Alteration of beta-amyloid clearance by the blood - cerebrospinal fluid barrier following lead exposure
Lead (Pb) toxicity remains a major public health concern due to its continued use in the modern industry. Recent evidence demonstrates an association between occupational Pb exposure and cognitive dysfunction. One of the major pathological hallmarks of Alzheimer’s disease (AD) is the extensive presence of beta-amyloid (Aβ) in the brain and cerebrospinal fluid (CSF). Autopsies of brains from AD patients reveal distinct Aβ stains in the choroid plexus (CP), a region where the blood-cerebrospinal fluid barrier (BCB) is located. The CP, in addition to being involved in the uptake and metabolism of Aβ from the CSF, has shown to be a target for Pb accumulation in the brain. ^ Currently there is no known link between Pb exposure and Aβ regulation at the BCB. Hence, this study was designed to investigate a possible link between Pb exposure and alteration in Aβ transport and clearance by the CP by potentially influencing: (i) alterations in the expression and/or subcellular localization of Aβ efflux transporter, Lipoprotein Receptor Protein 1 (LRP1), and/or (ii) alterations in the expression, localization and/or activitiy of Aβ degrading enzyme, Insulin Degrading Enzyme (IDE). Results from laser scanning cytometry studies revealed a significant increase in intracellular Aβ levels in Pb-exposed rats relative to controls (p<0.001). In vitro ELISA studies in choroid plexus epithelial (Z310) cells demonstrated a dose-dependent and time-related increase in Aβ accumulation following 24 h or 48 h of Pb exposure.^ To explore a mechanism for the above finding, the involvement of Aβ transporter, LRP1 was assessed. Results from real time RT-PCR and Western blot analysis revealed a significant decrease by 35% (p<0.05) in mRNA and 31.8% (p<0.05) in protein expression in the CP of rats exposed to Pb compared to controls. In vitro studies revealed a 33.1% and 33.4% decrease in the protein expression of LRP1 (p<0.05) following Pb exposure (10 μM) for 24 h or 48 h respectively. Knocking down LRP1 by siRNA resulted in an increase in the cellular accumulation of Aβ, from 31% to 72% (p<0.05). ^ A distinct subcellular relocalization of LRP1 was observed from the cytosol in control tissues to the apical surface and microvilli following Pb exposure. Double immunostaining of LRP1 with PKC-δ revealed a co-localization of both in the cytosol of controls with a distinct relocalization following Pb treatment. Interestingly, pre-incubation with PKC-δ inhibitor rottlerin, abolished the Pb-induced relocalization of LRP1 as well as the Pb-induced Aβ accumulation. ^ Next, we assessed the involvement of IDE, an Aβ degrading enzyme. Results from the IDE enzymatic activity assay revealed a significant decrease in activity in cells exposed to Pb compared to controls with a corresponding increase in Aβ accumulation. There was no visible relocalization of IDE either in the CP tissues or in Z310 cells following Pb exposure, as indicated by immunohistochemistry and confocal imaging. Additionally, there was no significant difference in IDE mRNA or protein expression as determined by real time RT-PCR and Western blot analysis. ^ Based on our findings, we conclude that Pb exposure results in Aβ dysregulation at the BCB by a combination of mechanisms involving LRP1 and IDE. These alterations in Aβ clearance in the CP may have significant implications for the subsequent build up of Aβ in surrounding brain regions like the hippocampus and cortex, and may potentially result in exacerbating AD-like pathology.^
Wei Zheng, Purdue University.
Health Sciences, Toxicology