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The pivotal role of micro-environmental cells in a human blood-brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis.

ABSTRACT: BACKGROUND:The blood-brain barrier (BBB) is altered in several diseases of the central nervous system. For example, the breakdown of the BBB during cerebral ischemia in stroke or traumatic brain injury is a hallmark of the diseases' progression. This functional damage is one key event which is attempted to be mimicked in in vitro models. Recent studies showed the pivotal role of micro-environmental cells such as astrocytes for this barrier damage in mouse stroke in vitro models. The aim of this study was to evaluate the role of micro-environmental cells for the functional, paracellular breakdown in a human BBB cerebral ischemia in vitro model accompanied by a transcriptional analysis. METHODS:Transwell models with human brain endothelial cell line hCMEC/D3 in mono-culture or co-culture with human primary astrocytes and pericytes or rat glioma cell line C6 were subjected to oxygen/glucose deprivation (OGD). Changes of transendothelial electrical resistance (TEER) and FITC-dextran 4000 permeability were recorded as measures for paracellular tightness. In addition, qPCR and high-throughput qPCR Barrier chips were applied to investigate the changes of the mRNA expression of 38 relevant, expressed barrier targets (tight junctions, ABC-transporters) by different treatments. RESULTS:In contrast to the mono-culture, the co-cultivation with human primary astrocytes/pericytes or glioma C6 cells resulted in a significantly increased paracellular permeability after 5 h OGD. This indicated the pivotal role of micro-environmental cells for BBB breakdown in the human model. Hierarchical cluster analysis of qPCR data revealed differently, but also commonly regulated clustered targets dependent on medium exchange, serum reduction, hydrocortisone addition and co-cultivations. CONCLUSIONS:The co-cultivation with micro-environmental cells is necessary to achieve a functional breakdown of the BBB in the cerebral ischemia model within an in vivo relevant time window. Comprehensive studies by qPCR revealed that distinct expression clusters of barrier markers exist and that these are regulated by different treatments (even by growth medium change) indicating that controls for single cell culture manipulation steps are crucial to understand the observed effects properly.

SUBMITTER: Gerhartl A

PROVIDER: S-EPMC7059670 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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The pivotal role of micro-environmental cells in a human blood-brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis.

Gerhartl Anna A Pracser Nadja N Vladetic Alexandra A Hendrikx Sabrina S Friedl Heinz-Peter HP Neuhaus Winfried W

Fluids and barriers of the CNS 20200305 1

<h4>Background</h4>The blood-brain barrier (BBB) is altered in several diseases of the central nervous system. For example, the breakdown of the BBB during cerebral ischemia in stroke or traumatic brain injury is a hallmark of the diseases' progression. This functional damage is one key event which is attempted to be mimicked in in vitro models. Recent studies showed the pivotal role of micro-environmental cells such as astrocytes for this barrier damage in mouse stroke in vitro models. The aim ...[more]

PMID: 32138745

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Project description:Cerebral amyloid angiopathy (CAA) is a small vessel disease characterized by amyloid β (Aβ) peptide deposition within the walls of medium to small-caliber blood vessels, cerebral microhemorrhage, and blood-brain barrier (BBB) leakage. It is commonly associated with late-stage Alzheimer's disease. BBB dysfunction is indicated as a pathological substrate for CAA progression with hyperpermeability, enhancing the extravasation of plasma components and inducing neuroinflammation, further worsening BBB injury and contributing to cognitive decline. Although significant effort has been made in defining the gene mutations and risk factors involved in microvascular alterations with vascular dementia and Alzheimer's disease, the intra- and intercellular pathogenic mechanisms responsible for vascular hyperpermeability are still largely unknown. The present study aimed to elucidate the transcriptional profile of the cerebral microvessels (BBB) in a murine model with CAA vasculopathy to define potential causes and underlying mechanisms of BBB injury. A comprehensive RNA sequencing analysis was performed of CAA vasculopathy in Tg-SwDI mice at 6 and 18 months in comparison to age-matched wildtype controls to examine how age and amyloid accumulation impact the transcriptional signature of the BBB. Results indicate that Aβ has a critical role in triggering brain endothelial cell and BBB dysfunction in CAA vasculopathy, causing an intense proinflammatory response, impairing oxidative metabolism, altering the coagulation status of brain endothelial cells, and remodeling barrier properties. The proinflammatory response includes both adaptive and innate immunity, with pronounced induction of genes that regulate macrophage/microglial activation and chemokines/adhesion molecules that support T and B cell transmigration. Age has an important impact on the effects of Aβ, increasing the BBB injury in CAA vasculopathy. However, early inflammation, particularly microglia/macrophage activation and the mediators of B lymphocytes' activities are underlying processes of BBB hyperpermeability and cerebral microbleeds in the early stage of CAA vasculopathy. These findings reveal a specific profile of the CAA-associated BBB injury that leads to a full progression of CAA.

Dataset Information

The pivotal role of micro-environmental cells in a human blood-brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis.

Publications

The pivotal role of micro-environmental cells in a human blood-brain barrier in vitro model of cerebral ischemia: functional and transcriptomic analysis.

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