Bate Clive, Tayebi Mourad, Diomede Luisa, Salmona Mario, Williams Alun
Department of Pathology and Infectious Diseases, Royal Veterinary College, Hawkshead Lane, North Mymms, Herts, UK, AL9 7TA.
BMC Biol. 2008 Sep 12;6:39. doi: 10.1186/1741-7007-6-39.
The transmissible spongiform encephalopathies, otherwise known as prion diseases, occur following the conversion of the cellular prion protein (PrPC) to an alternatively folded, disease-associated isoform (PrPSc). Recent studies suggest that this conversion occurs via a cholesterol-sensitive process, as cholesterol synthesis inhibitors reduced the formation of PrPSc and delayed the clinical phase of scrapie infection. Since polyunsaturated fatty acids also reduced cellular cholesterol levels we tested their effects on PrPSc formation in three prion-infected neuronal cell lines (ScGT1, ScN2a and SMB cells).
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 PrPSc formation, both DHA and EPA significantly increased the amounts of PrPSc in these cells. Unlike simvastatin, the effects of DHA and EPA on PrPSc 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 A2 and prostaglandin E2 production. Finally, treatment of neuronal cells with DHA and EPA increased the amounts of PrPC expressed at the cell surface and significantly increased the half-life of biotinylated PrPC.
We report that although treatment with DHA or EPA significantly reduced the free cholesterol content of prion-infected cells they significantly increased PrPSc formation in three neuronal cell lines. DHA or EPA treatment of infected cells increased activation of phospholipase A2, a key enzyme in PrPSc formation, and altered the trafficking of PrPC. PrPC expression at the cell surface, a putative site for the PrPSc formation, was significantly increased, and the rate at which PrPC 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, PrPC trafficking, cell activation and PrPSc formation is required before cholesterol manipulation can be considered as a prion therapeutic.
传染性海绵状脑病，又称朊病毒病，是在细胞朊蛋白（PrPC）转变为另一种折叠形式的、与疾病相关的异构体（PrPSc）后发生的。最近的研究表明，这种转变通过一个对胆固醇敏感的过程发生，因为胆固醇合成抑制剂减少了PrPSc的形成并延迟了羊瘙痒症感染的临床阶段。由于多不饱和脂肪酸也降低了细胞胆固醇水平，我们测试了它们对三种朊病毒感染的神经元细胞系（ScGT1、ScN2a和SMB细胞）中PrPSc形成的影响。
我们报告，用二十二碳六烯酸（DHA）、二十碳五烯酸（EPA）或胆固醇合成抑制剂辛伐他汀处理可降低朊病毒感染的神经元细胞膜提取物中的游离胆固醇含量。辛伐他汀减少胆固醇生成，而DHA和EPA促进游离胆固醇向胆固醇酯的转化。至关重要的是，虽然辛伐他汀减少了PrPSc的形成，但DHA和EPA均显著增加了这些细胞中PrPSc的量。与辛伐他汀不同，用甲羟戊酸刺激胆固醇合成并不能逆转DHA和EPA对PrPSc含量的影响。用DHA和EPA处理ScGT1细胞也增加了细胞质磷脂酶A2的活性和前列腺素E2的产生。最后，用DHA和EPA处理神经元细胞增加了细胞表面表达的PrPC量，并显著增加了生物素化PrPC的半衰期。
我们报告，虽然用DHA或EPA处理显著降低了朊病毒感染细胞的游离胆固醇含量，但它们显著增加了三种神经元细胞系中PrPSc的形成。用DHA或EPA处理感染细胞增加了磷脂酶A2的活性，磷脂酶A2是PrPSc形成中的关键酶，并改变了PrPC的运输。细胞表面PrPC的表达是PrPSc形成的假定部位，显著增加，且PrPC的降解速率降低。胆固醇耗竭被视为朊病毒病的一种潜在治疗策略。然而，这些结果表明，在将胆固醇调控视为朊病毒治疗方法之前，需要更深入了解膜胆固醇分布、PrPC运输、细胞激活和PrPSc形成之间的精确关系。
