Abstract

Within the CNS, microglia play important roles in host
defence, regulating brain development, repairing tissue injury and maintaining
neuronal homeostasis. Unrestrained over activation of microglia can result in a
chronic pro-inflammatory environment which has been implicated in several
neurodegenerative diseases such as Alzheimer’s disease, Parkinson’s disease and
ALS. Hence, research to understand aberrant microglia function has become increasingly
important in identification of new CNS therapeutic targets. Here, we have characterised
a range of relevant cellular models of microglial function which can be used to
aid progress of microglia-targeted drug discovery projects.

Generally, microglia exhibit two functionally different
activation states, termed M1 and M2. The M1 state develops in response to IFN-γ
or lipopolysaccharide and produces neuronal injury by secreting
pro-inflammatory cytokines such as TNF-α, IL-1β, as well as reactive oxygen
species/reactive nitrogen species. In contrast, the M2 activation state develops
in response to IL-4 or IL-13, leading to an anti-inflammatory response through
production of IL-10 to promote tissue repair and resolution of inflammation. Phagocytic
activity is also a key feature of microglia, to clear apoptotic cells, protein
aggregates as well as bacteria from the CNS. Here, we describe a
multi-parametric analysis approach which enables us to maximise value from a
suite of cell-based assays and to better understand microglia biology. Cells have
been characterised using immuno-fluorescence staining and high content confocal
imaging of IBA-1, a specific marker of microglial cells. Live microglia imaging
using the IncuCyte™ ZOOM was used to monitor cellular morphology and capture
phagocytosis of dead cells in real time. 
Additionally, real-time qPCR and cytokine analysis was used to confirm
M1 and M2 phenotypes following stimulation with LPS and IL-4 respectively. In
conclusion, we present robust analyses of in
vitro models of different microglia functions, which can be used to improve
understanding of microglial biology per
se, but also as useful platform to accelerate microglia-targeted drug
discovery projects.