Role Of Endothelin-1 Research Articles

Sexual dimorphism of hypothalamic serotonin release during systemic inflammation: Role of endothelin-1

The hypothalamus receives serotonergic projections from the raphe nucleus in a sex-specific manner. During systemic inflammation, hypothalamic levels of serotonin (5-hydroxytryptamine [5-HT]) decrease in male rats. The present study evaluated the involvement of endothelin-1 (ET-1) in the febrile response, hypolocomotion, and changes in hypothalamic 5-HT levels during systemic inflammation in male and female rats. An intraperitoneal injection of lipopolysaccharide (LPS) induced a febrile response and hypolocomotion in both male and female rats. However, although LPS reduced hypothalamic levels of 5-HT and its metabolite 5-hydroxyindol acetic acid (5-HIAA) in male rats, it increased these levels in female rats. An intracerebroventricular injection of the endothelin-B receptor antagonist BQ788 significantly reduced LPS-induced fever and hypolocomotion and changes in hypothalamic 5-HT and 5-HIAA levels in both male and female rats. The i.c.v. administration of ET-1 induced a significant fever and hypolocomotion, but reduced the hypothalamic levels of 5-HT and 5-HIAA in both males and females. These results suggest an important sexual dimorphism during systemic inflammation regarding the release of 5-HT in the hypothalamus. Moreover, ET-1 arises as an important mediator involved in the changes in hypothalamic 5-HT levels in both male and female rats.

Activation of ETAR and ETBR in myocardial tissue characterizes heart failure induced by experimental autoimmune myocarditis

BackgroundEndothelial dysfunction is characterized by an imbalance between endothelium-derived vasodilatory and vasoconstrictive effects and may play an important role in the development of heart failure. An increasing number of studies have shown that endothelial-derived NO-mediated vasodilation is attenuated in heart failure patients. However, the role of endothelin-1 (ET-1) in heart failure remains controversial due to its different receptors including ET-1 receptor type A (ETAR) and ET-1 receptor type B (ETBR). The aim of this study was to determine whether ET-1 and its receptors are activated and to explore the role of ETAR and ETBR in heart failure induced by myocarditis.MethodsWe constructed an animal model of experimental autoimmune myocarditis (EAM) with porcine cardiac myosin. Twenty rats were randomized to the control group (3 weeks, n = 5), the extended control group (8 weeks, n = 5), the EAM group (3 weeks, n = 5), the extended EAM group (8 weeks, n = 5). HE staining was used to detect myocardial inflammatory infiltration and the myocarditis score, Masson’s trichrome staining was used to assess myocardial fibrosis, echocardiography was used to evaluate cardiac function, ELISA was used to detect serum NT-proBNP and ET-1 concentrations, and immunohistochemistry and western blotting were used to detect ETAR and ETBR expression in myocardial tissue of EAM-induced heart failure. Subsequently, a model of myocardial inflammatory injury in vitro was constructed to explore the role of ETAR and ETBR in EAM-induced heart failure.ResultsEAM rats tended to reach peak inflammation after 3 weeks of immunization and developed stable chronic heart failure at 8 weeks after immunization. LVEDd and LVEDs were significantly increased in the EAM group compared to the control group at 3 weeks and 8 weeks after immunization while EF and FS were significantly reduced. Serum NT-proBNP concentrations in EAM (both 3 weeks and 8 weeks) were elevated. Therefore, EAM can induce acute and chronic heart failure due to myocardial inflammatory injury. Serum ET-1 concentration and myocardial ETAR and ETBR protein were significantly increased in EAM-induced heart failure in vivo. Consistent with the results of the experiments in vivo, ETAR and ETBR protein expression levels were significantly increased in the myocardial inflammatory injury model in vitro. Moreover, ETAR gene silencing inhibited inflammatory cytokine TNF-α and IL-1β levels, while ETBR gene silencing improved TNF-α and IL-1β levels.ConclusionsET-1, ETAR, and ETBR were activated in both EAM-induced acute heart failure and chronic heart failure. ETAR may positively regulate EAM-induced heart failure by promoting myocardial inflammatory injury, whereas ETBR negatively regulates EAM-induced heart failure by alleviating myocardial inflammatory injury.

