The purpose of this study is to verify if the increase of renal resistance, along with the consequent reduction of glomerular filtrate, and the activation of renina-angiotensin system, could be attributed to not only neonatal acute hypoxia but to other factors as well. These factors could provoke an increase in renal vascular resistance (R.V.R.), a reduction of renal blood flow (R.B.F.) and renal glomerular filtrate (R.G.F.) condition. These components are present in angiotensin as well as in endothelin (ET1), a potent peptide from vascular endothel with vasoconstricting action and whose secretion increases during hypoxia. The Authors have studied and compared two groups of newborns. The first group of newborns was affected by perinatal asphyxia or hypoxia of variable seriousness. It included 24 newborns with gestational age between 37 and 41 weeks and with a birth weight between 3.200 gms and 3.500 gms. The second, control group, was made of an identical number of newborns of the same gestational age with a weight adequate to the birth age. The dosage of the ET1 was evaluated on the plasma, using the RIA method. The diagnostic criteria for this evaluation included clinical and biohumoral evidence (tab. 1). Between the two groups, group 1 with anoxic syndrome showed the more intense increase of ET1. This increased is due to either an increase value of plasmatic creatinine that is present in normal situations or in physiopathologic role played by the ET1 in course of acute renal failure after asphyxia.

Increased intrapericardial levels of endothelin-1 (ET-1) induce myocardial ischemia and concomitant release of the purine metabolites adenosine (ADO), inosine (INO) and hypoxanthine (HXA) into the pericardial fluid. However, the potential modulatory role of nitrogen monoxide in compensating the ET-1-induced ischemic stress is not fully elucidated. The pericardial elevations of purine metabolite concentrations in the pericardial fluid after ET-1 administration (150 pmol/kg intrapericardially) were measured in the in situ dog heart with (n = 6) or without (n = 5) systemic nitrogen monoxide synthase blockade (30 mg/kg (G)-nitro-L-arginine methyl ester, followed by 6 mg/min intravenously). After control sampling, three consecutive pericardial infusate samples (ET1, ET2, ET3) were obtained for purine metabolite determinations (high-performance liquid chromatography-ultraviolet). It was found that intrapericardial ET-1 elevated the pericardial purine metabolite concentrations significantly in both groups. No significant differences were detected between the control and (G)-nitro-L-arginine methyl ester-treated groups in ischemic changes of pericardial ADOmax (+3.27 +/- 1.13 microM versus +1.84 +/- 0.56 microM), INOmax (+15.21 +/- 2.3 microM versus +12.09 +/- 4.04 microM) and HXAmax (+16.34 +/- 2.98 microM versus +17.09 +/- 5.22 microM) levels and in the maximal ST elevations (0.43 +/- 0.05 mV versus 0.61 +/- 0.08 mV). The hemodynamic variables did not change with ET-1 administration. In conclusion, systemic nitrogen monoxide synthase blockade does not aggravate the ET-1-induced acute myocardial ischemia and the release of purine metabolites, suggesting that endogenous nitrogen monoxide is not a supplementary factor to purine metabolites in this type of coronary adaptive responses.

OBJECTIVE

The aim of this study was to evaluate the influence of resin luting cement's activation mode in the final shade of porcelain veneers after accelerated artificial aging (AAA).

METHODS

Porcelain veneers (IPS Empress Esthetic) were produced using a standardized shade (ET1) and thickness (0.6 mm). Twenty bovine teeth were collected, prepared, and divided into two groups: group I (n = 10)-light-cured group, only base paste was applied to the veneers; group II (n = 10)-dual-cured group, in which the same base paste used in group I and a transparent catalyst were proportionally mixed for 20 seconds and then applied to the veneers. The specimens were light-cured for 60 seconds each and were next subjected to AAA. They were submitted to color readings with a spectrophotometer in three instances: in the tooth surface (only the substrate), after the cementation and polymerization of the veneers, and after the AAA. The values of L*, a*, and b* were obtained and the total color change was calculated (∆E*). Values obtained were subjected to statistical analysis, with a significance of 0.05.

RESULTS

There were no significant differences between dual- and light-cured modes considering ∆E*, L*, a*, and b* values obtained after aging (p>> 0.05). Within the dual-cured mode there were no significant differences in ∆E*, L*, a*, and b* values (p>> 0.05).

CONCLUSIONS

No relevant differences were found between the two activation modes in color change. When submitted to aging, dual- and light-cured modes of the resin cement showed visually perceptible (∆E*>> 1.0) color changes; however, within the threshold of clinical acceptance (∆E*>> 3.3).

Background: Diabetic retinopathy (DR) has become a worldwide concern because of the rising prevalence rate of diabetes mellitus (DM). Despite much energy has been committed to DR research, it remains a difficulty for diabetic patients all over the world. Since apoptosis of retinal microvascular pericytes (RMPs) is the early characteristic of DR, this study aimed to reveal the mechanism of Shuangdan Mingmu (SDMM) capsule, a Chinese patent medicine, on oxidative stress-induced apoptosis of pericytes implicated with poly (ADP-ribose) polymerase (PARP) / glyceraldehyde 3-phosphate dehydrogenase (GAPDH) pathway.

