| Summary: | <p>Abstract</p> <p>Background</p> <p>The transmissible spongiform encephalopathies, otherwise known as prion diseases, occur following the conversion of the cellular prion protein (PrP<sup>C</sup>) to an alternatively folded, disease-associated isoform (PrP<sup>Sc</sup>). Recent studies suggest that this conversion occurs via a cholesterol-sensitive process, as cholesterol synthesis inhibitors reduced the formation of PrP<sup>Sc </sup>and delayed the clinical phase of scrapie infection. Since polyunsaturated fatty acids also reduced cellular cholesterol levels we tested their effects on PrP<sup>Sc </sup>formation in three prion-infected neuronal cell lines (ScGT1, ScN2a and SMB cells).</p> <p>Results</p> <p>We report that treatment with docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA) or the cholesterol synthesis inhibitor simvastatin reduced the amounts of free cholesterol in membrane extracts from prion-infected neuronal cells. Simvastatin reduced cholesterol production while DHA and EPA promoted the conversion of free cholesterol to cholesterol esters. Crucially, while simvastatin reduced PrP<sup>Sc </sup>formation, both DHA and EPA significantly increased the amounts of PrP<sup>Sc </sup>in these cells. Unlike simvastatin, the effects of DHA and EPA on PrP<sup>Sc </sup>content were not reversed by stimulation of cholesterol synthesis with mevalonate. Treatment of ScGT1 cells with DHA and EPA also increased activation of cytoplasmic phospholipase A<sub>2 </sub>and prostaglandin E<sub>2 </sub>production. Finally, treatment of neuronal cells with DHA and EPA increased the amounts of PrP<sup>C </sup>expressed at the cell surface and significantly increased the half-life of biotinylated PrP<sup>C</sup>.</p> <p>Conclusion</p> <p>We report that although treatment with DHA or EPA significantly reduced the free cholesterol content of prion-infected cells they significantly increased PrP<sup>Sc </sup>formation in three neuronal cell lines. DHA or EPA treatment of infected cells increased activation of phospholipase A<sub>2</sub>, a key enzyme in PrP<sup>Sc </sup>formation, and altered the trafficking of PrP<sup>C</sup>. PrP<sup>C </sup>expression at the cell surface, a putative site for the PrP<sup>Sc </sup>formation, was significantly increased, and the rate at which PrP<sup>C </sup>was degraded was reduced. Cholesterol depletion is seen as a potential therapeutic strategy for prion diseases. However, these results indicate that a greater understanding of the precise relationship between membrane cholesterol distribution, PrP<sup>C </sup>trafficking, cell activation and PrP<sup>Sc </sup>formation is required before cholesterol manipulation can be considered as a prion therapeutic.</p>
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