Endothelin System in Hypertension and Chronic Kidney Disease.

ET (endothelin) is a powerful vasoconstrictor 21-amino acid peptide present in many tissues, which exerts many physiological functions across the body and participates as a mediator in many pathological conditions. ETs exert their effects through ETA and ETB receptors, which can be blocked by selective receptor antagonists. ETs were shown to play important roles among others, in systemic hypertension, particularly when resistant or difficult to control, and in pulmonary hypertension, atherosclerosis, cardiac hypertrophy, subarachnoid hemorrhage, chronic kidney disease, diabetic cardiovascular disease, scleroderma, some cancers, etc. To date, ET antagonists are only approved for the treatment of primary pulmonary hypertension and recently for IgA nephropathy and used in the treatment of digital ulcers in scleroderma. However, they may soon be approved for the treatment of patients with resistant hypertension and different types of nephropathy. Here, the role of ETs is reviewed with a special emphasis on participation in and treatment of hypertension and chronic kidney disease.

Endothelin-1 in Health and Disease

Discovered almost 40 years ago, the potent vasoconstrictor peptide endothelin-1 (ET-1) has a wide range of roles both physiologically and pathologically. In recent years, there has been a focus on the contribution of ET-1 to disease. This has led to the development of various ET receptor antagonists, some of which are approved for the treatment of pulmonary arterial hypertension, while clinical trials for other diseases have been numerous yet, for the most part, unsuccessful. However, given the vast physiological impact of ET-1, it is both surprising and disappointing that therapeutics targeting the ET-1 pathway remain limited. Strategies aimed at the pathways influencing the synthesis and release of ET-1 could provide new therapeutic avenues, yet research using cultured cells in vitro has had little follow up in intact ex vivo and in vivo preparations. This article summarises what is currently known about the synthesis, storage and release of ET-1 as well as the role of ET-1 in several diseases including cardiovascular diseases, COVID-19 and chronic pain. Unravelling the ET-1 pathway and identifying therapeutic targets has the potential to treat many diseases whether through disease prevention, slowing disease progression or reversing pathology.

The Role of Endothelin-1 and Oxidative Stress In Pregnancy Induced Hypertension


 Pregnancy Induced HypertensionPIH is one of the most frequent complications of pregnancy , however little is known about its etiology. Endothelial dysfunction serves as a causative factor in the initiation of the maternal pathophysiological changes of PIH & is not just a result of this disorder. Endothelin-1 may play an important role in the pathophysiology of PIH, either by acting on vascular smooth muscle directly to induce contraction or by increasing the formation of angiotensin II. Pregnancy induced hypertension is associated with endothelial dysfunction & could be caused by oxidative stress. Recent evidence suggest the role of oxidative stress in PIH. The high level of malondiahyde MDA may reflect the excessive oxidative damage in PIH. The study sample consist of 50 normal non pregnant women, 50 normotensive pregnant women, and 50 preeclamptic pregnant women in their third trimester in Mosul city. The aim of this study was designed to evaluate the role of endothelial dysfunction and oxidative stress in pathogenesis of PIH. The results of this study showed that there was a highly significant elevation (P0.000) in the serum level of endothelin-1 in the preeclamptic pregnant women in comparison with normotensive pregnant & the control group. Also there was a highly significant elevation (P0.000) in the level of serum MDA in the preeclamptic pregnant women in comparison with normotensive pregnancy & the control group. The serum endothelin-1 has a significant negative correlation with both maternal age & gestational age, while endothelin-1 has a significant positive correlation with diastolic blood pressure. The MDA showed a significant negative correlation with maternal age, while there was a significant positive correlation between MDA & diastolic blood pressure.
 
 

Research progress on the role of ET-1 in diabetic kidney disease.

Diabetic kidney disease (DKD) is one of the common complications of diabetes mellitus, which usually progresses to end-stage renal disease and causes great damage to the health of patients. Endothelin-1 (ET-1), a molecule closely associated with the progression of DKD, has increased expression in response to high glucose stimulation and is involved in hemodynamic changes, inflammation, glomerular and tubular dysfunction in the kidney, causing an increase in proteinuria and a decrease in glomerular filtration function, ultimately leading to glomerulosclerosis and renal failure. This paper aims to review the molecular level changes, regulatory mechanisms, and mechanisms of action of ET-1 under DKD, clinical trials of ET-1 receptor antagonists in recent years and current problems, to provide basic information and new research directions and ideas for the treatment of DKD and ET-1-related research.

