gels showed that the antibody bound to a single protein of molecular mass of 75 kDa in eel liver, gill and kidney and to a doublet of molecular mass of about 74 and 75 kDa in intestinal membrane preparations. Immunocytochemistry of paraffin-embedded and cryostat sections of eel liver, kidney, intestine and gill showed that antibody 6313/G2 bound to uniformly distributed intracellular sites and cell surface membranes in proximal tubular cells, absorptive intestinal cells, hepatocytes and chloride cells. It also stained endothelium and both the longitudinal and circular layers of smooth muscle cells in the intestine. The data suggest that the previously described Ang II-R from eel liver, kidney and intestine may be similar to the mammalian AT1 subtype.

scholarly journals Design, synthesis and evaluation of novel angiotensin II receptor 1 antagonists with antihypertensive activities

RSC Advances ◽  
2017 ◽  
Vol 7 (42) ◽  
pp. 26401-26410 ◽  
Author(s):  
Xiao-Lu Bao ◽  
Wei-Bo Zhu ◽  
Tian-Li Shan ◽  
Zhuo Wu ◽  
Rui-Jing Zhang ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Antihypertensive Drug ◽  
Drug Candidate ◽  
Ang Ii ◽  
Angiotensin Ii Receptor ◽  
Design Synthesis ◽  
Long Acting

A novel Ang II receptor 1 antagonist 1f was found to be an efficient, long-acting and safe antihypertensive drug candidate.

scholarly journals Molecular targeting therapy with angiotensin II receptor blocker for prostatic cancer

2011 ◽  
pp. 3-13
Author(s):  
Hiroji Uemura ◽  
Hitoshi Ishiguro ◽  
Yoshinobu Kubota
Keyword(s):  
Prostate Cancer ◽  
Angiotensin Ii ◽  
Cancer Cells ◽  
Stromal Cells ◽  
Angiotensin Ii Receptor Blocker ◽  
Ang Ii ◽  
Angiotensin Ii Receptor ◽  
Targeting Therapy ◽  
Receptor Blockers ◽  
Prostatic Diseases

Angiotensin II (Ang-II) plays a key role as a vasoconstrictor in controlling blood pressure and electrolyte/fluid homeostasis. Recently it has also been shown that this peptide is a cytokine, acting as a growth factor in cardiovascular and stromal cells. In addition, the physiological function of Ang-II seems to be similar in prostate cancer and stromal cells. It is widely assumed that Ang-II facilitates the growth of both cells, and its receptor blockers (ARBs) have the potential to inhibit the growth of various cancer cells and tumors through the Ang-II receptor type 1 (AT1 receptor). The mechanism of cell growth inhibition by ARBs has been considered to be that of suppression of the signal transduction systems activated by growth factors or cytokines in prostate cancer cells, and suppression of angiogenesis. This review highlights the possible use of ARBs as novel agents for prostatic diseases including prostate cancer and benign hypertrophy, and covers related literature.

Embryonic lethality in Dear gene-deficient mice: new player in angiogenesis

2005 ◽  
Vol 23 (3) ◽  
pp. 257-268 ◽  
Author(s):  
Victoria L. M. Herrera ◽  
Lorenz R. B. Ponce ◽  
Pia D. Bagamasbad ◽  
Benjamin D. VanPelt ◽  
Tamara Didishvili ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Embryonic Lethality ◽  
Female Rats ◽  
Ang Ii ◽  
Sprague Dawley ◽  
Angiotensin Ii Receptor ◽  
Endothelin 1 ◽  
Receptor Isoforms ◽  
Age Range ◽  
Deficient Mice

The dual endothelin-1/angiotensin II receptor (Dear) binds endothelin-1 (ET-1) and angiotensin II (ANG II) with equal affinities in the Dahl S/JRHS rat strain. To elucidate its physiological significance within the context of multiple receptor isoforms and diverse ET-1 and ANG II functions spanning blood pressure regulation, tumor proliferation, and angiogenesis, we characterized mouse Dear and Dear-deficient mice. Unlike null mutant models of ET-1, ANG II, and all other ET-1 and ANG II receptors, Dear−/− deficiency results in impaired angiogenesis, dysregulated neuroepithelial development, and embryonic lethality by embryonic day 12.5. Interestingly, mouse Dear does not bind ANG II, similar to Dahl R/JRHS rat Dear, but binds ET-1 and vascular endothelial growth factor (VEGF) signal peptide (VEGFsp) with equal affinities, suggesting a putative novel multifunction for VEGFsp and a parsimonious mechanism for coordination of VEGF-induced and Dear-mediated pathways. Consistent with its developmental angiogenic role, Dear inhibition results in decreased tumor growth in B16-F10 melanoma cell-induced subcutaneous tumor in female Dear+/−/C57BL6BC10 mice, but not in males (age 3.5 mo), and in 127Cs radiation-induced orthotopic mammary tumors in Sprague-Dawley female rats (age range 3–6.5 mo). Altogether, the data identify Dear as a new player in angiogenesis during development downstream to, and nonredundant with, VEGF-mediated pathways, as well as a putative modulator of tumor angiogenesis acting within a gender-specific paradigm.

