Cardiovascular diseases (CVDs) like hypertension are a major cause for death worldwide. In the cardiovascular tissue, the endothelin system-consisting of the receptor subtypes A (ETA R) and B (ETB R) and the mixed agonist endothelin 1 (ET-1)-is a major key player in the regulation of vascular tone and blood pressure. Tight control of this system is required to maintain homeostasis; otherwise, the endothelin system can cause severe CVDs like pulmonary artery hypertension. The high sequence homology between both receptor subtypes limits the development of novel and selective ligands. Identification of small differences in receptor-ligand interactions and determination of selectivity constraints are crucial to fine-tune ligand properties and subsequent signaling events.
Here, we report on novel ET-1 analogs and their detailed pharmacological characterization. We generated simplified ET-1-derived monocyclic peptides to provide an accessible synthesis route. By detailed in vitro characterization, we demonstrated that both G protein signaling and the subsequent arrestin recruitment of activated ETB R remain intact, whereas activation of the ETA R depends on the intramolecular ring size. Increasing of the intramolecular ring structure reduces activity at the ETA R and shifts the peptide toward ETB R selectivity. All ET-1 analogs displayed efficient ETB R-mediated signaling by G protein activation and arrestin 3 recruitment. Our study provides in-depth characterization of the ET-1/ETA R and ET-1/ETB R interactions, which has the potential for future development of endothelin-based drugs for CVD treatment. By identification of Lys9 for selective labeling, novel analogs for peptide-mediated shuttling by ET-1 are proposed.
<em>Endothelin</em>-<em>1</em> Decreases the Expression of Ephrin-A and B Subtypes in Cultured Rat Astrocytes through <em>ET</em> <sub>B</sub> Receptors
Ephrin family proteins are cell surface molecules that regulate several cellular functions through cell-cell interactions. During nervous tissue repair after injury, the expression of ephrin subtypes in astrocytes is altered, affecting the axonal elongation and migration of neuronal precursors. However, the mechanism regulating the expression of ephrin subtypes in astrocytes has not been investigated. Herein, we studied the effects of endothelin-1 (ET-1) on the expression of ephrin subtypes in cultured rat astrocytes. Our results showed that ET-1 (100 nM) treatment for 1-24 h reduced the expression of ephrin-A2, -A4, -B2, and -B3 mRNA and protein in astrocytes, whereas the expression of ephrin-A1, -A3, -A5, and -B1 mRNA were not affected.
Sarafotoxin S6c, a selective ETB receptor agonist, decreased the expression of ephrin-A2, -A4, -B2, and -B3 in cultured astrocytes. The decrease in ephrin-A2, -A4, -B2, and -B3 expression by ET-1 treatment was reduced in the presence of BQ788, an ETB receptor antagonist, while FR139317, an ETA receptor antagonist, had no effects. These results suggest that ET-1 is a signaling molecule that downregulates ephrin-A2, -A4, -B2, and -B3 expression in astrocytes.
C-type natriuretic peptide (CNP)/Guanylate cyclase B (GC-B) system and <em>endothelin</em>-<em>1</em>(<em>ET</em>-<em>1</em>)/<em>ET</em> receptor A and B system in human vasculature.
In order to assess the physiological and clinical implications of C-type natriuretic peptide (CNP)/guanylyl cyclase B (GC-B) system in the human vasculature, we have examined gene expressions of CNP and its receptor, GC-B, in human vascular endothelial cells (ECs) and smooth muscle cells (SMCs) and have also compared endothelin-1(ET-1)/endothelin receptor-A (ETR-A) and endothelin receptor-B (ETR-B) system in human aortic ECs (HAECs) and vascular SMCs (HSMCs) in vitro.
We also examined these gene expressions in human embryonic stem (ES)/induced pluripotent stem cell (iPS)-derived ECs and mural cells (MCs). A little but significant amount of mRNA encoding CNP was detected in both human ES-derived ECs and HAECs. Substantial amount of GC-B was expressed in both ECs (iPS-derived ECs and HAECs) and SMCs (iPS-derived MCs and HSMCs). ET-1 was expressed solely in ECs. ETR-A was expressed in SMCs, while ETR-B was expressed in ECs. These results indicate the existence of vascular CNP/GC-B system in the human vascular wall, indicating the evidence for clinical implication of CNP/GC-B system in concert with ET-1/ETR-A and ETR-B system in the human vasculature.
Downregulation of endothelial nitric oxide synthase (eNOS) and <em>endothelin</em>-<em>1</em> (<em>ET</em>-<em>1</em>) in a co-culture system with human stimulated X-linked CGD neutrophils.
Phagocytes in patients with chronic granulomatous disease (CGD) do not generate reactive oxidative species (ROS), whereas nitric oxide (NO) production is increased in response to the calcium ionophore A23187 in CGD phagocytes compared with healthy phagocytes. Recently, patients with X-linked CGD (X-CGD) have been reported to show higher flow-mediated dilation, suggesting that endothelial cell function is affected by NO production from phagocytes. We studied NOS3 and EDN1 mRNA and protein expression in human umbilical vein endothelial cells (HUVECs) in a co-culture system with neutrophils from X-CGD patients. HUVECs were co-cultured for 30 minutes with human neutrophils from X-CGD or healthy participants in response to A23187 without cell-to-cell contact.
