Abstract 66: Endothelial α5β1 Integrin is Acutely Upregulated and Destabilizes the Blood-brain Barrier After Stroke

Stroke ◽  
2016 ◽  
Vol 47 (suppl_1) ◽  
Author(s):  
Danielle Edwards ◽  
Jill Roberts ◽  
Leon de Hoog ◽  
Greg J Bix
Keyword(s):  
Ischemic Stroke ◽  
Endothelial Cell ◽  
Brain Injury ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Fibronectin Receptor ◽  
Post Stroke ◽  
Blood Brain ◽  
Α5β1 Integrin

Repeated opening of the blood-brain barrier (BBB) after ischemic stroke significantly contributes to brain injury. As the β1 integrin receptor family has been linked to changes in BBB permeability via changes to tight junction (TJ) protein expression and function, we hypothesized that the pro-angiogenic fibronectin receptor α5β1 integrin is acutely upregulated in cerebrovasculature after ischemic stroke and contributes to BBB breakdown via affects on the TJ protein claudin-5. First, α5β1 brain immunohistochemistry after transient middle cerebral artery occlusion (MCAo) in mice demonstrated an increase in core infarct and peri-infarct vascular expression of α5β1 as early as 24 hours after stroke that became even more prominent by post stroke day 2. Next, in vitro studies demonstrated an increase in barrier permeability where stroke was simulated using oxygen-glucose deprivation or TNF-α treatment, as measured by trans-endothelial cell electrical resistance and FITC-dextran permeability, that correlated with an increase in α5β1 expression and changes in claudin-5 expression and localization away from the cell-surface (i.e. away from TJ complexes). Importantly, this increased permeability could be completely prevented by addition of the α5β1 inhibitor ATN-161. Finally, MCAo performed in endothelial cell selective α5β1 knockout mice demonstrated significantly increased post-stroke BBB stability, stable claudin-5 expression, and little to no ischemic injury. Collectively, our results demonstrate that endothelial cell α5β1 integrin expression is increased acutely after stroke, may contribute to BBB breakdown and subsequent expansion of brain injury, and therefore could represent a novel stroke therapeutic target worthy of further investigation.

scholarly journals In vitro models of the blood–brain barrier: An overview of commonly used brain endothelial cell culture models and guidelines for their use

2016 ◽  
Vol 36 (5) ◽  
pp. 862-890 ◽  
Author(s):  
Hans C Helms ◽  
N Joan Abbott ◽  
Malgorzata Burek ◽  
Romeo Cecchelli ◽  
Pierre-Olivier Couraud ◽  
...  
Keyword(s):  
Cell Culture ◽  
Endothelial Cell ◽  
Blood Brain Barrier ◽  
Brain Parenchyma ◽  
Brain Barrier ◽  
Blood Brain ◽  
Culture Models ◽  
Cell Culture Models ◽  
The Brain

The endothelial cells lining the brain capillaries separate the blood from the brain parenchyma. The endothelial monolayer of the brain capillaries serves both as a crucial interface for exchange of nutrients, gases, and metabolites between blood and brain, and as a barrier for neurotoxic components of plasma and xenobiotics. This “blood-brain barrier” function is a major hindrance for drug uptake into the brain parenchyma. Cell culture models, based on either primary cells or immortalized brain endothelial cell lines, have been developed, in order to facilitate in vitro studies of drug transport to the brain and studies of endothelial cell biology and pathophysiology. In this review, we aim to give an overview of established in vitro blood–brain barrier models with a focus on their validation regarding a set of well-established blood–brain barrier characteristics. As an ideal cell culture model of the blood–brain barrier is yet to be developed, we also aim to give an overview of the advantages and drawbacks of the different models described.

scholarly journals CoQ10 Deficient Endothelial Cell Culture Model for the Investigation of CoQ10 Blood–Brain Barrier Transport

2020 ◽  
Vol 9 (10) ◽  
pp. 3236
Author(s):  
Luke Wainwright ◽  
Iain P. Hargreaves ◽  
Ana R. Georgian ◽  
Charles Turner ◽  
R. Neil Dalton ◽  
...  
Keyword(s):  
Cell Culture ◽  
Endothelial Cell ◽  
Blood Brain Barrier ◽  
Advanced Glycation Endproducts ◽  
Brain Barrier ◽  
High Dose ◽  
Blood Brain ◽  
Coq10 Deficiency ◽  
The Brain

