A SIMPLIFIED METHOD FOR EFFICIENT ISOLATION OF FUNCTIONAL MICROGLIAL CELLS: APPLICATION FOR MODELLING NEUROINFLAMMATORY RESPONSES IN VITRO

SEPULVEDA DIAZ, J.; OUIDJA O.; SOCIAS, S.B.; HAMADAT S.; GUERREIRO, S.; RAISMAN VOZARI R.; MICHEL, P.

Copenhague

Congreso; 10th Forum of Neuroscience; 2016

Federation of European Neuroscience Societies (FENS)

Microglial cells represent the major component of the innate immune system of the brain. When inappropriate and persistent, the activation of microglial cells leads to inflammatory processes that participate actively to the progression of a number of pathological conditions. As a consequence, purified microglial cells in culture are frequently used to model brain inflammatory responses but obtaining bulk quantities of these cells on a routine basis can be quite challenging.Here, we show that it is possible to achieve high-yield isolation of pure microglial cells frompost-natal mouse brain through a procedure that essentially relies onthe adhesionpreference of these cells for the polycationpolyethyleneimine (PEI).Accordingly, the replacement of PEI by other synthetic or biological substrates failed to promote microglial cell enrichment.When generated in large flasks coated with PEI, cultures of microglial cells were easily recovered by trypsin proteolysis to produce subcultures for functional studies. These cultures responded to lipopolysaccharide (LPS, 1-10 ng/mL) treatment by secreting proinflammatory cytokines such as TNF-𝛼, IL-6, IL-1and by generating nitric oxide and reactive oxygen species. Most interestingly, this response was curtailed by appropriate reference drugs. Microglial cells were also strongly responsive to the mitogenic cytokine GM-CSF, which confirms that the functional repertoire of these cells is well preserved. We believe that the present method will prove to be especially convenient to implement for mechanistic studies or screening assays.