Liver X receptor agonists and altered cellular cholesterol metabolism modulate APP expression in cerebromicrovascular endothelial cells

Abstract

Several lines of evidence suggest a close relationship between dysregulated lipid and lipoprotein metabolism and the development of Alzheimer's disease (AD). Impaired clearance of cerebral amyloid beta (Aß) across the blood-brain barrier (BBB) may facilitate the onset and progression of AD. However, regulatory pathways governing amyloid precursor protein (APP) synthesis, secretion, and processing by brain capillary endothelial cells (BCEC; the anatomical basis of the BBB) are largely unknown. Therefore, we aimed to investigate whether BCEC are capable of endogenous APP synthesis and whether to what extent APP synthesis is under control of cellular cholesterol homeostasis. Primary porcine brain capillary endothelial cells (pBCEC) were used as an in vitro model of the BBB. Gene and protein expression levels of APP and cholesterol metabolism genes were determined by real-time PCR and immunoblotting, respectively. Effects of either liver X receptor (LXR) activation or altered cholesterol biosynthesis on APP metabolism were investigated in pBCEC treated with 24(S), 27-hydroxycholesterol (24OH-C, 27OH-C), non steroidal LXR ligand TO901317, cholesterol or simvastatin. Cholesterol biosynthesis and cholesterol ester formation from radiolabeled precursor [14C]-acetate was examined in pBCEC in the absence or presence of the conditions described above. APP secretion was investigated in a polarized in vitro model mimicking the BBB. Using the in vitro model of the BBB consisting of pBCEC, we demonstrate that endogenous APP synthesis by pBCEC is significantly increased in response to 24OH-C or 27OH-C, TO901317, cholesterol, or simvastatin, while secretion of APP into the culture medium was attenuated (25-43%). In parallel, LXR agonists reduced cholesterol biosynthesis by 30-80% while stimulating its esterification up to 2.5-fold by modifying HMG-CoA reductase (HMGCR), sterol regulatory element-binding protein (SREBP-2), andacyl-coenzyme A: cholesterol acyltransferase (ACAT-2) expression levels. In a polarized in vitro model mimicking the BBB, pBCEC secreted APP/Aß peptides in a polarized manner - preferentially to the basolateral compartment. Our data demonstrate the BBB as an additional source of cerebral APP/Aß. An imbalance in its cholesterol and APP metabolism might therefore contribute to the pathogenesis of AD, whereas modulation of cellular cholesterol metabolism could contribute to regulate APP synthesis by cerebromicrovascular endothelial cells.