Methods: Network pharmacology approach was performed to predict biofunction of components of SDMM capsule dissolved in plasma on DR. Both PARP1 and GAPDH were found involved in the hub network of protein-protein interaction (PPI) of potential targets and were found to take part in many bioprocesses, including responding to the regulation of reactive oxygen species (ROS) metabolic process, apoptotic signaling pathway, and response to oxygen levels through enrichment analysis. Therefore, in vitro research was carried out to validate the prediction. Human RMPs cultured with media containing 0.5 mM hydrogen oxide (H₂O₂) for 4 h was performed as an oxidative-damage model. Different concentrations of SDMM capsule, PARP1 inhibitor, PARP1 activation, and GAPDH inhibitor were used to intervene the oxidative-damage model with N-Acetyl-L-cysteine (NAC) as a contrast. Flow cytometry was performed to determine the apoptosis rate of cells and the expression of ROS. Cell counting kit 8 (CCK8) was used to determine the activity of pericytes. Moreover, nitric oxide (NO) concentration of cells supernatant and expression of endothelial nitric oxide synthase (eNOS), superoxide dismutase (SOD), B cell lymphoma 2 (BCL2), vascular endothelial growth factor (VEGF), endothelin 1 (ET1), PARP1, and GAPDH were tested through RT-qPCR, western blot (WB), or immunocytochemistry (ICC).

Results: Overproduction of ROS, high apoptotic rate, and attenuated activity of pericytes were observed after cells were incubated with media containing 0.5 mM H₂O₂. Moreover, downregulation of SOD, NO, BCL2, and GAPDH, and upregulation of VEGFA, ET1, and PARP1 were discovered after cells were exposed to 0.5 mM H₂O₂ in this study, which could be improved by PARP1 inhibitor and SDMM capsule in a dose-dependent way, whereas worsened by PARP1 activation and GAPDH inhibitor.

Conclusions: SDMM capsule may attenuate oxidative stress-induced apoptosis of pericytes through downregulating PARP expression and upregulating GAPDH expression.

Keywords: Apoptosis; Chinese herbal formula; Diabetic retinopathy; Network pharmacology; Oxidative stress; PARP/GAPDH; Pericytes.

OBJECTIVE

To study the interrelation between nitric oxide (NO) and endothelin-1 (ET1) in experimental acute hypoxic rats, and to evaluate the mechanism of acute hypoxic pulmonary hypertension affected by NO and ET1 and its intervention.

METHODS

Nicotinamide adenine dinucleotide phosphate (NADPH) diaphorase histochemical staining method, Griess biochemical assay and radioimmune assay were applied to investigate the changes of nitric oxide syntheses (NOS), NO and ET1 in normal, hypoxic, and L-Arginine (L-Arg) and dexamethasone treated hypoxic rats.

RESULTS

In normal rats, the NOS stain was localized in pulmonary vascular endothelium, and in the hypoxic rats, the activity of NOS was significantly lower. The level of plasma NO was significantly lower during acute hypoxia, but L-Arg as well as dexamethasone could prevent the drop of plasma NO. The level of plasma ET1 rose up significantly in the acute hypoxic rats, but after L-Arg therapy, it was significantly reduced, however, dexamethasone could not affect plasma ET1. The level of plasma cyclic guanosine monophosphate (cGMP) was significantly lower in the acute hypoxic rats, and L-Arg could prevent the drop of plasma cGMP, but dexamethasone could not prevent the drop of plasma cGMP.

CONCLUSIONS

NO and ET1 may modulate hypoxic pulmonary hypertension and acute hypoxia can result in acute hypoxic pulmonary hypertension. L-Arg can reverse the acute hypoxic pulmonary hypertension. Further study is needed if dexamethasone is beneficial in acute hypoxic diseases. NO may play an important role in physiology of the lung and acute hypoxic diseases.

OBJECTIVE

To explore the mechanisms underlying endothelin-1 (ET-1) elevations induced by excessive fluoride exposure.

METHODS

We measured serum and bone fluoride ion content and plasma ET-1 levels and compared these parameters among different groups in an animal model. We also observed morphological changes in the aorta and endothelium of rabbits. In cell experiments, human umbilical vein endothelial cells (HUVECs) were treated with varying concentrations of NaF for 24h, with or without 10 µM U0126 pretreatment for 1 h. ET-1 levels in culture fluid and intracellular reactive oxygen species (ROS) levels, as well as ET1 gene, endothelin-converting enzyme-1 (ECE-1), extracellular signal-regulating kinase 1/2 (ERK1/2), pERK1/2 expression levels and RAS activation were measured and compared among the groups.

RESULTS

Plasma ET-1 levels of rabbits increased significantly in fluorinated groups compared with those in the control group. The rabbit thoracic aortas became slightly hardened in fluorinated groups compared with those in the control group, and some vacuoles were present in the endothelial cell cytoplasm of the rabbits in fluorinated groups. In our cell experiments, ET1 gene and ECE-1 expression levels in HUVECs and ET-1 expression levels in the cell culture supernatants increased significantly in some experimental groups compared with those in the control group. These trends paralleled the changes in intracellular ROS levels, RAS activation, and the pERK1/2-to-ERK1/2 ratio. After U0126 was added, ECE-1 expression and ET-1 levels decreased significantly.