Skin Temperature Circadian Rhythms and Dysautonomia in Myalgic Encephalomyelitis/Chronic Fatigue Syndrome: The Role of Endothelin-1 in the Vascular Tone Dysregulation.

There is accumulating evidence of autonomic dysfunction in myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS); however, little is known about its association with circadian rhythms and endothelial dysfunction. This study aimed to explore the autonomic responses through an orthostatic test and analysis of the peripheral skin temperature variations and vascular endothelium state in ME/CFS patients. Sixty-seven adult female ME/CFS patients and 48 healthy controls were enrolled. Demographic and clinical characteristics were assessed using validated self-reported outcome measures. Postural changes in blood pressure, heart rate, and wrist temperature were recorded during the orthostatic test. Actigraphy during one week was used to determine the 24-h profile of peripheral temperature and activity. Circulating endothelial biomarkers were measured as indicators of endothelial functioning. Results showed that ME/CFS patients presented higher blood pressure and heart rate values than healthy controls in the supine and standing position (p < 0.05 for both), and also a higher amplitude of the activity rhythm (p < 0.01). Circulating levels of endothelin-1 (ET-1) and vascular cell adhesion molecule-1 (VCAM-1) were significantly higher in ME/CFS (p < 0.05). In ME/CFS, ET-1 levels were associated with the stability of the temperature rhythm (p < 0.01), and also with the self-reported questionnaires (p < 0.001). This suggests that ME/CFS patients exhibited modifications in circadian rhythm and hemodynamic measures, which are associated with endothelial biomarkers (ET-1 and VCAM-1). Future investigation in this area is needed to assess dysautonomia and vascular tone abnormalities, which may provide potential therapeutic targets for ME/CFS.

Blockade of endothelin-1 receptor B regulates molecules of the major histocompatibility complex in sickle cell disease.

Major Histocompatibility Complex (MHC) molecules have been proposed to play a role in Sickle Cell Disease (SCD) pathophysiology. Endothelial cells express MHC molecules following exposure to cytokines. SCD is characterized, in part, by vascular endothelial cell activation, increased oxidative stress, sickle cell adhesion, and excess levels of endothelin-1 (ET-1) contributing to vaso-occlusive crises. ET-1 activates endothelial cells, induces oxidative stress and inflammation, and alters erythrocyte volume homeostasis. However, the role of ET-1 on MHC regulation in SCD is unclear. We first studied two sickle transgenic knockout mouse models of moderate to severe disease phenotype, βS-Antilles and Berkeley (BERK) mice. We observed significant increases in H2-Aa mRNA levels in spleens, lungs, and kidneys from transgenic sickle mice when compared to transgenic knockout mice expressing human hemoglobin A (HbA). Mice treated for 14 days with ET-1 receptor antagonists significantly reduced H2-Aa mRNA levels. We characterized the effect of ET-1 on MHC class II expression in the human endothelial cell line EA.hy926. We observed dose-dependent increases in the expression of MHC class II (HLA-DRA) and MHC transcription factor (CIITA) that were significantly blocked by treatment with BQ788, a selective blocker of ET-1 type B receptors. Chromatin immunoprecipitation studies in EA.hy926 cells showed that ET-1 increased Histone H3 acetylation of the HLA-DRA promoter, an event blocked by BQ788 treatment. These results implicate ET-1 as a novel regulator of MHC class II molecules and suggest that ET-1 receptor blockade represents a promising therapeutic approach to regulate both immune and vascular responses in SCD.

