scholarly journals Gold nanocarriers for transport of oligonucleotides across brain endothelial cells

2020 ◽  
Author(s):  
Nayab Fatima ◽  
Radka Gromnicova ◽  
Jane Loughlin ◽  
Basil Sharrack ◽  
David Male
Keyword(s):  
Gold Nanoparticles ◽  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Target Cells ◽  
Brain Endothelium ◽  
Mobility Shift ◽  
Dna Oligonucleotides ◽  
Blood Brain

AbstractTreatment of diseases that affect the CNS by gene therapy requires delivery of oligonucleotides to target cells within the brain. As the blood brain barrier prevents movement of large biomolecules, current approaches involve direct injection of the oligonucleotides, which is invasive and may have only a localised effect. The aim of this study was to investigate the potential of 2 nm galactose-coated gold nanoparticles (NP-Gal) as a delivery system of oligonucleotides across brain endothelium.DNA oligonucleotides of different types were attached to NP-Gal by the place exchange reaction and were characterised by EMSA (electrophoretic mobility shift assay). Several nanoparticle formulations were created, with single-or double-stranded (20nt or 40nt) DNA oligonucleotides, or with different amounts of DNA attached to the carriers. These nanocarriers were applied to transwell cultures of human brain endothelium in vitro (hCMEC/D3 cell-line) or to a 3D-hydrogel model of the blood-brain barrier including astrocytes. Transfer rates were measured by quantitative electron microscopy for the nanoparticles and qPCR for DNA.Despite the increase in nanoparticle size caused by attachment of oligonucleotides to the NP-Gal carrier, the rates of endocytosis and transcytosis of nanoparticles were both considerably increased when they carried an oligonucleotide cargo. Carriers with 40nt dsDNA were most efficient, accumulating in vesicles, in the cytosol and beneath the basal membrane of the endothelium. The oligonucleotide cargo remained attached to the nanocarriers during transcytosis and the transport rate across the endothelial cells was increased at least 50fold compared with free DNA. The nanoparticles entered the extracellular matrix and were taken up by the astrocytes in biologically functional amounts.Attachment of DNA confers a strong negative charge to the nanoparticles which may explain the enhanced binding to the endothelium and transcytosis by both vesicular transport and the transmembrane/cytosol pathway. These gold nanoparticles have the potential to transport therapeutic amounts of nucleic acids into the CNS.

scholarly journals Endothelin-1 as a Mediator of Endothelial Cell–Pericyte Interactions in Bovine Brain Capillaries

1997 ◽  
Vol 17 (4) ◽  
pp. 464-469 ◽  
Author(s):  
Marie-Pierre Dehouck ◽  
Paul Vigne ◽  
Gérard Torpier ◽  
Jean Philippe Breittmayer ◽  
Roméo Cecchelli ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Brain Cortex ◽  
Bovine Brain ◽  
Brain Barrier ◽  
Target Cells ◽  
Brain Capillaries ◽  
Endothelin 1 ◽  
Blood Brain

Endothelial cells and pericytes are closely associated in brain capillaries. Together with astrocytic foot processes, they form the blood–brain barrier. Capillaries were isolated from bovine brain cortex. Pure populations of endothelial cells and pericytes were isolated and cultured in vitro. Polarized monolayers of endothelial cells preferentially secreted immunoreactive endothelin-1 (Et-1) at their abluminal (brain-facing) membrane. They did not express receptors for Et-1. Pericytes expressed BQ-123-sensitive ETA receptors for endothelins as evidenced by 125I-Et-1 binding experiments. These receptors were coupled to phospholipase C as demonstrated by intracellular calcium measurements using indo-1-loaded cells. Addition of Et-1 to pericytes induced marked changes in the cell morphology that were associated with a reorganization of F-actin and intermediate filaments. It is concluded that Et-1 is a paracrine mediator at the bovine blood–brain barrier and that capillary pericytes are target cells for endothelium-derived Et-1.