CONCLUSIONS

Excessive fluoride exposure leads to characteristic endothelial damage (vacuoles), thoracic aorta hardening, and plasma ET-1 level elevations in rabbits. In addition, the ROS-RAS-MEK1/2-pERK1/2/ERK1/2 pathway plays a crucial-and at least partial-role in ET-1 over-expression, which is promoted by excessive fluoride exposure.

A novel maltogenic amylase from Bacillus stearothermophilus ET1, which has a dual activity of alpha-1,4- and alpha-1,6-glycosidic bond cleavages and alpha-1,6-glycosidic bond formation, was crystallized by using the hanging-drop vapor-diffusion method. The best crystals were obtained by employing a high concentration of protein (56 mg ml-1) and a precipitant containing 22% glycerol, 1.6 M ammonium sulfate in 0.1 M Tris-HCl (pH 8.5). Native diffraction data to 2.66 A resolution have been obtained from crystals flash-frozen at 110 K. The crystals belong to the space group P212121 with unit-cell dimensions of a = 77.62, b = 121.23, c = 244. 29 A, and contain three or four protomers per asymmetric unit. Structure determination by multiple isomorphous replacement is in progress.

The purpose of this study was to evaluate the relationship between the circadian profile of the vasorelaxing substances calcitonin gene-related peptide (CGRP) and epoxyeicosatrienoic acids (EETs) and the vasconstrictive agent endothelin-1 (ET1) and the daily rhythms of cardiac hemodynamic indices (CHI) and baroreflex (BRS) in Wistar rats with 1 kidney-1 clip model of arterial hypertension (1K-1C AH). The animals were divided into 3 groups: I- sham-operated (SO), II- 4-week and III- 8-week 1K-1C AH rats. Plasma concentration of ET1, CGRP and EET's were investigated every 4 h. In conscious freely moving 1K-1C AH rats unlike SO animals blood pressure (BP), heart period (HP) and BRS underwent significant circadian fluctuations, with more marked increase in mean values of BP in 8-week hypertensive rats in comparison to 4-week hypertensive rats (179 ± 5 vs. 162 ± 4 mm Hg, p < 0.05). These alterations correlated with more significant reduction in HP (138 ± 5 vs. 150 ± 6 ms, p < 0,05) and BRS (0.44 ± 0.04 vs. 0.58 ± 0.04 ms mm Hg^-1, p < 0.05) in 8-week 1K-1C AH rats. The acrophases of BP in 8-week 1K-1C AH rats in comparison with 4-week were shifted to more late night hours (1:58 a.m. vs. 11:32 p.m.) and in both groups of animals corresponded to lowest circadian plasma levels of CGRP and EETs and to greatest level of ET1. SO rats were characterized by lower values of BP (121 ± 3 mm Hg, p < 0,05) and higher indices of HP (158 ± 2 ms, p < 0,05) and BRS (0.86 ± 0.02 ms mmHg^-1, p < 0,001) in comparison with 1K-1C AH rats 4-week duration. The acrophases of BP, HP and BRS in hypertensive animals were revealed at 14.8 ± 0.5 h, 13.6 ± 0.4 h and 13.1 ± 0.2 h, which correlated with maximal circadian contents of ET1 and CGRP at 24:00 h and EETs at 12:00 h and were shifted in comparison to sham-operated group. In rats with 1K-1C AH, plasma levels of ET1, CGRP and EETs undergo circadian fluctuation with corresponding alterations in CHI and BRS which are more markedly expressed on the late stage of diseases and could be used in future for predictive, preventive, and personalized treatment of arterial hypertension.

Research question: Does uterine activity differ in patients who have undergone successful IVF treatment compared with patients who have undergone unsuccessful IVF treatment?

Design: Prospective study of 16 women who underwent fresh single embryo transfer. All patients underwent transvaginal ultrasound in three phases of the IVF treatment: ovarian stimulation 1 h before embryo transfer (ET1) and 5-7 days after embryo transfer (ET5-7). Uterine motion analysis was implemented by a dedicated speckle tracking algorithm; frequency- and amplitude-related features were extracted from the derived signals to characterize the uterine activity in relation to ongoing implantation (positive HCG after 6 weeks) and ongoing pregnancy at 11 weeks.

Results: Uterine activity in terms of frequency (ovarian stimulation ET1, P = 0.04; ovarian stimulation ET5-7, P = 0.002) and amplitude (ovarian stimulation ET1, P = 0.0003; ovarian stimulation ET5-7, P = 0.000008) is significantly higher in the ovarian stimulation phase compared with ET1 and ET5-7. Women with ongoing pregnancies showed significantly higher uterine contraction frequency compared with those with no ongoing pregnancies in all phases (ovarian stimulation, P = 0.006; ET1, P = 0.015; ET5-7, P = 0.007). Uterine contraction amplitude was significantly lower (P = 0.037) in women at ET5-7 in women with ongoing pregnancies.