Role of endothelin in hypertension: a review

Role of endothelin in hypertension: a review

Abstract 13344: Endothelin B Receptor Agonist, Sovateltide, Promotes Neural Regeneration by Improving Mitochondria Function in a Neonatal Rat Model of Hypoxic-Ischemic Encephalopathy

Introduction: Our preliminary studies have indicated the role of Endothelin B receptors (ETBR) stimulation in decreasing oxidative stress and cell death after sovateltide treatment following hypoxic-ischemic encephalopathy (HIE) in rat pups, which suggests the involvement of ETBR in neonatal hypoxic-ischemic injury. However, its mechanism of action is not yet established. Therefore, this study investigated the effect of sovateltide on mitochondrial dysfunction and neurogenesis in a neonatal rat model of HIE. Hypothesis : Sovateltide protects neuronal survival via preserving mitochondrial activity. Methods: Sprague-Dawley male and female rat pups grouped separately, were divided into 5 different subgroups (1) Control; (2) HIE + Vehicle; (3) HIE + Hypothermia; (4) HIE + sovateltide; and (5) HIE + sovateltide + hypothermia. HIE was induced by ligating the right carotid artery on a postnatal day (PND) 7, followed by hypoxia for 120 min and hypothermia (32-34°C) for 5 h. On PND 7, sovateltide (5 μg/kg, ICV) was injected after hypoxic-ischemic injury. Pups were euthanized on PND 10. Brains were isolated for analysis of markers for mitochondrial fission and fusion as well as neurogenesis in rat brain tissues using western blots. Results : Animals receiving sovateltide showed a significantly increased expression of neuronal differentiation marker, NeuroD1 (p&lt;0.0001) and DoubleCortin (p&lt;0.001) along with neural marker for mature neurons, NeuN (p&lt;0.001) compared to vehicle. Additionally, an upregulation (p&lt;0.0001) of mitochondrial fusion-related proteins, Mfn2, and downregulation of fission proteins, Drp1, in the sovateltide group compared to the vehicle was observed. Higher neuronal progenitor cell differentiation along with better mitochondrial biogenesis in the brain of sovateltide alone or as an adjunct to hypothermia in rat pups was noted. Similar findings were obtained in both male and female rats. Conclusion : Stimulation of ETBR by sovateltide promotes neural regeneration via maintaining mitochondrial dynamics, indicating a potential neuroprotective effect and may, therefore, offer a legitimate therapeutic target for the treatment of neonatal HI brain injury.

Role of Endothelin-1 in Right Atrial Arrhythmogenesis in Rabbits with Monocrotaline-Induced Pulmonary Arterial Hypertension.

Atrial arrhythmias are considered prominent phenomena in pulmonary arterial hypertension (PAH) resulting from atrial electrical and structural remodeling. Endothelin (ET)-1 levels correlate with PAH severity and are associated with atrial remodeling and arrhythmia. In this study, hemodynamic measurement, western blot analysis, and histopathology were performed in the control and monocrotaline (MCT, 60 mg/kg)-induced PAH rabbits. Conventional microelectrodes were used to simultaneously record the electrical activity in the isolated sinoatrial node (SAN) and right atrium (RA) tissue preparations before and after ET-1 (10 nM) or BQ-485 (an ET-A receptor antagonist, 100 nM) perfusion. MCT-treated rabbits showed an increased relative wall thickness in the pulmonary arterioles, mean cell width, cross-sectional area of RV myocytes, and higher right ventricular systolic pressure, which were deemed to have PAH. Compared to the control, the spontaneous beating rate of SAN–RA preparations was faster in the MCT-induced PAH group, which can be slowed down by ET-1. MCT-induced PAH rabbits had a higher incidence of sinoatrial conduction blocks, and ET-1 can induce atrial premature beats or short runs of intra-atrial reentrant tachycardia. BQ 485 administration can mitigate ET-1-induced RA arrhythmogenesis in MCT-induced PAH. The RA specimens from MCT-induced PAH rabbits had a smaller connexin 43 and larger ROCK1 and phosphorylated Akt than the control, and similar PKG and Akt to the control. In conclusion, ET-1 acts as a trigger factor to interact with the arrhythmogenic substrate to initiate and maintain atrial arrhythmias in PAH. ET-1/ET-A receptor/ROCK signaling may be a target for therapeutic interventions to treat PAH-induced atrial arrhythmias.

