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Generation of an Immortalized Murine Brain Microvascular Endothelial Cell Line as an In Vitro Blood Brain Barrier Model
Published on: August 29, 2012
Activation of glial cells induces proinflammatory properties in brain capillary endothelial cells in vitro
Annette Burkhart1, Steinunn Sara Helgudóttir1, Yahye A Mahamed1
1Neurobiology Research and Drug Delivery, Department of Health Science and Technology, Aalborg University, Selma Lagerlöfts Vej 249, 9260, Gistrup, Denmark.
Abstract:
Neurodegenerative diseases are often accompanied by neuroinflammation and impairment of the blood-brain barrier (BBB) mediated by activated glial cells through their release of proinflammatory molecules. To study the effects of glial cells on mouse brain endothelial cells (mBECs), we developed an in vitro BBB model with inflammation by preactivating mixed glial cells (MGCs) with lipopolysaccharide (LPS) before co-culturing with mBECs to study the influence of molecules released by activated MGCs. The response of the mBECs to activated MGCs was compared to direct stimulation with LPS. The cytokine profile of activated MGCs was analyzed together with their effects on the mBEC's integrity, expression of tight junction proteins, adhesion molecules, and BBB-specific transport proteins. Stimulation of MGCs significantly upregulated mRNA expression and secretion of several pro-inflammatory cytokines. Co-culturing mBECs with pre-stimulated MGCs significantly affected the barrier integrity of mBECs similar to direct stimulation with LPS. The gene expression levels of tight junction proteins were unaltered, but tight junction proteins revealed rearrangements with respect to subcellular distribution. Compared to direct stimulation with LPS, the expression of cell-adhesion molecules was significantly increased when mBECs were co-cultured with prestimulated MGCs and thus pre-activating MGCs transforms mBECs into a proinflammatory phenotype.
Insights
Activated glial cells release inflammatory molecules that impair the blood-brain barrier (BBB). This study shows that activated glial cells transform mouse brain endothelial cells (mBECs) into a proinflammatory state, affecting BBB integrity.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Neuroinflammation and blood-brain barrier (BBB) dysfunction are hallmarks of neurodegenerative diseases.
- Activated glial cells release proinflammatory molecules that can compromise BBB integrity.
Purpose of the Study:
- To investigate the effects of activated glial cells on mouse brain endothelial cells (mBECs) in an in vitro model.
- To compare the impact of glial cell-derived molecules versus direct lipopolysaccharide (LPS) stimulation on mBECs.
Main Methods:
- Developed an in vitro BBB model using co-cultured mBECs and lipopolysaccharide (LPS)-activated mixed glial cells (MGCs).
- Analyzed MGC cytokine profiles and their effects on mBEC integrity, tight junction proteins, adhesion molecules, and transport proteins.
- Compared mBEC responses to co-culture with activated MGCs versus direct LPS stimulation.
Main Results:
- LPS stimulation upregulated pro-inflammatory cytokine expression and secretion in MGCs.
- Co-culturing mBECs with activated MGCs significantly compromised mBEC barrier integrity, similar to direct LPS stimulation.
- Tight junction protein gene expression remained unchanged, but their subcellular distribution was altered.
- Cell-adhesion molecule expression was significantly increased in mBECs co-cultured with activated MGCs compared to direct LPS stimulation.
Conclusions:
- Pre-activating glial cells transforms mBECs into a proinflammatory phenotype.
- Glial cell-derived factors significantly impact BBB integrity and endothelial cell behavior, contributing to neuroinflammation in disease contexts.
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