Abstract
It is known that microglial morphology and function are related, but few studies have explored the subtleties of microglial morphological changes in response to specific pathogens. In the present report we quantitated microglia morphological changes in a monkey model of dengue disease with virus CNS invasion. To mimic multiple infections that usually occur in endemic areas, where higher dengue infection incidence and abundant mosquito vectors carrying different serotypes coexist, subjects received once a week subcutaneous injections of DENV3 (genotype III)-infected culture supernatant followed 24 h later by an injection of anti-DENV2 antibody. Control animals received either weekly anti-DENV2 antibodies, or no injections. Brain sections were immunolabeled for DENV3 antigens and IBA-1. Random and systematic microglial samples were taken from the polymorphic layer of dentate gyrus for 3-D reconstructions, where we found intense immunostaining for TNFα and DENV3 virus antigens. We submitted all bi- or multimodal morphological parameters of microglia to hierarchical cluster analysis and found two major morphological phenotypes designated types I and II. Compared to type I (stage 1), type II microglia were more complex; displaying higher number of nodes, processes and trees and larger surface area and volumes (stage 2). Type II microglia were found only in infected monkeys, whereas type I microglia was found in both control and infected subjects. Hierarchical cluster analysis of morphological parameters of 3-D reconstructions of random and systematic selected samples in control and ADE dengue infected monkeys suggests that microglia morphological changes from stage 1 to stage 2 may not be continuous.
Highlights
Microglia are often categorized as resting or activated based on a qualitative assessment of their morphology
Because we found frequent clusters of activated microglia and intense TNFα immunolabeling in the polymorphic layer of dentate gyrus of infected monkeys, we selected this layer as our target to investigate detailed microglial morphological changes
We found that a few microglial morphological features showed a multimodality index >0.55 and this index value indicates that the distribution is at least bimodal and may be multimodal, and these particular features were selected for cluster analysis as previously described (Schweitzer and Renehan, 1997)
Summary
Microglia are often categorized as resting or activated based on a qualitative assessment of their morphology. It is known that resting microglial cells are both motile and very active. In the disease free individual resting microglia are highly ramified and express very low levels of CD40, MHC class II, and B7-2. This morphology is just an extreme example of a dynamic process that appears to be a continuum of morphological changes. A variety of different models have been proposed to characterize microglial morphological changes after brain damage (Beynon and Walker, 2012; Walker et al, 2014). A novel classification following axotomy, based on hierarchical cluster analysis of 2D and 3D cell morphometric features has been proposed (Yamada and Jinno, 2013)
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