Renal effects of AT1 angiotensin receptor antagonists (AT1ra)

RENIN-ANGIOTENSIN ANTAGONISTS: The renal effects of angiotensin II receptor antagonists (AT1 blockers) can be compared with another class of drugs inhibiting the renin-angiotensin-aldosterone system, i.e. the angiotensin I converting enzyme inhibitors (ACE1). SIMILAR BUT SPECIFIC EFFECTS: The renal effects of these two classes of drugs are similar but each class has specific effects explained by several mechanisms. i) The system includes a large number of active peptides (angiotensin II, angiotensin III, angiotensin 1-7) which exert various effects according to their specific receptor(s): ii) several types of angiotensin II receptors have been identified (AT1, AT2, AT4 ...). Only AT1 blockers are available in clinical practice. iii) Receptor or enzyme blockade can produce varying effects; ACE inhibition is not specific since increased bradykinin activity is associated with the suppression of angiotensin peptide generation. EXPERIMENTAL AND CLINICAL TRIALS: Experimental and recent clinical studies have shown that AT1 blockers can induce, like ACE1, hypotension, renal vasodilation and natriuresis. The definite effects on discrete renal structures (vessels, glomeruli, tubules) differ however in magnitude which may suggest specific indications according to the pathophysiological background (renal disease, congestive heart failure, etc.).     

Regulation of angiotensin II type 1 (AT1) receptor function

We studied protein binding and structural features of perfect and imperfect composite (gt)n(ga)m blocks from different HLA-DRB1 alleles in their original genomic and artificial environments. The major retarded protein/DNA complex of the genomic (gt)n(ga)m fragments comprises a zinc-dependent protein present in nuclear extracts from different cell types. The protein binding is characterized by moderate affinities independent of the polymorphic form of the physiological microsatellite allele. The binding affinity depends on the 5' and 3' adjacent single copy parts. DNase I footprinting of genome-derived fragments revealed that the 5' adjacent sequence and the (gt)n repeat are preferentially protected on the (gt)n(ga)m strand. Comparing three alleles, a regular pattern of footprints was not detectable in the (gt)n part, indicating that the zinc-dependent protein recognizes structural rather than sequence-specific features in this region. Chemical probing resulted in a pattern characteristic for Z-DNA in the (gt)n tract of the fragments. However, EMSA experiments using the Z-DNA specific monoclonal antibody mABZ-22 did not prove the presence of Z-DNA. As demonstrated by chemical modifications of the different (ga)m targets, only one of three (gt)n(ga)m fragments formed intramolecular triplexes of the type H-y3 and H-y5. DNase I footprinting revealed only weak protection, if any, in the homopurine tract. Rather, the (tc)m strands are hypersensitive for DNase I. This is probably due to structural conversions into intramolecular *H-triplexes after binding of HIZP.     

Candesartan (CV-11974) dissociates slowly from the angiotensin AT1 receptor

The mechanisms of the insurmountable antagonism of 2-ethoxy-1-[[2'-(1H-tetrazol-5-yl)biphenyl-4-yl]methyl]1H-benzimid azole -7-carboxylic acid, candesartan (CV-11974), an angiotensin AT1 receptor antagonist, on angiotensin II-induced rabbit aortic contraction were examined in contraction and binding studies. Preincubation of the rabbit aorta with CV-11974 (0.1 nM) for 30 min reduced the maximal contractile response to angiostensin II by approximately 50%. This insurmountable antagonism of CV-11974 was reversed in the presence of losartan (1 microM), a surmountable angiotensin AT1 receptor antagonist. The inhibitory effect of CV-11974 on angiotensin II-induced contraction persisted longer after washing than did that of losartan but was irreversible. Scatchard analysis of [3H]CV-11974 binding in bovine adrenal cortical membranes indicated the existence of a single class of binding sites (Kd = 7.4 nM). Competition binding studies using angiotensin II receptor agonists and antagonists have demonstrated that [3H[CV-11974 binding sites may be identical to angiotensin AT1 receptors. The dissociation rate of [3H]CV-11974 binding (t1/2 = 66 min) was 5 times slower than that of [125I]angiotensin II binding (t1/2 = 12 min). These results suggest that the insurmountable antagonism by CV-11974 is due to its slow dissociation from angiotensin AT1 receptors.     

