Trottier M, Kallio P, Wang W, Lipton H L
Department of Neurology, Evanston Hospital, Northwestern University, Evanston, Illinois 60201-1782, USA.
J Virol. 2001 Aug;75(16):7420-8. doi: 10.1128/JVI.75.16.7420-7428.2001.
The low-neurovirulence Theiler's murine encephalomyelitis viruses (TMEV), such as BeAn virus, cause a persistent infection of the central nervous system (CNS) in susceptible mouse strains that results in inflammatory demyelination. The ability of TMEV to persist in the mouse CNS has traditionally been demonstrated by recovering infectious virus from the spinal cord. Results of infectivity assays led to the notion that TMEV persists at low levels. In the present study, we analyzed the copy number of TMEV genomes, plus- to minus-strand ratios, and full-length species in the spinal cords of infected mice and infected tissue culture cells by using Northern hybridization. Considering the low levels of infectious virus in the spinal cord, a surprisingly large number of viral genomes (mean of 3.0 x 10(9)) was detected in persistently infected mice. In the transition from the acute (approximately postinfection [p.i.] day 7) to the persistent (beginning on p.i. day 28) phase of infection, viral RNA copy numbers steadily increased, indicating that TMEV persistence involves active viral RNA replication. Further, BeAn viral genomes were full-length in size; i.e., no subgenomic species were detected and the ratio of BeAn virus plus- to minus-strand RNA indicated that viral RNA replication is unperturbed in the mouse spinal cord. Analysis of cultured macrophages and oligodendrocytes suggests that either of these cell types can potentially synthesize high numbers of viral RNA copies if infected in the spinal cord and therefore account for the heavy viral load. A scheme is presented for the direct isolation of both cell types directly from infected spinal cords for further viral analyses.
低神经毒力的泰勒氏鼠脑脊髓炎病毒（TMEV），如BeAn病毒，可在易感小鼠品系中引起中枢神经系统（CNS）的持续性感染，导致炎症性脱髓鞘。传统上，TMEV在小鼠中枢神经系统中持续存在的能力是通过从脊髓中回收感染性病毒来证明的。感染性测定结果导致了TMEV低水平持续存在的观点。在本研究中，我们通过Northern杂交分析了感染小鼠和感染组织培养细胞脊髓中TMEV基因组的拷贝数、正负链比率和全长种类。考虑到脊髓中感染性病毒水平较低，在持续感染的小鼠中检测到了数量惊人的病毒基因组（平均为3.0×10⁹）。在从急性感染（大约感染后[p.i.]第7天）到持续感染（从p.i.第28天开始）阶段的转变过程中，病毒RNA拷贝数稳步增加，表示TMEV的持续存在涉及活跃的病毒RNA复制。此外，BeAn病毒基因组大小为全长；即未检测到亚基因组种类，且BeAn病毒正负链RNA的比率表明病毒RNA复制在小鼠脊髓中未受干扰。对培养的巨噬细胞和少突胶质细胞的分析表明，如果在脊髓中被感染，这两种细胞类型中的任何一种都可能潜在地合成大量病毒RNA拷贝，因此可解释病毒载量高的现象。本文提出了一种直接从感染脊髓中分离这两种细胞类型以进行进一步病毒分析的方案。