Internalisation of the type I angiotensin II receptor (ATI) and angiotensin II function in the rat adrenal zona glomerulosa cell.

1994 ◽  
Vol 141 (2) ◽  
pp. R5-R9 ◽  
Author(s):  
G. P. Vinson ◽  
M. M. Ho. ◽  
J.R. Puddefoot ◽  
R. Teja ◽  
S. Barker
Keyword(s):  
Monoclonal Antibody ◽  
Plasma Membrane ◽  
Angiotensin Ii ◽  
Zona Glomerulosa ◽  
Type I ◽  
Specific Monoclonal Antibody ◽  
Angiotensin Ii Receptor ◽  
Cellular Localisation ◽  
Glomerulosa Cells

ABSTRACT Little is known about the cellular localisation of the angiotensin II (AII) type 1 receptor (ATI) in the rat adrenal glomerulosa cell, but some studies have suggested that receptor internalisation and recycling may occur. Using a specific monoclonal antibody (6313/G2) to the first extracellular domain, we show here that most of the receptor is internalised in the unstimulated cell. When viable glomerulosa cells are incubated with 6313/G2, the receptor is transiently concentrated on the cell surface, and aldosterone output is stimulated. This stimulated output is enhanced by neither threshold nor maximal stimulatory concentrations of All amide, although the antibody does not inhibit All binding to the receptor. Conversely, the stimulatory actions of the antibody and those of ACTH are additive. The data suggest that recycling to the plasma membrane is constitutive, or regulated by unknown factors. Retention of the ATI receptor in the membrane is alone enough to allow sufficient G protein interaction to generate maximal stimulatory events.

scholarly journals Changes in angiotensin II type 1 receptor signalling pathways evoked by a monoclonal antibody raised to the N-terminus

2008 ◽  
Vol 197 (1) ◽  
pp. 25-33 ◽  
Author(s):  
Fang Xiao ◽  
John R Puddefoot ◽  
Stewart Barker ◽  
Gavin P Vinson
Keyword(s):  
Monoclonal Antibody ◽  
Angiotensin Ii ◽  
Primary Cultures ◽  
Activator Protein 1 ◽  
Ang Ii ◽  
Reporter System ◽  
Conserved Sequence ◽  
N Terminus ◽  
Protein Kinase Cα

The extracellular N-terminus of G-protein-coupled receptors may be involved in signalling events. We examined this in the angiotensin II type 1 receptor (AT1-R) using monoclonal antibody 6313/G2, raised against a conserved sequence in the N-terminal domain, and found it evokes inhibitory and stimulatory responses. In rat aortic smooth muscle cell (RASMC) primary cultures, 6313/G2 (2.5 μg/ml) inhibited both basal and angiotensin II (Ang II; 10−7 mol/l)-stimulated [H3]thymidine incorporation. Exposure to 6313/G2 gave sustained increases in phosphorylated protein kinase Cα (PKCα) but gave a decrease in phosphorylated p44/42 extracellular signal-regulated kinases (ERK1/2) sustained from 10 min to 48 h compared with untreated control RASMC. In contrast, Ang II had no effect on PKCα, and, though it is acutely stimulatory (up to 5 min), it had no sustained effect on ERK1/2 either. Using Fura-2 and microfluorimetry, 6313/G2 added alone induced a transient increase in intracellular calcium ([Ca2+]i), with a characteristic response curve different from that of Ang II itself. The antibody was without effect on an Ang II-stimulated activator protein-1 reporter system, though it reduced unstimulated reporter activity. Such discriminatory effects on intracellular signalling suggest that the AT1-R N-terminus itself might be a target for therapeutic intervention in chronic vascular disease.

