Ex vivo propagation of infectious sheep scrapie agent in heterologous epithelial cells expressing ovine prion protein
Ex vivo propagation of infectious sheep scrapie agent in heterologous epithelial cells expressing ovine prion protein
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DOI：
10.1073/pnas.061337998
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发表时间：
2001-03-27
影响因子：
11.1
通讯作者：
Laude, H
中科院分区：
文献类型：
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作者：
Vilette, D;Andreoletti, O;Laude, H
传染性海绵状脑病或朊病毒病是影响人类和动物的致命中枢神经系统退行性疾病。朊病毒是非常规感染因子，其复制依赖于宿主朊病毒蛋白（PrP）。事实证明，将朊病毒传播到培养细胞是一项特别困难的任务，除了少数例外，它们的实验传播依赖于对实验动物的接种。在这里，我们报告了支持天然羊痒病繁殖的永久细胞系的开发，该模型是通过在兔上皮细胞系中稳定表达四环素调节的羊 PrP 基因而获得的。暴露于瘙痒病剂后，反复发现培养物积累了高水平的异常 PrP (PrPres)，细胞提取物在过度表达绵羊 PrP 的转基因小鼠中诱导了类似瘙痒病的疾病。这些培养物在传代时保持健康并稳定感染。这些数据表明（i）来自非神经元来源的培养细胞可以有效地复制朊病毒； (ii) 通过相关 PrP 基因的表达可以在体外跨越物种屏障。这种方法导致了天然传染性海绵状脑病病原体的离体繁殖（即，之前没有对啮齿动物进行实验适应），并且可能应用于人类或牛朊病毒。
Transmissible spongiform encephalopathies, or prion diseases, are fatal degenerative disorders of the central nervous system that affect humans and animals. Prions are nonconventional infectious agents whose replication depends on the host prion protein (PrP). Transmission of prions to cultured cells has proved to be a particularly difficult task, and with a few exceptions, their experimental propagation relies on inoculation to laboratory animals. Here, we report on the development of a permanent cell line supporting propagation of natural sheep scrapie, This model was obtained by stable expression of a tetracycline-regulatable ovine PrP gene in a rabbit epithelial cell line. After exposure to scrapie agent, cultures were repeatedly found to accumulate high revels of abnormal PrP (PrPres), Cell extracts induced a scrapie-like disease in transgenic mice overexpressing ovine PrP. These cultures remained healthy and stably infected upon subpassaging. Such data show that (i) cultivated cells from a nonneuronal origin can efficiently replicate prions; and (ii) species barrier can be crossed ex vivo through the expression of a relevant PrP gene. This approach led to the ex vivo propagation of a natural transmissible spongiform encephalopathy agent (i.e., without previous experimental adaptation to rodents) and might be applied to human or bovine prions.