Establishment of polarized microglial cells derived from mouse embryonic stem cells

Microglia are the resident immune cells of the central nervous system (CNS). Microglia undergo rapid activation in response to even minor pathological changes in the CNS. The activation state of microglia and tissue macrophages is characterized by two extremes that are known as M1 and M2. M1 is triggered upon pro-inflammatory stimuli and lead to the secretion of molecules involved in brain defense but also in neurotoxicity. M2 arises upon anti-inflammatory stimuli and is hallmarked by the secretion of molecules that mediate tolerance and neuroprotection. The aim of this study was to investigate the in vitro polarization capacity of microglia into M1 and M2 subtypes. Primary microglia are hard to obtain in sufficient number and therefore a recently developed protocol was applied to differentiate microglial precursors from mouse embryonic stem (ES) cells. The obtained ES cell derived microglial precursors (ESdM) were shown to be an appropriate substitute to primary microglia. Here we showed that ESdM stimulated with interferon-γ (IFN-gamma) and lipopolysaccharide (LPS) display typical M1 characteristics like higher transcription levels of pro-inflammatory cytokines as well as an increase in the expression level of M1 surface markers. They also secreted significantly higher levels of the chemokine CXCL10 and nitric oxide than untreated or interleukin-4 (IL-4) treated cells. These results indicate that ESdM can be polarized into a cytotoxic subtype in vitro using IFN-gamma and LPS. However, no evidence indicating that ESdM were polarized into a neuroprotective M2 subtype by IL-4 treatment has been found. Even though an increase in the transcription level of anti-inflammatory cytokines was observed upon IL-4 treatment, this increase was not significant. In addition, no increase in the expression levels of M2 markers or in the secretion of the anti-inflammatory cytokine IL-10 were found upon IL-4 treatment indicating that IL-4 might be insufficient to polarize ESdM into M2 subtype in vitro. Further experiments are required to better understand the mechanisms of microglial sub-differentiation and to determine the stability of the acquired M1 and M2 phenotypes over time