scholarly journals Deglycosylation and fragmentation of purified rat liver angiotensin II receptor: application to the mapping of hormone-binding domains

1993 ◽  
Vol 289 (1) ◽  
pp. 289-297 ◽  
Author(s):  
F Desarnaud ◽  
J Marie ◽  
C Lombard ◽  
R Larguier ◽  
R Seyer ◽  
...  
Keyword(s):  
Angiotensin Ii ◽  
Rat Liver ◽  
Transmembrane Domain ◽  
Selective Adsorption ◽  
Structural Data ◽  
Hormone Binding ◽  
Angiotensin Ii Receptor ◽  
Receptor Complexes ◽  
Binding Domains ◽  
Treatment Conditions

We report new structural data about the rat liver angiotensin II receptor, which belongs to the AT1 subclass. This receptor has been purified at analytical or semi-preparative levels by a previously described strategy involving its photolabelling with a biotinylated azido probe and selective adsorption of the covalent probe-receptor complexes to immobilized streptavidin [Marie, Seyer, Lombard, Desarnaud, Aumelas, Jard and Bonnafous (1990) Biochemistry 29, 8943-8950]. Chemical or enzymic deglycosylation of the purified receptor has shown a shift in its molecular mass from 65 kDa to 40 kDa. Fragmentation of the purified receptor was carried out with V8 protease from Staphylococcus aureus, CNBr and trypsin. It was possible to find trypsin-treatment conditions which allowed production of a 6 kDa probe-fragment complex with a satisfactory yield. Attempts to localize this small fragment (5 kDa after subtraction of the probe contribution) in the recently published rat AT1 receptor sequence are reported. As expected, this fragment is not glycosylated; moreover, its further fragmentation by CNBr induces a very slight decrease in its size. These data support the hypothesis that a receptor sequence comprising the third transmembrane domain and adjacent portions of extra- and intracellular loops is involved in photolabelling by the C-terminal azidophenylalanine of the angiotensin-derived probe. These preliminary results are discussed in terms of future prospects for the characterization of hormone-binding domains of angiotensin II receptors.

Angiotensin II receptors in the gill of sea water- and freshwater-adapted eel

1997 ◽  
Vol 18 (1) ◽  
pp. 67-76 ◽  
Author(s):  
S Marsigliante ◽  
A Muscella ◽  
G P Vinson ◽  
C Storelli
Keyword(s):  
Image Analysis ◽  
Angiotensin Ii ◽  
Atpase Activity ◽  
Sea Water ◽  
Anguilla Anguilla ◽  
Chloride Cell ◽  
Chloride Cells ◽  
Angiotensin Ii Receptors ◽  
Receptor Subtypes ◽  
Ang Ii

ABSTRACT Immunocytochemistry of paraffin-embedded and cryostat sections of eel (Anguilla anguilla) gill showed that angiotensin II receptors (Ang II-R) were present in chloride cells, uniformly distributed in the cytoplasm and on surface membranes. Computerised image analysis of these preparations showed that gills from sea water (SW)-adapted animals had a significantly (3-fold) higher Ang II-R concentration compared with freshwater (FW)-adapted eel gills. Isoelectric focusing gel electrophoresis revealed two Ang II-R isoforms with pI 6·5 and 6·6 that were differentially modulated by environmental salinity: they were equally abundant in SW while in FW the pI 6·6/pI 6·5 ratio was 1·66. Using catalytic cytochemistry with image analysis, gill chloride cell membrane Na+/K+ATPase activity was shown to increase 4-fold in response to SW adaptation. Additionally, perfusion of gills for 30 min with 0·1, 10 or with 100 nM Ang II provoked a dose-dependent increment in Na+/K+ATPase activity in FW, and a biphasic response in SW gills in which activity was significantly increased at low Ang II concentrations but was reduced to basal values at 100 nM. The data suggest that adaptation to sea water significantly increases Ang II-R concentration in the chloride cell and, together with the effects of Ang II on Na+/K+ATPase activity, suggest a role for this hormone in gill NaCl retention. The different responses of Na+/K+ATPase to Ang II stimulation in FW and SW may be attributed to the presence of two receptor subtypes that are differently modulated by salinity and that have opposing effects on Na+/K+ATPase.

