Premium
Expression of iNOS is reduced by an inhibitor of fatty acyl CoA synthetase.
Author(s) -
Weis Margaret Teresa,
Maliha George,
Chen Xiong
Publication year - 2010
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.24.1_supplement.784.6
Subject(s) - nitric oxide , nitrite , chemistry , metabolite , biochemistry , cytokine , fatty acid , nitric oxide synthase , citrulline , in vivo , in vitro , enzyme , microbiology and biotechnology , arginine , biology , amino acid , immunology , organic chemistry , nitrate
Objective To test the hypothesis that fatty acyl CoA derivatives of fatty acids modulate cytokine stimulated iNOS expression in C6 astrocytes. Rationale Triacsin C, a fungal metabolite, inhibits fatty acyl CoA synthetase in many cell types. The compound inhibits fatty acid induced iNOS expression and apoptosis in isolated islets of Langerhans (Shimabukuro et al 1996), suggesting that fatty acyl CoA derivatives regulate iNOS expression. Methods C6 cells were stimulated with a cytokine mix consisting of TNFα (60 ng/mL), ILβ (2 ng/mL) and IFNγ (100 units/mL) in the presence or absence of triacsin C. Expression of iNOS was estimated by measuring iNOS protein by immunoblotting. In vivo iNOS activity was estimated by measuring media nitrite (Greiss reaction) and in vitro iNOS activity was measured by conversion of [ 14 C]arginine to [ 14 C]citrulline in the absence of added calcium. Results In C6 cell homogenates, triacin C inhibited FACS activity in a concentration dependent manner, ranging from 31.9 ± 1.5% at 300 nM to 71.1 ± 3.8% at 5 μM. The effect of 5 μM triacsin C on cytokine‐stimulated media nitrite, catalytic activity and immunoreactive protein were measured at 18 and 24 hours:% of Cytokine Stimulated ActivityMedia Nitrite Catalytic Activity Immunoreactivity18 hours 62.5 ± 6.9% 103 ± 11.1% 25.1 ± 13.2% 24 hours 80.6 ± 1.39% * 373 ± 72% * 36.6 ± 5.81%* = p< 0.05 as compared to 18 hours by Student's ttest; n=3 independent experiments for each parameter, each done in duplicate or triplicate.Conclusions These results suggest that triacsin C inhibits cytokine stimulated NO formation, and that iNOS protein is significantly less in triacsin C treated cells. Surprisingly, not all of the immunoreactive iNOS protein seems to be catalytically active. The data suggest that triacsin C may delay cytokine‐stimulated expression of iNOS and thus its overall activity in C6 cells.