Blood-brain barrier properties in vitro depend on composition and assembly of endogenous extracellular matrices

2016 ◽  
Vol 365 (2) ◽  
pp. 233-245 ◽  
Author(s):  
Kathrin Zobel ◽  
Uwe Hansen ◽  
Hans-Joachim Galla
Keyword(s):  
Blood Brain Barrier ◽  
Barrier Properties ◽  
Brain Barrier ◽  
Extracellular Matrices ◽  
Blood Brain

scholarly journals VLA-4 mediated adhesion of melanoma cells on the blood–brain barrier is the critical cue for melanoma cell intercalation and barrier disruption

2018 ◽  
Vol 39 (10) ◽  
pp. 1995-2010 ◽  
Author(s):  
Ana B García-Martín ◽  
Pascale Zwicky ◽  
Thomas Gruber ◽  
Christoph Matti ◽  
Federica Moalli ◽  
...  
Keyword(s):  
Brain Metastasis ◽  
Blood Brain Barrier ◽  
Barrier Properties ◽  
Melanoma Cells ◽  
Brain Barrier ◽  
Adhesive Interaction ◽  
Inflammatory Conditions ◽  
Blood Brain ◽  
In Vitro Bbb Model

Melanoma is the most aggressive skin cancer in humans. One severe complication is the formation of brain metastasis, which requires extravasation of melanoma cells across the tight blood–brain barrier (BBB). Previously, VLA-4 has been assigned a role for the adhesive interaction of melanoma cells with non-BBB endothelial cells. However, the role of melanoma VLA-4 for breaching the BBB remained unknown. In this study, we used a mouse in vitro BBB model and imaged the shear resistant arrest of melanoma cells on the BBB. Similar to effector T cells, inflammatory conditions of the BBB increased the arrest of melanoma cells followed by a unique post-arrest behavior lacking immediate crawling. However, over time, melanoma cells intercalated into the BBB and compromised its barrier properties. Most importantly, antibody ablation of VLA-4 abrogated melanoma shear resistant arrest on and intercalation into the BBB and protected the BBB from barrier breakdown. A tissue microarray established from human brain metastasis revealed that indeed a majority of 92% of all human melanoma brain metastases stained VLA-4 positive. We propose VLA-4 as a target for the inhibition of brain metastasis formation in the context of personalized medicine identifying metastasizing VLA-4 positive melanoma.

Abstract TP86: The Ischemic Blood-brain Barrier In A Dish: Use Of Patient-derived Stem Cells To Model The Response Of The Neurovascular Unit To Oxygen-glucose Deprivation Stress

Stroke ◽  
2017 ◽  
Vol 48 (suppl_1) ◽  
Author(s):  
Shyanne Page ◽  
Ronak Patel ◽  
Abraham Alahmad
Keyword(s):  
Cell Death ◽  
Ischemic Stroke ◽  
Blood Brain Barrier ◽  
Barrier Function ◽  
Neurovascular Unit ◽  
Barrier Properties ◽  
Cell Types ◽  
Brain Barrier ◽  
Blood Brain

The blood-brain barrier (BBB) constitutes a component of the neurovascular unit formed by specialized brain microvascular endothelial cells (BMECs) surrounded by astrocytes, pericytes and neurons. During ischemic stroke injury, the BBB constitutes the first responding element resulting in the opening of the BBB and eventually neural cell death by excitotoxicity. A better understanding of the cellular mechanisms underlying the opening of the BBB during ischemic stroke is essential to identify targets to restore such barrier function after injury. Current in vitro models of the human BBB, based on primary or immortalized BMECs monocultures, display poor barrier properties but also lack one or two cellular components of the neurovascular unit.In this study, we designed an integrative in vitro model of the BBB by generating BMECs, astrocytes and neurons using patient-derived BMECs from two iPSC lines (IMR90-c4 and CTR66M). We were able to obtain all three cell types from these two cell lines. iPSC-derived BMECs showed barrier properties similar or better barrier function than hCMEC/D3 monolayer (an immortalized adult somatic BMEC). Furthermore, iPSC—derived astrocytes were capable to induce barrier properties in BMECs upon co-cultures. whereas iPSC-derived neurons were capable to form extensive and branched neurites. Upon OGD stress, iPSC-derived BMECs showed a disruption of their barrier function as early as 6 hours of OGD stress and showed a complete disruption by 24 hours. Such disruption was reversed by reoxygenation. Interestingly such barrier disruption occurs through a VEGF-independent mechanism. In the other hand, iPSC-derived neurons showed a significant decrease in cell metabolic activity preceding neurites pruning. Finally, astrocytes showed the most robust phenotype, as we noted no cell death by 24 hours OGD.In this study, we demonstrated the ability to differentiate three cell types from the same patient in two iPSC lines. We also demonstrated the ability of these cells to respond to OGD/reoxygenation stress in agreement with the current literature. We are currently investigating the molecular mechanisms by which OGD/reoxygenation drive the cellular response in these cell types.

