Notch-1 signaling regulates microglia activation via NF-κB pathway after hypoxic exposure in vivo and in vitro

PLoS One. 2013 Nov 6;8(11):e78439. doi: 10.1371/journal.pone.0078439. eCollection 2013.

Abstract

Neuroinflammation mediated by the activated microglia is suggested to play a pivotal role in the pathogenesis of hypoxic brain injury; however, the underlying mechanism of microglia activation remains unclear. Here, we show that the canonical Notch signaling orchestrates microglia activation after hypoxic exposure which is closely associated with multiple pathological situations of the brain. Notch-1 and Delta-1 expression in primary microglia and BV-2 microglial cells was significantly elevated after hypoxia. Hypoxia-induced activation of Notch signaling was further confirmed by the concomitant increase in the expression and translocation of intracellular Notch receptor domain (NICD), together with RBP-Jκ and target gene Hes-1 expression. Chemical inhibition of Notch signaling with N-[N-(3,5-difluorophenacetyl)-1-alany1- S-phenyglycine t-butyl ester (DAPT), a γ-secretase inhibitor, effectively reduced hypoxia-induced upregulated expression of most inflammatory mediators. Notch inhibition also reduced NF-κB/p65 expression and translocation. Remarkably, Notch inhibition suppressed expression of TLR4/MyD88/TRAF6 pathways. In vivo, Notch signaling expression and activation in microglia were observed in the cerebrum of postnatal rats after hypoxic injury. Most interestingly, hypoxia-induced upregulation of NF-κB immunoexpression in microglia was prevented when the rats were given DAPT pretreatment underscoring the interrelationship between Notch signaling and NF-κB pathways. Taken together, we conclude that Notch signaling is involved in regulating microglia activation after hypoxia partly through the cross talk between TLR4/MyD88/TRAF6/NF-κB pathways. Therefore, Notch signaling may serve as a prospective target for inhibition of microglia activation known to be implicated in brain damage in the developing brain.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amyloid Precursor Protein Secretases / antagonists & inhibitors
  • Amyloid Precursor Protein Secretases / genetics
  • Amyloid Precursor Protein Secretases / metabolism
  • Animals
  • Animals, Newborn
  • Basic Helix-Loop-Helix Transcription Factors / genetics
  • Basic Helix-Loop-Helix Transcription Factors / metabolism
  • Cell Hypoxia / genetics
  • Cell Line, Tumor
  • Dipeptides / pharmacology
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation, Developmental
  • Homeodomain Proteins / genetics
  • Homeodomain Proteins / metabolism
  • Hypoxia / genetics*
  • Hypoxia / metabolism
  • Hypoxia / pathology
  • Immunoglobulin J Recombination Signal Sequence-Binding Protein / genetics
  • Immunoglobulin J Recombination Signal Sequence-Binding Protein / metabolism
  • Intracellular Signaling Peptides and Proteins / genetics
  • Intracellular Signaling Peptides and Proteins / metabolism
  • Membrane Proteins / genetics
  • Membrane Proteins / metabolism
  • Mice
  • Microglia / metabolism*
  • Microglia / pathology
  • Myeloid Differentiation Factor 88 / genetics
  • Myeloid Differentiation Factor 88 / metabolism
  • NF-kappa B / genetics*
  • NF-kappa B / metabolism
  • Rats
  • Receptor, Notch1 / antagonists & inhibitors
  • Receptor, Notch1 / genetics*
  • Receptor, Notch1 / metabolism
  • Signal Transduction*
  • TNF Receptor-Associated Factor 6 / genetics
  • TNF Receptor-Associated Factor 6 / metabolism
  • Toll-Like Receptor 4 / genetics
  • Toll-Like Receptor 4 / metabolism
  • Transcription Factor HES-1

Substances

  • Basic Helix-Loop-Helix Transcription Factors
  • Dipeptides
  • Enzyme Inhibitors
  • Hes1 protein, rat
  • Homeodomain Proteins
  • Immunoglobulin J Recombination Signal Sequence-Binding Protein
  • Intracellular Signaling Peptides and Proteins
  • Membrane Proteins
  • Myd88 protein, rat
  • Myeloid Differentiation Factor 88
  • N-(N-(3,5-difluorophenacetyl)alanyl)phenylglycine tert-butyl ester
  • NF-kappa B
  • Notch1 protein, rat
  • Receptor, Notch1
  • TNF Receptor-Associated Factor 6
  • Tlr4 protein, rat
  • Toll-Like Receptor 4
  • Transcription Factor HES-1
  • delta protein
  • Amyloid Precursor Protein Secretases

Grants and funding

This study was supported by grant No. R-181-000-140-592 from the National University of Singapore. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.