Conclusions: This study is a first step towards assessing uterine activity during IVF objectively and non-invasively. It is an essential step to understanding the previously suggested effect of contractions on IVF failure. Uterine activity after embryo transfer characterized by high frequency and low amplitude may favour embryo implantation. Research with larger patient cohorts is needed to build on current evidence and knowledge of uterine contractions during IVF.

Keywords: Embryo implantation; IVF; Ongoing pregnancy; Quantitative ultrasound imaging; Uterine contractility.

BACKGROUND

Our objective was to determine whether adenosine-induced ischemia exerts a delayed cardiac protective effect in patients with stable effort angina ischemic heart disease.

METHODS

The study group was comprised of 32 patients (men) with symptoms of stable effort angina, aged 38-65 years (Group 1), and 18 clinically healthy subjects (3 women, 15 men), aged 35-55 years (Control group). The study protocol included baseline ECG and treadmill echocardiogram (ET1); ECG and adenosine echocardiogram performed 7 days after ET1; repeated exercise test exactly 24 h after adenosine infusion (ET2). Increases in heart rate (HR), systolic (SBP) and diastolic (DBP) blood pressure, maximum ST-segment depression (max IST) and total ST-segment depression (SIST) on ECG were compared, as well as left ventricular end-diastolic volume (LVEDV), end-systolic (LVESV) volume, ejection fraction (EF), and wall motion synergy index (WMSI).

RESULTS

No statistically significant differences were found in the increased values of the investigated electrocardiographic and echocardiographic parameters in either group on either exercise test. The only positive trend was observed in LVEDV. In Group 1 LVEDV increased significantly from rest values during ET1, whereas during ET2 LVEDV did not change.

CONCLUSIONS

Adenosine-induced ischemia does not exert a delayed protective effect in respect to cardiac bioelectrical and mechanical functions in patients with ischemic heart disease in the form of stable effort angina.

The key enzymes involved in modification of the steroid nucleus of sterol-transforming mycobacteria--3beta-hydroxysteroid oxidase (3-OH-SO, EC 1.13.1.2) and 17beta-hydroxysteroid dehydrogenase (17-OH-SDH, EC 1.1.1)--were isolated and characterized. It is shown that 3-OH-SO is a multifunctional enzyme catalyzing oxidation of the 3beta-OH group, delta5 ->> delta4 isomerization, and 6-hydroxylation. Two forms of intracellular 17-OH-SDH that catalyze redox reactions at C17 were found, and their properties were determined. The presence of an extracellular 17-OH-SDH in Mycobacterium spp. (VKM Ac-1815 D and Et1) was demonstrated for the first time.

OBJECTIVE

To investigate the effects of salvianolic acid B (Sal B) on endothelin-1 (ET1)-induced contraction and cytoskeleton reorganization of rat hepatic stellate cells (HSCs).

METHODS

HSCs were collected from Sprague-Dawley rats by in situ perfusion with pronase E and isolated by density-gradient centrifugation with Nycodenz. Cells were treated with ET-1, with or without Sal B or Y-27632 (a specific inhibitor of rho-associated protein kinases) pretreatment. HSC contraction was evaluated by collagen gel contraction assay. Cytoskeletal reorganization in response to ET-1 was evaluated by detecting changes in phosphorylation of myosin light chain 2 (MLC2) using glycerol-urea PAGE and the Odyssey Infrared Imaging System. Changes in actin stress fiber polymerization were detected by FITC-labeled phalloidin. Differences between the various cell treatment/pretreatment groups were statistically analyzed.

RESULTS

Compared to the untreated control cells, the lattice area of ET-1-treated cells showed significant shrinkage (76.89% ± 3.84% vs. 37.10% ± 5.10%; P less than 0.01). Pretreatment with 105 M Sal B or 105 M Y-27632 significantly reduced ET-1-induced contraction (67.01% ± 4.14% and 77.28% ± 2.00%, respectively; bothP less than 0.01 vs. the ET-1-treated cells). The untreated control cells showed a basal MLC2 phosphorylation of (0.35 ± 0.05) mol PO4/mol MLC2. In contrast, ET-1 treatment elicited a rapid and sustained MLC2 phosphorylation, which was (0.87 ± 0.04) mol PO₄/mol MLC2 at 5 min post-treatment and with the maximal level of (0.96 ± 0.04) mol PO₄/mol MLC2 detected at 30 min post-treatment. The Sal B pretreatment led to a significant decrease in ET-1-induced MLC2 phosphorylation (by 63.1%) and an obvious disassembly of actin stress fibers.

CONCLUSIONS

Sal B effectively inhibits ET-1-induced rat HSC contraction, through its suppressive effects on MLC2 phosphorylation and promotion of the disassembly of actin stress fibers.

