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Generation and characterization of immortalized human microglial cell lines: expression of cytokines and chemokines.

Authors
Nagai, A; Nakagawa, E; Hatori, K; Choi, HB; McLarnon, JG; Lee, MA; Kim, SU
Citation
Neurobiology of disease, 8(6):1057-1068, 2001
Journal Title
Neurobiology of disease
ISSN
0969-99611095-953X
Abstract
Microglia are a major glial component of the central nervous system (CNS), play a critical role as resident immunocompetent and phagocytic cells in the CNS, and serve as scavenger cells in the event of infection, inflammation, trauma, ischemia, and neurodegeneration in the CNS. Studies of human microglia have been hampered by the difficulty of obtaining sufficient numbers of human microglia. One way to circumvent this difficulty is to establish permanent cell lines of human microglia. In the present study we report the generation of immortalized human microglial cell line, HMO6, from human embryonic telencephalon tissue using a retroviral vector encoding myc oncogene. The HMO6 cells exhibited cell type-specific antigens for microglia-macrophage lineage cells including CD11b (Mac-1), CD68, CD86 (B7-2), HLA-ABC, HLA-DR, and ricinus communis agglutinin lectin-1 (RCA), and actively phagocytosed latex beads. In addition, HMO6 cells showed ATP-induced responses similar to human primary microglia in Ca2+ influx spectroscopy. Both human primary microglia and HMO6 cells showed the similar cytokine gene expression in IL-1beta, IL-6, IL-8, IL-10, IL-12, IL-15, and TNF-alpha. Using HMO6 cells, we investigated whether activation was induced by Amyloid-beta fragments or lipopolysaccharide (LPS). Treatment of HMO6 cells with Amyloid-beta 25-35 fragment (Abeta(25-35)) or Amyloid-beta 1-42 fragment (Abeta(1-42)) led to increased expression of mRNA levels of cytokine/chemokine IL-8, IL-10, IL-12, MIP-1beta MIP-1, and MCP-1, and treatment with LPS produced same results. Expression of TNF-alpha and MIP1-alpha was not detected in unstimulated HMO6 cells, but their expression was later induced by long-term exposure to Abeta(25-35) or Abeta(1-42.) ELISA assays of spent culture media showed increased protein levels of TNF-alpha and IL-8 in HMO6 cells following treatment with Abeta(25-35) or LPS. Taken together, our results demonstrate that treatment of human primary microglia and HMO6 immortalized human microglia cell line with Abeta(25-35), Abeta(1-42) and LPS upregulate gene expression and protein production of proinflammatory cytokines and chemokines in these cells. The human microglial cell line HMO6 exhibits similar properties to those documented in human microglia and should have considerable utility as an in vitro model for the studies of human microglia in health and disease.
MeSH terms
Adenosine Triphosphate/metabolismAdenosine Triphosphate/pharmacologyAmyloid beta-Peptides/pharmacologyCalcium Signaling/drug effectsCalcium Signaling/physiologyCell Culture Techniques/methodsCell Line, Transformed/drug effectsCell Line, Transformed/immunologyCell Line, Transformed/metabolism*Central Nervous System/cytology*Central Nervous System/immunologyCentral Nervous System/metabolismChemokines/geneticsChemokines/metabolism*Coloring Agents/diagnostic useCytokines/geneticsCytokines/metabolism*FetusFura-2/diagnostic useGene Expression Regulation/physiology*Genetic VectorsHumansImmunohistochemistryLipopolysaccharides/pharmacologyMicroglia/drug effectsMicroglia/immunologyMicroglia/metabolism*Peptide Fragments/pharmacologyRNA, Messenger/drug effectsRNA, Messenger/metabolismReceptors, Cell Surface/drug effectsReceptors, Cell Surface/metabolismReverse Transcriptase Polymerase Chain ReactionTransfection/methods
DOI
10.1006/nbdi.2001.0437
PMID
11741401
Appears in Collections:
Journal Papers > School of Medicine / Graduate School of Medicine > Brain Science
Journal Papers > School of Medicine / Graduate School of Medicine > Neurology
AJOU Authors
이, 명애김, 승업
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