Journal
JOURNAL OF NEUROSCIENCE
Volume 24, Issue 26, Pages 5942-5954Publisher
SOC NEUROSCIENCE
DOI: 10.1523/JNEUROSCI.1271-04.2004
Keywords
iNOS; astrocytes; lactosylceramide; lipopolysaccharide; interferon-gamma; spinal cord injury
Categories
Funding
- NINDS NIH HHS [NS-22576, NS-37766, R01 NS040144, NS-40810, NS-34741, R01 NS022576, R01 NS037766, NS-40144, R01 NS034741, R37 NS022576, R01 NS040810] Funding Source: Medline
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In the present study a possible role of glycosphingolipids (GSLs) in inducible nitric oxide synthase ( iNOS) gene expression and nitric oxide ( NO) production after spinal cord injury (SCI) in rats has been established. In primary rat astrocytes lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma) treatment increased the intracellular levels of lactosylceramide ( LacCer) and induced iNOS gene expression. D-Threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol . HCI ( PDMP), a glucosylceramide synthase and LacCer synthase ( galactosyltransferase, GalT-2) inhibitor, inhibited LPS/IFN-gamma induced iNOS expression, which was reversed by exogenously supplied LacCer, but not by other glycosphingolipids. LPS/IFN-gamma caused a rapid increase in the activity of GalT-2 and synthesis of LacCer. Silencing of GalT-2 gene with the use of antisense oligonucleotides resulted in decreased LPS/IFN-gamma-induced iNOS, TNF-alpha, and IL-1beta gene expression. The PDMP-mediated reduction in LacCer production and inhibition of iNOS expression correlated with decreased Ras and ERK1/2 activation along with decreased IkappaB phosphorylation, NF-kappaB DNA binding activity, and NF-kappaB-luciferase reporter activity. LacCer-mediated Ras activation was redox-mediated and was attenuated by antioxidants N-acetyl cysteine (NAC) and pyrrolidine dithiocarbamate ( PDTC). In vivo administration of PDMP after SCI resulted in improved functional outcome ( Basso, Beattie, Bresnahan score); inhibition of iNOS, TNF-alpha, and IL-1beta expression; decreased neuronal apoptosis; and decreased tissue necrosis and demyelination. The in vivo studies supported the conclusions drawn from cell culture studies and provided evidence for the possible role of GalT-2 and LacCer in SCI-induced inflammation and pathology. To our knowledge this is the first report of a role of LacCer in iNOS expression and the advantage of GSL depletion in attenuating post-SCI inflammation to improve the outcome of SCI.
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