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Phosphatidylcholine-specific phospholipase C regulates activation of RAW264.7 macrophage-like cells by lipopeptide

F Zhang1, G Zhao, Z Dong

  • 1Department of Cancer Biology, The University of Texas M. D. Anderson Cancer Center, Houston, Texas, USA.

Journal of Leukocyte Biology
|June 19, 2001
PubMed

Insights

The synthetic lipopeptide JBT3002 activates macrophages by requiring phosphatidylcholine-specific phospholipase C (PC-PLC) activity. This enzyme is crucial for JBT3002-induced nitric oxide and TNF-alpha production in macrophages.

Area of Science:

  • Immunology
  • Cell Biology
  • Biochemistry

Background:

  • Phospholipase activities are implicated in macrophage activation by lipopolysaccharide (LPS).
  • Synthetic lipopeptide JBT3002 may activate macrophages through LPS-like signaling pathways.

Purpose of the Study:

  • To investigate if phospholipase activities are essential for JBT3002-mediated macrophage activation.
  • To elucidate the specific phospholipase involved in JBT3002 signaling.

Main Methods:

  • Treatment of RAW264.7 murine macrophage-like cells with JBT3002.
  • Assessment of inducible nitric oxide synthase (iNOS) and tumor necrosis factor-alpha (TNF-alpha) expression.
  • Inhibition studies using tricyclodecan-9-yl xanthogenate (D609), a PC-PLC inhibitor, and butanol, a PC-PLC inhibitor.
  • Measurement of diacylglycerol (DAG) and phosphatidic acid production.
  • Stimulation with a DAG analogue in the presence of interferon-gamma.

Main Results:

  • JBT3002 dose-dependently stimulated iNOS and TNF-alpha expression.
  • D609 significantly inhibited JBT3002-induced nitric oxide and TNF-alpha production and mRNA expression.
  • JBT3002 treatment increased diacylglycerol (DAG) production, which was blocked by D609.
  • Butanol did not affect JBT3002-induced activation, and phosphatidic acid formation was not altered.
  • A DAG analogue induced TNF-alpha production in the presence of interferon-gamma.

Conclusions:

  • Activation of RAW264.7 cells by JBT3002 necessitates phosphatidylcholine-specific phospholipase C (PC-PLC) activity.
  • PC-PLC mediated diacylglycerol (DAG) generation appears to be a key step in JBT3002-induced macrophage activation.

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