Angiotensin II-induced nuclear targeting of the angiotensin type 1 (AT1) receptor in brain neurons

Angiotensin II (Ang II) interaction with the neuronal AT1 receptor results in a chronic stimulation of neuromodulation that involves the expression of norepinephrine transporter (NET) and tyrosine hydroxylase (TH). In view of this unique property and the presence of putative nuclear localization signal (NLS) consensus sequence in the AT1 receptor, this study was conducted to investigate the hypothesis that Ang II would induce nuclear sequestration of this G protein-coupled receptor and that the sequestration may have implications on Ang II-induced expression of NET and TH genes. Incubation of neuronal cultures with Ang II caused a time- and dose-dependent increase in the levels of AT1 receptor immunoreactivity in the nucleus. A 6.7-fold increase was observed with 100 nM Ang II, in 15 min, that was blocked by losartan, an AT1 receptor-specific antagonist. Ang II-induced nuclear sequestration was specific for AT1 receptor, because Ang II failed to produce a similar effect on neuronal AT2 receptors. The presence of the putative NLS sequence in the cytoplasmic tail of the AT1 receptor seems to be the key in nuclear targeting because: 1) nuclear targeting was attenuated by a peptide of the AT1 receptor that contained the putative NLS sequence; and 2) Ang II failed to cause nuclear translocation of the AT2 receptor, which does not contain the putative NLS. Ang II also caused a time- and dose-dependent stimulation of P62 phosphorylation, a glycoprotein of the nuclear pore complex. A 6-fold stimulation of phosphorylation was observed with 100 nM Ang II, in 15 min, that was completely blocked by losartan and not by PD123,319, an AT2 receptor specific antagonist. Preloading of neurons with p62-pep (a peptide containing consenses of mitogen-activated protein kinase in p62) resulted in a loss of Ang II-induced p62 phosphorylation and stimulation of NET and TH messenger RNA levels. In conclusion, these data demonstrate that Ang II induces nuclear sequestration of AT1 receptor involving NLS in the AT1 receptor and p62 of the nuclear pore complex in brain neurons. A possible role of such a nuclear targeting of the AT1 receptor on chronic neuromodulatory actions of Ang II has been discussed.     

Angiotensin II inhibits glomerular adenylate cyclase via the angiotensin II receptor subtype 1 (AT1)

We examined the role of angiotensin II (AII) receptor subtypes in the regulation of hormone-stimulated cyclic AMP (cAMP) accumulation in isolated rat glomeruli. All inhibited cAMP formation induced by histamine, serotonin and parathyroid hormone, but not by prostaglandin E2 or calcitonin gene-related peptide. Angiotensin III but not the angiotensin fragments (1-7) and (3-8) also showed inhibitory activity. The inhibition of histamine-induced cAMP accumulation by AII was concentration-dependent and was absent in glomeruli isolated from pertussis toxin-treated rats. The effect of AII on histamine-induced cAMP levels was not mimicked by the protein kinase C activator, phorbol-12-myristate-13-acetate, nor was the effect of AII inhibited by the protein kinase C inhibitors, staurosporine and H-7. The angiotensin II receptor subtype 1 (AT1) antagonists, SK&F 108566 and losartan, attenuated the inhibitory effect of AII on histamine-stimulated cAMP accumulation, whereas the AT2 selective antagonists, CGP 42112A, WL-19 and PD 123319, had no effect. Displacement of [125I]AII from glomerular membrane using the subtype-selective antagonists confirmed that the glomerular AII receptor has characteristics of an AT1 subtype. The results suggest that AII, through activation of the AT1 receptor, may act to maintain the contractile state of glomerular mesangial cells by attenuating the increase in cAMP levels induced by some hormones.     

