Cholesterol and Aβ Production: Methods for Analysis of Altered Cholesterol De Novo Synthesis

Abstract

The main characteristics of Alzheimer’s disease (AD) are massive cerebral accumulation of amyloid composed of fibrillary aggregates of the amyloid beta peptide (Ab) and intracellular accumulation of abnormally phosphorylated tau protein, associated with widespread neurodegeneration. The clinical picture is characterized by progressive and irreversible dementia, which is eventually fatal (for reviews see refs. [1–4]). Up to now four genes have been identified, harboring point mutations that significantly affect AD pathogenesis [5]. All of these mutations result in increased b-amyloid 42 (Ab42) levels [6]. Three of these genes-the amyloid precursor protein (APP), presenilin 1 (PS1), and presenilin 2 (PS2)—are involved in the molecular pathway of AD. The fourth gene, apolipoprotein E (ApoE) suggests a possible link to lipid pathways. ApoE encodes a lipidbinding protein that transports lipids between cells and is therefore an important factor in lipid homeostasis [7]. HumanApoE exists in three major alleles. In epidemiological studies the 4 allele of ApoE increases the risk for hypercholesterolemia and decreases the age of onset of AD [8]. Moreover, ApoE transgenic and knockout mice show altered Ab deposition, indicating that ApoE and lipids might play a significant role in Ab pathology [9]. Alterations in lipid homeostasis have long been recognized to interfere severely with neuronal and glial functions and to cause neurodegenerative diseases [10]. Studies on the infrequent familial forms of the disease (FAD), especially with presenilin FAD, indicate increased production of Ab42 to be associated invariably with FAD and to cause AD [1,2,5,11,12]. Ab is a proteolytic fragment of APP, a protein of unclear function. The BACE I-catalyzed cleavage of APP initiates Ab generation; and the resulting C-terminal fragment C99—but not full-length APP—is a substrate for g-secretase, a ubiquitous multimeric