Abstract 388: Transcellular Transport of HDL Bearing Gold Nanoparticles Across the Blood Brain Barrier

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Skylar T Chuang ◽  
Siobanth Cruz ◽  
Julia Stab ◽  
Sylvia Wagner ◽  
Hagen Von Briesen ◽  
...  
Keyword(s):  
Gold Nanoparticles ◽  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Aqueous Solubility ◽  
Brain Barrier ◽  
Spherical Particles ◽  
Glioblastoma Cells ◽  
Transcellular Transport ◽  
Blood Brain

The overall objective of this study was to develop a HDL-based multifunctional platform for transport and delivery of highly hydrophobic gold nanoparticles (AuNP) bearing photothermic properties across the blood brain barrier (BBB). We exploited the ability of apolipoprotein E3 (apoE3) to act as a high affinity ligand for the low-density lipoprotein receptor to gain entry into endothelial and glioblastoma cells. The issue of poor aqueous solubility of AuNP of varying diameters (3, 10, or 10 nm) was overcome by integrating them with phospholipids and apoE3, yielding reconstituted rHDL bearing AuNP (rHDL-AuNP). Transmission electron microscopy (TEM) revealed the presence of AuNP embedded in spherical particles. Incubation of human brain microvasculature endothelial cells or glioblastoma cells with rHDL-AuNP bearing unlabeled or FITC-labeled apoE3 revealed robust uptake of particles that were localized in endocytic/lysosomal vesicles. The transport of rHDL-AuNP across an in vitro BBB model developed from primary porcine endothelial cells was examined. The addition of rHDL-AuNP to the luminal side of the cells did not affect the integrity of the BBB as assessed by the localization of key tight junction markers such as occludin, claudins and ZO-1 by immunofluorescence, and, by continual measurement of the transepithelial electrical resistance by impedance spectroscopy under physiological conditions. Lastly, the appearance of fluorescein fluorescence and AuNP in the abluminal side suggested transport of rHDL-AuNP across the neurovascular junction. These findings demonstrate that rHDL bearing apoE3 acts as a detergent in solubilizing and dramatically improving the aqueous solubility of AuNP, facilitates cellular uptake and transcellular transport of rHDL-AuNP across endothelial cells. They are significant since they present rHDL bearing apoE3 as an effective platform for delivering AuNP across the BBB.

scholarly journals Intravenous injection of cyclophilin A realizes the transient and reversible opening of barrier of neural vasculature through basigin in endothelial cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Narumi Nakada-Honda ◽  
Dan Cui ◽  
Satoshi Matsuda ◽  
Eiji Ikeda
Keyword(s):  
Endothelial Cells ◽  
Single Dose ◽  
Blood Brain Barrier ◽  
Intravenous Injection ◽  
Cyclophilin A ◽  
Brain Barrier ◽  
Blood Brain ◽  
Neural Tissues ◽  
Neural Diseases

AbstractNeural vasculature forms the blood–brain barrier against the delivery of systemically administered therapeutic drugs into parenchyma of neural tissues. Therefore, procedures to open the blood–brain barrier with minimal damage to tissues would lead to the great progress in therapeutic strategy for intractable neural diseases. In this study, through analyses with mouse in vitro brain microvascular endothelial cells and in vivo neural vasculature, we demonstrate that the administration of cyclophilin A (CypA), a ligand of basigin which is expressed in barrier-forming endothelial cells, realizes the artificial opening of blood–brain barrier. Monolayers of endothelial cells lost their barrier properties through the disappearance of claudin-5, an integral tight junction molecule, from cell membranes in a transient and reversible manner. Furthermore, the intravenous injection of a single dose of CypA into mice resulted in the opening of blood–brain barrier for a certain period which enabled the enhanced delivery of systemically administered doxorubicin into the parenchyma of neural tissues. These findings that the pre-injection of a single dose of CypA realizes an artificial, transient as well as reversible opening of blood–brain barrier are considered to be a great step toward the establishment of therapeutic protocols to overcome the intractability of neural diseases.

