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Blood–Brain Barrier Transwell Modeling

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Abstract

In vitro blood–brain barrier (BBB) modeling with the use of the brain endothelial cells grown on a transwell membrane is widely used to investigate BBB disorders and factors intended to ameliorate these pathologies. Endothelial cells, due to tight junction proteins, ensure selective permeability for a number of substances. The low integrity (i.e., high permeability) of the BBB model, as compared to the physiological one, complicates evaluation of the effects caused by different agents. Thus, the selection of conditions to improve barrier integrity is an essential task. In this study, mouse brain endothelial cells bEnd.3 are used in experiments on transwell modeling. To determine which factors enhance BBB integrity, the effects of the cultivation medium, the number of cells during seeding, the state of the transwell membrane, and cultivation in the presence or in the absence of primary mouse neurons and matrigel as a matrix on the passage of a fluorescent label through the cell monolayer were assessed. The effect of fetal bovine serum on the tight junction protein claudin-5 was analyzed by immunocytochemistry. The obtained cultivation parameter data facilitate the solution to the problem of low integrity of the BBB transwell model and bring the model closer to the physiologically relevant indicators.

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Funding

The study was supported by the Ministry of Science and Higher Education of the Russian Federation (Contract no. 075-15-2020-795 in the electronic budget system, agreement no. 13.1902.21.0027 dated September 29, 2020, project ID: RF-190220X0027).

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Authors and Affiliations

  1. Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, 119991, Moscow, Russia

    A. V. Petrovskaya, E. P. Barykin, A. M. Tverskoi, K. B. Varshavskaya, V. A. Mitkevich, I. Yu. Petrushanko & A. A. Makarov

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  1. A. V. Petrovskaya
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  2. E. P. Barykin
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  3. A. M. Tverskoi
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  4. K. B. Varshavskaya
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  5. V. A. Mitkevich
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  6. I. Yu. Petrushanko
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  7. A. A. Makarov
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Correspondence to A. A. Makarov.

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Conflict of interest. The authors declare no conflicts of interest.

Statement on the welfare of animals. All applicable international, national, and/or institutional principles for the care and use of animals were observed in the work.

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Translated by N. Onishchenko

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Petrovskaya, A.V., Barykin, E.P., Tverskoi, A.M. et al. Blood–Brain Barrier Transwell Modeling. Mol Biol 56, 1020–1027 (2022). https://doi.org/10.1134/S0026893322060140

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  • DOI: https://doi.org/10.1134/S0026893322060140

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