scholarly journals The SWELL1-LRRC8 complex regulates endothelial AKT-eNOS signaling and vascular function

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Ahmad F Alghanem ◽  
Javier Abello ◽  
Joshua M Maurer ◽  
Ashutosh Kumar ◽  
Chau My Ta ◽  
...  
Keyword(s):  
Blood Flow ◽  
Shear Flow ◽  
Vascular Function ◽  
Umbilical Vein ◽  
Anion Channel ◽  
Retinal Blood Flow ◽  
Cell Alignment ◽  
Signaling Complex ◽  
Flow Stimulation ◽  
Umbilical Vein Endothelial Cells

The endothelium responds to numerous chemical and mechanical factors in regulating vascular tone, blood pressure, and blood flow. The endothelial volume-regulated anion channel (VRAC) has been proposed to be mechanosensitive and thereby sense fluid flow and hydrostatic pressure to regulate vascular function. Here, we show that the leucine-rich repeat-containing protein 8a, LRRC8A (SWELL1), is required for VRAC in human umbilical vein endothelial cells (HUVECs). Endothelial LRRC8A regulates AKT-endothelial nitric oxide synthase (eNOS) signaling under basal, stretch, and shear-flow stimulation, forms a GRB2-Cav1-eNOS signaling complex, and is required for endothelial cell alignment to laminar shear flow. Endothelium-restricted Lrrc8a KO mice develop hypertension in response to chronic angiotensin-II infusion and exhibit impaired retinal blood flow with both diffuse and focal blood vessel narrowing in the setting of type 2 diabetes (T2D). These data demonstrate that LRRC8A regulates AKT-eNOS in endothelium and is required for maintaining vascular function, particularly in the setting of T2D.

scholarly journals The SWELL1-LRRC8 complex regulates endothelial AKT-eNOS-mTOR signaling and vascular function

2020 ◽  
Author(s):  
Ahmad F. Alghanem ◽  
Chau Ta ◽  
Joshua M. Maurer ◽  
Susheel K. Gunasekar ◽  
Ashutosh Kumar ◽  
...  
Keyword(s):  
Blood Flow ◽  
Vascular Function ◽  
Vascular Reactivity ◽  
Ex Vivo ◽  
Umbilical Vein ◽  
Anion Channel ◽  
Aortic Ring ◽  
Tube Formation ◽  
Mtor Signaling ◽  
Retinal Blood Flow

AbstractThe endothelium responds to a multitude of chemical and mechanical factors in regulating vascular tone, angiogenesis, blood pressure and blood flow. The endothelial volume regulatory anion channel (VRAC) has been proposed to be mechano-sensitive, to activate in response to fluid flow/hydrostatic pressure and putatively regulate vascular reactivity and angiogenesis. Here, we show that the Leucine Rich Repeat Containing Protein 8a, LRRC8a (SWELL1) functionally encodes VRAC in human umbilical vein endothelial cells (HUVECs). Endothelial SWELL1 (SWELL1) expression positively regulates AKT-eNOS signaling while negatively regulating mTOR signaling, via a SWELL1-GRB2-Cav1-eNOS signaling complex. Endothelium-restricted SWELL1 KO (SWELL1 KO) mice exhibit enhanced tube formation from ex-vivo aortic ring explants in matrigel angiogenesis assays, develop hypertension in response to chronic angiotensin II infusion and have impaired retinal blood flow with both diffuse and focal blood vessel narrowing in the setting of Type 2 diabetes (T2D). These data demonstrate that SWELL1 antithetically regulates AKT-eNOS and mTOR signaling in endothelium and is required for maintaining vascular function, particularly in the setting of T2D.

scholarly journals Metformin prevented high glucose-induced endothelial reactive oxygen species via OGG1 in an AMPKα-Lin-28 dependent pathway