Anthropogenic Iron Oxide Nanoparticles Induce Damage to Brain Microvascular Endothelial Cells Forming the Blood-Brain Barrier

2021 ◽  
Author(s):  
Lorena Gárate-Vélez ◽  
Claudia Escudero-Lourdes ◽  
Daniela Salado-Leza ◽  
Armando González-Sánchez ◽  
Ildemar Alvarado-Morales ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Microvascular Endothelial Cells ◽  
Brain Microvascular Endothelial Cells ◽  
Blood Brain ◽  
Fe3o4 Nps ◽  
Microvascular Endothelial ◽  
The Brain

Background: Iron nanoparticles, mainly in magnetite phase (Fe3O4 NPs), are released to the environment in areas with high traffic density and braking frequency. Fe3O4 NPs were found in postmortem human brains and are assumed to get directly into the brain through the olfactory nerve. However, these pollution-derived NPs may also translocate from the lungs to the bloodstream and then, through the blood-brain barrier (BBB), into the brain inducing oxidative and inflammatory responses that contribute to neurodegeneration. Objective: To describe the interaction and toxicity of pollution-derived Fe3O4 NPs on primary rat brain microvascular endothelial cells (rBMECs), main constituents of in vitro BBB models. Methods: Synthetic bare Fe3O4 NPs that mimic the environmental ones (miFe3O4) were synthesized by co-precipitation and characterized using complementary techniques. The rBMECs were cultured in Transwell® plates. The NPs-cell interaction was evaluated through transmission electron microscopy and standard colorimetric in vitro assays. Results: The miFe3O4 NPs, with a mean diameter of 8.45 ± 0.14 nm, presented both magnetite and maghemite phases, and showed super-paramagnetic properties. Results suggest that miFe3O4 NPs are internalized by rBMECs through endocytosis and that they are able to cross the cells monolayer. The lowest miFe3O4 NPs concentration tested induced mid cytotoxicity in terms of 1) membrane integrity (LDH release) and 2) metabolic activity (MTS transformation). Conclusion: Pollution-derived Fe3O4 NPs may interact and cross the microvascular endothelial cells forming the BBB and cause biological damage.

scholarly journals Modulation of Wnt/β-catenin signaling promotes blood-brain barrier phenotype in cultured brain endothelial cells

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Marlyn D. Laksitorini ◽  
Vinith Yathindranath ◽  
Wei Xiong ◽  
Sabine Hombach-Klonisch ◽  
Donald W. Miller
Keyword(s):  
Endothelial Cells ◽  
Wnt Signaling ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Brain Endothelial Cells ◽  
Cancer Resistance ◽  
Blood Brain ◽  
Culture Model ◽  
External Activation

AbstractWnt/β-catenin signaling is important for blood-brain barrier (BBB) development and is implicated in BBB breakdown under various pathophysiological conditions. In the present study, a comprehensive characterization of the relevant genes, transport and permeability processes influenced by both the autocrine and external activation of Wnt signaling in human brain endothelial cells was examined using hCMEC/D3 culture model. The hCMEC/D3 expressed a full complement of Wnt ligands and receptors. Preventing Wnt ligand release from hCMEC/D3 produced minimal changes in brain endothelial function, while inhibition of intrinsic/autocrine Wnt/β-catenin activity through blocking β-catenin binding to Wnt transcription factor caused more modest changes. In contrast, activation of Wnt signaling using exogenous Wnt ligand (Wnt3a) or LiCl (GSK3 inhibitor) improved the BBB phenotypes of the hCMEC/D3 culture model, resulting in reduced paracellular permeability, and increased P-glycoprotein (P-gp) and breast cancer resistance associated protein (BCRP) efflux transporter activity. Further, Wnt3a reduced plasmalemma vesicle associated protein (PLVAP) and vesicular transport activity in hCMEC/D3. Our data suggest that this in vitro model of the BBB has a more robust response to exogenous activation of Wnt/β-catenin signaling compared to autocrine activation, suggesting that BBB regulation may be more dependent on external activation of Wnt signaling within the brain microvasculature.

