AT1 Receptor mRNA Research Articles

Expression of angiotensin II type 1 and type 2 receptor mRNAs in the gastric mucosa of Helicobacter pylori-infected Mongolian gerbils

The renin-angiotensin system (RAS) plays an important role in normal homeostasis, carcinogenesis-related angiogenesis, and cell proliferation. Helicobacter pylori infection causes infiltration of inflammatory cells into the gastric mucosa and is considered the major cause of gastric cancer. Whether RAS plays a role in H. pylori infection-related gastric diseases remains unclear. We investigated the changes in gastric mucosal angiotensin II type 1 receptor (AT1R) and type 2 receptor (AT2R) mRNA levels throughout the time course of H. pylori infection in Mongolian gerbils. Mongolian gerbils were infected with wild-type H. pylori (for 12 months) or with its isogenic oipA mutant (for 3 months). Gastric mucosal AT1R and AT2R mRNA levels were assessed using real-time reverse transcription-polymerase chain reaction. The gastric mucosal AT1R mRNA level was significantly associated with the severity of inflammatory cell infiltration into the gastric mucosa that reached maximal levels at 12 months after infection in both the antrum and body. Inflammatory cell infiltration scores and AT1R and AT2R mRNA levels were significantly lower in oipA mutant than wild-type infections. Mucosal AT1R and AT2R mRNA expressions in wild-type H. pylori-infected gerbils with gastric ulcers were significantly higher than in those without ulcers (P < 0.01). Gastric mucosal ATR expression gradually increases during the course of H. pylori infection. Up-regulation of the RAS in association with progressive gastric inflammation suggests a potential role of the RAS in gastric carcinogenesis. OipA appears to play a role in AT1R and AT2R expression and the resulting inflammation.

Enhanced Angiotensin II type 1 receptor expression in leukocytes of patients with chronic kidney disease

Previously, we demonstrated increased Angiotensin II type I receptor expression in leukocytes from patients with untreated, but not in treated, essential hypertension (essential hypertension). We hypothesized that the Angiotensin II AT1 receptor is also increased in leukocytes from patients with chronic kidney disease, however and can still be corrected with combined anti-hypertensive treatment with renin–angiotensin system (RAS) blockers and statins. Blood pressure, cholesterol, renal function oxidative stress parameters, inflammation, and leukocyte Angiotensin II AT1 receptor mRNA expression were measured both on and (6 weeks) off treatment. Data were compared to data of 10 healthy control subjects. Untreated chronic kidney disease patients (n = 20) had higher blood pressure, cholesterol and leukocyte Angiotensin II AT1 receptor mRNA expression, but no different ox-LDL, thiobarbituric acid reactive substances, paraoxonase activity or hs-CRP. OxLDL and Lipoprotein(a) were increased in untreated chronic kidney disease. Angiotensin II AT1 receptor expression inversely correlated with renal function (R 2 = 0.15, P < 0.03) and Lipoprotein(a) but not with the other parameters. Treatment with RAS blockers and statins normalized blood pressure and cholesterol, however it did not correct enhanced leukocyte Angiotensin II AT1 receptor expression. Leukocyte Angiotensin II AT1 receptor expression is inappropriately high in chronic kidney disease, correlates inversely with renal function and does not depend on antihypertensive and lipid-lowering treatment. The uremic environment seems to dominate over previously reported actions of high blood pressure and cholesterol to enhance leukocyte Angiotensin II AT1 receptor expression.

