title: Transcriptome sequencing reveals that LPS-triggered transcriptional responses in established microglia BV2 cell lines are poorly representative of primary microglia.
abstract: 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.
authors.affiliation: Institute of Natural Science and Technology, Hanyang University, Ansan, 15588, Republic of Korea.
authors.forename: Amitabh
authors.initials: A
authors.lastname: Das
authors.affiliation: Department of Molecular and Life Sciences, Hanyang University, Ansan, 15588, Republic of Korea.
authors.forename: Sun Hwa
authors.initials: SH
authors.lastname: Kim
authors.affiliation: Department of Bionanotechnology, Hanyang University, Seoul, 04673, Republic of Korea.
authors.forename: Sarder
authors.initials: S
authors.lastname: Arifuzzaman
authors.affiliation: Department of Molecular and Life Sciences, Hanyang University, Ansan, 15588, Republic of Korea.
authors.forename: Taeho
authors.initials: T
authors.lastname: Yoon
authors.affiliation: Department of Molecular and Life Sciences, Hanyang University, Ansan, 15588, Republic of Korea.
authors.forename: Jin Choul
authors.initials: JC
authors.lastname: Chai
authors.affiliation: Department of Molecular and Life Sciences, Hanyang University, Ansan, 15588, Republic of Korea.
authors.forename: Young Seek
authors.initials: YS
authors.lastname: Lee
authors.affiliation: Institute of Natural Science and Technology, Hanyang University, Ansan, 15588, Republic of Korea.
authors.forename: Kyoung Sun
authors.initials: KS
authors.lastname: Park
authors.affiliation: Institute of Natural Science and Technology, Hanyang University, Ansan, 15588, Republic of Korea. khjung2@gmail.com.
authors.forename: Kyoung Hwa
authors.initials: KH
authors.lastname: Jung
authors.affiliation: Department of Molecular and Life Sciences, Hanyang University, Ansan, 15588, Republic of Korea. ygchai@hanyang.ac.kr.|Department of Bionanotechnology, Hanyang University, Seoul, 04673, Republic of Korea. ygchai@hanyang.ac.kr.
authors.forename: Young Gyu
authors.initials: YG
authors.lastname: Chai
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D001921:Brain / Q000166:cytology; D002478:Cells, Cultured; D016207:Cytokines / Q000235:genetics / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005786:Gene Expression Regulation / Q000187:drug effects*; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D017628:Microglia / Q000187:drug effects*; D012333:RNA, Messenger / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013997:Time Factors; D051197:Toll-Like Receptor 4 / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D059467:Transcriptome / Q000187:drug effects* / Q000502:physiology
chemical_list: D016207:Cytokines; D008070:Lipopolysaccharides; D012333:RNA, Messenger; C493487:Tlr4 protein, mouse; D051197:Toll-Like Receptor 4; D014157:Transcription Factors
keywords: Gene regulation; Innate immunity; Microglia; RNA sequencing; Transcription factors