The Effect of Normoxic and Hypoxic U-87 Glioblastoma Paracrine Secretion on the Modulation of Brain Endothelial Cells

Background: Glioblastoma multiforme (GBM) is a highly invasive brain tumour, characterized by its ability to secrete factors promoting its virulence. Brain endothelial cells (BECs) in the GBM environment are physiologically modulated. The present study investigated the modulatory effects of normoxically and hypoxically induced glioblastoma U-87 cell secretions on BECs. Methods: Conditioned media (CM) were derived by cultivating U-87 cells under hypoxic incubation (5% O2) and normoxic incubation (21% O2). Treated bEnd.3 cells were evaluated for mitochondrial dehydrogenase activity, mitochondrial membrane potential (ΔΨm), ATP production, transendothelial electrical resistance (TEER), and endothelial tight-junction (ETJ) gene expression over 96 h. Results: The coculture of bEnd.3 cells with U-87 cells, or exposure to either hypoxic or normoxic U-87CM, was associated with low cellular viability. The ΔΨm in bEnd.3 cells was hyperpolarized after hypoxic U-87CM treatment (p < 0.0001). However, normoxic U-87CM did not affect the state of ΔΨm. BEC ATP levels were reduced after being cocultured with U-87 cells, or with hypoxic and normoxic CM (p < 0.05). Suppressed mitochondrial activity in bEnd.3 cells was associated with increased transendothelial permeability, while bEnd.3 cells significantly increased the gene expression levels of ETJs (p < 0.05) when treated with U-87CM. Conclusions: Hypoxic and normoxic glioblastoma paracrine factors differentially suppressed mitochondrial activity in BECs, increasing the BECs’ barrier permeability.

Download Full-text

Differential Effects of Normoxic versus Hypoxic Derived Breast Cancer Paracrine Factors on Brain Endothelial Cells

Biology ◽

10.3390/biology10121238 ◽

2021 ◽

Vol 10 (12) ◽

pp. 1238

Author(s):

Mariam Rado ◽

Brian Flepisi ◽

David Fisher

Keyword(s):

Breast Cancer ◽

Gene Expression ◽

Endothelial Cells ◽

Mitochondrial Activity ◽

Toxic Substances ◽

Brain Endothelial Cells ◽

Paracrine Factors ◽

Hypoxic Conditions ◽

Dehydrogenases Activity ◽

The Brain

Background: The blood-brain barrier (BBB) is a central nervous system protective barrier formed primarily of endothelial cells that regulate the entry of substances and cells from entering the brain. However, the BBB integrity is disrupted in disease, including cancer, allowing toxic substances, molecules, and circulating cells to enter the brain. This study aimed to determine the mitochondrial changes in brain endothelial cells co-cultured with cancer cells. Method: Brain endothelial cells (bEnd.3) were co-cultivated with various concentrations of breast cancer (MCF7) conditioned media (CM) generated under normoxic (21% O2) and hypoxic conditions (5% O2). The mitochondrial activities (including; dehydrogenases activity, mitochondrial membrane potential (ΔΨm), and ATP generation) were measured using Polarstar Omega B.M.G-Plate reader. Trans-endothelial electrical resistance (TEER) was evaluated using the EVOM system, followed by quantifying gene expression of the endothelial tight junction (ETJs) using qPCR. Results: bEnd.3 cells had reduced cell viability after 72 h and 96 h exposure to MCF7CM under hypoxic and normoxic conditions. The ΔΨm in bEnd.3 cells were hyperpolarized after exposure to the hypoxic MCF7CM (p < 0.0001). However, the normoxic MCF7CM did not significantly affect the state of ΔΨm in bEnd.3 cells. ATP levels in bEnd.3 co-cultured with hypoxic and normoxic MCF7CM was significantly reduced (p < 0.05). The changes in brain endothelial mitochondrial activity were associated with a decrease in TEER of bEnd.3 monolayer co-cultured with MCF7CM under hypoxia (p = 0.001) and normoxia (p < 0.05). The bEnd.3 cells exposed to MCF7CM significantly increased the gene expression level of ETJs (p < 0.05). Conclusions: MCF7CM modulate mitochondrial activity in brain endothelial cells, affecting the brain endothelial barrier function.