The effect of contractions elicited with ET1 and AVP after preincubating rat aortic and tail artery rings with a hyperinsulinemic dose (3 nM) of insulin were studied. Insulin preincubation (120 min), in the presence of 0.1 mM L-NAME, depressed contraction of aortic rings to 0.01 microM ET1 (132 +/- 6 vs. 161 +/- 9 mg/mm2 in control, n = 25; p < 0.05) and to 1 microM AVP (84 +/- 7 vs. 110 +/- 9 mg/mm2 in control, n = 16; p < 0.05), but did not modify 45Ca influx to the cell. Insulin-induced relaxation was inhibited by indomethacin 10 microM, an antagonist of prostaglandin synthesis, and also by blockade of insulin receptors with 30 microM genistein. A short insulin preincubation (15 min) did not modify ET1 contractions. In rat tail artery, insulin preincubation (120 min) increased the force developed by ET1 (847 +/- 45 vs. 596 +/- 99 mgF/mgW in controls, n = 14) by stimulating TXA2 release and/or actions. In summary, the present results suggest that endothelial factors are involved in both the vasoconstrictor and vasodilator effects of insulin on rat vessels.

Functional non-HLA antibodies (antibodies to non-human leukocyte antigens) targeting the G protein-coupled receptors angiotensin II type 1 receptor (AT1R) and endothelin-1 type A receptor (ETAR) are implicated in the pathogenesis of transplant vasculopathy. While ERK signaling (a regulator of cell growth) may represent a general cellular response to agonist stimulation, the molecular link between receptor stimulation and development of vascular obliteration has not been fully established. Here we hypothesize involvement of the versatile adaptor proteins, β-arrestins, and the major regulator of cell growth, PI3K/mTOR signaling, in impaired endothelial repair. To test this, human microvascular endothelial cells were treated with AT1R-/ETAR-antibodies isolated from patients with kidney transplant vasculopathy. These antibodies activated both mTOR complexes via AT1R and ETAR in a PI3K-dependent and ERK-independent manner. The mTOR inhibitor, rapamycin, completely abolished activation of mTORC1 and mTORC2 after long term treatment with receptor antibodies. Imaging studies revealed that β2- but not β1-arrestin was recruited to ETAR in response to ET1 and patient antibodies but not with antibodies isolated from healthy individuals. Silencing of β2-arrestin by siRNA transfection significantly reduced ERK1/2 and mTORC2 activation. Non-HLA antibodies impaired endothelial repair by AT1R and ETAR-induced mTORC2 signaling. Thus, we provide evidence that functional AT1R-/ETAR antibodies induce ERK1/2 and mTOR signaling involving β2-arrestin in human microvascular endothelium. Hence, our data may provide a translational rational for mTOR inhibitors in combination with receptor blockers in patients with non-HLA receptor recognizing antibodies.

Keywords: Endothelial cells; arrestin; mTOR; non-HLA antibodies; signal transduction; transplant vasculopathy.

Increasing water scarcity and rapid socio-economic development are driving farmers in Asia to transform traditionally flooded rice cropping systems into non-flooded crop production. The management of earthworms in non-flooded rice fields appears to be a promising strategy to support residue recycling and mitigate greenhouse gas (GHG) emissions triggered by residue amendment. We conducted a field experiment on non-flooded rainfed rice fields, with and without residue amendment. In-situ mesocosms were inoculated with endogeic earthworms (Metaphire sp.), with either low (ET1: 150 individuals m^-2), or high density (ET2: 450 individuals m^-2), and a control (ET0: no earthworms). We measured GHG emissions (methane (CH₄); nitrous oxide (N₂O); carbon dioxide (CO₂)) twice a week during the cropping season with static chambers. Effects of earthworms on yield and root growth were additionally assessed. Earthworms offset the enormous increase of CH₄ emissions induced by straw amendment (from 4.6 ± 5 to 75.3 ± 46 kg CH₄-C ha^-1 in ET0). Earthworm activity significantly reduced CH₄ release, particularly at ET2, by more than one-third (to 22 ± 15 kg CH₄-C ha^-1). In contrast, earthworm inoculation did not affect N₂O emission. Straw amendment more than doubled the global warming potential (GWP). Earthworms reduced GWP by 39% at low (ET1) and 55% at high densities (ET2). Earthworm activity reduced root mass density under conditions of straw amendment but did not affect yield. Earthworms can significantly reduce detrimental effects of rice crop residue amendment on GHG release under upland rice production. Organic carbon (C) might be preserved in earthworm casts and thereby limit C availability for CH₄ production. At the same time, earthworm activity might increase methanotrophic CH₄ consumption, due to improved soil aeration or less root exudates. Consequently, earthworms have a strong potential for regulating ecosystem functions related to rice straw decomposition, nutrient allocation and thus GHG reduction.

Background: Emerging evidence from animal studies and clinical trials indicates that systemic inhibition of endothelin1 (ET1) signaling by endothelin receptor antagonists improves pathological features of diabetes and its complications. It is indicated that endothelin type A receptor (ETAR) plays a major role in ET1-mediated pathophysiological actions including diabetic pathology. However, the effects as well as the mechanistic targets of hepatic ET1/ETAR signaling inhibition on the pathology of metabolic diseases remain unclear. This study aimed to investigate the beneficial effects as well as the underlying mechanisms of hepatic ETAR knockdown on metabolism abnormalities in high-fat diet (HFD)-fed mice.