Intracerebroventricular administration of angiotensin type 1 (AT1) receptor antisense oligonucleotides attenuate thirst in the rat

Chinese hamster ovary cells were pulse-labeled with 125I-iododeoxyuridine during early S phase, and cell samples were harvested 30 min or 5 h after labeling. The samples were frozen (with or without 25 mM cysteamine) and stored at -196 degrees C for accumulation of 125I decays. X-ray control experiments were performed at 37 degrees C and -196 degrees C. Aliquots of cells were plated for evaluating micronucleus formation and cell survival. The results demonstrated a striking shift in micronucleus formation and cell death with time after labeling. Cells frozen 30 min after labeling exhibited effects typical of low-LET radiation, but cells frozen 5 h after labeling showed a response characteristic of high-LET radiation. Cysteamine provided protection against the effects of 125I during the initial phase of effects characteristic of low-LET radiation, but no protection was seen during the phase characteristic of high-LET radiation. When cell survival was evaluated as a function of micronucleus frequency rather than dose in decays/cell, the survival curves for all treatment groups became superimposed. Previous work using the same experimental system had failed to show a direct link between 125I-induced DNA double-strand breaks and cell death. These findings are consistent with the hypothesis that DNA damage may not be the sole mechanism for cell killing and that damage to higher-order structures in the cell nucleus may contribute to (or modify) radiation-induced cell death.     

AT1 angiotensin receptor inhibition as a new therapeutic possibility

The octapeptide hormone, angiotensin II, binds to two major subtypes of cell surface receptors: the AT1 and the AT2 angiotensin receptors. The important physiological and pathophysiological effects of angiotensin II on cardiovascular regulation and salt-water balance are mediated by the AT1 receptor subtype. As a consequence of the outstanding clinical success of angiotensin-converting enzyme inhibitors, the appearance of AT1 receptor inhibitors in the therapy of hypertension and other cardiovascular diseases was preceded with great expectations. The available experimental and clinical data indicate that the first AT1 receptor inhibitor, losartan, has the same therapeutic potential as angiotensin-converting enzyme inhibitors, but it does not evoke the angiotensin-independent side-effects of ACE inhibitors, such as dry cough or angioedema. The physiological importance and the biochemical, molecular biological and pharmacological properties of AT1 and AT2 receptors are reviewed in this paper, and a summary of the available clinical data is presented.     

Regulation of the angiotensin AT1 receptor expression by hypercholesterolemia

The AT1 receptor mediates most of the biological effects of angiotensin II and has therefore been implicated in the pathogenesis of hypertension as well as arteriosclerosis. Hypercholesterolemia is a prominent risk factor for the development of these cardiovascular diseases. Since experimental results from hyperlipidemic animal models suggested an interaction of the renin-angiotensin system and hypercholesterolemia, the effects of low-density lipoprotein (LDL) and hypercholesterolemia has recently been investigated in vitro and in vivo. LDL causes in cultured vascular smooth muscle cells an up-regulation of AT1 receptor gene expression which is followed by an enhanced functional response upon stimulation with angiotensin II. This effect is also evident in vivo, as assessed in hypercholesterolemic rabbits. The vascular AT1 receptor expression is increased approximately twofold in hypercholesterolemic rabbits in comparison to normocholesterolemic animals. This leads ultimately to an enhanced angiotensin II-induced vasoconstriction. Thus, hypercholesterolemia and elevated concentrations of LDL enhance AT1 receptor expression in the vasculature causing an enhanced biological effectiveness of the renin-angiotensin system. This interaction may explain that blockade of the renin-angiotensin system attenuates the progress of arteriosclerosis in the hypercholesterolemic organismen and indicates a pathophysiologically important role for the lipid-induced AT1 receptor up-regulation.     