Role of endothelin in the pathophysiology of migraine: A new view on an old player

There is cumulating evidence that endothelin-1 (ET-1) may play a role in migraine, however controversial findings still impede a conclusion to be drawn. Herein we tested the hypothesis that endothelin ETB receptors are major contributors to migraine-like responses. ET-1, IRL-1620 (selective ETB receptor agonist) or CGRP were injected into the trigeminal ganglion (TG) of female Wistar rats, and the development of periorbital mechanical allodynia was assessed hourly with von Frey hairs. Twenty-four hours later, rats were exposed to an aversive light for 1 h, after which the reactivation of periorbital mechanical allodynia (indicating photic sensitivity) was assessed up to 4 h. Moreover, the effect of systemic Bosentan (ETA/ETB receptors antagonist) or the selective antagonists of ETA (BQ-123) and ETB (BQ-788) receptors injected into the TG were evaluated against CGRP-induced responses. ET-1 and IRL-1620 injection into the TG induced periorbital mechanical allodynia and photic sensitivity. Bosentan attenuated periorbital mechanical allodynia but failed to affect photic sensitivity induced by CGRP. Selective blockade of ETB receptors in the TG fully prevented the development of periorbital mechanical allodynia and photic sensitivity induced by CGRP, but ETA receptor blockade caused only a slight reduction of periorbital mechanical allodynia without affecting photic sensitivity. ETB receptor-operated mechanisms in the TG may contribute to migraine-like responses in female rats.

Endothelin-1 as a novel target for the prevention of metabolic dysfunction with intermittent hypoxia in male participants.

We examined the effect of intermittent hypoxia (IH, a hallmark feature of sleep apnea) on adipose tissue lipolysis and the role of endothelin-1 (ET-1) in this response. We hypothesized that IH can increase ET-1 secretion and plasma free fatty acid (FFA) concentrations. We further hypothesized that inhibition of ET-1 receptor activation with bosentan could prevent any IH-mediated increase in FFA. To test this hypothesis, 16 healthy male participants (32 ± 5 yr, 26 ± 2 kg/m2) were exposed to 30 min of IH in the absence (control) and presence of bosentan (62.5 mg oral twice daily for 3 days prior). Arterial blood samples for ET-1, epinephrine, and FFA concentrations, as well as abdominal subcutaneous adipose tissue biopsies (to assess transcription of cellular receptors/proteins involved in lipolysis), were collected. Additional proof-of-concept studies were conducted in vitro using primary differentiated human white preadipocytes (HWPs). We show that IH increased circulating ET-1, epinephrine, and FFA (P < 0.05). Bosentan treatment reduced plasma epinephrine concentrations and blunted IH-mediated increases in FFA (P < 0.01). In adipose tissue, bosentan had no effect on cellular receptors and proteins involved in lipolysis (P > 0.05). ET-1 treatment did not directly induce lipolysis in differentiated HWP. In conclusion, IH increases plasma ET-1 and FFA concentrations. Inhibition of ET-1 receptors with bosentan attenuates the FFA increase in response to IH. Based on a lack of a direct effect of ET-1 in HWP, we speculate the effect of bosentan on circulating FFA in vivo may be secondary to its ability to reduce sympathoadrenal tone.