Long-term systemic angiotensin II type 1 receptor blockade regulates mRNA expression of dorsomedial medulla renin-angiotensin system components

In Fischer 344 (F344) rats, renin-angiotensin system (RAS) blockade for 1 yr with the angiotensin II type 1 (AT(1)) receptor blocker L-158,809 prevents age-related impairments in metabolic function, similar to transgenic rats with low glial angiotensinogen (Aogen). Brain RAS regulation may contribute to the benefits of long-term systemic AT(1) antagonism. We assessed the mRNA of RAS components in the dorsomedial medulla of F344 rats at 3 (young; n = 8) or 15 mo of age (old; n = 7) and in rats treated from 3 to 15 mo of age with 20 mg/l of the AT(1) receptor antagonist L-158,809 (Old+L; n = 6). Aogen and renin mRNA were lower in the young compared with old group. Angiotensin-converting enzyme (ACE) mRNA was lower in the old and Old+L compared with the young group. ACE2 and neprilysin expression were significantly higher in Old+L compared with young or old rats. AT(1b), AT(2), and Mas receptor mRNA were higher with treatment. Leptin receptor mRNA was lower in the old rats and this was prevented by L-158,809 treatment. Dual-specificity phosphatase 1 (DUSP1) mRNA was highest in the Old+L group. Aggregate correlate summation revealed a positive relationship for Mas receptor mRNA with food intake. The findings provide evidence for regulation of dorsomedial medullary renin and Aogen mRNA during aging. Long-term AT(1) receptor blockade increases the mRNA of the enzymes ACE2 and neprilysin and the MAS receptor, which could potentially shift the balance from ANG II to ANG-(1-7) and prevent age-related declines in the leptin receptor and its signaling pathway.

Heart Rate Contributes to the Vascular Effects of Chronic Mental Stress

Vascular effects of mental stress are only partially understood. Therefore, we studied effects of chronic stress and heart rate (HR) on endothelial function and cerebral ischemia. 129S6/SvEv mice were randomized to the I(f)-channel inhibitor ivabradine (10 mg/kg per day) or vehicle and underwent a chronic stress protocol for 28 days. Stress increased HR from 514 ± 10 bpm to 570 ± 14 bpm, this was prevented by ivabradine (485 ± 7 bpm). Endothelium-dependent relaxation of aortic rings was impaired in mice exposed to stress. HR reduction restored endothelial function to the level of naive controls. Vascular lipid hydroperoxides were increased to 333% ± 24% and vascular NADPH oxidase activity was upregulated to 223 ± 38% in stressed mice, which was prevented by ivabradine. Stress reduced aortic endothelial nitric oxide synthase mRNA expression to 84% ± 3% and increased AT1 receptor mRNA to 168% ± 18%. Both effects were attenuated by HR reduction. In brain tissue, stress resulted in an upregulation of lipid hydroperoxides to 140% ± 11%, which was attenuated by HR reduction. Ivabradine increased brain capillary density in naive and in stressed mice. Mice exposed to chronic stress before induction of ischemic stroke by transient middle cerebral artery occlusion exhibited increased lesion size (33.7 ± 2.3 mm3 versus 23.9 ± 2.4 mm3). HR reduction led to a marked reduction of the infarct volume to 12.9 ± 3.3 mm3. Chronic stress impairs endothelial function and aggravates ischemic brain injury. HR reduction protects from cerebral ischemia via improvement of endothelial function and reduction of oxidative stress. These results identify heart rate as a mediator of vascular effects induced by chronic stress.

Human GRK4 variants regulate renal angiotensin AT1 receptor expression

Variants of the GRK4γ gene are associated with essential hypertension. Constitutively active GRK4 variants are linked with decreased function of the renal dopamine D1 receptor and increased expression of the angiotensin II type 1 receptor (AT1R). Renal AT1R mRNA and protein expressions are greater in transgenic mice expressing human (h) GRK4γ 142V variant than hGRK4γ wild-type (WT) (240±50 vs 100±15%, qRT-PCR, n=5, P<0.05, 150±8 vs 100±5 8%, western blot, n=5, P<0.05). Luciferase assays performed in human renal proximal tubule cells (hRPTC) co-transfected with the hAT1R promoter and hGRK4γ WT or hGRK4γ 142V showed that hGRK4γ WT decreases hAT1R promoter activity (60%, n=6, P<0.01) while hGRK4γ 142V increases it (400%, n=6, P<0.01). We hypothesized that GRK4 gene variants act in the nucleus to phosphorylate histone deacetylases (HDAC) and increase AT1R transcription. We found nuclear localization of GRK4 (laser confocal microscopy) and the presence of GRK4 and class I HDACs, HDAC1, HDAC2 and HDAC3, in purified nuclear extracts from hRPTCs. Nuclear GRK4 expression was greater in hRPTCs with GRK4 gene variants than those with WT GRK4. Co-immunoprecipitation experiments demonstrated physical interaction between GRK4 and HDAC1 and between GRK4 and HDAC2. Our results suggest that GRK4 may regulate AT1R expression by regulating HDAC activity.

