Inflammation leads to distinct populations of extracellular vesicles from microglia

Yiyi Yang, Antonio Boza-Serrano, Christopher J.R. Dunning, Bettina Hjelm Clausen, Kate Lykke Lambertsen, Tomas Deierborg

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Abstract

Background: Activated microglia play an essential role in inflammatory responses elicited in the central nervous system (CNS). Microglia-derived extracellular vesicles (EVs) are suggested to be involved in propagation of inflammatory signals and in the modulation of cell-to-cell communication. However, there is a lack of knowledge on the regulation of EVs and how this in turn facilitates the communication between cells in the brain. Here, we characterized microglial EVs under inflammatory conditions and investigated the effects of inflammation on the EV size, quantity, and protein content. Methods: We have utilized western blot, nanoparticle tracking analysis (NTA), and mass spectrometry to characterize EVs and examine the alterations of secreted EVs from a microglial cell line (BV2) following lipopolysaccharide (LPS) and tumor necrosis factor (TNF) inhibitor (etanercept) treatments, or either alone. The inflammatory responses were measured with multiplex cytokine ELISA and western blot. We also subjected TNF knockout mice to experimental stroke (permanent middle cerebral artery occlusion) and validated the effect of TNF inhibition on EV release. Results: Our analysis of EVs originating from activated BV2 microglia revealed a significant increase in the intravesicular levels of TNF and interleukin (IL)-6. We also observed that the number of EVs released was reduced both in vitro and in vivo when inflammation was inhibited via the TNF pathway. Finally, via mass spectrometry, we identified 49 unique proteins in EVs released from LPS-activated microglia compared to control EVs (58 proteins in EVs released from LPS-activated microglia and 37 from control EVs). According to Gene Ontology (GO) analysis, we found a large increase of proteins related to translation and transcription in EVs from LPS. Importantly, we showed a distinct profile of proteins found in EVs released from LPS treated cells compared to control. Conclusions: We demonstrate altered EV production in BV2 microglial cells and altered cytokine levels and protein composition carried by EVs in response to LPS challenge. Our findings provide new insights into the potential roles of EVs that could be related to the pathogenesis in neuroinflammatory diseases.

Original languageEnglish
Article number168
JournalJournal of Neuroinflammation
Volume15
Number of pages19
ISSN1742-2094
DOIs
Publication statusPublished - 28. May 2018

Keywords

  • Extracellular vesicles (EVs)
  • Microglia
  • Neuroinflammation
  • TNF
  • Microglia/drug effects
  • Protein Transport/drug effects
  • Nitric Oxide Synthase Type II/metabolism
  • Tumor Necrosis Factor-alpha/deficiency
  • Etanercept/pharmacology
  • Mice, Inbred C57BL
  • Infarction, Middle Cerebral Artery/complications
  • Male
  • Lipopolysaccharides/pharmacology
  • Signal Transduction/drug effects
  • Mice, Knockout
  • Animals
  • Inflammation/etiology
  • Immunosuppressive Agents/pharmacology
  • Mice
  • Cytokines/metabolism
  • Extracellular Vesicles/pathology
  • Cell Line, Transformed
  • Gene Expression Regulation/drug effects
  • Disease Models, Animal

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