CD40 ligand-dependent tyrosine nitration of prostacyclin synthase in vivo.
BACKGROUND
Cells in human atherosclerotic lesions express the immune mediator CD40 and its ligand, CD40L, but the mechanisms and the mediators by which CD40L contributes to atherosclerosis are poorly defined. Here, we show how CD40L increases vascular inflammation and thrombosis via tyrosine nitration and inhibition of prostacyclin synthase (PGIS), an enzyme with antithrombotic, antiproliferative, and dilatory functions in the normal vasculature.


RESULTS
Exposure of cultured human aortic endothelial cells to clinically relevant concentrations of CD40L (20 to 80 ng/mL) dose-dependently increased the production of superoxide (O2*-), decreased nitric oxide (NO) bioactivity, and increased PGIS nitration. Furthermore, inhibition of CD40 expression by small interfering RNA blocked the effects of CD40L on O2*-, NO bioactivity, and PGIS nitration, which indicates a specific effect of CD40L. In addition, either depletion of mitochondria (rho0 cells, ie, mitochondria-depleted cells, to prevent mitochondrial O2*-) or adenoviral overexpression of superoxide dismutase, as well as inhibition of NO synthase, abolished the CD40L-enhanced PGIS nitration, which implies that the mitochondria might be the source of O2*- and thus peroxynitrite (ONOO-). Furthermore, SQ29548, a thromboxane A2/prostaglandin H2 receptor antagonist, significantly reduced CD40L-enhanced expression of intercellular adhesion molecule-1. Finally, administration of CD40L resulted in PGIS inhibition and nitration in the aortas of C57BL6 mice but less in mice overexpressing human superoxide dismutase, which suggests that ONOO- might be required for CD40L-enhanced PGIS nitration in vivo.


CONCLUSIONS
We conclude that CD40L might contribute to the initiation and progression of atherosclerosis by increasing O2*(-)- and ONOO(-)-dependent PGIS nitration and thromboxane A2/prostaglandin H2 receptor stimulation.
Davis
Bradley
B
Vascular Research Laboratory, Graduate School of Medicine, University of Tennessee, Knoxville, Tennessee, USA.
Zou
Ming-Hui
MH
D000818:Animals; D001011:Aorta; D023201:CD40 Ligand / Q000494:pharmacology / Q000502:physiology*; D002455:Cell Division; D002478:Cells, Cultured; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology / Q000502:physiology*; D006801:Humans; D019746:Intramolecular Oxidoreductases / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000378:metabolism*; D013482:Superoxide Dismutase / Q000235:genetics
D023201:CD40 Ligand; D019000:Phosphotyrosine; D003577:Cytochrome P-450 Enzyme System; D013482:Superoxide Dismutase; D019746:Intramolecular Oxidoreductases; C022461:prostacyclin synthetase