Chronic angiotensin II infusion modulates angiotensin II type I receptor expression in the subfornical organ and the rostral ventrolateral medulla in hypertensive rats

Blood-borne angiotensin II (Ang II) has profound effects on the central nervous system, including regulation of vasopressin secretion and modulation of sympathetic outflow. However, the mechanism by which circulating Ang II affects the central nervous system remains largely unknown. We tested the hypothesis that increased circulating levels of Ang II activate angiotensin type I (AT1) receptors in the subfornical organ (SFO), increasing the Ang II signalling in the rostral ventrolateral medulla (RVLM). Male Wistar rats were subcutaneously implanted with two 14-day osmotic minipumps filled with Ang II (150 ng/kg/minute), Losartan (10mg/kg/day), or saline. In addition, AT1 receptor mRNA levels in the SFO and RVLM were detected by reverse transcription polymerase chain reaction (RT-PCR). Infusion of Ang II-induced hypertension (134 ± 10 mmHg vs 98 ± 9 mmHg, n = 9, p < 0.05), which was blunted by concomitant infusion of Losartan (105 ± 8 vs 134 ± 10 mmHg, n = 9, p < 0.05). In addition, hexamethonium produced a greater decrease in blood pressure in Ang II-infused rats. Real time PCR revealed that chronic Ang II infusion induced an increase in AT1 receptor mRNA levels in the RVLM and a decrease in the SFO. Taken together, using combined in vivo and molecular biology approaches, our data suggest that Ang II-induced hypertension is mediated by an increase in sympathetic nerve activity, which seems to involve up-regulation of AT1 receptors in the RVLM and down-regulation of AT1 receptors in the SFO.

Vascular, renal and placental effects on pregnant offspring of protein‐restricted rat dams

Our previous study showed that a maternal low-protein diet induced hypertension and vascular dysfunction in rat offspring after day 175. In the present study, we hypothesized that these female offspring would develop hypertension in their own pregnancies even at ages less than 175 days because potential vascular dysfunction is exacerbated by the circulatory demands of pregnancy. Wistar rats were fed an isocaloric diet containing either 18% (control group) or 9% (low-protein group) casein throughout pregnancy. The female offspring were fed standard chow and mated between days 70 and 125. At the end of pregnancy, blood pressure was measured, and the uterine arteries were dissected and investigated with a wire myograph. Placental weights were significantly lower in offspring of the low-protein group versus control. There were no significant differences in blood pressure. Renal expression of AT1 receptor mRNA was greater, and of AT2 receptor was less, in the low-protein group versus control. Vasoconstriction of uterine arteries to phenylephrine and U46619 was increased in the low-protein group, and vasodilatation to sodium nitroprusside was also increased. Low-protein diet induces vascular effects on female offspring in their pregnancy, in terms of increased uterine artery vasoconstriction. This may be compensated for by increased sensitivity to nitric oxide (NO), maintaining blood pressure normal in the face of the demands of pregnancy. Such renal and vascular effects, combined with placental size, may transmit risk of vascular dysfunction to subsequent generations.

