Neurobiology of Disease (Dec 2001)
Generation and Characterization of Immortalized Human Microglial Cell Lines: Expression of Cytokines and Chemokines
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 aggulutinin 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-1β, IL-6, IL-8, IL-10, IL-12, IL-15, and TNF-α. Using HMO6 cells, we investigated whether activation was induced by Amyloid-β fragments or lipopolysaccharide (LPS). Treatment of HMO6 cells with Amyloid-β 25–35 fragment (Aβ25–35) or Amyloid-β 1–42 fragment (Aβ1–42) led to increased expression of mRNA levels of cytokine/chemokine IL-8, IL-10, IL-12, MIP-1β MIP-1, and MCP-1, and treatment with LPS produced same results. Expression of TNF-α and MIP1-α was not detected in unstimulated HMO6 cells, but their expression was later induced by long-term exposure to Aβ25–35 or Aβ1–42. ELISA assays of spent culture media showed increased protein levels of TNF-α and IL-8 in HMO6 cells following treatment with Aβ25–35 or LPS. Taken together, our results demonstrate that treatment of human primary microglia and HMO6 immortalized human microglia cell line with Aβ25–35, Aβ1–42 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.