Hypoxia Regulates Endothelin‐1 Production and Microglial Cell Activation

Oxidative stress and inflammatory responses play critical roles in hypoxic‐ischemic brain injury.Microglial cells are rapidly activated in response to injury and stressful stimuli, including hypoxia.Endothelin‐1 (ET‐1) is a potent vasoconstrictor that has been associated with cerebrovascular diseases. Hypoxia stimulates endothelial ET‐1 production. However, the role of ET‐1 in microglia under hypoxia is not clear. The aim of this project was to characterize the effect of hypoxia in a human microglial cell line, HMC3. We induced hypoxia using a chamber (1% O2, 5% CO2, and 92 % N2) at 37 °C for 4h. MUSE Oxidative Stress Assay was performed to measure reactive oxygen species (ROS) formation, ELISA to determine TNF, IL‐6, and ET‐1 levels, qPCR to measure gene expression of ET‐1, and immunofluorescence staining to visualize and compare the presence of ET‐1 in hypoxic and controlgroup cells. It was observed that, when compared to normoxic HMC3 cells, hypoxic HMC3 exposure significantly increased the ROS by a factor of 2.5 (p&amp;lt;0.001, n=3), the pro‐inflammatory cytokine TNF‐α increased 3.8 times (p&amp;lt;0.01, n=4), and IL‐6 increased by a factor of 1.6 (p&amp;lt;0.01, n=4). In addition, hypoxia stimulates ET‐1 gene expression 5.0‐fold (p&amp;lt;0.001, n=4) and increased protein production 1.3 times (p&amp;lt;0.01, n=4). Consequently, treatment with ET‐1 increased the amount of ROS, TNF‐α, and IL‐6 in HMC3 cells by a factor of 1.4 (p&amp;lt;0.05, n=4), 1.6 (p&amp;lt;0.001, n=4), and 1.9 (p&amp;lt;0.05, n=4), respectively. All these events were blocked by ET‐1 receptor A (ETRA) antagonist, BQ123. Our results suggest that hypoxic conditions create a cycle of microglial cell activation leading to increased ROS and ET‐1 production that further stimulate microglial cells. Thus, we posit that the ET‐1 receptor blockade represents a promising therapeutic approach to regulate microglial cell responses in hypoxic‐ischemic brain injury.

Evaluation of Cardiovascular Experiment Animal

Abstract: In vivo models of myocardial infarction induced by coronary artery ligation in rats usually suffer from high early mortality and a low rate of induction. This study investigated the time course initiation of chronic myocardial infarction in albino rats and the possibility of reducing early mortality rate due to myocardial infarction by modification of the surgical technique. CAL was carried out by passing the suture through the pericardial layer around the midway of the left anterior descending coronary artery including a small area of the myocardium to avoid mechanical damage to the heart geometry. In addition, the role of endothelin-1 in rat heart with congestive heart failure was critically assessed. Time course initiation experiments were designed by sacrificing the animals at different time intervals and by carrying out physiological, biochemical, histopathological, electron microscopical and immunohistochemical studies. Specific markers of myocardial injury, viz. cardiac troponin-T, high sensitivity C-reactive protein, lactate dehydrogenase and fibrinogen were measured at different time points. Serum marker enzymes and activities of lysosomal hydrolases were found to be elevated on the eighth day post-ligation. Histopathological studies demonstrated focal areas showing fibrovascular tissue containing fibroblasts, collagenous ground substance and numerous small capillaries replacing cardiac muscle fibers. Transmission electron micrographs exhibited mitochondrial changes of well-developed irreversible cardiac injury, viz. swelling, disorganization of cristae, appearance of mitochondrial amorphous matrix densities, and significant distortion of muscle fibers and distinct disruption of the intercalated discs. Immune blotting studies confirmed the presence of alpha 2-macroglobulin which supported the inflammatory response. The severity of the CMI was inferred by the measurement of the level of ET-1 in plasma and left ventricle which was significantly higher in the CMI rats than in the sham-operated rats. Immunohistochemical studies at different time intervals showed that there was a significant immunoexpression of ET-1 on the eighth day post-ligation. This study conclusively showed that ligation of left anterior descending artery minimised mortality and ET-1 was expressed during CMI.

Circulating Levels of Endothelin-1 and Big Endothelin-1 in Patients with Essential Hypertension.

The role of endothelin-1 (ET-1) in the pathogenesis of hypertension (HTN) is not clearly established. There is evidence that its circulating levels are elevated in some forms of experimental and human HTN, but this was not a consistent finding. Based on these controversial data, we tested serum levels of ET-1 and Big ET-1 (the precursor of ET-1) in patients with essential HTN, comparing the results with those of healthy normotensive controls. The levels of ET-1 and Big ET-1 were measured by ELISA. Our results in patients with essential HTN showed that the mean levels of ET-1 (5.01 ± 2.1 pg/mL) were significantly higher (F = 6.34, p = 0.0144) than the mean levels in the control group (3.2 ± 1.0 pg/mL). The levels of Big ET-1 in patients with essential HTN (0.377 ± 0.1 pmol/L) were similar to those in the control group (0.378 ± 0.07 pmol/L) and did not differ significantly (F = 0.00, p = 0.9531). These data suggest that ET-1, but not Big ET-1, may play an important role in the pathogenesis of primary HTN.