(Pro)renin receptors and angiotensin converting enzyme 2/angiotensin-(1-7)/Mas receptor axis in human aortic valve stenosis

BackgroundThere is increasing evidence that renin–angiotensin system (RAS) may play a major role in the actively regulated fibrocalcific process in aortic valve stenosis (AS), but the gene expression or function of (pro)renin receptor ((P)RR), prorenin and renin or angiotensin converting enzyme 2(ACE2)/angiotensin-(1-7)/Mas receptor axis in calcific aortic valve disease is not known. Methods and resultsWe characterized expression of (P)RR, ACE2 and Mas receptor as well as renin, prorenin and angiotensin II type 2 (AT2) receptors in human aortic valves, and compared normal control valves (n=11) with valves obtained from patients with aortic regurgitation (AR, n=14), AR with fibrosis (n=20) and AS (n=61). By immunohistochemistry (P)RR positive staining was seen in the valvular endothelial cells of control and in the neovessels of stenotic valves. By RT-PCR, renin mRNA levels were 72% (P=0.001) and prorenin mRNA levels 64% lower (P=0.002) in stenotic aortic valves compared to control valves. ACE2, Mas receptor and AT2-receptor mRNA levels were 69% (P<0.001), 58% (P=0.008) and 75% (P=0.001) lower, respectively, in stenotic valves. ACE2 positive staining, existing to lesser extent in stenotic aortic valves, was localized mainly to stromal area in spongiosa layer in control valves. Conclusions(P)RR, prorenin and renin are expressed in human aortic valves. We also report for the first time expression of ACE2/angiotensin-(1-7)/-Mas receptor axis in human aortic valve cusps. The downregulation of ACE2/angiotensin-(1-7)/-Mas receptor axis as well as AT2-receptors may promote fibrosis, proliferation and inflammation in patients with AS.

Effects of cytokines on expression of angiotensin II type 1 receptors in vascular smooth muscle cells in rats

Objective To investigate the effects of cytokines on the expression of angiotensin Ⅱ type 1 receptor (AT1R) in vascular smooth muscle cells (VSMCs) in rats. Methods Primary cultured VSMCs from SD rat thoracic aorta were cultured in serum-free DMEM for 24 h, and then in DMEM supplemented with 10% fetal bovine serum for another 12 h. The cultured VSMCs were randomly divided into 5 groups (n =6 each): control group (group C); 10% cytokine group (group L); 50% cytokine group (group N); 100% cytokine group (group H) and L-arginine methy ester (L-NAME), an inhibitor of nitric oxide synthase) group. In group C, the cellswere cultured continuously for 12 h. In L, N and H groups, 10%, 50% and 100% cytokines (IL-1β 50 ng/ml +TNF-α 100 ng/ml + IFN-γ 500 ng/ml) were added to the culture medium respectively and the cells were then incubated for 12 h. In group L-NAME, 100% cytokines + L-NAME 5 mmol/L were added to the culture medium and the cells were then incubated for 12 h. The expression of AT1R mPNA and protin was determined by RT-PCR and Western blot respectively.Results Cytokines down-regulated AT1R mRNA and protein expression in a concentration-dependent manner (P ＜ 0.05 or 0.01). L-NAME reversed cytokines-induced changes in AT1R mRNA and protein expression ( P ＜ 0.01). Conclusion Cytokines can down-regulate the expression of AT1R in rat VSMCs and the mechanism is related to the NO synthesis. Key words: Cytokines; Receptor, angiotensin, type 1; Muscle, smooth, vascular

Antenatal maternal hypoxic stress: Adaptations of the placental renin-angiotensin system in the mouse

