Abstract
Culture-based blood–brain barrier (BBB) models are crucial tools to enable rapid screening of brain-penetrating drugs. However, reproducibility of in vitro barrier properties and permeability remain as major challenges. Here, we report that self-assembling multicellular BBB spheroids display reproducible BBB features and functions. The spheroid core is comprised mainly of astrocytes, while brain endothelial cells and pericytes encase the surface, acting as a barrier that regulates transport of molecules. The spheroid surface exhibits high expression of tight junction proteins, VEGF-dependent permeability, efflux pump activity and receptor-mediated transcytosis of angiopep-2. In contrast, the transwell co-culture system displays comparatively low levels of BBB regulatory proteins, and is unable to discriminate between the transport of angiopep-2 and a control peptide. Finally, we have utilized the BBB spheroids to screen and identify BBB-penetrant cell-penetrating peptides (CPPs). This robust in vitro BBB model could serve as a valuable next-generation platform for expediting the development of CNS therapeutics.
Highlights
Culture-based blood–brain barrier (BBB) models are crucial tools to enable rapid screening of brain-penetrating drugs
It has been reported that direct cell–cell interaction of brain endothelial cells (ECs) with other components of the neurovascular unit such as pericytes and/or astrocytes is important for induction and maintenance of the specialized BBB properties in culture[1,8]
As a proof-of-principle, we demonstrate the versatility of this model in analysing the lipoprotein receptor-related protein-1 (LRP-1) receptor-mediated transport of angiopep-2, as well as the permeability of a BBB-penetrant small molecule that inhibits the phosphatidylinositol 3-kinase inhibitor, known as BKM12020
Summary
Culture-based blood–brain barrier (BBB) models are crucial tools to enable rapid screening of brain-penetrating drugs. One major challenge in developing representative models of the BBB is the fact that brain ECs rapidly dedifferentiate and lose their BBB characteristics when they are removed from their native microenvironment and grown in culture, giving rise to a more generic EC phenotype[6] This phenomenon, known as ‘phenotypic drift’, is especially likely to occur in ECs at higher passages, resulting in lack of expression of key BBB modulators and leaky paracellular barrier function[7]. It has been reported that direct cell–cell interaction of brain ECs with other components of the neurovascular unit such as pericytes and/or astrocytes is important for induction and maintenance of the specialized BBB properties in culture[1,8] This has led to the development of several in vitro BBB co-culture models with this modular organization to facilitate discovery and advancement of neuropharmaceuticals[9,10]. Given the facile construction and the ability of the BBB spheroids to reproduce key barrier activity and characteristics, this model is attractive for cost-effective and highthroughput drug permeability testing
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