Prevention of VEGF inhibitor-induced toxicity by salt restriction: the SUN-SALT study

Abstract Introduction Vascular endothelial growth factor (VEGF) inhibitors target the formation of new blood vessels required for growth and metastatic spread of a malignant tumor. Although this is an effective anticancer treatment, many patients develop cardiovascular side effects such as hypertension, requiring dose reduction or early termination of treatment. In animals, VEGF inhibitor-induced hypertension is salt-sensitive. Aim To prospectively study whether salt restriction can prevent or attenuate the rise in blood pressure in response to anti-cancer treatment with VEGF inhibitors. Method This is a single centre prospective open-label intervention study. Patients are eligible when treated with a VEGF inhibitor according to standard of care and developing hypertension or a blood pressure rises of 20 mmHg or more during the first treatment cycle. A salt restricted diet (&amp;lt;4 grams/day) including provided salt-less bread is started during the off-treatment period under guidance of a specialized dietitian. The primary endpoint is mean difference in blood pressure rise between the treatment cycle with and the treatment cycle without salt restriction. We aim for a total of 16 patients with a blood pressure rise of at least 20mmHg and/or development of hypertension undergoing the intervention. Results Between 28 November 2019 and 25 March 2021, 45 patients gave informed consent. Fourteen patients developed hypertension and/or a blood pressure rise of at least 20 mmHg after three- four weeks of treatment making them eligible for the intervention. In 10 patients, salt restriction was the only intervention to reduce the blood pressure rise during the following treatment cycle, leading to a reduction in blood pressure rise of 17 mmHg (10 vs 27 mmHg; p&amp;lt;0.001). In four patients antihypertensive treatment was started during the first treatment cycle due to blood pressure rise above 170 mmHg. Salt restriction did not appear to have an important further blood pressure lowering effect, although in one patient the antihypertensive treatment was interrupted during the stop week and salt restriction was sufficient to limit the blood pressure rise in the second cycle. Importantly, the intervention was well tolerated and most patients continued salt restriction after the study finished. Conclusion Applying salt restriction might be an effective and well tolerated intervention to decrease blood pressure rise during treated with VEGF inhibitors. More importantly, this gives important information about the pathogenesis. Further studies of collected blood and 24h urine samples will allow conclusions on the role of endothelin-1, the renin aldosterone system and prostacyclins. Funding Acknowledgement Type of funding sources: Foundation. Main funding source(s): De Merel (Charity aiming for research directly benefiting patients; yearly award, success rate ∼30% “Stichting De Merel”)

ET-1 Promotes Epithelial-Mesenchymal Transition in Oral Squamous Cell Carcinoma Cells via the microRNA-489-3p /TWIST Axis.

ObjectiveOral squamous cell carcinoma (OSCC) constitutes almost 90% of head and neck malignancies and has a poor prognosis. To improve the efficacy of OSCC therapy, it is of great significance to explore other therapy for OSCC. Endothelin-1 (ET-1), a potent vasoconstrictor peptide, is implicated in cancer pathogenesis. Moreover, ET-1 promotes epithelial-mesenchymal transition (EMT) during the development of human cancers. We further to found that ET-1 exposure induced EMT in human squamous cell carcinoma cell lines SCC4 and SAS, by enhancing the expression of EMT biomarkers N-cadherin and vimentin and reducing E-cadherin expression via upregulation of the transcription factor TWIST.Materials and MethodsCell motility was examined by migration, invasion and wound-healing assays. Quantitative real time polymerase chain reaction (q-PCR), and promoter assays confirmed the inhibitory effects of ET-1 on miRNAs expression in oral cancer cells. We demonstrate an intravenous injection model of lung metastasis followed by an advanced method for quantifying metastatic tumor using image analysis software.ResultsIn addition, ET-1/ETAR reduced levels of microRNA-489-3p (miR-489-3p), a transcriptional repressor of TWIST. We have identified a novel bypass mechanism through which ET-1/ETAR are involved in TWIST signaling and downregulate miR-489-3p expression, enabling OSCC cells to acquire the EMT phenotype. Notably, ET-1 knockdown dramatically decreased levels of EMT markers and cell migration potential.ConclusionThe role of ET-1 in OSCC progression is supported by our findings from an in vivo murine model of OSCC. ET-1 may therefore represent a novel molecular therapeutic target in OSCC metastasis.

