Microglia Morphological Categorization in a Rat Model of Neuroinflammation by Hierarchical Cluster and Principal Components Analysis

Abstract

It is known that microglia morphology and function are closely related, but only few studies have objectively described different morphological subtypes. To address this issue, morphological parameters of microglial cells were analyzed in a rat model of aseptic neuroinflammation. After the injection of a single dose of the enzyme neuraminidase (NA) within the lateral ventricle (LV) an acute inflammatory process occurs. Sections from NA-injected animals and sham controls were immunolabeled with the microglial marker IBA1, which highlights ramifications and features of the cell shape. Using images obtained by section scanning, individual microglial cells were sampled from various regions (septofimbrial nucleus, hippocampus and hypothalamus) at different times post-injection (2, 4 and 12 h). Each cell yielded a set of 15 morphological parameters by means of image analysis software. Five initial parameters (including fractal measures) were statistically different in cells from NA-injected rats (most of them IL-1β positive, i.e., M1-state) compared to those from control animals (none of them IL-1β positive, i.e., surveillant state). However, additional multimodal parameters were revealed more suitable for hierarchical cluster analysis (HCA). This method pointed out the classification of microglia population in four clusters. Furthermore, a linear discriminant analysis (LDA) suggested three specific parameters to objectively classify any microglia by a decision tree. In addition, a principal components analysis (PCA) revealed two extra valuable variables that allowed to further classifying microglia in a total of eight sub-clusters or types. The spatio-temporal distribution of these different morphotypes in our rat inflammation model allowed to relate specific morphotypes with microglial activation status and brain location. An objective method for microglia classification based on morphological parameters is proposed.Main points Microglia undergo a quantifiable morphological change upon neuraminidase induced inflammation.Hierarchical cluster and principal components analysis allow morphological classification of microglia.Brain location of microglia is a relevant factor.

Glucocorticoid‐mediated modulation of morphological changes associated with aging in microglia

“…As the microglial morphology is dynamically altered depending on its cellular function (Fernández‐Arjona, Grondona, Granados‐Durán, Fernández‐Llebrez & López‐Ávalos, 2017), we hypothesized that age‐associated increases in GC levels implicate hippocampal microglia cell morphology and therefore possibly their function. By both artificially increasing GC levels and using specific siRNAs to knockdown GR expression in vivo , we here describe a mechanism wherein GCs modulate hippocampal microglial morphology and may affect microglial amoebification during aging.…”

A Primary Neural Cell Culture Model for Neuroinflammation

“…The size of microglia also significantly increased in tri-cultures exposed to LPS as compared to vehicle control tri-cultures ( Figure 2C, p = 0.0052). Increased microglia area is not consistent with in vivo observations, as activated microglia in vivo typically exhibit reduced area as they lose their ramifications and take on an amoeboid morphology (Fernández-Arjona et al, 2017). However, the morphology of the microglia following exposure to LPS is consistent with previous in vitro experiments, with the microglia showing a distinct polarity and the formation of large lamellipodia (Abd-El-Basset and Fedoroff, 1995;Nakamura et al, 1999;Persson et al, 2005).…”

Sex and regional differences in microglia morphology and complement receptor 3 are independent of constitutive neuroinflammatory protein concentrations in healthy mice

“…Fractal analysis has been widely used as a morphometric analysis tool (Barreto et al, 2014;Fernandez-Arjona et al, 2017;Karperien et al, 2013;Karperien and Jelinek, 2015;Namjoshi et al, 2014;Orlowski et al, 2003;Soltys et al, 2001;Soltys et al, 2005) and added as a plugin to ImageJ for convenient use (Karperien, 1999(Karperien, -2013. In contrast with the skeleton analysis, fractal analysis is carried out on single cell and is therefore complementary to skeleton analysis.…”

“…In the healthy brain, microglia are observed as ramified cells with small somas (Kettenmann et al, 2011), however, with observable morphological distinctions among brain regions (Lawson et al, 1990), which have only begun to be assessed using quantitative methods (Fernandez-Arjona et al, 2017). A ramified cell is one that has a complicated network of processes that originate from the cell soma.…”