P4–073: BACE and C99 protein levels in primary astrocytes from the frontal cortex of Alzheimer's disease and normal control brains

The deposition of amyloid β peptide (Aβ) is one of the predominant pathological hallmarks of Alzheimer's disease (AD). Aβ derives from the sequential cleavage of amyloid precursor protein (APP) by β– and γ–secretases. BACE (beta–site APP–cleaving enzyme) was cloned and characterized as the most possible candidate of the long sought–after β–secretase. However, the mechanisms under the Aβ deposition in AD patients, particularly sporadic cases remain unclear. We have recently demonstrated that BACE mRNA and protein expression is elevated, and the elevation of BACE enzymatic activity is correlated with Aβ production in the temporal cortex and hippocampus of sporadic AD brain. Although the major cellular source of BACE in brain is extensively believed to be neurons, in vitro studies using rodent primary astrocytes and the human astrocytoma cell line have recently revealed that activated astrocytes can express BACE. The purpose of this study is to address the issue that glial cells, i.e. astrocytes, express BACE, and if do they, how glial BACE participates in Aβ production in AD brain. By using primary astrocytes cultured from brain autopsy samples of AD and non–demented elderly control (ND) subjects that enrolled in the Sun Health Research Institute Brain Donation Program, we conducted Western blot analysis, immunoprecipitation, ELISA, and enzymatic activity assay for comparative determination of BACE and C99 (a product cleaved by BACE) levels, and immunocytochemistry for comparison of subcellular localization of BACE, between AD and normal brains. Our initial experiments demonstrated that activated astrocytes from AD brain are more capable of participating in Aβ production than those from ND brain. The result suggests that glial BACE does play a role in Aβ production in AD brain.