Progress and limitations in the use of in vitro cell cultures to serve as a permeability screen for the blood-brain barrier

Central nervous system (CNS) diseases are one of the top causes of death worldwide. As there is a difficulty of drug penetration into the brain due to the blood–brain barrier (BBB), many CNS drugs treatments fail in clinical trials. Hence, there is a need to develop effective CNS drugs following strategies for delivery to the brain by better selecting them as early as possible during the drug discovery process. The use of in vitro BBB models has proved useful to evaluate the impact of drugs/compounds toxicity, BBB permeation rates and molecular transport mechanisms within the brain cells in academic research and early-stage drug discovery. However, these studies that require biological material (animal brain or human cells) are time-consuming and involve costly amounts of materials and plastic wastes due to the format of the models. Hence, to adapt to the high yields needed in early-stage drug discoveries for compound screenings, a patented well-established human in vitro BBB model was miniaturized and automated into a 96-well format. This replicate met all the BBB model reliability criteria to get predictive results, allowing a significant reduction in biological materials, waste and a higher screening capacity for being extensively used during early-stage drug discovery studies.

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Low nanogel stiffness favors nanogel transcytosis across an in vitro blood–brain barrier

Nanomedicine Nanotechnology Biology and Medicine ◽

10.1016/j.nano.2021.102377 ◽

2021 ◽

Vol 34 ◽

pp. 102377

Author(s):

Laís Ribovski ◽

Edwin de Jong ◽

Olga Mergel ◽

Guangyue Zu ◽

Damla Keskin ◽

...

Keyword(s):

Blood Brain Barrier ◽

Brain Barrier ◽

Blood Brain

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Bisphenol A Inhibits the Transporter Function of the Blood-Brain Barrier by Directly Interacting with the ABC Transporter Breast Cancer Resistance Protein (BCRP)

International Journal of Molecular Sciences ◽

10.3390/ijms22115534 ◽

2021 ◽

Vol 22 (11) ◽

pp. 5534

Author(s):

Elin Engdahl ◽

Maarten van Schijndel ◽

Dimitrios Voulgaris ◽

Michela Di Criscio ◽

Kerry Ramsbottom ◽

...

Keyword(s):

Breast Cancer ◽

Bisphenol A ◽

Blood Brain Barrier ◽

Breast Cancer Resistance Protein ◽

Brain Barrier ◽

Production Volume ◽

Cancer Resistance ◽

Blood Brain ◽

Resistance Protein

The breast cancer resistance protein (BCRP) is an important efflux transporter in the blood-brain barrier (BBB), protecting the brain from a wide range of substances. In this study, we investigated if BCRP function is affected by bisphenol A (BPA), a high production volume chemical used in common consumer products, as well as by bisphenol F (BPF) and bisphenol S (BPS), which are used to substitute BPA. We employed a transwell-based in vitro cell model of iPSC-derived brain microvascular endothelial cells, where BCRP function was assessed by measuring the intracellular accumulation of its substrate Hoechst 33342. Additionally, we used in silico modelling to predict if the bisphenols could directly interact with BCRP. Our results showed that BPA significantly inhibits the transport function of BCRP. Additionally, BPA was predicted to bind to the cavity that is targeted by known BCRP inhibitors. Taken together, our findings demonstrate that BPA inhibits BCRP function in vitro, probably by direct interaction with the transporter. This effect might contribute to BPA’s known impact on neurodevelopment.

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Trojan Horse Transit Contributes to Blood-Brain Barrier Crossing of a Eukaryotic Pathogen

mBio ◽

10.1128/mbio.02183-16 ◽

2017 ◽

Vol 8 (1) ◽

Cited By ~ 76

Author(s):

Felipe H. Santiago-Tirado ◽

Michael D. Onken ◽

John A. Cooper ◽

Robyn S. Klein ◽

Tamara L. Doering

Keyword(s):

Experimental Evidence ◽

Cryptococcus Neoformans ◽

Blood Brain Barrier ◽

Fungal Pathogen ◽

Trojan Horse ◽

Brain Barrier ◽

Barrier Crossing ◽

Blood Brain ◽

The Brain

ABSTRACT The blood-brain barrier (BBB) protects the central nervous system (CNS) by restricting the passage of molecules and microorganisms. Despite this barrier, however, the fungal pathogen Cryptococcus neoformans invades the brain, causing a meningoencephalitis that is estimated to kill over 600,000 people annually. Cryptococcal infection begins in the lung, and experimental evidence suggests that host phagocytes play a role in subsequent dissemination, although this role remains ill defined. Additionally, the disparate experimental approaches that have been used to probe various potential routes of BBB transit make it impossible to assess their relative contributions, confounding any integrated understanding of cryptococcal brain entry. Here we used an in vitro model BBB to show that a “Trojan horse” mechanism contributes significantly to fungal barrier crossing and that host factors regulate this process independently of free fungal transit. We also, for the first time, directly imaged C. neoformans-containing phagocytes crossing the BBB, showing that they do so via transendothelial pores. Finally, we found that Trojan horse crossing enables CNS entry of fungal mutants that cannot otherwise traverse the BBB, and we demonstrate additional intercellular interactions that may contribute to brain entry. Our work elucidates the mechanism of cryptococcal brain invasion and offers approaches to study other neuropathogens. IMPORTANCE The fungal pathogen Cryptococcus neoformans invades the brain, causing a meningoencephalitis that kills hundreds of thousands of people each year. One route that has been proposed for this brain entry is a Trojan horse mechanism, whereby the fungus crosses the blood-brain barrier (BBB) as a passenger inside host phagocytes. Although indirect experimental evidence supports this intriguing mechanism, it has never been directly visualized. Here we directly image Trojan horse transit and show that it is regulated independently of free fungal entry, contributes to cryptococcal BBB crossing, and allows mutant fungi that cannot enter alone to invade the brain. IMPORTANCE The fungal pathogen Cryptococcus neoformans invades the brain, causing a meningoencephalitis that kills hundreds of thousands of people each year. One route that has been proposed for this brain entry is a Trojan horse mechanism, whereby the fungus crosses the blood-brain barrier (BBB) as a passenger inside host phagocytes. Although indirect experimental evidence supports this intriguing mechanism, it has never been directly visualized. Here we directly image Trojan horse transit and show that it is regulated independently of free fungal entry, contributes to cryptococcal BBB crossing, and allows mutant fungi that cannot enter alone to invade the brain.

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