Iwata Naoko, Yoshida Hiroaki, Tobiume Minoru, Ono Fumiko, Shimazaki Takuya, Sata Tetsutaro, Nakajima Noriko
Department of Pathology, National Institute of Infectious Disease, Tokyo, Japan.
J Neurovirol. 2007;13(1):11-22. doi: 10.1080/13550280601086064.
The pathogenesis of neurologic dysfunctions caused by human immunodeficiency virus type 1 (HIV-1) infection is not yet well understood. Simian immunodeficiency virus (SIV) infection of macaques is an important animal model for HIV-1 infection. This is the first report to characterize brain progenitor cells (BPCs) isolated from embryonic brain of cynomolgus monkeys (Macaca fascicularis) by neurosphere assay and utilize BPC-derived cell culture for studying SIV infection. The self-renewal and multilineage differentiation properties of BPCs are convenient for planning viral infection experiments. The BPC-derived culture does not contain macrophage/microglial cells, fibroblasts, or endothelial cells. Thus, this culture is appropriate for studying direct relation between SIV infection and neuronal and glial cells. First, the authors characterized undifferentiated and differentiated simian BPCs by immunocytochemistry, flow cytometry analysis, real-time polymerase chain reaction (PCR), and reverse transcriptase (RT)-PCR. The BPCs induced to differentiate by the addition of 1% fetal bovine serum (FBS) were composed of heterogeneous cells expressing nestin, glial fibrillary acidic protein (GFAP), and/or tubulin beta III isoform (Tuj). None of them expressed the monocyte/macrophage/microglial marker. mRNA expression of CD4, CXCR4, CCR5, GPR1, STRL33, and APJ in both undifferentiated and differentiated BPCs were shown by RT-PCR method, suggesting that SIV would infect and replicate in this culture system. Then, it was confirmed that the neurotropic SIV strain, SIV17/E-Fr, replicated productively in BPC-derived cells. The SIV/17E-FrDelta nefGFP was inoculated to identify the infected cells and immunocytochemistry analysis revealed that green fluorescent protein (GFP)-expressing cells were mostly GFAP positive and coexpressed with SIV p27 antigen. Thus, BPC-derived cell culture system is applicable for studying SIV infection in glial and neuronal cells.
1型人类免疫缺陷病毒（HIV-1）感染所致神经功能障碍的发病机制尚未完全明确。猕猴感染猿猴免疫缺陷病毒（SIV）是HIV-1感染的重要动物模型。这是首篇通过神经球分析法对从食蟹猴（猕猴属）胚胎脑中分离出的脑祖细胞（BPC）进行特性描述，并利用BPC来源的细胞培养物研究SIV感染的报告。BPC的自我更新和多谱系分化特性便于规划病毒感染实验。BPC来源的培养物不含巨噬细胞/小胶质细胞、成纤维细胞或内皮细胞。因此，这种培养物适合研究SIV感染与神经元和神经胶质细胞之间的直接关系。首先，作者通过免疫细胞化学、流式细胞术分析、实时聚合酶链反应（PCR）和逆转录酶（RT）-PCR对未分化和分化的猿猴BPC进行了特性描述。添加1%胎牛血清（FBS）诱导分化的BPC由表达巢蛋白、胶质纤维酸性蛋白（GFAP）和/或βIII微管蛋白异构体（Tuj）的异质性细胞组成。它们均未表达单核细胞/巨噬细胞/小胶质细胞标志物。通过RT-PCR方法显示了未分化和分化的BPC中CD4、CXCR4、CCR5、GPR1、STRL33和APJ的mRNA表达，表明SIV会在该培养系统中感染和复制。然后，证实了嗜神经SIV毒株SIV17/E-Fr在BPC来源的细胞中高效复制。接种SIV/17E-FrDelta nefGFP以鉴定感染细胞，免疫细胞化学分析显示，表达绿色荧光蛋白（GFP）的细胞大多为GFAP阳性，并与SIV p27抗原共表达。因此，BPC来源的细胞培养系统适用于研究SIV在神经胶质细胞和神经元细胞中的感染。
