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Characterizing microglial signaling dynamics during inflammation using single-cell mass cytometry.

Sushanth Kumar1,2, August D Kahle1, Austin B Keeler1

  • 1Department of Biology, College of Arts and Sciences, University of Virginia, Charlottesville, VA 22908, USA.

Biorxiv : the Preprint Server for Biology
|August 12, 2024
PubMed
Summary

Single-cell mass cytometry reveals distinct microglial signaling pathways activated by lipopolysaccharide (LPS) and polyinosinic-polycytidylic acid (Poly(I:C)). Astrocytes significantly dampen microglial inflammatory responses, highlighting the importance of cellular context in neuroinflammation.

Keywords:
AstrocytesCyTOFLipopolysaccharide (LPS)Mass CytometryMicrogliaNeuroinflammationPoly(I:C)Signaling Pathways

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Area of Science:

  • Neuroscience
  • Immunology
  • Cell Biology

Background:

  • Microglia are crucial for central nervous system (CNS) homeostasis but their inflammatory signaling profiles are not fully understood.
  • Traditional methods like transcriptomics miss dynamic post-translational signaling changes.

Purpose of the Study:

  • To investigate dynamic microglial signaling responses to bacterial (LPS) and viral (Poly(I:C)) mimetics using time-resolved single-cell mass cytometry (CyTOF).
  • To assess the immunomodulatory effect of astrocytes on microglial signaling during inflammation.

Main Methods:

  • Neonatal mouse microglia or mixed glial-astrocyte cultures were stimulated with LPS or Poly(I:C) for 48 hours.
  • A 33-antibody panel was used for CyTOF analysis of signaling and identity markers.
  • High-dimensional clustering identified signaling modules.

Main Results:

  • LPS induced stronger early activation of p38, ERK, RSK, and CREB compared to Poly(I:C).
  • Both stimuli upregulated classical activation markers CD40 and CD86 at later time points.
  • Astrocytes significantly reduced microglial responses, notably CD40 upregulation.

Conclusions:

  • CyTOF effectively captures dynamic microglial signaling in neuroinflammation.
  • Astrocyte presence modulates microglial responses, emphasizing the need to consider cellular context in neuroinflammatory research.
  • Findings support CyTOF as a tool for investigating neuroinflammatory disorders.