Reconstitution of the blood-brain barrier in cell culture for studies of drug transport and metabolism

doi:10.1016/S0169-409X(96)00421-8

Advanced Drug Delivery Reviews

Volume 22, Issues 1–2, 15 November 1996, Pages 251-264

https://doi.org/10.1016/S0169-409X(96)00421-8 Get rights and content

Abstract

In this paper the anatomy of the blood-brain barrier and the properties of peripheral versus brain-microvessel-endothelial cells (BMEC) are outlined and the isolation and application of microvessels are discussed. In addition, the isolation and culture of BMEC are described and discussed and the application of this system in drug transport-and metabolism-studies, visualization of drug transport routes, and in the study of changes in BBB functionality during inflammation, is illustrated.

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Cited by (14)

In vitro screening of nanomedicines through the blood brain barrier: A critical review
2016, Biomaterials
Citation Excerpt :
However, as also happens with PAMPA, this model has only proven useful for the prediction of transcellular passive diffusion. To overcome limitations of in vitro non-cell based models, cell-based models are often integrated as screening tools to give a more comprehensive insight into the mechanisms involved in the passage through the BBB [138–140]. Emerged in the 1980s, cell-based models to screen BBB permeability represent a more physiologically relevant tool over PAMPA to obtain mechanistic information since they account for both transcellular passive diffusion and active transport devoid of additional physiological factors that exist in vivo [141,142].
The blood-brain barrier accounts for the high attrition rate of the treatments of most brain disorders, which therefore remain one of the greatest health-care challenges of the twenty first century. Against this background of hindrance to brain delivery, nanomedicine takes advantage of the assembly at the nanoscale of available biomaterials to provide a delivery platform with potential to raising brain levels of either imaging or therapeutic agents. Nevertheless, to prevent later failure due to ineffective drug levels at the target site, researchers have been endeavoring to develop a battery of in vitro screening procedures that can predict earlier in the drug discovery process the ability of these cutting-edge drug delivery platforms to cross the blood-brain barrier for biomedical purposes.
This review provides an in-depth analysis of the currently available in vitro blood-brain barrier models (both cell-based and non-cell-based) with the focus on their suitability for understanding the biological brain distribution of forthcoming nanomedicines. The relationship between experimental factors and underlying physiological assumptions that would ultimately lead to a more predictive capacity of their in vivo performance, and those methods already assayed for the evaluation of the brain distribution of nanomedicines are comprehensively discussed.
Functional role of adenosine receptor subtypes in the regulation of blood-brain barrier permeability: Possible implications for the design of synthetic adenosine derivatives
2003, European Journal of Pharmaceutical Sciences
The objective of this investigation was to determine the functional role of adenosine receptor subtypes in the regulation of blood–brain barrier (BBB) permeability. The presence of the equilibrative es and ei nucleoside transporters at the BBB was also determined. Studies were conducted in an experimental in vitro BBB model comprising bovine brain capillary endothelial cells (BCECs) and rat astrocytes (RAs). The presence of the receptors and transporters was investigated by a combination of RT-PCR and radioligand binding assays. Changes in paracellular permeability were investigated on basis of changes in trans-endothelial-electrical-resistance (TEER) and transport of paracellular markers. In BCECs the presence of A_2A and A₃ receptors and the es nucleoside transporter was demonstrated. The A₁ receptor was absent, while the presence of the A_2B receptor and the ei nucleoside transporter remained uncertain. In RAs the presence of all four receptor subtypes and the es and ei nucleoside transporters was demonstrated. Upon application of selective agonists no significant changes in TEER or the transport of the paracellular markers were observed. The functional role of adenosine receptor subtypes in regulating the paracellular permeability of the BBB is probably small. It is unlikely therefore that the BBB transport of synthetic adenosine analogues is modified by permeability changes. The es nucleoside transporter might play a role in the BBB transport of synthetic adenosine analogues.
Establishment and functional characterization of an in vitro model of the blood-brain barrier, comprising a co-culture of brain capillary endothelial cells and astrocytes