Methods: Mice were fed a HFD to induce insulin resistance and metabolism abnormalities. L02 cells were treated with ET1 to assess the action of ET1/ETAR signaling in vitro. Liver-selective knockdown of ETAR was achieved by tail vein injection of adeno-associated virus 8 (AAV8). Systemic and peripheral metabolism abnormalities were determined in vivo and in vitro. Mitochondrial fragmentation was observed by transmission electron microscope (TEM) and mitoTracker red staining.

Results: Here we provided in vivo and in vitro evidence to demonstrate that liver-selective knockdown of ETAR effectively ameliorated hepatic insulin resistance and hyperglycemia in HFD-fed mice. Mechanistically, hepatic ETAR knockdown alleviated mitochondrial fragmentation and dysfunction via inactivating 66-kDa Src homology 2 domain-containing protein (p66Shc) to recover mitochondrial dynamics, which was mediated by inhibiting protein kinase Cδ (PKCδ), in the livers of HFD-fed mice. Ultimately, hepatic ETAR knockdown attenuated mitochondria-derived oxidative stress and related liver injuries in HFD-fed mice. These ETAR knockdown-mediated actions were confirmed in ET1-treated L02 cells.

Conclusion: This study defined an ameliorative role of hepatic ETAR knockdown in HFD-induced metabolism abnormalities by alleviating p66Shc-mediated mitochondrial fragmentation and consequent oxidative stress-related disorders and indicated that hepatic ETAR knockdown may be a promising therapeutic strategy for metabolic diseases.

Keywords: 66-kDa Src homology 2 domain-containing protein; endothelin type A receptor; endothelin-1; metabolism abnormality; mitochondrial fragmentation.

Objectives: Patients with heart failure and reduced left ventricular ejection fraction (HFrEF) are prone for ventricular tachyarrhythmias. We tested if assessment of the biomarkers C-terminal Endothelin 1 (CT-ET1), midregional pro atrial natriuretic peptide (MR-proANP) and midregional pro adrenomedullin (MR-proADM) might improve risk stratification for arrhythmic death.

Methods: This prospective observational study included 160 heart failure patients with ischemic cardiomyopathy (ICM) or non-ischemic, dilated cardiomyopathy (DCM) and 30 control patients without heart disease. Primary endpoint was arrhythmic death (ArD) or resuscitated cardiac arrest (resCA).

Results: A total of 61 patients died during the median follow-up of 7.0 [5.2-8.4] years. An ArD or resCA was observed in 48 patients. Plasma levels of CT-ET1 (p=0.002), MR-proANP (p<0.001) and MR-proADM (p=0.013) were significantly higher in ICM or DCM patients compared to controls. MR-proANP levels in ICM patients were associated with a significantly increased risk for ArD or resCA (hazard ratio (HR)=1.42, [95%CI: 1.08-1.85], p=0.011) in a multivariable Cox regression model. Plasma levels of CT-ET1 (HR=1.07 [0.98-1.17], p=0.113) and MR-proADM (HR=1.80 [0.92-3.55], p=0.087) were not associated with ArD or resCA in ICM patients. No significant association with ArD or resCA was found in DCM patients. Multivariable Cox regression showed that CT-ET1 (HR=1.14 [1.07-1.22], p<0.001), MR-proANP (HR=1.64 [1.29-2.08], p<0.001) and MR-pro ADM (HR=2.06 [1.12-3.77], p=0.020) were associated with a higher risk for overall mortality.

Conclusion: Patients with HFrEF had elevated levels of CT-ET1, MR-proANP and MR-proADM. Plasma levels of MR-proANP are useful as predictor for arrhythmic death in patients with ICM.

Keywords: Sudden cardiac death; arrhythmic death; biomarker; overall mortality; risk stratification; ventricular tachycardia.

Background: Mechanisms of exercise intolerance in patients with heart failure with preserved ejection fraction (HFpEF) are not well understood. Pulmonary hypertension, a common accompaniment in patients with HFpEF, is associated with poor outcomes. While Endothelin -1 (ET-1) plays a mechanistic role in pulmonary hypertension, its role in exercise intolerance in HFpEF is not well established.

Objective: To explore the association between plasma ET-1 levels and maximal oxygen consumption (pVO2), and their changes over 24 weeks in HFpEF.

Methods: This is a post-hoc analysis of the Phosphodiesterase-5 Inhibition to Improve Clinical Status and Exercise Capacity in Heart Failure with Preserved Ejection Fraction (RELAX) trial. We performed linear regressions to assess the relationship between plasma ET-1 and pVO2. We also used linear regressions to determine whether ET-1 was associated with change in peak VO2 (ΔpVO₂).

Results: A total of 210 patients were included. Baseline plasma ET-1 levels were associated with older age, higher NT-proBNP levels, higher serum creatinine levels, and higher prevalence of atrial fibrillation. Patients with higher ET1 levels also had higher plasma galectin-3 and CITP levels. After multiple adjustments, baseline ET1 levels were associated with lower pVO₂ (β -0.927, SE 0.196, p < 0.001). Over 24 weeks, the change in ET1 levels was associated with the change in pVO2 (multivariable adjusted β -0.415, SE 0.115, p = 0.018). Baseline ET1 levels did not modify the effect of sildenafil on change in peak VO₂.