Differential roles of AT1 and AT2 receptor subtypes in vascular trophic and phenotypic changes in response to stimulation with angiotensin II

The essential reaction in the widely accepted proton-motive Q-cycle mechanism of the bc1 complex is the bifurcation of the electron flow during hydroquinone oxidation at the hydroquinone oxidation (Q(P)) site formed by the 'Rieske' iron sulfur protein and by the heme bL domain of cytochrome b. The 'Rieske' [2Fe-2S] cluster has a unique structure containing two exposed histidine ligands, which are the binding site for quinones. The affinity of the 'Rieske' cluster for quinones increases several orders of magnitude upon reduction; this will stabilize semiquinone at the Q(P) site. Based on this affinity change, a reaction scheme is presented which can explain the bifurcation of the electron flow without invoking highly unstable semiquinone species.     

Retrovirus-mediated transfer of an angiotensin type I receptor (AT1-R) antisense sequence decreases AT1-Rs and angiotensin II action in astroglial and neuronal cells in primary cultures from the brain

The AT1-R has been implicated in many cellular and physiological actions of angiotensin II (AII) in the brain. A retrovirus vector (LNSV) containing an AT1B-R antisense sequence (AT1B-AS) (termed LNSV-AT1B-AS) was constructed and used to determine the feasibility of using viral-mediated gene transfer to control AT1-Rs and AII actions in astroglial and neuronal cells in primary cultures from rat brain. Briefly, a 1.26-kb antisense sequence corresponding to nt -132 to +1128 of AT1-R cDNA was cloned into the LNSV vector, the vector was transfected into PA317 cells, and transfected cells were selected in G418. Incubation of brain cells with culture medium containing LNSV-AT1B-AS viral particles showed that AT1B-AS was integrated into the genome and transcribed in brain cells. This was associated with a significant decrease in AT1-Rs and in the AII-stimulated increase of c-fos mRNA, a measure of AT1-R function. These observations show that the AT1B-AS gene can be transferred into astroglial cells in culture by LNSV and that such a transfer inhibits AT1-Rs and the AII stimulation of cellular activities. In addition, the usefulness of this approach to study AII-dependent pathophysiology in primary neuronal cultures from brain, in particular, is established.     

In vivo pharmacology of an angiotensin AT1 receptor antagonist with balanced affinity for AT2 receptors

L-163,017 (6-[benzoylamino]-7-methyl-2-propyl-3-[[2'-(N-(3-methyl-1-butoxy) carbonylaminosulfonyl)[1,1']-biphenyl-4-yl]methyl]-3H-imidazo[4,5- b]pyridine) is a potent, orally active, nonpeptide angiotensin II receptor antagonist. Conscious rats and dogs were dosed p.o. and i.v.; in both species the plasma bioequivalents are similar at the angiotensin AT1 and AT2 receptor sites indicating balanced activity is maintained in vivo. L-163,017 prevents the pressor response to intravenous (i.v.) angiotensin II in the conscious rat, dog, and rhesus monkey. L-163,017 also significantly reduces blood pressure in a renin-dependent model of hypertension, similar to an angiotensin converting enzyme inhibitor (Enalapril) and an angiotensin AT1 receptor-selective antagonist (L-159,282). These studies indicate that neither the angiotensin AT2 receptor nor bradykinin is important in the acute antihypertensive activity of angiotensin converting enzyme inhibitors or angiotensin II receptor antagonists.     

The subtype-2 (AT2) angiotensin receptor regulates renal cyclic guanosine 3', 5'-monophosphate and AT1 receptor-mediated prostaglandin E2 production in conscious rats