2020 ◽  
Author(s):  
Liangliang Tao ◽  
Xiucai Fan ◽  
Jing Sun ◽  
Zhu zhang
Keyword(s):  
High Glucose ◽  
Vascular Function ◽  
Umbilical Vein ◽  
Vascular Complications ◽  
Diabetic Patients ◽  
Type 2 Diabetic Patients ◽  
Umbilical Vein Endothelial Cells ◽  
Dependent Pathway

AbstractMetformin improved vascular function in obese type 2 diabetic patients. 8-oxoguanine glycosylase (OGG1), a main DNA glycosylase, was involved in vascular complications in diverse diseases. However, whether metformin suppressed endothelial ROS via OGG1 pathway was unclear. Human umbilical vein endothelial cells (HUVECs) were exposed to HG (high glucose) or HG with metformin. OGG1 and AMPfα levels were measured after metformin treatment while HG-caused ROS was measured by DHE prober. Diabetic mice were induced by daily intraperitoneal injections of streptozotocin (STZ). Metformin reduced Endothelial ROS caused by HG via upregulating OGG1. Additionally, OGG1 protein expression was dependent on its mRNA stability, which was reversed by genetic inhibition of AMPKα and Lin-28. The role of OGG1 on ROS stimulated by HG was partially dependent on NFKB/NOX4 pathway in HUVECs. These results suggested that metformin contacted HG-induced endothelial ROS via AMPKα/Lin-28/OGG1 pathway.

scholarly journals Longitudinal stability of retinal blood flow regulation in response to flicker stimulation and systemic hyperoxia in mice assessed with laser speckle flowgraphy

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Junya Hanaguri ◽  
Harumasa Yokota ◽  
Masahisa Watanabe ◽  
Lih Kuo ◽  
Satoru Yamagami ◽  
...  
Keyword(s):  
Blood Flow ◽  
Vascular Function ◽  
Systemic Blood Pressure ◽  
Laser Speckle ◽  
Retinal Blood Flow ◽  
Laser Speckle Flowgraphy ◽  
Adult Mice ◽  
Ocular Perfusion ◽  
Vascular Area ◽  
Flicker Stimulation

AbstractThis study aimed to evaluate longitudinal changes in retinal blood flow in response to flicker stimulation and systemic hyperoxia in mice using a laser speckle flowgraphy (LSFG-Micro). The retinal blood flow in vascular area surrounding the optic nerve head was measured in 8-week-old male mice every 2 weeks until age 20-week. The coefficient of variation of retinal blood flow under resting condition was analyzed every 2 weeks to validate the consistency of the measurement. On day 1 of the experiment, retinal blood flow was assessed every 20 s for 6 min during and after 3 min flicker light (12 Hz) stimulation; on day 2, retinal blood flow was measured every minute for 20 min during and after 10 min systemic hyperoxia; and on day 3, electroretinography (ERG) was performed. Body weight, systemic blood pressure, and ocular perfusion pressure increased significantly with age, but the resting retinal blood flow and ERG parameters remained unchanged. Retinal blood flow significantly increased with flicker stimulation and decreased with systemic hyperoxia, independent of age. The LSFG-Micro provides consistent and reproducible retinal blood flow measurement in adult mice. Longitudinal assessments of retinal blood flow in response to flicker stimulation and systemic hyperoxia may be useful indexes for noninvasive monitoring of vascular function in retinas.

scholarly journals Therapeutic Angiogenesis by a “Dynamic Duo”: Simultaneous Expression of HGF and VEGF165 by Novel Bicistronic Plasmid Restores Blood Flow in Ischemic Skeletal Muscle

Pharmaceutics ◽  
2020 ◽  
Vol 12 (12) ◽  
pp. 1231
Author(s):  
Ekaterina Slobodkina ◽  
Maria Boldyreva ◽  
Maxim Karagyaur ◽  
Roman Eremichev ◽  
Natalia Alexandrushkina ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Blood Flow ◽  
Growth Factor ◽  
Umbilical Vein ◽  
Limb Ischemia ◽  
Bidirectional Promoter ◽  
Capillary Density ◽  
Therapeutic Angiogenesis ◽  
Angiogenic Growth Factors ◽  
Umbilical Vein Endothelial Cells