AbstractThe stress of Antenatal Maternal Hypoxia (AMH) can lead to a number of physiological and pathological changes in both mother and fetus, changes which can be linked to alterations in placental morphology and gene regulation. Recently, in the Brown Norway rat “model” of placental insufficiency, we reported alterations in placental renin-angiotensin system (RAS) genes. Moreover, AMH can lead to reduced oxygen availability to the fetus, similar to a state of placental insufficiency. Thus, in pregnant mice dams we tested the hypothesis that antenatal maternal hypoxic stress leads to alterations in the placental RAS. These alterations may, in part, account for the phenotypic changes in both pregnant mice dams as well as fetus and adult offspring. MethodsPregnant FVB/NJ mice dams were either maintained as controls, or exposed to 10.5% O2 for 48 h from 15.5 to 17.5 day post coitum. We then measured placental mRNA and protein expression of several RAS genes (n = 4 to 5; P < 0.05 was considered significant). ResultIn murine placenta: (1) angiotensinogen (AGT) mRNA was undetectable; however, AGT protein was detectable and increased significantly with AMH. (2) In AMH, although renin mRNA was reduced protein expression increased, in association with decreased microRNA (miRNA) 199b, which can lead to increased renin translation. (3) Also in AMH placenta, angiotensin converting enzyme (ACE) -1 mRNA was unaltered; however, protein expression increased significantly, in association with decreased miRNA 27a, which can result in increased ACE-1 translation. (4) In AMH placenta, ACE-2 mRNA was reduced significantly, whereas protein expression was significantly greater, in association with reduced miRNA 429. (5) In AMH placenta, angiotensin II type (AT) -1a receptor mRNA expression was unaltered while AT-1b receptor mRNA was undetectable in both groups. Moreover, AT-1 receptor protein expression was unchanged in response to AMH. (6) AT-2 receptor mRNA and proteins were undetectable in both groups. ConclusionThe normal murine placenta possesses several components of RAS, and in response to AMH several of these elements undergo important changes. In addition, differential expression of RAS mRNA, miRNA and protein, indicate post-transcriptional regulatory mechanisms involved with hypoxic stress, and necessitate further investigation.

Endogenous angiotensin II suppresses stretch-induced ANP secretion via AT1 receptor pathway

Angiotensin II (Ang II) is released by stretch of cardiac myocytes and has paracrine and autocrine effects on cardiac myocytes and fibroblasts. However, the direct effect of Ang II on the secretion of atrial natriuretic peptide (ANP) is unclear. The aim of the present study is to test whether Ang II affects stretch-induced ANP secretion. The isolated perfused beating atria were used from control and two-kidney one-clip hypertensive (2K1C) rats. The volume load was achieved by elevating the height of outflow catheter connected with isolated atria from 5 cmH 2O to 7.5 cmH 2O. Atrial stretch by volume load caused increases in atrial contractility by 60% and in ANP secretion by 100%. Ang II suppressed stretch-induced ANP secretion and tended to increase atrial contractility whereas losartan stimulated stretch-induced ANP secretion. Neither PD123319 nor A779 had direct effect on stretch-induced ANP secretion. The suppressive effect of Ang II on stretch-induced ANP secretion was blocked by the pretreatment of losartan but not by the pretreatment of PD123319 or A779. In hypertrophied atria from 2K1C rats, stretch-induced ANP concentration attenuated and atrial contractility augmented. The response of stretch-induced ANP secretion to Ang II and losartan augmented. The expression of AT1 receptor protein and mRNA increased but AT2 and Mas receptor mRNA did not change in 2K1C rat atria. Therefore, we suggest that Ang II generated endogenously by atrial stretch suppresses stretch-induced ANP secretion through the AT1 receptor and alteration of Ang II effect in 2K1C rat may be due to upregulation of AT1 receptor.

Interplay Between miR-155, AT1R A1166C Polymorphism, and AT1R Expression in Young Untreated Hypertensives

