Macrophage oxidation of L-arginine to nitrite and nitrate: nitric oxide is an intermediate

Abstract

Previous studies have shown that murine macrophages immunostimulated with interferon gamma and Escherichia coli lipopolysaccharide synthesize NO2-, NO3-, and citrulline from L-arginine by oxidation of one of the two chemically equivalent guanido nitrogens. The enzymatic activity for this very unusual reaction was found in the 100,000g supernatant isolated from activated RAW 264.7 cells and was totally absent in unstimulated cells. This activity requires NADPH and L-arginine and is enhanced by Mg2+. When the subcellular fraction containing the enzyme activity was incubated with L-arginine, NADPH, and Mg2+, the formation of nitric oxide was observed. Nitric oxide formation was dependent on the presence of L-arginine and NADPH and was inhibited by the NO2-/NO3- synthesis inhibitor NG-monomethyl-L-arginine. Furthermore, when incubated with L-[guanido-15N2]arginine, the nitric oxide was 15N-labeled. The results show that nitric oxide is an intermediate in the L-arginine to NO2-, NO3-, and citrulline pathway. L-Arginine is required for the activation of macrophages to the bactericidal/tumoricidal state and suggests that nitric oxide is serving as an intracellular signal for this activation process in a manner similar to that very recently observed in endothelial cells, where nitric oxide leads to vascular smooth muscle relaxation [Palmer, R. M. J., Ashton, D. S., & Moncada, S. (1988) Nature (London) 333, 664-666].

Inhibition of nitric oxide synthase by nasal decongestants

“…Hydrogen peroxide was purchased from J.T. Baker (Deventer, Holland); [2,3,4,5] The macrophage cell line, NR8383 was kindly provided by R.J. Helmke, (Dept of Pediatrics, University of Texas Health Science Center, San Antonio, Texas, USA). Klebsiella Pneumoniae were a kind gift from J. van Amsterdam (Rijksinstituut voor Volksgezondheid en Milieu, Bilthoven, the Netherlands).…”

“…Nitric oxide (NO) is a free radical that is known to have an important function as mediator in several biological processes amongst which, vasorelaxation [1,2], cellmediated immune responses [3,4], inhibition of platelet aggregation [5] and neurotransmission [6] are extensively investigated. The formation of NO has been described in many mammalian cell types and can be ascribed to three isoforms of nitric oxide synthase (NOS), all of which oxidize L-arginine to L-citrulline.…”

Selective Targeting of Nitric Oxide Synthase Inhibitors to System y+ in Activated Macrophages

“…In activated macrophages generation of NO by inducible NO synthase (iNOS) is critically dependent on extracellular arginine (2,4,5), despite reports that activated cells contain high intracellular arginine levels (1,6). N G -derivatized analogues of L-arginine are potent inhibitors of iNOS, and their rank order of potency has been attributed to different NOS isoforms and/or their differential rates of cellular uptake (7,8).…”

Synthesis and biological effects of NO in malignant glioma cells: modulation by cytokines including CD95L and TGF-β dexamethasone, and p53 gene transfer

“…Inducible NOS (iNOS) is calcium-independent and produces large amounts of NO which may mediate cGMP-independent cytotoxicity against microorganisms and tumor cells (Stuehr and Nathan, 1989). iNOS is found in a variety of cells such as macrophages (Marletta et al, 1988), endothelial cells (Kilbourn and Belloni, 1990) and glial cells . While all three types of NOS may be expressed in astrocytomas (Cobbs et al, 1995), expression of iNOS was only observed in high-grade tumors and mainly restricted to endothelial cells within the tumor.…”