Part 3/7: NTNeurons_6hpi_inactivatedvirus_dendrimer

Differentiated NT2 cells are a potentially useful model system to study herpes simples virus (HSV) replication in human neurons. These cells can be irreversibly differentiated into NT-neurons in the presence of retinoic acid and non-dividing cultures NT-neurons can be maintained for up to several months without retinoic acid. HSV is capable of infecting and replicating in differentiated NT-neurons and thus differentiated NT-neurons provide a ready source of human post-mitotic cells which can be useful to study certain aspects of the interaction of HSV and the neuron. Microarray analysis was used to examine how HSV-1 infection modulates cellular transcription in human NT-neurons, focusing on changes that take place during the first 24 hours following infection and compared to changes resulting from infection with inactivated virus. In addition, the transcriptional changes resulting from virus infection of neurons were compared to those observed in primary human fibroblasts. At early times after HSV infection a small number of cellular transcripts in NT-Neurons appear to be increased in abundance. Most of these transcripts that are up-regulated after infection are not unique to neuronal cells, and possibly represent a common cellular response to HSV infection. A few transcripts were not detected in infected human fibroblasts and may represent part of a transcriptional profile specific to this type of human neuronal cell. In contrast to the situation at early times after infection, analysis of cellular transcripts in NT-neurons at late times after infection is complicated by the overall profound decrease in cellular transcription. Thus, microarray analysis appeared to be most useful as a screening method for transcription changes following infection at early times after virus infection of NT-neurons. Two-condition experiment, NTNeurons infected with inactivated HSV-1 versus uninfected NTNeurons, 4 independent biological replicate sets (uninfected/infected) in dual channel array setup (Cy3/Cy5).

分化型NT2细胞（NT2 cells）是研究人类神经元内单纯疱疹病毒（herpes simplex virus, HSV）复制过程的潜在理想模型系统。该类细胞可在视黄酸（retinoic acid）存在的条件下被不可逆地诱导分化为NT神经元（NT-neurons），且无需添加视黄酸即可将处于非增殖状态的NT神经元培养物维持长达数月之久。HSV可在分化后的NT神经元中完成感染与复制过程，因此分化型NT2细胞可提供易于获取的人类有丝分裂后细胞来源，为研究HSV与神经元的相互作用机制提供了便利。本研究采用微阵列分析（microarray analysis），探究HSV-1感染对人类NT神经元细胞转录的调控作用，重点关注感染后最初24小时内发生的转录变化，并与灭活病毒感染所诱导的转录变化进行对照分析。此外，本研究还将神经元病毒感染引发的转录变化，与原代人成纤维细胞中观察到的转录变化进行了对比。在HSV感染早期，NT神经元中仅有少量细胞转录本的丰度出现上调。此类感染后上调的转录本中，绝大多数并非神经元细胞所特有，可能代表了细胞对HSV感染的通用应答反应。另有少量转录本在受感染的人成纤维细胞中未被检测到，可能属于这类人类神经元细胞特有的转录谱的一部分。与感染早期的情况相反，感染后期对NT神经元细胞转录本的分析会因细胞转录整体发生显著下调而变得复杂。因此，微阵列分析作为NT神经元病毒感染早期转录变化的筛查手段，展现出最佳应用价值。本实验为双分组对照：灭活HSV-1感染的NT神经元与未感染NT神经元，采用Cy3/Cy5双通道芯片检测体系，包含4组独立生物学重复样本（未感染/感染组）。