Conclusions: Plasma ET1 levels are significantly associated with lower exercise oxygen consumption both at baseline and longitudinally over 24 weeks. Future studies should explore Endothelin-1 antagonism to improve exercise tolerance in HFpEF.

Keywords: Endothelin-1; Exercise intolerance; HFpEF.

The current preventive treatment for iron deficiency in pigs is inefficient, resulting in a high prevalence of iron-deficient or anemic postweaned pigs. The aim of this study was to develop and characterize edible toys (ETs) to be used as oral iron supplements, and to assess their effect on feeding behavior and iron status of postweaned pigs. Three types of ETs, varying in sweetness, were produced by ionic gelation, using whey, sodium alginate, ferrous sulfate and atomized bovine erythrocytes. ET control (ETC) was developed without sweetener, ET1 contained 15% w/v sucrose and ET2 contained 0.03% w/v of Sucram (98% sodium saccharin, 1% neosperidine dihydrocalcone and 1% maltol). ETs were mainly composed of carbohydrates and protein, with a similar concentration of iron (2.2-2.7 mg/g). The ETs were offered to 24 postweaned pigs to measure acceptability and preference. The animals preferred ETC and ET2 over ET1. To assess the nutritional benefit of the ETs, 24 postweaned pigs were distributed into three groups: ETC (without iron), ETC-Fe (ETC with iron) and ET2-Fe (with iron and Sucram). Iron-loaded ET (ETC-Fe and ET2-Fe) significantly increased the concentration of red blood cells (from 6.1 to 7.5·10⁶ x mm³ for ETC-Fe and from 6.2 to 7.8 for ET2-Fe), hematocrit (from 32.8 to 37.9% for ETC-Fe and from 32.3 to 35.1 for ET2-Fe), serum iron (from 28.6 to 120.6 µmol/L for ETC-Fe and from 34.9 to 145.4 for ET2-Fe) and serum ferritin (from 7.8 to 18.5 µg/L for ETC-Fe and from 8.1 to 20.2 for ET2-Fe). In conclusion, the ETs developed in this study were accepted by the pigs and provided adequate iron to improve the iron status of postweaned pigs.

Keywords: Anemia; Encapsulation; Heme-iron; Piglets; Supplement.

Objective: Growth factors [transforming growth factor-β (TGF-β), epidermal growth factor (EGF), endothelin-1 (ET1)] stimulate proteoglycan synthesis resulting in retention and accumulation of low density lipoprotein (LDL) in vessel intima and leading to atherosclerosis development. This study investigated the role of ET-1 on the expression of CHSY1, proteoglycan synthesizing enzyme, through both EGF and TGF-β receptor transactivation in human vascular smooth muscle cells (VSMCs). Also, we explored the involvement of NADPH oxidase (NOX), an important intermediate of redox signaling, in ET-1 transactivated EGF receptor (EGFR) through endothelin receptors.

Materials and methods: In this experimental study, phosphorylated ERK1/2 and CHSY1 protein levels in the human VSMCs were measured by Western blot analysis using anti phospho-ERK1/2 (Thr202/Tyr204) and anti CHSY1 antibodies.

Results: ET-1 (100 nM) and EGF (100 ng/ml) stimulated ERK1/2 phosphorylation and inhibited in the presence of bosentan (ET receptor inhibitor), AG1478 (EGFR inhibitor), and DPI (NOX antagonist). Also, ET-1 treatment increased CHSY1 enzyme level; this response was suppressed by bosentan, AG1478, DPI, and SB431542, TGF-β receptor antagonist. This study revealed that ET-1 increases expression of CHSY1 through transactivation of EGF and TGF-β receptors.

Conclusion: Transactivation through the EGF receptor mediated by phospho-ERK1/2 leads to expression of CHSY1 protein. EGF receptor transactivation by ET-1 is shown for the first time, to be dependent on NOX enzymes.

Keywords: CHSY1 Enzyme; Endothelin-1; Epidermal Growth Factor; NADPH Oxidase.

Brain capillaries are crucial for cognitive functions by supplying oxygen and other nutrients to and removing metabolic wastes from the brain. Recent studies have demonstrated that constriction of brain capillaries is triggered by beta-amyloid (Aβ) oligomers via endothelin-1 (ET1)-mediated action on the ET1 receptor A (ETRA), potentially exacerbating Aβ plaque deposition, the primary pathophysiology of Alzheimer's disease (AD). However, direct evidence is still lacking whether changes in brain capillaries are causally involved in the pathophysiology of AD. Using APP/PS1 mouse model of AD (AD mice) relative to age-matched negative littermates, we identified that reductions of density and diameter of hippocampal capillaries occurred from 4 to 7 months old while Aβ plaque deposition and spatial memory deficit developed at 7 months old. Notably, the injection of ET1 into the hippocampus induced early Aβ plaque deposition at 5 months old in AD mice. Conversely, treatment of ferulic acid against the ETRA to counteract the ET1-mediated vasoconstriction for 30 days prevented reductions of density and diameter of hippocampal capillaries as well as ameliorated Aβ plaque deposition and spatial memory deficit at 7 months old in AD mice. Thus, these data suggest that reductions of density and diameter of hippocampal capillaries are crucial for initiating Aβ plaque deposition and spatial memory deficit at the early stages, implicating the development of new therapies for halting or curing memory decline in AD.

