title: An efficient method to limit microglia-dependent effects in astroglial cultures.
abstract: Microglia, the CNS resident macrophages, and astrocytes, the most abundant glial cell population, are both implicated in brain pathologies and can exhibit a pro-inflammatory phenotype. Microglial cells are known to rapidly and strongly react to brain insults. They will promote astrocyte activation and may lead to a vicious, self-perpetuating cycle of chronic inflammation. To obtain a better understanding of the individual role of both cell types, primary cells are frequently used in in vitro studies, but the purity of specific cell cultures remains rarely investigated. The aim of this study is to determine the effect of specific removal of microglial cells on the inflammatory properties of different glial cultures. Here, the removal of microglial contamination from mixed glial cultures to obtain astrocyte-enriched cultures was achieved using a magnetic cell sorting approach. Compared to mixed cultures, we clearly showed that these enriched cultures are only weakly activated by pro-inflammatory agents (lipopolysaccharide, interferon-γ or beta-amyloid peptide). This finding was confirmed using twice-sorted astrocyte-enriched cultures and microglia-free cultures composed of neurosphere-derived astrocytes. Thus, we present evidence that the magnitude of the pro-inflammatory response is linked to the percentage of microglia in cultures. Due to their high reactivity to various insults or pro-inflammatory stimuli, microglia-derived effects could be credited to astrocytes in mixed glial cultures. Therefore, we highlight the importance of monitoring the presence of microglia in glial cultures since they can affect the interpretation of the results, especially when inflammatory processes are studied.
authors.affiliation: Life Sciences Research Unit, Faculty of Science, Technology and Communication, University of Luxembourg, Campus Limpertsberg, 162A Avenue de la Faïencerie, L-1511 Luxembourg.
authors.forename: Sophie
authors.initials: S
authors.lastname: Losciuto
authors.forename: Gauthier
authors.initials: G
authors.lastname: Dorban
authors.forename: Sébastien
authors.initials: S
authors.lastname: Gabel
authors.forename: Audrey
authors.initials: A
authors.lastname: Gustin
authors.forename: Claire
authors.initials: C
authors.lastname: Hoenen
authors.forename: Luc
authors.initials: L
authors.lastname: Grandbarbe
authors.forename: Paul
authors.initials: P
authors.lastname: Heuschling
authors.forename: Tony
authors.initials: T
authors.lastname: Heurtaux
mesh_terms: D000818:Animals; D001253:Astrocytes / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D002469:Cell Separation / Q000379:methods*; D002478:Cells, Cultured; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D020869:Gene Expression Profiling; D007249:Inflammation / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000166:cytology* / Q000276:immunology / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction