Systemic outcomes of (Pyr(1))-apelin-13 infusion at mid-late pregnancy in a rat model with preeclamptic features

Preeclampsia is a syndrome with diverse clinical presentation that currently has no cure. The apelin receptor system is a pleiotropic pathway with a potential for therapeutic targeting in preeclampsia. We established the systemic outcomes of (Pyr(1))-apelin-13 administration in rats with preeclamptic features (TGA-PE, female transgenic for human angiotensinogen mated to male transgenic for human renin). (Pyr(1))-apelin-13 (2 mg/kg/day) or saline was infused in TGA-PE rats via osmotic minipumps starting at day 13 of gestation (GD). At GD20, TGA-PE rats had higher blood pressure, proteinuria, lower maternal and pup weights, lower pup number, renal injury, and a larger heart compared to a control group (pregnant Sprague-Dawley rats administered vehicle). (Pyr(1))-apelin-13 did not affect maternal or fetal weights in TGA-PE. The administration of (Pyr(1))-apelin-13 reduced blood pressure, and normalized heart rate variability and baroreflex sensitivity in TGA-PE rats compared to controls. (Pyr(1))-apelin-13 increased ejection fraction in TGA-PE rats. (Pyr(1))-apelin-13 normalized proteinuria in association with lower renal cortical collagen deposition, improved renal pathology and lower immunostaining of oxidative stress markers (4-HNE and NOX-4) in TGA-PE. This study demonstrates improved hemodynamic responses and renal injury without fetal toxicity following apelin administration suggesting a role for apelin in the regulation of maternal outcomes in preeclampsia

Chymase-Dependent Generation of Angiotensin II from Angiotensin-(1-12) in Human Atrial Tissue

Since angiotensin-(1-12) [Ang-(1-12)] is a non-renin dependent alternate precursor for the generation of cardiac Ang peptides in rat tissue, we investigated the metabolism of Ang-(1-12) by plasma membranes (PM) isolated from human atrial appendage tissue from nine patients undergoing cardiac surgery for primary control of atrial fibrillation (MAZE surgical procedure). PM was incubated with highly purified 125I-Ang-(1-12) at 37°C for 1 h with or without renin-angiotensin system (RAS) inhibitors [lisinopril for angiotensin converting enzyme (ACE), SCH39370 for neprilysin (NEP), MLN-4760 for ACE2 and chymostatin for chymase; 50 µM each]. 125I-Ang peptide fractions were identified by HPLC coupled to an inline γ-detector. In the absence of all RAS inhibitor, 125I-Ang-(1-12) was converted into Ang I (2±2%), Ang II (69±21%), Ang-(1-7) (5±2%), and Ang-(1-4) (2±1%). In the absence of all RAS inhibitor, only 22±10% of 125I-Ang-(1-12) was unmetabolized, whereas, in the presence of the all RAS inhibitors, 98±7% of 125I-Ang-(1-12) remained intact. The relative contribution of selective inhibition of ACE and chymase enzyme showed that 125I-Ang-(1-12) was primarily converted into Ang II (65±18%) by chymase while its hydrolysis into Ang II by ACE was significantly lower or undetectable. The activity of individual enzyme was calculated based on the amount of Ang II formation. These results showed very high chymase-mediated Ang II formation (28±3.1 fmol×min−1×mg−1, n = 9) from 125I-Ang-(1-12) and very low or undetectable Ang II formation by ACE (1.1±0.2 fmol×min−1×mg−1). Paralleling these findings, these tissues showed significant content of chymase protein that by immunocytochemistry were primarily localized in atrial cardiac myocytes. In conclusion, we demonstrate for the first time in human cardiac tissue a dominant role of cardiac chymase in the formation of Ang II from Ang-(1-12)

Uptake and Metabolism of the Novel Peptide Angiotensin-(1-12) by Neonatal Cardiac Myocytes

Angiotensin-(1-12) [Ang-(1-12)] functions as an endogenous substrate for the productions of Ang II and Ang-(1-7) by a non-renin dependent mechanism. This study evaluated whether Ang-(1-12) is incorporated by neonatal cardiac myocytes and the enzymatic pathways of ¹²⁵I-Ang-(1-12) metabolism in the cardiac myocyte medium from WKY and SHR rats.The degradation of ¹²⁵I-Ang-(1-12) (1 nmol/L) in the cultured medium of these cardiac myocytes was evaluated in the presence and absence of inhibitors for angiotensin converting enzymes 1 and 2, neprilysin and chymase. In both strains uptake of ¹²⁵I-Ang-(1-12) by myocytes occurred in a time-dependent fashion. Uptake of intact Ang-(1-12) was significantly greater in cardiac myocytes of SHR as compared to WKY. In the absence of renin angiotensin system (RAS) enzymes inhibitors the hydrolysis of labeled Ang-(1-12) and the subsequent generation of smaller Ang peptides from Ang-(1-12) was significantly greater in SHR compared to WKY controls. ¹²⁵I-Ang-(1-12) degradation into smaller Ang peptides fragments was significantly inhibited (90% in WKY and 71% in SHR) in the presence of all RAS enzymes inhibitors. Further analysis of peptide fractions generated through the incubation of Ang-(1-12) in the myocyte medium demonstrated a predominant hydrolytic effect of angiotensin converting enzyme and neprilysin in WKY and an additional role for chymase in SHR.These studies demonstrate that neonatal myocytes sequester angiotensin-(1-12) and revealed the enzymes involved in the conversion of the dodecapeptide substrate to biologically active angiotensin peptides

