Using primary hippocampal neurons transfected with APP by means of the Semliki-forest-virus expression system Christine Bergmann (319.2) demonstrated that lovastatin (which inhibits de novo cholesterol synthesis) and methyl-b-cyclodextrin (which can form inclusion complexes with plasma membrane cholesterol) cause a dramatic decrease in both intracellular and secreted Aβ40 and 42 and an increase in C99. By fluorescence microscopy of filipin-cholesterol complexes, she nicely demonstrated that even when cholesterol levels were reduced by 60%-70% that the neurons were still viable and that the Golgi network managed to retain some cholesterol. To determine if γ-secretase processing was also affected by cholesterol she transfected neurons with SPA4CT (C99, an APP construct that does not require beta-secretase cleavage) and assessed the effect of lovastatin and β-cyclodextrin. In contrast to her results using FLAPP she found that Aβ42 generation is resistant to extracellular cholesterol depletion, whereas Aβ40 is dramatically reduced. This differential effect on Aβ40 production probably results from Aβ40 being produced in a compartment (the TGN) which is more accessible to the effects of cyclodextrin than the IC/ER where Aβ42 is generated. Together these results demonstrate that both β- and γ-secretase are strongly influenced by their lipid environment and that modulation of cholesterol levels may be a useful therapeutic strategy.
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