A silent polymorphism (+1166 A/C single-nucleotide polymorphism) localized in the 3'-UTR (untranslated region) of the human angiotensin II type-1 receptor (AT1R) has been associated with hypertension and cardiovascular complications. The +1166 A/C is recognized by a specific microRNA-155 (miR-155), which is base-pairing complementary with the +1166 A-allele but not with the mutant +1166 C allele. Aim of our study was to investigate the interplay between miR-155 and AT1R protein expression. Sixty-four subjects were selected for the +1166 A/C from the cohort of hypertensives (n = 573) of the Hypertension and Ambulatory Recording Venetia Study (HARVEST): 25 were homozygous for the 1166 A allele, 20 heterozygous, and 19 homozygous for the 1166 C allele. miR-155 expression was significantly decreased in subjects with CC genotype in comparison to AA and AC genotype. AT1R protein expression was significantly increased in the CC group in comparison to AA and AC (P < 0.01) although AT1R mRNA expression was not significantly different in the three groups. AT1R protein expression was positively correlated with systolic and diastolic blood pressure and negatively correlated with miR-155 expression level. Plasma transforming growth factor-β1 (TGF-β1) may have a modulator role in the interplay between miR-155 and AT1R protein expression as it was correlated negatively with miR-155 expression and positively with AT1R protein expression in subjects with CC genotype. The interplay between miR-155 expression, +1166C polymorphism, and AT1R protein expression may have a role in the regulation of blood pressure.

E0348 The Effects of valsartan on angiotensin ii type 1 and type 2 receptor in isolated reperfused ischaemic rat hearts

ObjectTo determine the effects of Angiotensin II Type 1 receptor blockade valsartan on AT1 and AT2 receptor during ischaemia reperfusion.MethodsThe hearts of 24 SD rats were isolated, linked to Langedorff...

T6.4 Plasma 25-hydroxyvitamin D in early-onset severe preeclampsia

The stress of Antenatal Maternal Hypoxia (AMH) can lead to a number of physiological and pathological changes in both mother and fetus, changes which can be linked to alterations in placental morphology and gene regulation. Recently, in the Brown Norway rat “model” of placental insufficiency, we reported alterations in placental renin-angiotensin system (RAS) genes. Moreover, AMH can lead to reduced oxygen availability to the fetus, similar to a state of placental insufficiency. Thus, in pregnant mice dams we tested the hypothesis that antenatal maternal hypoxic stress leads to alterations in the placental RAS. These alterations may, in part, account for the phenotypic changes in both pregnant mice dams as well as fetus and adult offspring.Pregnant FVB/NJ mice dams were either maintained as controls, or exposed to 10.5% O2 for 48 h from 15.5 to 17.5 day post coitum. We then measured placental mRNA and protein expression of several RAS genes (n = 4 to 5; P < 0.05 was considered significant).In murine placenta: (1) angiotensinogen (AGT) mRNA was undetectable; however, AGT protein was detectable and increased significantly with AMH. (2) In AMH, although renin mRNA was reduced protein expression increased, in association with decreased microRNA (miRNA) 199b, which can lead to increased renin translation. (3) Also in AMH placenta, angiotensin converting enzyme (ACE) -1 mRNA was unaltered; however, protein expression increased significantly, in association with decreased miRNA 27a, which can result in increased ACE-1 translation. (4) In AMH placenta, ACE-2 mRNA was reduced significantly, whereas protein expression was significantly greater, in association with reduced miRNA 429. (5) In AMH placenta, angiotensin II type (AT) -1a receptor mRNA expression was unaltered while AT-1b receptor mRNA was undetectable in both groups. Moreover, AT-1 receptor protein expression was unchanged in response to AMH. (6) AT-2 receptor mRNA and proteins were undetectable in both groups.The normal murine placenta possesses several components of RAS, and in response to AMH several of these elements undergo important changes. In addition, differential expression of RAS mRNA, miRNA and protein, indicate post-transcriptional regulatory mechanisms involved with hypoxic stress, and necessitate further investigation.

E0053 Electroporation-mediated angiotensin II type 2 receptor gene transfected into rat carotid arteries and the effects of AT2R gene transfer on neointimal hyperplasia in rat carotid arteries after balloon angioplasty

AimTo study the effects of Electroporation on the angiotensin II type 2 receptor (AT2R) transfected into rat carotid arteries and study the effects of AT2R gene transfer on neointimal hyperplasia...

