Transcriptome sequencing reveals that LPS-triggered transcriptional responses in established microglia BV2 cell lines are poorly representative of primary microglia.
Transcriptome sequencing reveals that LPS-triggered transcriptional responses in established microglia BV2 cell lines are poorly representative of primary microglia.
复制标题
DOI：
10.1186/s12974-016-0644-1
复制
发表时间：
2016-07-11
影响因子：
9.3
通讯作者：
Chai YG
中科院分区：
文献类型：
--
作者：
Das A;Kim SH;Arifuzzaman S;Yoon T;Chai JC;Lee YS;Park KS;Jung KH;Chai YG
小胶质细胞是驻留在中枢神经系统的髓样细胞，可被感染、神经元损伤和炎症激活。已建立的BV2小胶质细胞系十多年来一直是用于研究神经炎症的主要体外模型，因为它们减少了对持续维持细胞准备和动物实验模型的要求。然而，最近有人对BV2细胞系作为模型系统的价值提出了质疑。我们使用三倍体RNA测序技术(RNA-seq)研究了原代和BV2小胶质细胞系的分子特征，采用了两种转录转录技术：全局转录生物学三倍体RNA-seq和实时定量PCR。我们分析了差异表达基因(DIG)以确定转录因子(TF)基序(5‘上游启动子的−950到+50碱基)和表观遗传机制。测序分析和质量评估表明，原代小胶质细胞具有明显的转录特征，并表达一组独特的转录物，以响应内毒素。在BV2小胶质细胞系中没有观察到这种小胶质细胞的特征。重要的是，我们观察到以前未知的转录因子(即IRF2、IRF5、IRF8、STAT1、STAT2和STAT5A)以及表观遗传调节因子KDM1A、NSD3和SETDB2都在原代小胶质细胞(PM)中显著和选择性地表达。虽然在PM中发现了BV2小胶质细胞系中已知的转录转录改变，但我们也在PM中观察到了几个在BV2小胶质细胞系中不常见的新的转录转录改变。总而言之，这些史无前例的发现表明，已建立的BV2小胶质细胞株可能不是PM的不良代表，我们为未来的神经炎症研究建立了一个资源。本文的在线版本(doi：10.1186/s12974-0160644-1)包含补充材料，授权用户可以使用。
Microglia are resident myeloid cells in the CNS that are activated by infection, neuronal injury, and inflammation. Established BV2 microglial cell lines have been the primary in vitro models used to study neuroinflammation for more than a decade because they reduce the requirement of continuously maintaining cell preparations and animal experimentation models. However, doubt has recently been raised regarding the value of BV2 cell lines as a model system. We used triplicate RNA sequencing (RNA-seq) to investigate the molecular signature of primary and BV2 microglial cell lines using two transcriptomic techniques: global transcriptomic biological triplicate RNA-seq and quantitative real-time PCR. We analyzed differentially expressed genes (DEGs) to identify transcription factor (TF) motifs (−950 to +50 bp of the 5′ upstream promoters) and epigenetic mechanisms. Sequencing assessment and quality evaluation revealed that primary microglia have a distinct transcriptomic signature and express a unique cluster of transcripts in response to lipopolysaccharide. This microglial signature was not observed in BV2 microglial cell lines. Importantly, we observed that previously unidentified TFs (i.e., IRF2, IRF5, IRF8, STAT1, STAT2, and STAT5A) and the epigenetic regulators KDM1A, NSD3, and SETDB2 were significantly and selectively expressed in primary microglia (PM). Although transcriptomic alterations known to occur in BV2 microglial cell lines were identified in PM, we also observed several novel transcriptomic alterations in PM that are not frequently observed in BV2 microglial cell lines. Collectively, these unprecedented findings demonstrate that established BV2 microglial cell lines are probably a poor representation of PM, and we establish a resource for future studies of neuroinflammation. The online version of this article (doi:10.1186/s12974-016-0644-1) contains supplementary material, which is available to authorized users.