The expression of NOS3 and EDN1 mRNA in HUVECs was quantified by real-time polymerase chain reaction. Moreover, we demonstrated the protein expression of eNOS, ET-1, and NFκB p65, including phosphorylation at Ser1177 of eNOS and Ser536 of NFκB p65. Neutrophils from X-CGD patients showed significantly higher NO and lower H2O2 production in response to A23187 than healthy neutrophils in vitro. Compared with healthy neutrophils, X-CGD neutrophils under A23187 stimulation exhibited significantly increased NO and decreased H2O2, and promoted downregulated NOS3 and EDN1 expression in HUVECs. The total expression and phosphorylation at Ser1177 of eNOS and ET-1 expression were significantly decreased in HUVECs co-cultures with stimulated X-CGD neutrophils.
Also, phosphorylation at Ser536 of NFκB p65 were significantly decreased. In conclusions, eNOS and ET-1 significantly down-regulated in co-culture with stimulated X-CGD neutrophils through their excessive NO and the lack of ROS production. These findings suggest that ROS generated from neutrophils may mediate arterial tone affecting eNOS and ET-1 expression via their NO and ROS production.
Endothelin 1 (ET-1) Antibody |
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abx024086-100ug | Abbexa | 100 ug | 1128 EUR |
Endothelin-1 (ET-1), big (Rat) |
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023-32 | PHOENIX PEPTIDE | 100 μg | 206.28 EUR |
ET-1(Endothelin-1) ELISA Kit |
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EKF60092-48T | Biomatik Corporation | 48T | 396.9 EUR |
ET-1(Endothelin-1) ELISA Kit |
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EKF60092-5x96T | Biomatik Corporation | 5x96T | 2693.25 EUR |
ET-1(Endothelin-1) ELISA Kit |
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EKF60092-96T | Biomatik Corporation | 96T | 567 EUR |
ET-1(Endothelin-1) ELISA Kit |
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EU0205 | FN Test | 96T | 628.92 EUR |
Endothelin-1 (ET-1), big (Human) |
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023-10 | PHOENIX PEPTIDE | 100 μg | 154.44 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS606359-01mg | MyBiosource | 0.1mg | 580 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS606359-5x01mg | MyBiosource | 5x0.1mg | 2470 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6248913-01mL | MyBiosource | 0.1(mL | 865 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6248913-5x01mL | MyBiosource | 5x0.1mL | 3750 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6249840-01mL | MyBiosource | 0.1(mL | 865 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6249840-5x01mL | MyBiosource | 5x0.1mL | 3750 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6250834-01mL | MyBiosource | 0.1(mL | 865 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6250834-5x01mL | MyBiosource | 5x0.1mL | 3750 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6251787-01mL | MyBiosource | 0.1(mL | 865 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6251787-5x01mL | MyBiosource | 5x0.1mL | 3750 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6247892-01mL | MyBiosource | 0.1(mL | 865 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6247892-5x01mL | MyBiosource | 5x0.1mL | 3750 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6253840-01mL | MyBiosource | 0.1(mL | 865 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6253840-5x01mL | MyBiosource | 5x0.1mL | 3750 EUR |
Endothelin, pan (Endothelin 1, Endothelin-1, EDN1, ET1, ET-1, Endothelin 2, Endothelin-2, EDN2, ET2, ET-2, Endothelin 3, Endothelin-3, EDN3, ET3, ET-3, HDLCQ7, MGC15067, MGC61498, Preproendothelin-1, PPET1, Preproendothelin-2, PPET2, Preproendothelin-3, P |
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MBS6254880-01mL | MyBiosource | 0.1(mL | 865 EUR |
<em>Endothelin</em>-<em>1</em> (<em>ET</em>-<em>1</em>) Promotes a Proinflammatory Microglia Phenotype in Diabetic Conditions.
Diabetes increases the risk and severity of cognitive impairment, especially after ischemic stroke. It is also known that the activation of ET system is associated with cognitive impairment and microglia around periinfarct area produce ET-1. However, little is known about the effect of ET-1 on microglial polarization, especially under diabetic conditions. We hypothesized that a) ET-1 activates microglia to proinflammatory M-1 like phenotype, and b) Hypoxia/LPS activates the microglial ET system and promotes microglial activation towards M-1 phenotype in diabetic conditions.
Microglial cells (C8B4) cultured under normal glucose (25mM) and diabetes-mimicking high glucose (50mM) conditions for 48 hours were stimulated with ET-1, cobalt chloride (200µM) or lipopolysaccharide (100ng/ml) for 24 hours. PPET-1, ET receptor subtypes and M1/M2 marker genes mRNA expression measured by RT-PCR. Secreted ET-1 was measured by ELISA. High dose of ET-1 (1µM), increases the mRNA levels of ET receptors and activates the microglia towards M1 phenotype. Hypoxia or LPS activates the ET system in microglial cells and shifts the microglia towards M1 phenotype in diabetic conditions. These in vitro observations warrant further investigation into the role of ET-1-mediated activation of proinflammatory microglia in post-stroke cognitive impairment in diabetes.