Therapeutic angiogenesis is a promising strategy for relief of ischemic conditions, and gene delivery was used to stimulate blood vessels’ formation and growth. We have previously shown that intramuscular injection of a mixture containing plasmids encoding vascular endothelial growth factor (VEGF)165 and hepatocyte growth factor (HGF) leads to restoration of blood flow in mouse ischemic limb, and efficacy of combined delivery was superior to each plasmid administered alone. In this work, we evaluated different approaches for co-expression of HGF and VEGF165 genes in a panel of candidate plasmid DNAs (pDNAs) with internal ribosome entry sites (IRESs), a bidirectional promoter or two independent promoters for each gene of interest. Studies in HEK293T culture showed that all plasmids provided synthesis of HGF and VEGF165 proteins and stimulated capillary formation by human umbilical vein endothelial cells (HUVEC), indicating the biological potency of expressed factors. Tests in skeletal muscle explants showed a dramatic difference and most plasmids failed to express HGF and VEGF165 in a significant quantity. However, a bicistronic plasmid with two independent promoters (cytomegalovirus (CMV) for HGF and chicken b-actin (CAG) for VEGF165) provided expression of both grow factors in skeletal muscle at an equimolar ratio. Efficacy tests of bicistronic plasmid were performed in a mouse model of hind limb ischemia. Intramuscular administration of plasmid induced significant restoration of perfusion compared to an empty vector and saline. These findings were supported by increased CD31+ capillary density in animals that received pHGF/VEGF. Overall, our study reports a first-in-class candidate gene therapy drug to deliver two pivotal angiogenic growth factors (HGF and VEGF165) with properties that provide basis for future development of treatment for an unmet medical need—peripheral artery disease and associated limb ischemia.

scholarly journals Fractalkine-induced microglial vasoregulation occurs within the retina and is altered early in diabetic retinopathy

2021 ◽  
Vol 118 (51) ◽  
pp. e2112561118
Author(s):  
Samuel A. Mills ◽  
Andrew I. Jobling ◽  
Michael A. Dixon ◽  
Bang V. Bui ◽  
Kirstan A. Vessey ◽  
...  
Keyword(s):  
Blood Flow ◽  
Diabetic Retinopathy ◽  
Vascular Function ◽  
Immune Cell ◽  
Neurovascular Unit ◽  
Retinal Blood Flow ◽  
Proper Function ◽  
Early Diabetes ◽  
Vascular Regulation ◽  
The Brain

Local blood flow control within the central nervous system (CNS) is critical to proper function and is dependent on coordination between neurons, glia, and blood vessels. Macroglia, such as astrocytes and Müller cells, contribute to this neurovascular unit within the brain and retina, respectively. This study explored the role of microglia, the innate immune cell of the CNS, in retinal vasoregulation, and highlights changes during early diabetes. Structurally, microglia were found to contact retinal capillaries and neuronal synapses. In the brain and retinal explants, the addition of fractalkine, the sole ligand for monocyte receptor Cx3cr1, resulted in capillary constriction at regions of microglial contact. This vascular regulation was dependent on microglial Cx3cr1 involvement, since genetic and pharmacological inhibition of Cx3cr1 abolished fractalkine-induced constriction. Analysis of the microglial transcriptome identified several vasoactive genes, including angiotensinogen, a constituent of the renin-angiotensin system (RAS). Subsequent functional analysis showed that RAS blockade via candesartan abolished microglial-induced capillary constriction. Microglial regulation was explored in a rat streptozotocin (STZ) model of diabetic retinopathy. Retinal blood flow was reduced after 4 wk due to reduced capillary diameter and this was coincident with increased microglial association. Functional assessment showed loss of microglial–capillary response in STZ-treated animals and transcriptome analysis showed evidence of RAS pathway dysregulation in microglia. While candesartan treatment reversed capillary constriction in STZ-treated animals, blood flow remained decreased likely due to dilation of larger vessels. This work shows microglia actively participate in the neurovascular unit, with aberrant microglial–vascular function possibly contributing to the early vascular compromise during diabetic retinopathy.