scholarly journals Cerebromicrovascular endothelial cells are resistant to l-glutamate

2008 ◽  
Vol 295 (4) ◽  
pp. R1099-R1108 ◽  
Author(s):  
Ferenc Domoki ◽  
Béla Kis ◽  
Tamás Gáspár ◽  
Ferenc Bari ◽  
David W. Busija
Keyword(s):  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Primary Cultures ◽  
Glucose Deprivation ◽  
Brain Barrier ◽  
Cranial Window ◽  
Blood Brain ◽  
Microvascular Cells

Cerebral microvascular endothelial cells (CMVECs) have recently been implicated as targets of excitotoxic injury by l-glutamate (l-glut) or N-methyl-d-aspartate (NMDA) in vitro. However, high levels of l-glut do not compromise the function of the blood-brain barrier in vivo. We sought to determine whether primary cultures of rat and piglet CMVECs or cerebral microvascular pericytes (CMVPCs) are indeed sensitive to l-glut or NMDA. Viability was unaffected by 8-h exposure to 1–10 mM l-glut or NMDA in CMVECs or CMVPCs isolated from both species. Furthermore, neither 1 mM l-glut nor NMDA augmented cell death induced by 12-h oxygen-glucose deprivation in rat CMVECs or by 8-h medium withdrawal in CMVPCs. Additionally, transendothelial electrical resistance of rat CMVEC-astrocyte cocultures or piglet CMVEC cultures were not compromised by up to 24-h exposure to 1 mM l-glut or NMDA. The Ca2+ ionophore calcimycin (5 μM), but not l-glut (1 mM), increased intracellular Ca2+ levels in rat CMVECs and CMVPCs assessed with fluo-4 AM fluorescence and confocal microscopy. CMVEC-dependent pial arteriolar vasodilation to hypercapnia and bradykinin was unaffected by intracarotid infusion of l-glut in anesthetized piglets by closed cranial window/intravital microscopy. We conclude that cerebral microvascular cells are insensitive and resistant to glutamatergic stimuli in accordance with their in vivo role as regulators of potentially neurotoxic amino acids across the blood-brain barrier.

scholarly journals Mechanisms of Glutamate Efflux at the Blood—Brain Barrier: Involvement of Glial Cells

2011 ◽  
Vol 32 (1) ◽  
pp. 177-189 ◽  
Author(s):  
Katayun Cohen-Kashi-Malina ◽  
Itzik Cooper ◽  
Vivian I Teichberg
Keyword(s):  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Glial Cells ◽  
Excitatory Amino Acid ◽  
Active Role ◽  
In Vitro Models ◽  
Brain Barrier ◽  
Amino Acid Transporters ◽  
Blood Brain

At high concentrations, glutamate (Glu) exerts potent neurotoxic properties, leading to irreversible brain damages found in numerous neurological disorders. The accepted notion that Glu homeostasis in brain interstitial fluid is maintained primarily through the activity of Glu transporters present on glial cells does not take into account the possible contribution of endothelial cells constituting the blood-brain barrier (BBB) to this process. Here, we present evidence for the presence of the Glu transporters, excitatory amino-acid transporters (EAATs) 1 to 3, in porcine brain endothelial cells (PBECs) and show their participation in Glu uptake into PBECs. Moreover, transport of Glu across three in vitro models of the BBB is investigated for the first time, and evidence for Glu transport across the BBB in both directions is presented. Our results provide evidence that the BBB can function in the efflux mode to selectively remove Glu, via specific transporters, from the abluminal side (brain) into the luminal compartment (blood). Furthermore, we found that glial cells lining the BBB have an active role in the efflux process by taking up Glu and releasing it, through hemichannels, anion channels, and possibly the reversal of its EAATs, in close proximity to ECs, which in turn take up Glu and release it to the blood.

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