The Role of Endothelins, IL-18, and NGAL in Kidney Hypothermic Machine Perfusion.

Ischemia-reperfusion injury (IRI) occurring after renal transplantation is a complex biochemical process that can be monitored by specific biomarkers. The roles of those are not yet fully elucidated. The aim of this study was to analyze the concentrations of endothelins (ET-1, ET-2, and ET-3), interleukin-18 (IL-18), and neutrophil gelatinase-associated lipocalin (NGAL) during the reperfusion of human kidneys grafted from brain dead donors and later transplanted. The study group (n = 44) was analyzed according to the method of kidney storage: Group 1 underwent hypothermic machine perfusion (HMP) in the LifePort perfusion pump (n = 22), and Group 2 underwent static cold storage (SCS) (n = 22). The analysis of kidney function was performed daily during the first seven days after transplantation. The kidneys in Group 1 were characterized by higher absolute concentrations of ET-1, IL-18, and NGAL, as well as a lower concentration of ET-2 (p = 0.017) and ET-3. The relative increase of ET-1 (p = 0.033), ET-2, and ET-3 during reperfusion was lower in this group, while the relative decrease of NGAL was higher. Group 1 was also characterized by significant decrease of IL-18 (p = 0.026) and a tendency for better kidney function based on the higher total diuresis, higher glomerular filtration rate (GFR), higher potassium level, lower serum creatinine, and lower urea concentration during the seven-day postoperative observation period. The long-term beneficial impact of hypothermic machine perfusion on the outcome of transplanted kidneys may rely on the early modified proceedings and intensity of ischemia-reperfusion injury reflected by the dynamics of the concentrations of examined biomarkers.

Abstract P148: Role of Endothelin B Receptors in Neuronal Progenitor Cell Differentiation and Mitochondrial Function in Ischemic Stroke

Neuronal progenitor cells (NPCs) and mitochondria are well known to play roles in regeneration and functions of the brain, respectively. However, stimulation of endothelin-B receptors (ETBR) using an agonist, sovateltide (IRL-1620) on their fate in the brain after stroke remains elusive. We evaluated the effect of sovateltide mediated ETBR stimulation on differentiation of NPCs and fate of mitochondria after stroke. Ischemic stroke was induced by performing permanent right middle cerebral artery occlusion (MCAO) in rats. Sovateltide (5 μg/kg) or saline (equal volume) was injected intravenously; neurological and motor function evaluation was done at 24 hrs and day 7 post MCAO. Brain tissues were analyzed for NPCs differentiation and mitochondrial fate using techniques such as western blots, immunofluorescence, transmission electron microscopy and in situ PCR. In vitro hypoxia experiment was carried out to confirm sovateltide mediated neuronal differentiation. Neurological and motor functions were significantly improved in sovateltide treated rats at 24 hrs and day 7 post MCAO. Differentiation of NPCs was evident with upregulation of neuronal differentiation markers Neuro D1 (p=0.00002) as well as HuC/HuD (p=0.0037) along with neuronal marker Doublecortin (DCX) (p=0.00011) at 24 hrs post MCAO. However, significant upregulation only in HuC/HuD (p=0.043) was observed at day 7. Better preserved mitochondrial fate was observed in sovateltide rat brains with downregulation of mitochondrial fission marker, DRP1 (p&lt;0.001), increase in fusion marker, MFN2 (p&lt;0.0001) and increase in cross-sectional area x number (p&lt;0.05) as well as mitochondrial/tissue area (p&lt;0.05) at 24 hrs and day 7 post MCAO. In situ PCR analysis showed increased mitochondrial DNA (p=0.0418), indicating better mitochondrial biogenesis at day 7 post MCAO. In vitro exposure of sovateltide and hypoxia to cultured NPCs showed higher NeuroD1 and NeuN (a mature neuronal marker) expression confirming NPCs differentiation. Sovateltide mediated ETBR stimulation promotes differentiation of NPCs and mitochondrial fusion as well as biogenesis and helps in neuronal regeneration and function restoration in acute ischemic brains.