The renal effects of angiotensin II(AII) are attributed to AT1 receptors. In contrast, the function of renal AT2 receptors in unknown. Using a microdialysis technique, we monitored changes in renal interstitial fluid (RIF) prostaglandin E2 (PGE2) and cyclic guanosine 3', 5'-monophosphate (cGMP) in response to dietary sodium (Na) depletion alone, or Na depletion or normal Na diet combined with the AT1 receptor blocker, Losartan, the AT2 receptor blocker, PD 123319 (PD), or angiotensin II, individually or combined in conscious rats. Na depletion significantly increased PGE2 and cGMP. During Na depletion, Losartan decreased PGE2 and did not change cGMP. In contrast, PD significantly increased PGE2 and decreased cGMP. Combined administration of Losartan and PD decreased PGE2 and cGMP. During normal Na diet, RIF PGE2 and cGMP increased in response to angiotensin II. Neither Losartan nor PD, individually or combined, changed RIF PGE2 or cGMP. Combined administration of angiotensin II and Losartan or PD produced a significant decrease in response of PGE2 and cGMP to angiotensin II, respectively. These data demonstrate that activation of the reninangiotensin system during Na depletion increases renal interstitial PGE2 and cGMP. The AT1 receptor mediates renal production of PGE2. The AT2 receptor mediates cGMP. AT2 blockade potentiates angiotensin-induced PGE2 production at the AT1 receptor.     

Growth response of human coronary smooth muscle cells to angiotensin II and influence of angiotensin AT1 receptor blockade

BACKGROUND: Adolescents and young adults are four times more likely to be victims of sexual assault than women in all other age groups. In the vast majority of these cases, the perpetrator is an acquaintance of the victim. Date rape is a subset of acquaintance rape where nonconsensual sex occurs between two people who are in a romantic relationship. METHODS: We conducted a MEDLINE and Current Concepts search for articles relating to date rape and then systematically reviewed all relevant articles. RESULTS: Lifetime prevalence of date or acquaintance rape ranges from 13% to 27% among college-age women and 20% to a high of 68% among adolescents. Demographic characteristics that increase vulnerability to date rape include younger age at first date, early sexual activity, earlier age of menarche, a past history of sexual abuse or prior sexual victimization, and being more accepting of rape myths and violence toward women. Other risk factors include date-specific behaviors such as who initiated, who paid expenses, who drove, date location and activity, as well as the use of alcohol or illicit drugs such as flunitrazepam (Rohypnol). Alcohol use that occurs within the context of the date can lead to: the misinterpretation of friendly cues as sexual invitations, diminished coping responses, and the female's inability to ward off a potential attack. CONCLUSIONS: Longitudinal research designs are needed to further our understanding of sexual violence among adolescents and young adults and the most effective ways to eliminate it. Understanding and comparing research findings would be easier if consensus regarding the definitions of date rape, sexual aggression, and sexual assault was obtained. Finally, primary and secondary date and acquaintance rape prevention programs must be developed and systematically evaluated.     

Cardiac-specific overexpression of angiotensin II AT2 receptor causes attenuated response to AT1 receptor-mediated pressor and chronotropic effects

BACKGROUND & AIMS: The gastroduodenal epithelium is protected from acid-peptic damage, in part, by its ability to secrete bicarbonate. Patients with duodenal ulcer disease have impaired proximal duodenal mucosal bicarbonate secretion. We have shown in vitro that histamine inhibits prostaglandin-stimulated bicarbonate secretion in rabbit duodenal mucosa via histamine H2 receptors and enteric nerves. In this study we examined whether the proulcerogenic compounds aspirin or ethanol regulate duodenal bicarbonate secretion and the involvement of histamine. METHODS: Bicarbonate secretion by rabbit proximal duodenal mucosa was examined in vitro in Ussing chambers. RESULTS: Aspirin and ethanol decreased basal and prostaglandin-stimulated bicarbonate secretion; the latter effect was specific for prostaglandin. The inhibitory effects of the two ulcerogenic compounds were at least additive. Ranitidine and tetrodotoxin abolished the inhibitory effects on stimulated, but not basal, secretion. Aspirin and ethanol also induced release of duodenal histamine. CONCLUSIONS: Aspirin and ethanol act by two distinct pathways to impair duodenal bicarbonate secretion. Both agents inhibit basal secretion via a histamine-independent and neurally independent pathway while they inhibit prostaglandin E2-stimulated secretion via histamine release, likely from mast cells, and actions on enteric nerves. Our findings may be of relevance to the understanding and potential treatment of nonsteroidal anti-inflammatory drug-associated mucosal injury.