title: RNA-sequencing reveals oligodendrocyte and neuronal transcripts in microglia relevant to central nervous system disease.
abstract: Expression profiling of distinct central nervous system (CNS) cell populations has been employed to facilitate disease classification and to provide insights into the molecular basis of brain pathology. One important cell type implicated in a wide variety of CNS disease states is the resident brain macrophage (microglia). In these studies, microglia are often isolated from dissociated brain tissue by flow sorting procedures [fluorescence-activated cell sorting (FACS)] or from postnatal glial cultures by mechanic isolation. Given the highly dynamic and state-dependent functions of these cells, the use of FACS or short-term culture methods may not accurately capture the biology of brain microglia. In the current study, we performed RNA-sequencing using Cx3cr1(+/GFP) labeled microglia isolated from the brainstem of 6-week-old mice to compare the transcriptomes of FACS-sorted versus laser capture microdissection (LCM). While both isolation techniques resulted in a large number of shared (common) transcripts, we identified transcripts unique to FACS-isolated and LCM-captured microglia. In particular, ∼50% of these LCM-isolated microglial transcripts represented genes typically associated with neurons and glia. While these transcripts clearly localized to microglia using complementary methods, they were not translated into protein. Following the induction of murine experimental autoimmune encephalomyelitis, increased oligodendrocyte and neuronal transcripts were detected in microglia, while only the myelin basic protein oligodendrocyte transcript was increased in microglia after traumatic brain injury. Collectively, these findings have implications for the design and interpretation of microglia transcriptome-based investigations.
authors.affiliation: Department of Neurology, Washington University School of Medicine, St. Louis MO.
authors.forename: Anne C
authors.initials: AC
authors.lastname: Solga
authors.affiliation: Department of Neurology, Washington University School of Medicine, St. Louis MO.
authors.forename: Winnie W
authors.initials: WW
authors.lastname: Pong
authors.affiliation: The Genome Institute, Washington University School of Medicine.
authors.forename: Jason
authors.initials: J
authors.lastname: Walker
authors.affiliation: The Genome Institute, Washington University School of Medicine.
authors.forename: Todd
authors.initials: T
authors.lastname: Wylie
authors.affiliation: The Genome Institute, Washington University School of Medicine.
authors.forename: Vincent
authors.initials: V
authors.lastname: Magrini
authors.affiliation: Department of Neurology, Washington University School of Medicine, St. Louis MO.
authors.forename: Anthony J
authors.initials: AJ
authors.lastname: Apicelli
authors.affiliation: The Genome Institute, Washington University School of Medicine.
authors.forename: Malachi
authors.initials: M
authors.lastname: Griffith
authors.affiliation: The Genome Institute, Washington University School of Medicine.
authors.forename: Obi L
authors.initials: OL
authors.lastname: Griffith
authors.affiliation: Department of Neurochemistry, National Institute of Neuroscience, Kodaira, Tokyo, Japan.
authors.forename: Shinichi
authors.initials: S
authors.lastname: Kohsaka
authors.affiliation: Department of Neurology, Washington University School of Medicine, St. Louis MO.
authors.forename: Gregory F
authors.initials: GF
authors.lastname: Wu
authors.affiliation: Department of Neurology, Washington University School of Medicine, St. Louis MO.
authors.forename: David L
authors.initials: DL
authors.lastname: Brody
authors.affiliation: The Genome Institute, Washington University School of Medicine.
authors.forename: Elaine R
authors.initials: ER
authors.lastname: Mardis
authors.affiliation: Department of Neurology, Washington University School of Medicine, St. Louis MO.
authors.forename: David H
authors.initials: DH
authors.lastname: Gutmann
mesh_terms: D000818:Animals; D001933:Brain Stem / Q000166:cytology; D000073883:CX3C Chemokine Receptor 1; D002493:Central Nervous System Diseases / Q000235:genetics*; D003584:Cytological Techniques / Q000379:methods; D005434:Flow Cytometry / Q000379:methods; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D017628:Microglia / Q000166:cytology / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D009836:Oligodendroglia / Q000166:cytology / Q000378:metabolism*; D012313:RNA / Q000302:isolation & purification*; D019707:Receptors, Chemokine / Q000172:deficiency / Q000235:genetics; D017423:Sequence Analysis, RNA
chemical_list: D000073883:CX3C Chemokine Receptor 1; C471885:Cx3cr1 protein, mouse; D019707:Receptors, Chemokine; D012313:RNA
keywords: fluorescence-activated cell sorting; laser-capture microdissection; macrophage