Transient blood–brain barrier disruption is induced by low pulsed electrical fields in vitro: an analysis of permeability and trans-endothelial electric resistivity

Shirley Sharabi,Yael Bresler,Orly Ravid,Chen Shemesh,Dana Atrakchi,Michal Schnaider-Beeri,Fabien Gosselet,Lucie Dehouck,David Last,David Guez,Dianne Daniels,Yael Mardor,Itzik Cooper

doi:10.1080/10717544.2019.1571123

Abstract

The blood–brain barrier (BBB) is limiting transcellular and paracellular movement of molecules and cells, controls molecular traffic, and keeps out toxins. However, this protective function is the major hurdle for treating brain diseases such as brain tumors, Parkinson’s disease, Alzheimer’s disease, etc. It was previously demonstrated that high pulsed electrical fields (PEFs) can disrupt the BBB by inducing electroporation (EP) which increases the permeability of the transcellular route. Our goal was to study the effects of low PEFs, well below the threshold of EP on the integrity and function of the BBB. Ten low voltage pulses (5–100 V) were applied to a human in vitro BBB model. Changes in permeability to small molecules (NaF) were studied as well as changes in impedance spectrum and trans-endothelial electric resistivity. Viability and EP were evaluated by Presto-Blue and endogenous Lactate dehydrogenase release assays. The effect on tight junction and adherent junction protein was also studied. The results of low voltage experiments were compared to high voltage experiments (200–1400 V). A significant increase in permeability was found at voltages as low as 10 V despite EP only occurring from 100 V. The changes in permeability as a function of applied voltage were fitted to an inverse-exponential function, suggesting a plateau effect. Staining of VE-cadherin showed specific changes in protein expression. The results indicate that low PEFs can transiently disrupt the BBB by affecting the paracellular route, although the mechanism remains unclear.

Highlights

The blood–brain barrier (BBB) is composed of brain endothelial cells, pericytes, and astrocytes and is located at the brain microvessel level
Claudin-5 and Occludin are linked via zonula occludens (ZO) protein complexes to scaffolding proteins ZO-1, ZO-2, and ZO-3 that bind to the actin/myosin cytoskeletal system, resulting in modification of the tight junctions (TJs) properties (Obermeier et al, 2013; van Tellingen et al, 2015)
Since the viability assays revealed no cell death, we measured the lactate dehydrogenase (LDH) levels in the medium after pulsed electrical fields (PEFs) application in order to assess whether BBB disruption can be explained by EP of the brain-like endothelial cells (BLECs)’s membranes

Summary

Introduction

The blood–brain barrier (BBB) is composed of brain endothelial cells, pericytes, and astrocytes and is located at the brain microvessel level. TJ associated proteins include occludin, claudin-5, and junctional adhesion molecules (Abbott et al, 2006; Abbott, 2013). Claudin-5 and Occludin are linked via zonula occludens (ZO) protein complexes to scaffolding proteins ZO-1, ZO-2, and ZO-3 that bind to the actin/myosin cytoskeletal system, resulting in modification of the TJs properties (Obermeier et al, 2013; van Tellingen et al, 2015). The endothelium acts as a dynamic barrier limiting transcellular and paracellular movement of molecules and cells and its main functions include control of molecular traffic and keeping out toxins (Abbott, 2013)

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