Tetrahydrobiopterin-dependent formation of nitrite and nitrate in murine fibroblasts.
The present study demonstrates that murine dermal fibroblasts produce nitrite (NO2-) and nitrate (NO3-) upon treatment with interferon gamma (IFN-gamma). This formation is dependent on L-arginine and can be inhibited by the L-arginine analogue NG-monomethyl-L-arginine. The effect of IFN-gamma is drastically increased by cotreatment with tumor necrosis factor alpha (TNF-alpha), interleukin 1 (IL-1), or lipopolysaccharide (LPS). The tested cytokines also induce formation of tetrahydrobiopterin in murine fibroblasts. Inhibition of guanosine triphosphate-cyclohydrolase I, the key enzyme of tetrahydrobiopterin de novo synthesis with 2,4-diamino-6-hydroxy-pyrimidine, leads to decreased formation of NO2- and NO3-. This effect can be reversed by addition of sepiapterin, which provides tetrahydrobiopterin via a salvage pathway. Methotrexate, which inhibits the salvage pathway, blocks the restoration of NO2- and NO3- production by sepiapterin. The cytotoxic effect of combinations of IFN-alpha with TNF-gamma, IL-1, or LPS is attenuated by inhibition of tetrahydrobiopterin synthesis. These results show that intracellular concentrations of tetrahydrobiopterin control the amount of NO2- and NO3- produced in situ and suggest that the role of cytokine-induced tetrahydrobiopterin synthesis is to provide cells with the active cofactor for production of nitrogen oxides.
Werner-Felmayer
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Department of Medical Chemistry and Biochemistry, University of Innsbruck, Austria.
Werner
E R
ER
Fuchs
D
D
Hausen
A
A
Reibnegger
G
G
Wachter
H
H
D000818:Animals; D001708:Biopterins / Q000031:analogs & derivatives* / Q000378:metabolism; D002478:Cells, Cultured; D016207:Cytokines / Q000494:pharmacology*; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D007371:Interferon-gamma / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D007700:Kinetics; D008727:Methotrexate / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D009566:Nitrates / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism*; D011621:Pteridines / Q000494:pharmacology; D011622:Pterins; D011994:Recombinant Proteins / Q000494:pharmacology; D012867:Skin / Q000187:drug effects / Q000378:metabolism*; D013400:Sugar Acids / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology
C017182:5-dehydro-3-deoxy-D-arabino-heptulosonic acid-7-phosphate; D016207:Cytokines; D007375:Interleukin-1; D009566:Nitrates; D009573:Nitrites; D011621:Pteridines; D011622:Pterins; D011994:Recombinant Proteins; D013400:Sugar Acids; D014409:Tumor Necrosis Factor-alpha; D001708:Biopterins; D007371:Interferon-gamma; C016727:sepiapterin; C003402:sapropterin; D008727:Methotrexate