Primary coenzyme Q10 (CoQ10) deficiency is unique among mitochondrial respiratory chain disorders in that it is potentially treatable if high-dose CoQ10 supplements are given in the early stages of the disease. While supplements improve peripheral abnormalities, neurological symptoms are only partially or temporarily ameliorated. The reasons for this refractory response to CoQ10 supplementation are unclear, however, a contributory factor may be the poor transfer of CoQ10 across the blood–brain barrier (BBB). The aim of this study was to investigate mechanisms of CoQ10 transport across the BBB, using normal and pathophysiological (CoQ10 deficient) cell culture models. The study identifies lipoprotein-associated CoQ10 transcytosis in both directions across the in vitro BBB. Uptake via SR-B1 (Scavenger Receptor) and RAGE (Receptor for Advanced Glycation Endproducts), is matched by efflux via LDLR (Low Density Lipoprotein Receptor) transporters, resulting in no “net” transport across the BBB. In the CoQ10 deficient model, BBB tight junctions were disrupted and CoQ10 “net” transport to the brain side increased. The addition of anti-oxidants did not improve CoQ10 uptake to the brain side. This study is the first to generate in vitro BBB endothelial cell models of CoQ10 deficiency, and the first to identify lipoprotein-associated uptake and efflux mechanisms regulating CoQ10 distribution across the BBB. The results imply that the uptake of exogenous CoQ10 into the brain might be improved by the administration of LDLR inhibitors, or by interventions to stimulate luminal activity of SR-B1 transporters.

scholarly journals MicroRNA-30a regulates acute cerebral ischemia-induced blood–brain barrier damage through ZnT4/zinc pathway

2020 ◽  
pp. 0271678X2092678 ◽  
Author(s):  
Peng Wang ◽  
Rong Pan ◽  
John Weaver ◽  
Mengjie Jia ◽  
Xue Yang ◽  
...  
Keyword(s):  
Ischemic Stroke ◽  
Cerebral Ischemia ◽  
Tight Junction ◽  
Blood Brain Barrier ◽  
Brain Barrier ◽  
Tight Junction Proteins ◽  
Blood Brain ◽  
Acute Cerebral Ischemia ◽  
Junction Proteins

The mechanism of early blood–brain barrier (BBB) disruption after stroke has been intensively studied but still not fully understood. Here, we report that microRNA-30a (miR-30a) could mediate BBB damage using both cellular and animal models of ischemic stroke. In the experiments in vitro, inhibition of miR-30a decreased BBB permeability, prevented the degradation of tight junction proteins, and reduced intracellular free zinc in endothelial cells. We found that the zinc transporter ZnT4 was a direct target of negative regulation by miR-30a, and ZnT4/zinc signaling pathway contributed significantly to miR-30a-mediated BBB damage. Consistent with these in vitro findings, treatment with miR-30a inhibitor reduced zinc accumulation, increased the expression of ZnT4, and prevented the loss of tight junction proteins in microvessels of ischemic animals. Furthermore, inhibition of miR-30a, even at 90 min post onset of middle cerebral artery occlusion, prevented BBB damage, reduced infarct volume, and ameliorated neurological deficits. Together, our findings provide novel insights into the mechanisms of cerebral ischemia-induced BBB disruption and indicate miR-30a as a regulator of BBB function that can be an effective therapeutic target for ischemic stroke.

Immortalized endothelial cell lines for in vitro blood–brain barrier models: A systematic review

2016 ◽  
Vol 1642 ◽  
pp. 532-545 ◽  
Author(s):  
Nurul Adhwa Rahman ◽  
Alifah Nur’ain Haji Mat Rasil ◽  
Uta Meyding-Lamade ◽  
Eva Maria Craemer ◽  
Suwarni Diah ◽  
...  
Keyword(s):  
Systematic Review ◽  
Endothelial Cell ◽  
Blood Brain Barrier ◽  
Cell Lines ◽  
Brain Barrier ◽  
Blood Brain

Development of immortalized human cerebromicrovascular endothelial cell line as an in vitro model of the human blood–brain barrier

1997 ◽  
Vol 11 (13) ◽  
pp. 1187-1197 ◽  
Author(s):  
Arumugam Muruganandam ◽  
Leonie Moorhouse Herx ◽  
Robert Monette ◽  
Jon P. Durkin ◽  
Danica B. Stanimirovic
Keyword(s):  
Endothelial Cell ◽  
Cell Line ◽  
Blood Brain Barrier ◽  
Human Blood ◽  
In Vitro Model ◽  
Brain Barrier ◽  
Endothelial Cell Line ◽  
Blood Brain ◽  
Vitro Model

scholarly journals The effect of nanoparticle size on the probability to cross the blood-brain barrier: an in-vitro endothelial cell model

nano Online ◽  
2016 ◽  
Author(s):  
Malka Shilo ◽  
Anat Sharon ◽  
Koby Baranes ◽  
Menachem Motiei ◽  
Jean-Paul M Lellouche ◽  
...  
Keyword(s):  
Endothelial Cell ◽  
Blood Brain Barrier ◽  
Cell Model ◽  
Brain Barrier ◽  
Nanoparticle Size ◽  
Blood Brain

scholarly journals A Novel lncRNA ENST00000530525 Affects ANO1 Contributing to Blood-Brain Barrier Injury in Cultured hCMEC/D3 Cells Under OGD/R Conditions.