2001, European Journal of Pharmaceutical Sciences
Objective: The aim was to establish a flexible, abundantly available, reproducible and functionally characterized in vitro model of the blood–brain barrier (BBB). Methods: In a first step, bovine brain capillaries and newborn rat astrocytes were isolated. Subsequently, a co-culture of primary brain capillary endothelial cells (BCEC) on semi-permeable filter inserts, with astrocytes on the bottom of the filter was established. The cell material was characterized on the basis of specific cell-type properties and (functional expression of) specific BBB properties. Results: BCEC displayed: (1) characteristic endothelial cell morphology; (2) expression of endothelial cell markers (i.e., CD51, CD62P, CD71 and cadherin 5); (3) marginal F-actin localization; (4) tight junction formation between the cells; (5) expression of γ-glutamyl-transpeptidase (γ-GTP); (6) expression of P-glycoprotein (Pgp); (7) functional transendothelial transferrin transport and uptake; (8) restriction of paracellular transport; and (9) high transendothelial electrical resistance (TEER). Astrocytes displayed characteristic astrocyte morphology and expressed glial fibrillary acidic protein (GFAP). Co-culture with astrocytes increased TEER and decreased paracellular transport. In addition, expression of the glucocorticoid receptor (GR) was demonstrated in the endothelial cells of the BBB, while no expression of the mineralocorticoid receptor (MR) was found. Conclusions: A high quality and mass-production in vitro BBB model was established in which experiments with physiological (e.g., regulation of BBB permeability), pharmacological (e.g., pharmacokinetics and pharmacodynamics) and pathophysiological (e.g., disease influence on BBB permeability) objectives can be reproducibly performed.
Relationship between permeability status of the blood-brain barrier and in vitro permeability coefficient of a drug
2000, European Journal of Pharmaceutical Sciences
Objective: The aim was to test the hypothesis that the assessment of basal and drug-induced changes in permeability of the blood–brain barrier (BBB) during in vitro drug transport assays is essential for an accurate estimation of the permeability coefficient of a drug. Methods: An in vitro BBB model was used, comprising of brain capillary endothelial cells (BCEC) and astrocytes co-cultured on semi-permeable filter inserts. Experiments were performed under control and challenged experimental circumstances, induced to simulate drug effects. The apparent BBB permeability coefficient for two markers for paracellular drug transport, sodium fluorescein (P_app,FLU, M_w 376 Da) and FITC-labeled dextran (P_app,FD4, M_w 4 kDa), was determined. Transendothelial electrical resistance (TEER) was used to quantify basal and (simulated) drug-induced changes in permeability of the in vitro BBB. The relationship between P_app and TEER was determined. Drug effects were simulated by exposure to physiologically active endogenous and exogenous substances (i.e., histamine, deferroxamine mesylate, adrenaline, noradrenaline, bradykinin, vinblastine, sodium nitroprusside and lipopolysaccharide). Results: P_app,FLU and P_app,FD4 in control experiments varied from 1.6 up to 17.6 (10⁻⁶cm/s) and 0.3 up to 7.3 (10⁻⁶cm/s), respectively; while for individual filters P_app,FLU was 4 times higher than P_app,FD4 (R²=0.97). As long as TEER remained above 131·Ω cm² for FLU or 122·Ω cm² for FD4 during the transport assay, P_app remained independent from the basal permeability of the in vitro BBB. Below these TEER values, P_app increased exponentially. This nonlinear relationship between basal BBB permeability and P_app was described by a one-phase exponential decay model. From this model the BBB permeability status independent permeability coefficients for FLU and FD4 (P_FLU and P_FD4) were estimated to be 2.2±0.1 and 0.48±0.03 (10⁻⁶cm/s), respectively. In the experimentally challenged experiments, a reliable indication for P_FLU and P_FD4 could be estimated only after the (simulated) drug-induced change in BBB permeability was taken into account. Conclusions: The assessment of basal BBB permeability status during drug transport assays was essential for an accurate estimation of the in vitro permeability coefficient of a drug. To accurately extrapolate the in vitro permeability coefficient of a drug to the in vivo situation, it is essential that drug-induced changes in the in vitro BBB permeability during the drug transport assay are determined.
Brainpeps: The blood-brain barrier peptide database
2012, Brain Structure and Function
Lipophilicity and its relationship with passive drug permeation
2011, Pharmaceutical Research

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Reconstitution of the blood-brain barrier in cell culture for studies of drug transport and metabolism

Abstract

Neurochem. Int.