HNF4alpha Dysfunction as a Molecular Rational for Cyclosporine Induced Hypertension

Induction of tolerance against grafted organs is achieved by the immunosuppressive agent cyclosporine, a prominent member of the calcineurin inhibitors. Unfortunately, its lifetime use is associated with hypertension and nephrotoxicity. Several mechanism for cyclosporine induced hypertension have been proposed, i.e. activation of the sympathetic nervous system, endothelin-mediated systemic vasoconstriction, impaired vasodilatation secondary to reduction in prostaglandin and nitric oxide, altered cytosolic calcium translocation, and activation of the renin-angiotensin system (RAS). In this regard the molecular basis for undue RAS activation and an increased signaling of the vasoactive oligopeptide angiotensin II (AngII) remain elusive. Notably, angiotensinogen (AGT) is the precursor of AngII and transcriptional regulation of AGT is controlled by the hepatic nuclear factor HNF4alpha. To better understand the molecular events associated with cyclosporine induced hypertension, we investigated the effect of cyclosporine on HNF4alpha expression and activity and searched for novel HNF4alpha target genes among members of the RAS cascade. Using bioinformatic algorithm and EMSA bandshift assays we identified angiotensin II receptor type 1 (AGTR1), angiotensin I converting enzyme (ACE), and angiotensin I converting enzyme 2 (ACE2) as genes targeted by HNF4alpha. Notably, cyclosporine represses HNF4alpha gene and protein expression and its DNA-binding activity at consensus sequences to AGT, AGTR1, ACE, and ACE2. Consequently, the gene expression of AGT, AGTR1, and ACE2 was significantly reduced as evidenced by quantitative real-time RT-PCR. While RAS is composed of a sophisticated interplay between multiple factors we propose a decrease of ACE2 to enforce AngII signaling via AGTR1 to ultimately result in vasoconstriction and hypertension. Taken collectively we demonstrate cyclosporine to repress HNF4alpha activity through calcineurin inhibitor mediated inhibition of nuclear factor of activation of T-cells (NFAT) which in turn represses HNF4alpha that leads to a disturbed balance of RAS

Myocardial extravascular extracellular volume fraction measurement by gadolinium cardiovascular magnetic resonance in humans: slow infusion versus bolus

<p>Abstract</p> <p>Background</p> <p>Myocardial extravascular extracellular volume fraction (Ve) measures quantify diffuse fibrosis not readily detectable by conventional late gadolinium (Gd) enhancement (LGE). Ve measurement requires steady state equilibrium between plasma and interstitial Gd contrast. While a constant infusion produces steady state, it is unclear whether a simple bolus can do the same. Given the relatively slow clearance of Gd, we hypothesized that a bolus technique accurately measures Ve, thus facilitating integration of myocardial fibrosis quantification into cardiovascular magnetic resonance (CMR) workflow routines. Assuming equivalence between techniques, we further hypothesized that Ve measures would be reproducible across scans.</p> <p>Methods</p> <p>In 10 volunteers (ages 20-81, median 33 yr, 3 females), we compared serial Ve measures from a single short axis slice from two scans: first, during a constant infusion, and second, 12-50 min after a bolus (0.2 mmol/kg gadoteridol) on another day. Steady state during infusion was defined when serial blood and myocardial T1 data varied <5%. We measured T1 on a 1.5 T Siemens scanner using a single-shot modified Look Locker inversion recovery sequence (MOLLI) with balanced SSFP. To shorten breath hold times, T1 values were measured with a shorter sampling scheme that was validated with spin echo relaxometry (TR = 15 sec) in CuSO4-Agar phantoms. Serial infusion vs. bolus Ve measures (n = 205) from the 10 subjects were compared with generalized estimating equations (GEE) with exchangeable correlation matrices. LGE images were also acquired 12-30 minutes after the bolus.</p> <p>Results</p> <p>No subject exhibited LGE near the short axis slices where Ve was measured. The Ve range was 19.3-29.2% and 18.4-29.1% by constant infusion and bolus, respectively. In GEE models, serial Ve measures by constant infusion and bolus did not differ significantly (difference = 0.1%, p = 0.38). For both techniques, Ve was strongly related to age (p < 0.01 for both) in GEE models, even after adjusting for heart rate. Both techniques identically sorted older individuals with higher mean Ve values.</p> <p>Conclusion</p> <p>Myocardial Ve can be measured reliably and accurately 12-50 minutes after a simple bolus. Ve measures are also reproducible across CMR scans. Ve estimation can be integrated into CMR workflow easily, which may simplify research applications involving the quantification of myocardial fibrosis.</p

