In Vitro Blood-Brain Barrier Models for Ischemic Stroke
At a glance
Introduction of In Vitro Blood-Brain Barrier Models for Ischemic Stroke
The blood-brain barrier (BBB) is a dynamic element of the cerebrovascular interface, playing a crucial role in maintaining cerebral homeostasis by restricting the circulation of harmful substances while facilitating nutrient transport. Damage to the BBB due to ischemic stroke can significantly impair its function and integrity, which directly impacts the central nervous system. Additionally, the high selectivity of the BBB poses a substantial challenge for the effective administration of drugs and therapies for ischemic stroke. The reason for that is that a great deal of research has been conducted into permeability control of BBB.
Given the complexity of the BBB in vivo, several simplified in vitro BBB models have been developed and investigated, including monolayer models, co-culture models, dynamic models, and microfluidic BBB models. These systems serve as valuable tools for conducting more robust mechanistic studies and preclinical drug screening, potentially paving the way for the development of effective treatments for ischemic stroke in the future.
Based on an advanced cellular technology platform, Ace Therapeutics is committed to developing 3D in vitro ischemic stroke models to better reproduce the brain environment, helping you to achieve high-throughput screening and improve your chances of finding new and successful therapies.
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Monolayer Models of BBB in Transwell
A transwell device-based BBB model was originally designed from animal brain microvascular endothelial cells (BMECs). This model is capable of imitating to some degree the physiological and biochemical aspects of the BBB, including the flow of ions, glucose and amino acids, for example, or the behaviour of endothelial antigen transporters and metabolic enzymes. Since the advent of oxygen-glucose deprivation (OGD) induction, the monoculture blood-brain barrier model has become commonplace to characterize the pathological development and molecular physiology of ECs after ischemic stroke. However, the BBB formed by immortalized mouse cell lines lacks blood-brain barrier-specific proteins, resulting in poor BBB performance.
Therefore, researchers have established blood-brain barrier models using human cerebral endothelial cells (hCMEC/D3). In summary, these monoculture models are two-dimensional static systems that involve only endothelial cells and lack cellular communication between the brain's multiple cell types. They fail to fully replicate the human anatomical complexity of the blood-brain barrier, leading to inaccurate estimates of in vivo permeability.
Coculture Models of BBB in Transwell
Various co-culture models have been established to better mimic the cellular diversity of the BBB.
| BMEC-astrocyte/pericyte coculture models |
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| BMEC-pericyte-astrocyte triple-culture model | BMECs are seeded on the top surface of transwell insert, human pericytes and human astrocytes are seeded on the undersurface of transwell inserts and the bottom of the plate respectively. |
| BMEC-astrocyte-pericyteneuron quadruple-culture models |
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These coculture models of the BBB can be utilized to screen neuroprotective drugs for ischemic stroke and to investigate changes in BBB permeability, as well as tight junction (TJ) protein expressions and localizations following ischemic events. However, a significant limitation of transwell-based models is the absence of physiological shear stress from blood flow, which is crucial for the induction and maintenance of BBB barrier properties.
Fig.1. In vitro BBB models based on transwell devices. (Liu, et al., 2023)
Dynamic DIV-BBB System
Under physiological circumstances, blood pressure puts shear stress on the ECs of the wall of a blood vessel, which has ramifications for endothelial function. Such as the expression of tight junction proteins like occludin and ZO-1, as well as cells' division, apoptosis, and transport proteins. To replicate the shear stress experienced in blood vessels within BBB models, dynamic DIV-BBB models have been developed.
The DIV-BBB model features a hollow porous fiber made of polypropylene, which allows for the exchange of gases and nutrients between luminal compartments while preventing cell migration. hCMEC/D3 are embedded within these fibers, with astrocytes on the luminal surface. This model demonstrates higher transendothelial electrical resistance (TEER) and enhanced barrier function compared to static systems, enabling controlled overflow rates and mimicking physiological shear stress.
The DIV-BBB model is particularly useful in studying BBB pathology under ischemia-reperfusion conditions. Ischemia-reperfusion is induced by manipulating fluid flow and oxygen levels, effectively reproducing changes in permeability, inflammatory factor release, and immune cell extravasation that occur after ischemic stroke.
To investigate immune cell migration, the traditional hollow fibers were modified to include transcapillary pores, allowing immune cells to extravasate and facilitating the study of their interactions with brain cells. This modification led to earlier and prolonged disruption of BBB integrity compared to the conventional model.
While the DIV-BBB model enables controlled studies of BBB dynamics and cell migration, its applications are limited by a restricted variety of cell types, a thicker "basement membrane" than typical vascular anatomy, challenges in visualization, lengthy stabilization of TEER values, and low throughput capacity.
Microfluidic-based Brain-on-Chips
Organ-on-a-chips are microdevices based on microfluidics technology that are used to create microchannels and chambers in a microenvironment for multi-cell culture. Brain chips composed of various neurovascular unit cells have been used to mimic the BBB and play a key role in exploring barrier function, drug delivery, and neuroinflammation.
Microfluidic BBB (µBBB) Model
The Microfluidic BBB (µBBB) model was one of the first microfluidic systems to be used in neurological research. µBBB successfully replicates BBB-specific features such as TJ protein expression, high TEER values, and low permeability to compounds. The μBBB offers many advantages over existing in vitro static and DIV-BBB models, such as shear stress stimulation, thinner basement membranes, smaller functional volumes, and lower cost.
Fig.2. Examples of experimental microfluidic devices designed to model the BBB. (Holloway, et al., 2016)
syM-BBB
For real-time visualization of in vitro BBB models, a new microfluidic BBB system, defined as a synthetic microvascular model of the BBB (syM-BBB), was created. syM-BBB devices can be assembled so that the apical and basolateral sides are separated by columns with a gap of 3 μm, thus eliminating the upper and lower configurations of the traditional μBBB model and allowing for the real-time monitoring of the fluorescein isothiocyanate- dextran (FITC-dextran) diffusion through the BBB in real time.
- Liu, Z., et al. (2023). Engineering Neurovascular Unit and Blood–Brain Barrier for Ischemic Stroke Modeling. Advanced Healthcare Materials, 12(19), 2202638.
- Holloway, P. M., & Gavins, F. N. (2016). Modeling ischemic stroke in vitro: status quo and future perspectives. Stroke, 47(2), 561-569.
