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Journal of Neuroinflammation
Published in: Journal of Neuroinflammation 1/2016

Open Access 01-12-2016 | Research

Phenotypic clustering: a novel method for microglial morphology analysis

Authors: Franck Verdonk, Pascal Roux, Patricia Flamant, Laurence Fiette, Fernando A. Bozza, Sébastien Simard, Marc Lemaire, Benoit Plaud, Spencer L. Shorte, Tarek Sharshar, Fabrice Chrétien, Anne Danckaert

Published in: Journal of Neuroinflammation | Issue 1/2016

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Abstract

Background

Microglial cells are tissue-resident macrophages of the central nervous system. They are extremely dynamic, sensitive to their microenvironment and present a characteristic complex and heterogeneous morphology and distribution within the brain tissue. Many experimental clues highlight a strong link between their morphology and their function in response to aggression. However, due to their complex “dendritic-like” aspect that constitutes the major pool of murine microglial cells and their dense network, precise and powerful morphological studies are not easy to realize and complicate correlation with molecular or clinical parameters.

Methods

Using the knock-in mouse model CX3CR1GFP/+, we developed a 3D automated confocal tissue imaging system coupled with morphological modelling of many thousands of microglial cells revealing precise and quantitative assessment of major cell features: cell density, cell body area, cytoplasm area and number of primary, secondary and tertiary processes. We determined two morphological criteria that are the complexity index (CI) and the covered environment area (CEA) allowing an innovative approach lying in (i) an accurate and objective study of morphological changes in healthy or pathological condition, (ii) an in situ mapping of the microglial distribution in different neuroanatomical regions and (iii) a study of the clustering of numerous cells, allowing us to discriminate different sub-populations.

Results

Our results on more than 20,000 cells by condition confirm at baseline a regional heterogeneity of the microglial distribution and phenotype that persists after induction of neuroinflammation by systemic injection of lipopolysaccharide (LPS). Using clustering analysis, we highlight that, at resting state, microglial cells are distributed in four microglial sub-populations defined by their CI and CEA with a regional pattern and a specific behaviour after challenge.

Conclusions

Our results counteract the classical view of a homogenous regional resting state of the microglial cells within the brain. Microglial cells are distributed in different defined sub-populations that present specific behaviour after pathological challenge, allowing postulating for a cellular and functional specialization. Moreover, this new experimental approach will provide a support not only to neuropathological diagnosis but also to study microglial function in various disease models while reducing the number of animals needed to approach the international ethical statements.
Appendix
Available only for authorised users
Footnotes
1
Directive 2010/63/EU of the European Parliament and of the Council.
 
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Metadata
Title
Phenotypic clustering: a novel method for microglial morphology analysis
Authors
Franck Verdonk
Pascal Roux
Patricia Flamant
Laurence Fiette
Fernando A. Bozza
Sébastien Simard
Marc Lemaire
Benoit Plaud
Spencer L. Shorte
Tarek Sharshar
Fabrice Chrétien
Anne Danckaert
Publication date
01-12-2016
Publisher
BioMed Central
Published in
Journal of Neuroinflammation / Issue 1/2016
Electronic ISSN: 1742-2094
DOI
https://doi.org/10.1186/s12974-016-0614-7

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