Involvement of DMT1 +IRE in the transport of lead in an in vitro BBB model

Abstract

Homeostasis of the central nervous system (CNS) microenvironment is maintained by the blood-brain barrier (BBB). The BBB is particularly vulnerable to lead (Pb) insults. This study was designed to test the hypothesis that divalent metal transporter 1 (DMT1), which is a divalent cation membrane transporter, was involved in transcellular transport across the BBB. An in vitro BBB model, which was a co-culture system of human umbilical vascular endothelial cells (ECV304) and rat glioma cells (C6), was established. Transendothelial electrical resistance (TEER) and fluoresceinisothiocyanate (FITC)-dextran permeability results showed that Pb exposure at the tested concentrations had no significant effects on intercellular tightness. Pb transport displayed properties that were associated with iron response element (IRE) positive isoform of DMT1. Accordingly, Pb transport was significantly blocked by iron (Fe). Moreover, ECV304 cells that were depleted of Fe with the chelator deferoxamine (DFO) demonstrated increased Pb transport. By transfecting ECV-304 cells with a DMT1 expression vector, overexpression of DMT1 promoted an increase in Pb transport. Treatment of ECV304 cells with DMT1 antisense oligonucleotides (ASONs) MA1 significantly inhibited the transport of Pb. Our results suggest that Pb is transported in the in vitro BBB model by a transporter with biochemical properties similar to those of the DMT1 IRE-positive isoform.