Download Full-text

NF-κB is activated and ICAM-1 gene expression is upregulated during reoxygenation of human brain endothelial cells

Neuroscience Letters ◽

10.1016/s0304-3940(98)00239-0 ◽

1998 ◽

Vol 248 (3) ◽

pp. 199-203 ◽

Cited By ~ 57

Author(s):

Eugene F Howard ◽

Qiang Chen ◽

Charles Cheng ◽

James E Carroll ◽

David Hess

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Human Brain ◽

Brain Endothelial Cells

Download Full-text

Flow-Mediated Susceptibility and Molecular Response of Cerebral Endothelia to SARS-CoV-2 Infection

Stroke ◽

10.1161/strokeaha.120.032764 ◽

2021 ◽

Vol 52 (1) ◽

pp. 260-270 ◽

Cited By ~ 1

Author(s):

Naoki Kaneko ◽

Sandro Satta ◽

Yutaro Komuro ◽

Sree Deepthi Muthukrishnan ◽

Visesha Kakarla ◽

...

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Human Brain ◽

Middle Cerebral Artery ◽

Monolayer Culture ◽

Umbilical Vein ◽

Brain Endothelial Cells ◽

S Protein ◽

3 Dimensional ◽

Cerebrovascular Events

Background and Purpose: Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection is associated with an increased rate of cerebrovascular events including ischemic stroke and intracerebral hemorrhage. The mechanisms underlying cerebral endothelial susceptibility and response to SARS-CoV-2 are unknown yet critical to understanding the association of SARS-CoV-2 infection with cerebrovascular events. Methods: Endothelial cells were isolated from human brain and analyzed by RNA sequencing. Human umbilical vein and human brain microvascular cells were used in both monolayer culture and endothelialized within a 3-dimensional printed vascular model of the middle cerebral artery. Gene expression levels were measured by quantitative polymerase chain reaction and direct RNA hybridization. Recombinant SARS-CoV-2 S protein and S protein–containing liposomes were used to measure endothelial binding by immunocytochemistry. Results: ACE2 (angiotensin-converting enzyme-2) mRNA levels were low in human brain and monolayer endothelial cell culture. Within the 3-dimensional printed vascular model, ACE2 gene expression and protein levels were progressively increased by vessel size and flow rates. SARS-CoV-2 S protein–containing liposomes were detected in human umbilical vein endothelial cells and human brain microvascular endothelial cells in 3-dimensional middle cerebral artery models but not in monolayer culture consistent with flow dependency of ACE2 expression. Binding of SARS-CoV-2 S protein triggered 83 unique genes in human brain endothelial cells including upregulation of complement component C3. Conclusions: Brain endothelial cells are susceptible to direct SARS-CoV-2 infection through flow-dependent expression of ACE2. Viral S protein binding triggers a unique gene expression profile in brain endothelia that may explain the association of SARS-CoV-2 infection with cerebrovascular events.

Download Full-text

Short regulatory DNA sequences to target brain endothelial cells for gene therapy

Journal of Cerebral Blood Flow & Metabolism ◽

10.1177/0271678x211039617 ◽

2021 ◽

pp. 0271678X2110396 ◽

Cited By ~ 1

Author(s):

Hanna Graßhoff ◽

Helge Müller-Fielitz ◽

Godwin K Dogbevia ◽

Jakob Körbelin ◽

Jacqueline Bannach ◽

...

Keyword(s):

Gene Expression ◽

Gene Therapy ◽

Endothelial Cells ◽

Blood Brain Barrier ◽

Regulatory Elements ◽

Brain Barrier ◽

Regulatory Sequences ◽

Brain Endothelial Cells ◽

Enhancer Element ◽

Blood Brain

Gene vectors targeting CNS endothelial cells allow to manipulate the blood-brain barrier and to correct genetic defects in the CNS. Because vectors based on the adeno-associated virus (AAV) have a limited capacity, it is essential that the DNA sequence controlling gene expression is short. In addition, it must be specific for endothelial cells to avoid off-target effects. To develop improved regulatory sequences with selectivity for brain endothelial cells, we tested the transcriptional activity of truncated promoters of eleven (brain) endothelial-specific genes in combination with short regulatory elements, i.e., the woodchuck post-transcriptional regulatory element (W), the CMV enhancer element (C), and a fragment of the first intron of the Tie2 gene (S), by transfecting brain endothelial cells of three species. Four combinations of regulatory elements and short promoters ( Cdh5, Ocln, Slc2a1, and Slco1c1) progressed through this in-vitro pipeline displaying suitable activity. When tested in mice, the regulatory sequences C- Ocln-W and C- Slc2a1-S-W enabled a stronger and more specific gene expression in brain endothelial cells than the frequently used CAG promoter. In summary, the new regulatory elements efficiently control gene expression in brain endothelial cells and may help to specifically target the blood-brain barrier with gene therapy vectors.