Angiotensin II-receptor stimulation of cytosolic calcium concentration in cultured renal resistance arterioles

1996 ◽  
Vol 271 (6) ◽  
pp. F1239-F1247 ◽  
Author(s):  
Z. Zhu ◽  
W. J. Arendshorst
Keyword(s):  
Smooth Muscle ◽  
Angiotensin Ii ◽  
Vascular Reactivity ◽  
Calcium Concentration ◽  
Morphological Properties ◽  
Free Calcium ◽  
Functional Coupling ◽  
Receptor Subtypes ◽  
Ang Ii ◽  
Angiotensin Ii Receptor

This study provides an initial characterization of basic morphological properties of cultures of vascular smooth muscle cells (VSMC) from rat preglomerular resistance vessels and of the functional coupling of angiotensin II (ANG II) receptors to cytosolic free calcium concentration ([Ca2+]i (fura 2 fluorescence photometry). Renal VSMC were isolated from interlobular arteries and afferent arterioles (< 50 microns) using an iron oxide sieving method and compared with rat aortic VSMC cultured under similar conditions. Quiescent monolayers maintained uniform morphology and [Ca2+]i signaling profile between passages 3 and 10. Arteriolar and aortic VSMC were spindle shaped and expressed smooth muscle-specific alpha-actin and myosin heavy chains SM-1 and SM-2. ANG II caused a rapid increase in [Ca2+]i, followed by a sustained plateau phase at 50-60% of the peak value. The initial maximum [Ca2+]i responses were dose dependent and of similar magnitude in renal arteriolar and aortic VSMC. ANG II (10(-7) M) increased [Ca2+]i from 50 to 240 nM in arteriolar and from 57 to 201 nM in aortic VSMC (P < 0.001 for both). Inhibition of ANG II effects on [Ca2+]i revealed significant signaling through distinct AT-receptor subtypes (losartan and PD-123319 sensitive) in renal arteriolar VSMC. In contrast, only losartan was effective in aortic VSMC. The AT2-receptor ligand CGP-42112 had no effect in either vessel type. Our results demonstrate that cultured arteriolar VSMC have anatomical similarities to aortic VSMC and functional differences in AT-receptor signaling in response to ANG II. This novel preparation should provide a useful approach with which to investigate cellular mechanisms concerning receptor coupling to signaling pathways involved in vascular reactivity of arteriolar VSMC in the microcirculation in general and the kidney in particular.

Characterization of angiotensin II receptor subtypes in human glomeruli and mesangial cells

1992 ◽  
Vol 262 (3) ◽  
pp. F432-F441 ◽  
Author(s):  
D. Chansel ◽  
S. Czekalski ◽  
P. Pham ◽  
R. Ardaillou
Keyword(s):  
Angiotensin Ii ◽  
Mesangial Cells ◽  
Receptor Subtypes ◽  
Ang Ii ◽  
Glomerular Mesangial Cells ◽  
Angiotensin Ii Receptor ◽  
Functional Responses ◽  
At1 Antagonists ◽  
Angiotensin Ii Receptor Subtypes ◽  
High Concentrations

This study was designed to identify the subtypes of angiotensin II (ANG II) receptors present on glomeruli and glomerular mesangial cells and establish their functional significance. Dup 753 and its metabolite EXP 3174, two nonpeptide ANG II-1 receptor (AT1) antagonists, displaced 125I-ANG II and its analogue 125I-[Sar1,Ala8]ANG II from their binding sites in rat and human glomeruli and cultured human mesangial cells, whereas CGP 42112 A and PD 123177, two ANG II-2 receptor (AT2) antagonists, exhibited little displacing activity. Dup 753 and EXP 3174 did not modify the dissociation constant (Kd) value but markedly decreased the number of sites of 125I-[Sar1,Ala8]ANG II binding. The addition of PD 123177 did not further inhibit binding when all AT1 sites were occupied by Dup 753. Binding was markedly reduced by dithiothreitol. EXP 3174 and Dup 753 inhibited the main biological functions of ANG II in mesangial cells including increases in intracellular calcium concentration, PGE2 production, and protein synthesis. PD 123177 was also active but at concentrations 1,000- to 10,000-fold greater than those of AT1 antagonists. These results indicate that 1) only AT1 receptors are present in glomeruli and glomerular mesangial cells; 2) these receptors mediate the functional responses to ANG II; 3) the nonpeptide AT1 antagonists behave as noncompetitive inhibitors; and 4) high concentrations of the nonpeptide AT2 antagonists can recognize AT1 sites.

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