Central Pressor Response by Olmesartan in the Rostral Ventrolateral Medulla of Spontaneously Hypertensive Rats

Objectives: The major aims of the present study were to evaluate the effect of chronic oral treatment with olmesartan on cardiovascular response to exogenous angiotensin II (Ang II), and its relationship with the AT1 receptor (AT1R) mRNA in rostral ventrolateral medulla (RVLM) of spontaneously hypertensive rats (SHR). Methods: SHR (12 weeks old) were treated with olmesartan, 30 mg/(kg . day), in drinking water. 4 weeks later, Ang II (100 pmol) was microinjected into the RVLM, and arterial pressure and the heart rate were observed. Results: The blood pressure of all SHR-olme rats reach near normal level during breeding period. Ang IIresulted in an increase in mean arterial pressure (MAP) of each group. While, the increase of MAP in olmesartan-treated SHR was significantly smaller than that in untreated SHR(26.3 ± 0.75 mmHg vs. 47.2 ± 1.41 mmHg, n = 10, P < 0.01 ); however, it was still greater than that in WKY rats(26.3 ± 0.75 mmHg vs. 21.5 ± 0.72 mmHg, n = 10, P < 0.05). The heart rate (HR) of the each group was also increased, but there was no statistical significance among groups. In addition, AT1R mRNA in RVLM was measured by RT-PCR and analyzed with optic density. It demonstrated that the mean optic density (MOD) of AT1R mRNA in olmesartan-treated SHR was significantly reduced compared with untreated SHR(0.94 ± 0.41 vs. 2.41 ± 0.37, n = 10, P < 0.01); however, it was still higher than that in WKY rats(0.94 ± 0.41 vs. 0.81 ± 0.22, n = 10, P < 0.05). Conclusions: These results demonstrated that chronic oral treatment with olmesartan could obviously attenuate the exaggerated pressor response to exogenous Ang II and inhibit the excessive expression of AT1R in RVLM of SHR, so we presume olmesartan may attenuate central pressor response by reducing AT1R in RVLM of SHR.

High-salt diet during pregnancy and angiotensin-related cardiac changes

High-salt intake has been demonstrated in link to hypertension, and cardiovascular diseases could be programmed in fetal origins. We determined the influence of high-salt diet during pregnancy on the development of the heart. Fetal cardiac structures, cell cycle, renin-angiotensin system (RAS), and epigenetic alternations in the heart following maternal high salt intake during pregnancy were examined. Following exposure to high salt, disorganized myofibrillae and mitochondria cristae loss were found in the fetus, S-phase for cardiac cells was enhanced, plasma angiotensin II decreased, and cardiac angiotensin II increased in the fetus. Angiotensin II-increased S-phase in the fetal cardiac cells was primarily via AT1 receptor mechanisms. AT2 receptor mRNA and protein in the fetal heart were not affected, whereas AT1 receptor protein, AT1a, and AT1b mRNA were increased. DNA methylation was found at the CpG sites that were related to AT1b receptors in the fetal heart. Cardiac AT1 receptor protein in the adult offspring was also higher following exposure to prenatal high salt. The results suggest a relationship between high-salt diet in pregnancy and developmental changes of the cardiac cells and renin-angiotensin system.

Histone Deacetylases Are Critical Regulators of the Renin-Angiotensin System During Ureteric Bud Branching Morphogenesis

Mutations in the genes encoding components of the renin-angiotensin system (RAS) in mice or humans cause congenital abnormalities of the kidney and urinary tract. We hypothesized that absence of angiotensin (Ang) II in angiotensinogen (AGT)-deficient mice leads to defects in ureteric bud (UB) branching and that RAS genes are critically dependent on histone deacetylase (HDAC) activity. The number of UB tips was lower in AGT-/- compared with AGT+/+ embryonic (E) day E13.5 metanephroi (24+/-1.5 versus 36+/-3.7, p<0.05). Real-time RT-PCR demonstrated that pharmacological inhibition of HDAC activity with Scriptaid increases AGT, renin, angiotensin-converting enzyme (ACE), and AT1 receptor (AT1R) mRNA levels in E12.5 mouse metanephroi and early mesenchymal (MK3) cells. Furthermore, Scriptaid enhanced Ang II induced decrease in Sprouty (Spry) 1 gene expression in cultured UB cells. Treatment of intact E12.5 mouse metanephroi grown ex vivo with Ang II (10(-5) M, 24 h) increased HDAC-1 and decreased total acetylated histone H3 protein levels. These findings indicate that lack of endogenous Ang II in AGT-deficient mice inhibits UB branching. We conclude that intact RAS is critical in structural integrity of the renal collecting system and that UB morphogenetic program genes, such as AGT, renin, ACE, AT1R, or Spry1, are epigenetically controlled via HDACs.