Download Full-text

P4-220 β-amyloid-induced inflammatory gene expression in human brain endothelial cells

Neurobiology of Aging ◽

10.1016/s0197-4580(04)81778-0 ◽

2004 ◽

Vol 25 ◽

pp. S537-S538

Author(s):

Wandong Zhang ◽

Sabina Giuseppin ◽

Danica Stanimirovic

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Human Brain ◽

Brain Endothelial Cells ◽

Inflammatory Gene Expression ◽

Inflammatory Gene ◽

Β Amyloid

Download Full-text

Gene expression profiles of brain endothelial cells during embryonic development at bulk and single-cell levels

Science Signaling ◽

10.1126/scisignal.aag2476 ◽

2017 ◽

Vol 10 (487) ◽

Author(s):

Mike Hupe ◽

Minerva Xueting Li ◽

Susanne Kneitz ◽

Daria Davydova ◽

Chika Yokota ◽

...

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Embryonic Development ◽

Single Cell ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Brain Endothelial Cells

Download Full-text

Platelets Alter Gene Expression Profile in Human Brain Endothelial Cells in an In Vitro Model of Cerebral Malaria

PLoS ONE ◽

10.1371/journal.pone.0019651 ◽

2011 ◽

Vol 6 (5) ◽

pp. e19651 ◽

Cited By ~ 26

Author(s):

Mathieu Barbier ◽

Dorothée Faille ◽

Béatrice Loriod ◽

Julien Textoris ◽

Claire Camus ◽

...

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Human Brain ◽

Cerebral Malaria ◽

Gene Expression Profile ◽

Expression Profile ◽

In Vitro Model ◽

Brain Endothelial Cells ◽

Vitro Model

Download Full-text

P2-126 Gene expression of brain endothelial cells in response to Alzheimer disease-associated stresses

Neurobiology of Aging ◽

10.1016/s0197-4580(04)80873-x ◽

2004 ◽

Vol 25 ◽

pp. S260

Author(s):

Cheryl G. Pfeifer ◽

George Su ◽

Sarah McCalmon ◽

Wilfred A. Jefferies

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Alzheimer Disease ◽

Brain Endothelial Cells

Download Full-text

Gene expression profiling of brain endothelial cells after experimental subarachnoid haemorrhage

Scientific Reports ◽

10.1038/s41598-021-87301-z ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Michael K. Tso ◽

Paul Turgeon ◽

Bert Bosche ◽

Charles K. Lee ◽

Tian Nie ◽

...

Keyword(s):

Gene Expression ◽

Endothelial Cells ◽

Brain Injury ◽

Subarachnoid Haemorrhage ◽

Gene Expression Profiling ◽

Expression Profiling ◽

Delayed Cerebral Ischemia ◽

High Morbidity ◽

Brain Endothelial Cells ◽

Cox 2

AbstractSubarachnoid haemorrhage (SAH) is a type of hemorrhagic stroke that is associated with high morbidity and mortality. New effective treatments are needed to improve outcomes. The pathophysiology of SAH is complex and includes early brain injury and delayed cerebral ischemia, both of which are characterized by blood–brain barrier (BBB) impairment. We isolated brain endothelial cells (BECs) from mice subjected to SAH by injection of blood into the prechiasmatic cistern. We used gene expression profiling to identify 707 unique genes (2.8% of transcripts, 403 upregulated, 304 downregulated, 24,865 interrogated probe sets) that were significantly differentially expressed in mouse BECs after SAH. The pathway involving prostaglandin synthesis and regulation was significantly upregulated after SAH, including increased expression of the Ptgs2 gene and its corresponding COX-2 protein. Celecoxib, a selective COX-2 inhibitor, limited upregulation of Ptgs2 in BECs. In this study, we have defined the gene expression profiling of BECs after experimental SAH and provide further insight into BBB pathophysiology, which may be relevant to other neurological diseases such as traumatic brain injury, brain tumours, ischaemic stroke, multiple sclerosis, and neurodegenerative disorders.

Download Full-text