scholarly journals P2Y2 receptor modulates shear stress-induced cell alignment and actin stress fibers in human umbilical vein endothelial cells

2016 ◽  
Vol 74 (4) ◽  
pp. 731-746 ◽  
Author(s):  
Ramasri Sathanoori ◽  
Paulina Bryl-Gorecka ◽  
Christa E. Müller ◽  
Laurie Erb ◽  
Gary A. Weisman ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Umbilical Vein ◽  
Stress Fibers ◽  
Cell Alignment ◽  
P2y2 Receptor ◽  
Human Umbilical Vein ◽  
Umbilical Vein Endothelial Cells ◽  
Actin Stress Fibers

ERYTHROCYTES(RBC) AS SUICIDAL ENDOGENOUS SCAVENGERS IN IMMUNE TRIGGERED GRANULOCYTE(PMN)MEDIATED VASCULAR DAMAGE

1987 ◽  
Author(s):  
M A Boogaerts ◽  
P Zachée ◽  
M P Emonds ◽  
W Goossens ◽  
R L Verwilghen ◽  
...  
Keyword(s):  
Umbilical Vein ◽  
Anion Channel ◽  
Hemodialysis Patients ◽  
Endothelial Damage ◽  
Sodium Ascorbate ◽  
High Dose ◽  
Chronic Anemia ◽  
Umbilical Vein Endothelial Cells ◽  
Clinical Conditions

PMN produced toxic oxygen radicals(TOR)have been implicated in the generation of endothelial injury in a number of clinical conditions e.g. in apheresis,hemodialysis, ARDS and atherosclerosis. RBC have shown to inhibit TOR induced damage in a number of hyper-oxic lung injury models.We surmised RBC may serve as endogenous TOR scavengers in those in vivo situations where PMN are immunologically triggered(e.g. complement-activation in hemodialysis)to produce endothelial damage.However, RBC in their role as scavengers may became more vulnerable to further oxydant stress and display a reduced life span.Confluent monolayers of 51cr-labeled human umbilical vein endothelial cells will release 7.5 ± 1.3% of their label upon incubation with complement triggered PMN.When these PMN (l)are premixed with RBC(10),the endothelial damage can be inhibited by 78.4 ± 3.2%.This inhibition can be reproduced by replacing intact RBC by their hemolysates,but not by red cell ghosts.In a 51cr-RBC cytotoxicity system,phorbolester stimulated PMN will lyse 52.6 ± 4.2% RBC.Addition of unlabeled RBC(1/5),inhibits cytotoxicity by 31.1%,their hemolysate by 100%.Pretreatment of added unlabeled RBC with the anion channel blocker DIDS,did not significantly block the scavenger effect.RBC-targets fron hemodialysis-patients(n=8),are more vulnerable to PMN mediated cytotoxicity than normal controls(+24.1%,p<0.001). This vulnerability is further increased(+16.3%,p<0.05)during the early stages of the hemodialysis-procedure,at the time when granulocyte counts are lowest and TOR-generation highest.The GSH of he-modialysis-RBC can be more rapidly depleted upon challenge byAPH, while they also display a significant higher methemoglobin production upon sodium ascorbate challenge,both indicative of their, increased oxidative sensitivity. High dose Vitamin C(10™3M)or desfe-rioxamine(10™3M)inhibit all RBC cytotoxicity.We conclude that RBC serve as endogenous scavengers of TOR,generated by PMN,triggered by corplement activation during hemodialysis. However,by doing so,they became themselves more vulnerable to further oxidative stress,which may contribute to the chronic anemia of hemodialysis-patients.