Cell

Cell

Cell

Dev. Brain Res.

Int. J. Dev. Neurosci.

Biochem. Pharmacol.

Neurochem. Int.

J. Vasc. Surg.

Biomaterials

J. Neuroimmunol.

Biochem. Pharmacol.

Microvascular Res.

Brain Res.

J. Biol. Chem.

Biochem. Pharmacol.

Biochem. Biophys. Acta

Brain Res. Rev.

Biochem. Biophys. Res. Commun.

Pathophysiological aspects of acute EAE

Acta Neurologica Scand.

Drug transport across the blood-brain barrier: II. Experimental techniques to study drug transport

Pharm. Weekblad Sci. Ed.

The cerebral microvessels in culture, an update

J. Neurochem.

Characterization of an in vitro blood-brain barrier model system for studying drug transport and metabolism

Pharm. Res.

Cultivation of endothelial cells: limitations and perspectives

Med. Biol.

Das sauerstoffbedurfenis des organismus; eine farbenanalytische studie

Die aussere und innere sekretion des gesunden und kranken organismus im lichte der vitalen farbung

Beitr. Klin. Chirurg.

Vitalfarbung am zentral-nervensystem

Abh. Preuss. Akad. Wiss., Phys.-Math. KL

Die bedeutung der vitalen farbung fur die lehre vom stofaustausch zwischen dem zentralnervensystem und dem ubrigen korper

Arch. Psychiatr. Nervenkrank.

Die allgemeinen grundlagen des stoffaustausches zwischen dem Zentralnervensystem und dem ubrigen korper

Arch. Psychiatr. Nervenkrank.

Fine structural localization of a blood-brain barrier to exogenous peroxidase

J. Cell. Biol.

Junctions between intimately apposed cell membranes in the vertebrate brain

J. Cell. Biol.

The concepts of the blood-brain barrier

Blood-brain barrier transport of nutrients

Drug transport into the central nervous system

Evolution in the concept of the blood-brain barrier phenomenon

Electronmikroskopischen befund an den Capillaren der Hirnrinde

Arch. Psychiatr. Nervenkrank.

Characterization of in vitro blood-brain barrier: effects of molecular size and lipophilicity on cerebrovascular endothelial transport rates of drugs

J. Pharmacol. Exp. Ther.

Multidrug-resistance gene (P-glycoprotein) is expressed by endothelial cells at the blood-brain barrier sites

Astrocytes induce blood-brain barrier properties in endothelial cells

Nature

In vitro interaction of astrocyte and pericytes with capillary-like structures of brain microvessel endothelium

Lab. Invest.

Peptide drug delivery to the brain

Differences in function and structure of the capillary endothelium in grey matter, white matter and a circumventricular organ of rat brain

Blood Vessels

Simultaneous determination of regional cerebral blood flow, glucose metabolism, and pH in acute experimental allergic encephalomyelitis

J. Cereb. Blood Flow Metab.

Microvessels isolated from rat brain: localization of astrocyte processes by immunohistochemical techniques

J. Neurochem.

The Blood-Brain Barrier. Transport across the cerebral endothelium

Circ. Res.

Characterization of the blood-brain barrier: protein composition of the capillary endothelial cell membrane

J. Neurochem.

Capillary depletion method for quantifying blood-brain barrier transcylosis of circulating peptides and plasma proteins

J. Neurochem.

Blood-brain Barrier: transport studies in isolated brain capillaries and in cultured brain endothelial cells

The susceptibility of cerebral endothelial cells to astroglial induction of blood-brain barrier enzymes depends on their proliferative state

J. Neurochem.