Clinical utility of fixed-dose combinations in hypertension: evidence for the potential of nebivolol/valsartan

Jasmina Varagic,1&ndash;3 Henry Punzi,4,5 Carlos M Ferrario2,3,61Hypertension and Vascular Research Center, 2Division of Surgical Sciences, 3Department of Physiology and Pharmacology, Wake Forest University, Winston-Salem, NC USA; 4Trinity Hypertension and Diagnostic Research Center, Carrollton, TX, USA; 5Department of Family and Community Medicine, University of Texas Southwestern Medical Center, Dallas, TX, USA; 6Department of Internal Medicine and Nephrology, Wake Forest University, Winston-Salem, NC, USAAbstract: Despite significant advances in pharmacologic approaches to treat hypertension during the last decades, hypertension- and hypertension-related organ damage are still a high health and economic burden because a large proportion of patients with hypertension do not achieve optimal blood pressure control. There is now general agreement that combination therapy with two or more antihypertensive drugs is required for targeted blood pressure accomplishment and reduction of global cardiovascular risk. The goals of combination therapies are to reduce long-term cardiovascular events by targeting different mechanism underlying hypertension and target organ disease, to block the counterregulatory pathways activated by monotherapies, to improve tolerability and decrease the adverse effects of up-titrated single agents, and to increase persistence and adherence with antihypertensive therapy. Multiple clinical trials provide evidence that fixed-dose combinations in a single pill offer several advantages when compared with loose-dose combinations. This review discusses the advances in hypertension control and associated cardiovascular disease as they relate to the prospect of combination therapy targeting a third-generation beta (&beta;) 1-adrenergic receptor (nebivolol) and an angiotensin II receptor blocker (valsartan) in fixed-dose single-pill formulations.Keywords: blood pressure control, hypertension, &beta;1-adrenergic receptor, renin angiotensin system, fixed-dose combination therapy, nebivolol, valsarta

Central and peripheral catecholamine stores in spontaneously hypertensive rats under immobilization stress

The effects of increasing duration of immobilization stress (from 30 min to 240 min) on catecholamine stores in some central (hypothalamus, corpus striatum) and peripheral tissues (suprarenal glands, heart auricles) in spontaneously hypertensive rats were investigated. Sufficiently long immobilization induced significant depletion of both central and peripehral catecholamine stores. The greatest depletion occurred in the suprarenal gland(adrenaline) and in the hypothalamus (noradrenaline and dopamine). The corpus striatum (dopamine) and the heart auricles (noradrenaline) were the most resistant to immobilization stress. These data indicate that catecholamine depletion in all investigated structures in spontaneously hypertensive rats depended on the duration of immobilization. If was concluded that after immobilization for more than 30 min the release of catecholamines was stronger that the processes of synthesis and reuptake, thus leading to progressive depletion of catecholamine stores, particularly when imobilization lasted up to 240 min. The present results indicate a difference in the effects of immobilization stress on catecholamine stores between some brain regions and peripheral tissues in spontaneously hypertensive rats

Effects of chronic ethanol administration on the serotonin-producing cells in rat gastric antral and duodenal mucosa

The present study describes our observations on optical and ultrastructural features of serotonincontaining cells in the rat antral and upper duodenal mucosa, utilizing optic morphometric measurements in a model of experimental chronic alcoholism of rat in which nutrition was well controlled. Male Wistar rats were given ethanol to provide 23 per cent of the total calories, while starch replaced ethanol isocalorically in controls. Twenty-five per cent of the calories were provided by protein in both groups. Blood levels of serotonin were significantly raised after chronic ethanol feeding (0.05910.06 vs. 0.15910.012 pglml, p<0.01). Decrease in the number of immunohistochemicallydetectable serotonin-containing cells was found in the pyloric gland mucosal area specimens of the chronically ethanol-treated rats (68.915.2 vs 43.313.0; p<0.001). The immunohistologically-evaluated number of the same cells in the duodenal mucosa specimens was significantly decreased by alcohol feeding. Although total villi and crypt count per whole circular section, and the number of crypts per villus were not significantly changed either in control animals or in chronically ethanol-fed rats, decreased number of these cells per whole circular section (289121.6 vs. 183f 10.5; p<O.OOI) per villus (2.5210.14 vs. 1.2110.10; p<0.001) and per crypts (0.97kO.08 vs. 0.7910.04; p<0.05) were reported after alcohol consumption. In both control and experimental rats the cells were predominantly found in the basa1 half of the antropyloric mucosa. Alcohol did not lead to any changes in normal distribution of the duodenal serotonin-producing cells. The above quantitative changes in serotonin-producing cells were not accompanied by changes in their subcellular appearance in stomach and duodenal mucosa of alcoholtreated rat