scholarly journals Tetragonia tetragonoides (Pall.) Kuntze Restores Blood Perfusion from Hind-Limb Ischemic Mice

2020 ◽  
Vol 10 (23) ◽  
pp. 8562
Author(s):  
Hyun Yang ◽  
Dong Ho Jung ◽  
Hye Won Lee ◽  
Dongoh Lee ◽  
Byoung Seob Ko
Keyword(s):  
Blood Flow ◽  
Mouse Model ◽  
Hind Limb ◽  
Umbilical Vein ◽  
Blood Perfusion ◽  
Blood Flow Rate ◽  
Capillary Density ◽  
Peripheral Circulation ◽  
Tube Formation ◽  
Umbilical Vein Endothelial Cells

Tetragonia tetragonoides (Pall.) Kuntze (TTK) is grown for the edible leaves, and can be used as food. And which commonly called Beonhaengcho in Republic of Korea. TTK is found along the seaside of the Jeju-Island and it has long been consumed as a food for women’s health. We investigated the effects of TTK on peripheral circulation disorder during menopausal transition and/or menopause in a hind-limb ischemic (HLI) mouse model. Chemotactic motility and tube formation of vascular epithelial cells were evaluated in human umbilical vein endothelial cells (HUVECs). Female C57BL/6 mice were fed a TTK (150 or 450 mg/kg/day) for four weeks and the rate of blood flow was assessed using a laser Doppler after HLI. TTK treatment significantly increased cell migration and the branch interval value of tubular structure in a dose-dependently. In the TTK treatment group, blood flow rate was significant induced at 7, 14, and 28 days after HLI, compared with the vehicle. TTK treatment also an increase in capillary density, and the highest levels of pERK(1/2), pAkt, pPLCγ1 and pFAK proteins compared to the vehicle control. These results suggest that extract of TTK may ameliorate the blood flow via improvement of peripheral angiogenesis under hind-limb ischemic stress in a menopausal mouse model.

scholarly journals KLF2 Is a Novel Transcriptional Regulator of Endothelial Proinflammatory Activation

2004 ◽  
Vol 199 (10) ◽  
pp. 1305-1315 ◽  
Author(s):  
Sucharita SenBanerjee ◽  
Zhiyong Lin ◽  
G. Brandon Atkins ◽  
Daniel M. Greif ◽  
Ravi M. Rao ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Adhesion Molecule ◽  
Proinflammatory Cytokines ◽  
Vascular Function ◽  
Cell Attachment ◽  
Umbilical Vein ◽  
Laminar Shear Stress ◽  
Endothelial Adhesion Molecule ◽  
Umbilical Vein Endothelial Cells

The vascular endothelium is a critical regulator of vascular function. Diverse stimuli such as proinflammatory cytokines and hemodynamic forces modulate endothelial phenotype and thereby impact on the development of vascular disease states. Therefore, identification of the regulatory factors that mediate the effects of these stimuli on endothelial function is of considerable interest. Transcriptional profiling studies identified the Kruppel-like factor (KLF)2 as being inhibited by the inflammatory cytokine interleukin-1β and induced by laminar shear stress in cultured human umbilical vein endothelial cells. Overexpression of KLF2 in umbilical vein endothelial cells robustly induced endothelial nitric oxide synthase expression and total enzymatic activity. In addition, KLF2 overexpression potently inhibited the induction of vascular cell adhesion molecule-1 and endothelial adhesion molecule E-selectin in response to various proinflammatory cytokines. Consistent with these observations, in vitro flow assays demonstrate that T cell attachment and rolling are markedly attenuated in endothelial monolayers transduced with KLF2. Finally, our studies implicate recruitment by KLF2 of the transcriptional coactivator cyclic AMP response element–binding protein (CBP/p300) as a unifying mechanism for these various effects. These data implicate KLF2 as a novel regulator of endothelial activation in response to proinflammatory stimuli.

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