scholarly journals A silicon nanomembrane platform for the visualization of immune cell trafficking across the human blood–brain barrier under flow

2018 ◽  
Vol 39 (3) ◽  
pp. 395-410 ◽  
Author(s):  
Adrien Mossu ◽  
Maria Rosito ◽  
Tejas Khire ◽  
Hung Li Chung ◽  
Hideaki Nishihara ◽  
...  
Keyword(s):  
Blood Brain Barrier ◽  
Immune Cell ◽  
Metastatic Cancer ◽  
Neurovascular Unit ◽  
Barrier Properties ◽  
Brain Barrier ◽  
Cell Trafficking ◽  
Physiological Flow ◽  
Blood Brain

Here we report on the development of a breakthrough microfluidic human in vitro cerebrovascular barrier (CVB) model featuring stem cell-derived brain-like endothelial cells (BLECs) and nanoporous silicon nitride (NPN) membranes (µSiM-CVB). The nanoscale thinness of NPN membranes combined with their high permeability and optical transparency makes them an ideal scaffold for the assembly of an in vitro microfluidic model of the blood–brain barrier (BBB) featuring cellular elements of the neurovascular unit (NVU). Dual-chamber devices divided by NPN membranes yield tight barrier properties in BLECs and allow an abluminal pericyte-co-culture to be replaced with pericyte-conditioned media. With the benefit of physiological flow and superior imaging quality, the µSiM-CVB platform captures each phase of the multi-step T-cell migration across the BBB in live cell imaging. The small volume of <100 µL of the µSiM-CVB will enable in vitro investigations of rare patient-derived immune cells with the human BBB. The µSiM-CVB is a breakthrough in vitro human BBB model to enable live and high-quality imaging of human immune cell interactions with the BBB under physiological flow. We expect it to become a valuable new tool for the study of cerebrovascular pathologies ranging from neuroinflammation to metastatic cancer.

scholarly journals In Vitro Blood-Brain Barrier Models Using Brain Capillary Endothelial Cells Isolated from Neonatal and Adult Rats Retain Age-Related Barrier Properties

PLoS ONE ◽  
2013 ◽  
Vol 8 (1) ◽  
pp. e55166 ◽  
Author(s):  
Fuyuko Takata ◽  
Shinya Dohgu ◽  
Atsushi Yamauchi ◽  
Junichi Matsumoto ◽  
Takashi Machida ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Brain Barrier ◽  
Barrier Properties ◽  
Brain Barrier ◽  
Adult Rats ◽  
Brain Capillary ◽  
Blood Brain ◽  
Age Related ◽  
Capillary Endothelial Cells

scholarly journals Wnt signaling mediates acquisition of blood-brain barrier properties in naïve endothelium derived from human pluripotent stem cells

2021 ◽  
Author(s):  
Benjamin D Gastfriend ◽  
Hideaki Nishihara ◽  
Scott G Canfield ◽  
Koji L Foreman ◽  
Britta Engelhardt ◽  
...  
Keyword(s):  
Stem Cells ◽  
Wnt Signaling ◽  
Blood Brain Barrier ◽  
Pluripotent Stem Cells ◽  
Barrier Properties ◽  
Human Pluripotent Stem Cells ◽  
Brain Barrier ◽  
Blood Brain ◽  
The Central Nervous System

Endothelial cells (ECs) in the central nervous system (CNS) acquire their specialized blood-brain barrier (BBB) properties in response to extrinsic signals, with Wnt/β-catenin signaling coordinating multiple aspects of this process. Our knowledge of CNS EC development has been advanced largely by animal models, and human pluripotent stem cells (hPSCs) offer the opportunity to examine BBB development in an in vitro human system. Here we show that activation of Wnt signaling in hPSC-derived naïve endothelial progenitors, but not in matured ECs, leads to robust acquisition of canonical BBB phenotypes including expression of GLUT-1, increased claudin-5, and decreased PLVAP. RNA-seq revealed a transcriptome profile resembling ECs with CNS-like characteristics, including Wnt-upregulated expression of LEF1, APCDD1, and ZIC3. Together, our work defines effects of Wnt activation in naïve ECs and establishes an improved hPSC-based model for interrogation of CNS barriergenesis.

scholarly journals Design of Experiment Based Optimization of an in Vitro Direct Contact Triculture Blood Brain Barrier Model for Permeability Screening

2021 ◽  
Author(s):  
Kelsey E Lubin ◽  
Gregory T. Knipp
Keyword(s):  
Endothelial Cell ◽  
Blood Brain Barrier ◽  
Direct Contact ◽  
Barrier Properties ◽  
Cell Types ◽  
Brain Barrier ◽  
Seeding Density ◽  
Blood Brain

