Abstract W P359: Activation of Prostacyclin Receptor (IP) in Human Brain Microvascular Endothelial Cells Stimulates Production of SAPPa

Abstract

Previous studies have established that cerebral blood vessels possess high capacity to synthesize prostacyclin (PGI 2 ). Substantial evidence indicates that amyloid precursor protein (APP) and soluble APPα (sAPPα) have neurotrophic and nueroprotective properties. We hypothesized that activation of IP-cAMP pathway stimulates non-amyloidogenic processing of APP in cerebral endothelial cells. Western blotting was used to detect protein expression; real time RT-PCR was employed to measure mRNA levels of APP, and a disintegrin and metalloprotease 10 (ADAM10, an α processing enzyme); sAPPα secretion was detected in condition medium derived from culturing human brain microvascular endothelial cells (BMVECs), by ELISA. After human BMVECs were treated with PGI 2 analog iloprost (0.01, 0.05, 0.1, or 0.5 μM) for 3 days, protein levels of ADAM10 (n=7-9, P<0.05) and APP (n=6, P<0.05) were significantly increased in a concentration dependent manner, while β amyloidogenic processing enzyme BACE1, and other α processing enzymes such as ADAM9 and ADAM17 were not changed. mRNA levels of APP (n=7, P<0.05) and ADAM10 (n=4, P<0.05), as well as sAPPα production in human BMVECs (n=6, P<0.05) were also significantly augmented in response to iloprost treatment. Moreover, stimulation of human BMVECs with forskolin (40μM) significantly enhanced APP and ADAM10 protein expression. Most notably, iloprost significantly augmented protein expression of peroxisome proliferation-activated receptor-δ (PPARδ) and SIRT1. We further explored the role of PPARδ in APP processing. ADAM10 expression (n=6, P<0.05), but not APP and BACE1, was significantly increased in human BMVECs treated with PPARδ ligand GW501516 (30 or 100 nM). GW501516 (100 nM) also significantly increase sAPPα production (n=9-10, P<0.05). siRNA against PPARδ significantly suppressed basal levels of ADAM10, and blocked iloprost-induced ADAM10 expression (n=4, P<0.05). These findings suggest that activation of IP-cAMP pathway enhances non-amyloidogenic processing of APP and consequently increases production of neurotrophic molecule sAPPα in human BMVECs. Activation of PPARδ in part mediates iloprost-induced α processing of APP.