Increased sympathoexcitation to angiotensin II (A II) and AT1 receptor mRNA in the paraventricular nucleus (PVN) of pregnant rats

Previously we reported that pressor and sympathetic nerve activity (SNA) responses due to AT1 receptor activation in the SFO are potentiated in term pregnant (P) compared to nonpregnant (NP) rats. The pathway requires ionotropic glutamate receptors (iGluRs) in the PVN and rostral ventrolateral medulla (RVLM) in both P and NP rats. AT1 receptors in the PVN and RVLM contribute to potentiated responses to SFO activation in P rats. Since the PVN is obligatory to this pathway, and PVN to RVLM projections are almost exclusively glutamatergic, we considered that the enhanced role of A II in the RVLM might be related to an A II dependent increase in excitatory drive from the PVN, which could then stimulate local production of A II in the RVLM. In sinoaortic denervated inactin anesthetized rats, increases in mean arterial pressure (MAP) and lumbar SNA (LSNA) due to microinjected A II (50 nl, 1.6 mM) in the PVN were greater in P (n = 4) compared to NP (n = 7) rats (ΔMAP: P= 25±5 mmHg; NP= 15±2 mmHg; ΔLSNA: P= 13±3%; NP= 6.5±2%). Real‐time RT‐PCR was used to assess relative expression of AT1 receptor mRNA (reference gene = GAPDH) in PVN tissue punches from 5 P and 5 NP rats. Expression of AT1 receptors in the PVN tended (P = 0.1) to be greater in P (2.4±0.82) compared to NP rats (1.0±0.22). These results are consistent with an increased role of PVN AT1 receptors in sympathoexcitation in pregnancy. (NIH HL 091164)

Hypothyroid dependent myocardial angiotensin receptor trafficking is involved in improved cardiac performance after heat acclimation

Aims The renin–angiotensin system (RAS) plays a key role in heat acclimation, a process which induces adaptive changes in cardiac function. These changes are mediated in part by reduced thyroid hormone activity and improve myocardial function during and following exposure to various (non-heat) stresses such as ischemia. The aim of this study was to examine the role of RAS in the development of the heat acclimated protected heart. Main methods Three treatment groups were used: (1) C, controls; (2) AC, heat acclimated rats (1 mo 34 °C,); and (3) HAEL, heat acclimated euthyroid rats treated with 3 ng/ml of eltroxine. A Langendorff perfusion apparatus was used to measure hemodynamic parameters at baseline and following administration of angiotensin-II, losartan and PD123319 in isolated hearts. Protein and mRNA levels of angiotensin receptors were measured. Key findings Both C and HAEL animals showed increased contractility and a drop in coronary flow during angiotensin II exposure whereas AC animals did not have an inotropic response or vasoconstriction. Significantly different patterns of AT1 and AT2 receptor densities (a 50% reduction and a 30% increase in outer cell membrane AT1 and AT2 receptors respectively) were observed in AC animals compared to the other two groups. AT receptor mRNA levels were similar in all treatment groups. Significance The attenuated response of heat acclimated hearts to angiotensin is mediated by reduced thyroxine levels and is associated with a shift in AT1 receptors from the outer to the inner membrane. This shift appears to be caused by modified posttranslational trafficking of AT receptors.