2021 ◽  
Author(s):  
Wen Jiang ◽  
Jie Li ◽  
Yuefang Cai ◽  
Wenchen Liu ◽  
Mei Chen ◽  
...  
Keyword(s):  
Ischemic Stroke ◽  
Cell Viability ◽  
Blood Brain Barrier ◽  
Differentially Expressed ◽  
Brain Barrier ◽  
Cell Counting ◽  
Blood Samples ◽  
Post Stroke ◽  
Blood Brain ◽  
Acute Cerebral Ischemia

Abstract Ischemic stroke (IS) is a major neurological disease with high fatality and residual disability burdens. Increasing amount of long noncoding RNAs (lncRNAs) have been revealed to play an important role in ischemic stroke. However, the roles and significances of most lncRNAs in ischemic stroke are still unknown.This study was performed to identify differentially expressed lncRNAs using a lncRNA microarray in whole blood samples of patients suffered from acute cerebral ischemia. Bioinformatics analyses including GO, KEGG pathway enrichment analysis, and proximity to putative stroke risk location analysis were performed. A novel lncRNA ENST00000530525 significantly decreased after ischemic stroke. Furthermore, we evaluated lncRNA ENST00000530525 expression in cultured hCMEC/D3 cells under oxygen-glucose deprivation/reoxygenation(OGD/R) conditions using fluorescent in situ hybridization (FISH) and quantitative real-time polymerase chain reaction (RT-qPCR) analysis. To investigate the function of lncRNA ENST00000530525, the plasmid of overexpression(OE) and negative control(NC) were transfected into hCMEC/D3, then cell viability was detected by cell counting kit-8 (CCK-8) Assay after OGD/R,lncRNA ENST00000530525 and ANO1 expression were investigated using RT-qPCR and Immunofluorescence. For blood-brain barrier(BBB) permeability,FITC-dextran transendothelial permeability assay and Tight junction(Tj) protein was detected.There were 3352 differentially expressed lncRNAs in blood samples of acute ischemic stroke patients. The validation results were consistent with gene chip data.GO and KEGG results showed these lncRNAs were mainly related to oxygen and glucose metabolism, leukocyte transendothelial migration,mitophagy and cellular senescence.Among these, lncRNA ENST00000530525 was the highly down-regulated lncRNA and mapped within the ischemic stroke associated gene anoctamin-1 (ANO1). We furtherly found lncRNA ENST00000530525 was down-regulated in hCMEC/D3 cells under 4h OGD and 20h reoxygenation(OGD4/R20) conditions. Up-regulating lncRNA ENST00000530525 decreased the cell viability while increased ANO1 expression and contributed to BBB injury of hCMEC/D3 cells after OGD4/R20.The lncRNA ENST00000530525 might plays deleterious roles in post-stroke pathogenesis. The results show light on some differentially expressed lncRNAs in human certainly participate through characteristic roles in post-stroke pathogenesis, thus, the roles and significances of some novel lncRNAs in ischemic stroke thereby warranting further study.

scholarly journals Yuzu and Hesperidin Ameliorate Blood-Brain Barrier Disruption during Hypoxia via Antioxidant Activity

Antioxidants ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 843
Author(s):  
Bo Kyung Lee ◽  
Soo-Wang Hyun ◽  
Yi-Sook Jung
Keyword(s):  
Endothelial Cell ◽  
Blood Brain Barrier ◽  
Cell Model ◽  
Antioxidant Properties ◽  
Brain Barrier ◽  
Blood Brain ◽  
In Vivo Animal Model ◽  
Bbb Permeability

Yuzu and its main component, hesperidin (HSP), have several health benefits owing to their anti-inflammatory and antioxidant properties. We examined the effects of yuzu and HSP on blood–brain barrier (BBB) dysfunction during ischemia/hypoxia in an in vivo animal model and an in vitro BBB endothelial cell model, and also investigated the underlying mechanisms. In an in vitro BBB endothelial cell model, BBB permeability was determined by measurement of Evans blue extravasation in vivo and in vitro. The expression of tight junction proteins, such as claudin-5 and zonula occludens-1 (ZO-1), was detected by immunochemistry and western blotting, and the reactive oxygen species (ROS) level was measured by 2′7′-dichlorofluorescein diacetate intensity. Yuzu and HSP significantly ameliorated the increase in BBB permeability and the disruption of claudin-5 and ZO-1 in both in vivo and in vitro models. In bEnd.3 cells, yuzu and HSP were shown to inhibit the disruption of claudin-5 and ZO-1 during hypoxia, and the protective effects of yuzu and HSP on claudin-5 degradation seemed to be mediated by Forkhead box O 3a (FoxO3a) and matrix metalloproteinase (MMP)-3/9. In addition, well-known antioxidants, trolox and N-acetyl cysteine, significantly attenuated the BBB permeability increase, disruption of claudin-5 and ZO-1, and FoxO3a activation during hypoxia, suggesting that ROS are important mediators of BBB dysfunction during hypoxia. Collectively, these results indicate that yuzu and HSP protect the BBB against dysfunction via maintaining integrity of claudin-5 and ZO-1, and these effects of yuzu and HSP appear to be a facet of their antioxidant properties. Our findings may contribute to therapeutic strategies for BBB-associated neurodegenerative diseases.

Sign in / Sign up

Export Citation Format

Share Document