title: Establishment and characterization of primary adult microglial culture in mice.
abstract: Microglial cells account for approximately 12-15 % of the cells in the central nervous system (CNS). Microglial cells are polarized by pathological stimuli such as cytokines, chemokines, and growth factors, and play important roles in the deterioration and repair of the CNS. Here, we established cultures of primary microglial cells isolated from the brains of adult C57/BL6 mice using Percoll density gradients. The cells were cultured and stained with antibodies against CD11b, glial fibrillary acidic protein, myelin basic protein and NeuN to determine microglial, astroglial, oligodendroglial, and neuronal cells respectively. Moreover, the cells were exposed to interferon-γ (IFNγ) plus interleukin-1β (IL-1β) or IL-4 for 24 h to demonstrate the activating phenotypes with inducible nitric oxide synthase (iNOS), Ym1, and Iba-1 immunoblotting. At least 95 % of the cultured cells were CD11b-positive and -negative for astroglial, neuronal, and oligodendrocyte markers. IFNγ plus IL-1β treatment resulted in classical activation, which was represented by an increase in iNOS. The cells also displayed alternative activation, which increased Ym1 when treated with IL-4. The present study indicates that the microglial cells isolated as described here are a useful tool for elucidating adult microglial function.
authors.affiliation: Department of Anatomy, Showa University School of Medicine, Shinagawa-Ku, Tokyo, Japan. taki@med.showa-u.ac.jp
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Ohtaki
authors.forename: Tomomi
authors.initials: T
authors.lastname: Tsumuraya
authors.forename: Dandan
authors.initials: D
authors.lastname: Song
authors.forename: Atsushi
authors.initials: A
authors.lastname: Sato
authors.forename: Kenji
authors.initials: K
authors.lastname: Ohara
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Miyamoto
authors.forename: Hiroyasu
authors.initials: H
authors.lastname: Nakano
authors.forename: Keisuke
authors.initials: K
authors.lastname: Kiriyama
authors.forename: Kenji
authors.initials: K
authors.lastname: Dohi
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Hiraizumi
authors.forename: Masaji
authors.initials: M
authors.lastname: Matsunaga
authors.forename: Seiji
authors.initials: S
authors.lastname: Shioda
mesh_terms: D000818:Animals; D001921:Brain / Q000166:cytology*; D039481:CD11b Antigen / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000378:metabolism; D016764:Cell Polarity / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology; D037102:Lectins / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008840:Microfilament Proteins / Q000378:metabolism; D017628:Microglia / Q000187:drug effects / Q000502:physiology*; D009419:Nerve Tissue Proteins / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D013997:Time Factors; D001619:beta-N-Acetylhexosaminidases / Q000378:metabolism
chemical_list: C491049:Aif1 protein, mouse; D039481:CD11b Antigen; D002135:Calcium-Binding Proteins; D016207:Cytokines; D037102:Lectins; D008840:Microfilament Proteins; D009419:Nerve Tissue Proteins; D052247:Nitric Oxide Synthase Type II; C429541:Chil3 protein, mouse; D001619:beta-N-Acetylhexosaminidases