Abstract Background: The in vivo restrictive properties of the blood brain barrier (BBB) largely arise from astrocyte and pericyte synergistic cell signaling interactions that underlie the brain microvessel endothelial cells (BMEC). In vivo relevant direct contact between astrocytes, pericytes, and BMECS, to our knowledge, has not been established in conventional Transwell® based in vitro screening models of the BBB. We hypothesize that a design of experiments (DOE) optimized direct contact layered triculture model will offer more in vivo relevance for screening in comparison to indirect models. Methods: Plating conditions including the seeding density of all three cell types, matrix protein, and culture time were assessed in DOEP. DOEP was followed by DOEM1 and DOEM2 to assess the influence of medium additives on barrier properties. The permeability of 4 kD dextran, a paracellular marker, was the measured response to arrive at the optimal plating conditions. The optimized model was further assessed for p-glycoprotein function using a substrate and inhibitor along with a set of BBB paracellular and transcellular markers at varying permeation rates.Results: DOEP revealed that length of culture post endothelial cell plating correlated highest with paracellular tightness. In addition, seeding density of the endothelial cell layer influenced paracellular tightness at earlier times of culture, and its impact decreased as culture is extended. Medium additives had varying effects on barrier properties as seen from DOEM1 and DOEM2. At optimal conditions, the model revealed P-gp function along with the ability to differentiate between BBB positive and negative permeants. Conclusions: We have demonstrated that the implementation of DOE based optimization for biologically based systems is an expedited method to establish multi-component in vitro cell models. The direct contact BBB triculture model reveals that the physiologically relevant layering of the three cell types is a practical method of culture to establish a screening model compared to indirect plating methods that incorporate physical barriers between cell types. Additionally, the ability of the model to differentiate between BBB positive and negative permeants suggests that this model may be an enhanced screening tool for potential neuroactive compounds.

scholarly journals Wnt/β-catenin signaling controls development of the blood–brain barrier

2008 ◽  
Vol 183 (3) ◽  
pp. 409-417 ◽  
Author(s):  
Stefan Liebner ◽  
Monica Corada ◽  
Thorsten Bangsow ◽  
Jane Babbage ◽  
Andrea Taddei ◽  
...  
Keyword(s):  
Blood Brain Barrier ◽  
Barrier Properties ◽  
Gene Signature ◽  
Brain Barrier ◽  
Neuronal Function ◽  
Tight Junction Formation ◽  
Blood Brain ◽  
Junction Formation

The blood–brain barrier (BBB) is confined to the endothelium of brain capillaries and is indispensable for fluid homeostasis and neuronal function. In this study, we show that endothelial Wnt/β-catenin (β-cat) signaling regulates induction and maintenance of BBB characteristics during embryonic and postnatal development. Endothelial specific stabilization of β-cat in vivo enhances barrier maturation, whereas inactivation of β-cat causes significant down-regulation of claudin3 (Cldn3), up-regulation of plamalemma vesicle-associated protein, and BBB breakdown. Stabilization of β-cat in primary brain endothelial cells (ECs) in vitro by N-terminal truncation or Wnt3a treatment increases Cldn3 expression, BBB-type tight junction formation, and a BBB characteristic gene signature. Loss of β-cat or inhibition of its signaling abrogates this effect. Furthermore, stabilization of β-cat also increased Cldn3 and barrier properties in nonbrain-derived ECs. These findings may open new therapeutic avenues to modulate endothelial barrier function and to limit the devastating effects of BBB breakdown.

scholarly journals Wnt signaling mediates acquisition of blood-brain barrier properties in naïve endothelium derived from human pluripotent stem cells

eLife ◽  
2021 ◽  
Vol 10 ◽  
Author(s):  
Benjamin D Gastfriend ◽  
Hideaki Nishihara ◽  
Scott G Canfield ◽  
Koji L Foreman ◽  
Britta Englehardt ◽  
...  
Keyword(s):  
Stem Cells ◽  
Wnt Signaling ◽  
Blood Brain Barrier ◽  
Pluripotent Stem Cells ◽  
Barrier Properties ◽  
Human Pluripotent Stem Cells ◽  
Brain Barrier ◽  
Blood Brain ◽  
The Central Nervous System

Endothelial cells (ECs) in the central nervous system (CNS) acquire their specialized blood-brain barrier (BBB) properties in response to extrinsic signals, with Wnt/β-catenin signaling coordinating multiple aspects of this process. Our knowledge of CNS EC development has been advanced largely by animal models, and human pluripotent stem cells (hPSCs) offer the opportunity to examine BBB development in an in vitro human system. Here we show that activation of Wnt signaling in hPSC-derived naïve endothelial progenitors, but not in matured ECs, leads to robust acquisition of canonical BBB phenotypes including expression of GLUT-1, increased claudin-5, decreased PLVAP and decreased permeability. RNA-seq revealed a transcriptome profile resembling ECs with CNS-like characteristics, including Wnt-upregulated expression of LEF1, APCDD1, and ZIC3. Together, our work defines effects of Wnt activation in naïve ECs and establishes an improved hPSC-based model for interrogation of CNS barriergenesis.

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