A novel in vitro human microglia model: Characterization of human monocyte-derived microglia

• Published in: Journal of neuroscience methods 2012-07, Vol.209 (1), p.79-89
• Main Authors: Etemad, Samar, Zamin, Rasheeda Mohd, Ruitenberg, Marc J., Filgueira, Luis
• Format: Article
• Language: English
• Subjects: CCL2; Cell Culture Techniques - methods; Cell Differentiation - immunology; Cell Lineage - immunology; Chemokine receptors; Flow Cytometry; GM-CSF; Human peripheral blood monocytes; Humans; Immunophenotyping; M-CSF; Microglia; Microglia - cytology; Microglia - immunology; Monocytes; Monocytes - cytology; Monocytes - immunology; Nervous system; NGF; Phagocytosis; Serum-free medium
• ISSN: 0165-0270; 1872-678X
• DOI: 10.1016/j.jneumeth.2012.05.025

[Display omitted] ► New culture protocol for the generation of human monocyte-derived microglia in vitro. ► Morphological, phenotypic and functional characterization of human monocyte-derived microglia. ► Expression of surface markers by human microglia. ► Chemokine receptors expression pattern in human microglia. ► Comparison of human microglia with monocytes and dendritic cells. Microglia are the innate immune cells of the central nervous system. They help maintaining physiological homeostasis and contribute significantly to inflammatory responses in the course of infection, injury and degenerative processes. To date, there is no standardized simple model available to investigate the biology of human microglia. The aim of this study was to establish a new human microglia model. For that purpose, human peripheral blood monocytes were cultured in serum free medium in the presence of M-CSF, GM-CSF, NGF and CCL2 to generate monocyte-derived microglia (M-MG). M-MG were clearly different in morphology, phenotype and function from freshly isolated monocytes, cultured monocytes in the absence of the cytokines and monocyte-derived dendritic cells (M-DC) cultured in the presence of GM-CSF and IL-4. M-MG acquired a ramified morphology with primary and secondary processes. M-MG displayed a comparable phenotype to the human microglia cell line HMC3, expressing very low levels of CD45, CD14 and HLA-DR, CD11b and CD11c; and undetectable levels of CD40, CD80 and CD83, and a distinct pattern of chemokine receptors (positive for CCR1, CCR2, CCR4, CCR5, CXCR1, CXCR3, CX3CR1; negative for CCR6 and CCR7). In comparison with M-DC, M-MG displayed lower T-lymphocyte stimulatory capacity, as well as lower phagocytosis activity. The described protocol for the generation of human monocyte-derived microglia is feasible, well standardized and reliable, as it uses well defined culture medium and recombinant cytokines, but no serum or conditioned medium. This protocol will certainly be very helpful for future studies investigating the biology and pathology of human microglia.