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Application of Genetically Encoded Fluorescent Nitric Oxide (NO•) Probes, the geNOps, for Real-time Imaging of NO• Signals in Single Cells
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Application of Genetically Encoded Fluorescent Nitric Oxide (NO•) Probes, the geNOps, for Real-time Imaging of NO• Signals in Single Cells

Published on: March 16, 2017

NO suppresses while peroxynitrite sustains NF-kappaB: a paradigm to rationalize cytoprotective and cytotoxic actions

Yoshiyuki Hattori1, Kikuo Kasai, Steven S Gross

  • 1Department of Endocrinology and Metabolism, Dokkyo University School of Medicine, Mibu, Tochigi, 321-0293, Japan. yhattori@dokkyomed.ac.jp

Cardiovascular Research
|June 15, 2004
PubMed
Abstract

Insights

Nitric oxide (NO) inhibits nuclear factor-kappaB (NF-kappaB) activation and suppresses inducible nitric oxide synthase (iNOS) gene expression, unlike peroxynitrite (ONOO-). This NO-mediated suppression of NF-kappaB is crucial for its anti-inflammatory effects.

Area of Science:

  • Molecular Biology
  • Immunology
  • Biochemistry

Background:

  • Nitric oxide (NO) exhibits dual cytoprotective and cytotoxic properties.
  • A key cytoprotective mechanism of NO involves inhibiting nuclear factor-kappaB (NF-kappaB)-mediated gene expression, conferring anti-inflammatory activity.
  • NO's own synthesis via inducible nitric oxide synthase (iNOS) is NF-kappaB-dependent, suggesting NO may limit its production.

Purpose of the Study:

  • To determine if peroxynitrite (ONOO-), a reactive product of NO, shares NO's ability to inhibit NF-kappaB activation and iNOS gene expression.
  • To compare the effects of NO donors and ONOO- donors on lipopolysaccharide (LPS)-induced responses in vascular smooth muscle cells (VSMC).

Main Methods:

  • Utilized NO donors (NOR3, SNAP) and an ONOO- donor (SIN-1) in VSMC cultures stimulated with LPS.
  • Assessed NF-kappaB activation using electrophoretic mobility shift assays (EMSA) and immunocytochemistry.
  • Quantified iNOS mRNA expression via Northern blot and RT-PCR.
  • Investigated the role of G-cyclase and IkappaB kinase (IKK) activity.

Main Results:

  • NO donors, but not ONOO-, suppressed LPS-induced NF-kappaB activation and iNOS gene expression.
  • NO inhibited the nuclear translocation of NF-kappaB and decreased iNOS mRNA levels.
  • NO, but not ONOO-, inhibited LPS-induced IkappaB-alpha phosphorylation and IKK activity.
  • The inhibitory effect of NO on iNOS transcription was mediated by a G-cyclase-independent mechanism.

Conclusions:

  • Only NO, not ONOO-, effectively inhibits NF-kappaB activation and suppresses iNOS gene expression.
  • This distinct mechanism of action provides a novel framework for understanding the differential cytoprotective and cytotoxic effects of NO.

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