Increased vascular permeability to Evans Blue dye in the hippocampus of PDAPPJ20 mice, model of Alzheimer's disease (AD): potential implication of ER stress mechanisms

Abstract

The blood-brain barrier (BBB) limits flux from and into the brain compartment that is essential for normal neuronal functioning and information processing. Post-mortem tissue analysis indicates BBB damage in AD. However, timing and mechanisms underlying BBB breakdown remain elusive. Endoplasmic reticulum (ER) stress is caused by disruption of homeostatic mechanisms that cause unfolded proteins to accumulate. The ER adapts to stress by activating the unfolded protein response (UPR). Chronic ER stress is increasingly being associated to neurodegeneration. We found augmented vascular permeability in the hippocampus of 12-month-old PDAPPJ20 transgenic mice, compared to the non-transgenic group, assessed by Evans Blue i.v. injections and analysis of coronal brain sections (D.O.= 5666±387 vs 12373±3082 n=5-6 p<0,04) and a higher number of microhemorrhages (four fold more in transgenic mice vs non-transgenic mice p<0,001). Human brain microvascular endothelial cells (hBMEC) proved to be a very suitable human cell line for an in vitro BBB model. We treated hBMEC with 1 ug/ul of Thapsigargin (Tg), a known inductor of ER stress. Treated cells showed increased mRNA expression of Ire1 and PERK by RT-qPCR (two and three fold increase n=3, p=0.01 respectively), evidencing UPR activation. We also employed a sealed monolayer of hBMEC on a transwell membrane, monitored by TEER. Tg exposure provoked loss of resistance and augmented permeability to Evans Blue and NaFl. In addition, AB1-40 peptide was able to induce changes in GRP78/BiP -a central central regulator for ER stress- in endothelial cells: we found not only increased cytoplasmic BiP area (p<0.05) but a different pattern distribution, with predominance of bright dots, suggesting the implication of ER in the brain vasculature exposed to amyloid peptides. Further experiments are in progress to elucidate the role of ER stress in endothelial cells and BBB integrity in vivo and in vitro models of AD.