Keywords: APP/PS1 mouse; Alzheimer’s disease; Aβ plaque; Endothelin-1; Ferulic acid (FA); Hippocampus.

The antihypertensive effect of wine lees powder (WLPW) from a Cabernet grape variety was related to its high content in flavanols and anthocyanins compounds. This study investigates the involvement of endothelial-derived factors and SIRT1 in its bioactivity. Spontaneously hypertensive rats (SHR) were orally administered water or WLPW (125 mg/kg bw). Posteriorly, both groups were intraperitoneally administered saline, Nω-nitro-L-arginine methyl ester (L-NAME), a nitric oxide (NO) synthesis inhibitor, indomethacin, a prostacyclin synthesis inhibitor, or sirtinol, an inhibitor of sirtuins. Blood pressure (BP) was recorded before and 6 h after WLPW administration. In an additional experiment, SHR were administered water or WLPW and endothelial expressions of eNos, Sirt1, Nox4, and Et1 were determined. The BP-lowering properties of WLPW were abolished by L-NAME and partially reduced by indomethacin, demonstrating that WLPW antihypertensive effect was mediated by changes in NO availability, although prostacyclin also contributed to this activity. Moreover, BP-lowering effect was reduced by sirtinol, indicating that WLPW decreased BP in a SIRT1-dependent manner. Furthermore, WLPW upregulated eNos and Sirt1 and downregulated Nox4 and Et1 endothelial gene expression. These results evidence the vasoprotective effect of WLPW and show that its antihypertensive effect in SHR is endothelium dependent and mediated by SIRT1.

Keywords: NADPH oxidase; endothelial function; endothelial nitric oxide synthase; endothelin-1; spontaneously hypertensive rats.

Background: Microvascular angina is a common clinical entity, with about a three-fold higher frequency in women. The pathogenesis of microvascular angina has not been much studied, but inflammation and endothelial dysfunction have been incriminated as the main mechanisms of this disease. Methoss: Our purpose was to analyze whether certain inflammatory markers, i.e., interleukin 6 (IL-6) and endothelin 1 (ET-1), can play a role in the diagnosis of microvascular angina in women.

Results: Ninety women with ischemic heart disease were divided into two groups, based on their affliction with either microvascular or macrovascular disease. In general, the levels of IL6 and ET1 were similar between the two groups. Analyzing these marker levels according to the number of coronary lesions, we obtained an increased IL6 value that was similar for patients with microvascular angina, one-vessel, and two-vessel coronary disease, but significantly lower than in women with three-vessel coronary lesions. Also, in microvascular angina, IL6 level was correlated with the NYHA IV functional class. Unexpectedly, the level of ET1 was correlated with left ventricular systolic dysfunction.

Conclusions: In women with an increased suspicion of microvascular angina, in whom microvascular dysfunction cannot be tested invasively, IL-6 level, unlike the ET-1 level, might be considered a diagnostic marker of this disease.

Keywords: ET1; IL6; ischemic heart disease in women; microvascular angina.

The study was aimed to evaluate the therapeutic effects of Zizyphus oxyphyla leaves methanolic (ZOX-LME), on serum liver, kidney and hematology along with other serum parameters in Carbon tetrachloride (CCl4) intoxicated rabbits. Experimental animals were divided into five groups, six rabbits in each. These were: group NC (normal control), group, TC (toxic control) and group ST i.e. silymarine administered group at dose rate (50) mg/kg body weight (BW). Group ET1 and group ET2 treated with (ZOX-LME) at dose 200 mg/kg BW and 400 mg/kg BW. CCl4 administration caused significant (P> 0.05) impairment in serum liver enzymes, blood factors and other serum indices. Treatment with (ZOX-LME) significantly (P<0.05) reduced and normalized the levels of serum alanine transaminase (ALT) aspartate transaminase (AST) and alkaline phosphatase (ALP) and hematological indices. Also significant (P< 0.05) reduction was observed in creatinine, urea, uric acid, blood urea nitrogen (BUN), and albumin and glucose concentrations. The altered levels of lipid profile and serum electrolytes (Ca, Mg, Cl, Na, K, and P) were significantly (P<0.05) change toward normal levels with (ZOX-LME) feeding. In addition (ZOX-LME) ingestion caused significant improvement in GSH, GST and CAT levels, while reducing the TBARS levels, exhibited antioxidant capacity. Also (ZOX-LME) showed increase inhibition against percent scavenging of 2, 2-diphenile-1-picrylehydrazyle (DPPH) free radical. Significant (P<0.05) normalizing effects were observed with high dose 400 mg/kg BW of (ZOX-LME and were equivalent to silymarine administered groups. The histological study of liver supported the hepatoprotective and renal curative activity of (ZOX-LME).