Identification of N‐iminoethyl‐<scp>l</scp>‐ornithine as an irreversible inhibitor of nitric oxide synthase in phagocytic cells

McCall, Therese; Feelisch, Martin; Palmer, Richard; Moncada, Salvador

doi:10.1111/j.1476-5381.1991.tb12159.x

Cited by 246 publications

(154 citation statements)

References 17 publications

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“…Differences in the rank order potency of inhibitors of iNOS in J774 cells have been reported and McCall et al (8) hypothesized that these differences may reflect uptake of NOS inhibitors by different transport systems. Our findings have confirmed this hypothesis and identified system y + as a potential target for inhibitors of iNOS.…”

Section: Discussionmentioning

confidence: 99%

Section: ____________________________________________________________mentioning

confidence: 99%

“…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). In this context, we reported previously that N G -monomethyl-L-arginine (L-NMMA) and N G -iminoethyl-L-ornithine (L-NIO), but not N G -nitro-L-arginine (L-NNA) nor its methyl ester L-NAME inhibit transport of arginine into cultured porcine aortic endothelial cells via system y + (9).…”

Section: ____________________________________________________________mentioning

confidence: 99%

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

Baydoun

Mann

1994

Biochemical and Biophysical Research Communications

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SUMMARY:Amino acid transport systems mediating uptake of nitric oxide (NO) synthase inhibitors were characterized in the murine macrophage cell line J774. Treatment of J774 cells with bacterial endotoxin (LPS, 1 μg ml -1 , 24 h) selectively increased the transport capacity forInhibition studies established that the cationic transport system y + mediates uptake of L-arginine, L-NMMA and N G -iminoethyl-L-ornithine (L-NIO). A neutral transporter, with low substrate specificity and insensitive to LPS, mediates uptake of L-citrulline, L-NNA and its methyl ester L-NAME. We conclude that enhanced expression of the y + transporter in LPS-stimulated macrophages (1) may facilitate the targeting of selective inhibitors of inducible NO synthase to activated cells generating NO in endotoxin shock. ________________________________________________________________________ Activation of murine macrophage J774 cells with lipopolysaccharide (LPS) results in acycloheximide-dependent induction of NO synthase and arginine transport via system y + (1-3).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). In this context, we reported previously that N G -monomethyl-L-arginine (L-methyl ester L-NAME inhibit transport of arginine into cultured porcine aortic endothelial cells via system y + (9). Recent studies of radiolabelled L-NMMA and arginine uptake in porcine aortic endothelial cells have confirmed these findings (10).

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Section: Discussionmentioning

confidence: 99%

Section: ____________________________________________________________mentioning

confidence: 99%

Section: ____________________________________________________________mentioning

confidence: 99%

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

Baydoun

Mann

1994

Biochemical and Biophysical Research Communications

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“…L-Arginine analogs in conjunction with spectroscopic studies to determine the factors contributing to the Arg-NOS interaction yielded conflicting results. Some reports suggested that positive charge is required for the tight binding of L-Arg to NOS (33,34). Others showed that positive charge may not be critical for the Arg-NOS interaction (35,36).…”

Section: Discussionmentioning

confidence: 99%

Mutation of Glu-361 in Human Endothelial Nitric-oxide Synthase Selectively Abolishes L-Arginine Binding without Perturbing the Behavior of Heme and Other Redox Centers

Chen

Tsai

Berka

et al. 1997

Journal of Biological Chemistry

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Nitric oxide (NO) and L-citrulline are formed from the oxidation of L-arginine by three different isoforms of NO synthase (NOS). Defining amino acid residues responsible for L-arginine binding and oxidation is a primary step toward a detailed understanding of the NOS reaction mechanisms and designing strategies for the selective inhibition of the individual isoform. We have altered Glu-361 in human endothelial NOS to Gln or Leu by site-directed mutagenesis and found that these mutations resulted in a complete loss of L-citrulline formation without disruption of the cytochrome c reductase and NADPH oxidase activities. Optical and EPR spectroscopic studies demonstrated that the Glu-361 mutants had similar spectra either in resting state or reduced CO-complex as the wild type. The heme ligand, imidazole, could induce a low spin state in both wild-type and Glu-361 mutants. However, unlike the wild-type enzyme, the low spin imidazole complex of Glu-361 mutants was not reversed to a high spin state by addition of either L-arginine, acetylguanidine, or 2-aminothiazole. Direct L-arginine binding could not be detected in the mutants either. These results strongly indicate that Glu-361 in human endothelial NOS is specifically involved in the interaction with L-arginine. Mutation of this residue abolished the L-arginine binding without disruption of other functional characteristics.Nitric oxide (NO) 1 has been identified as an important signal mediator that is involved in many physiological or pathophysiological processes. NO is produced together with L-citrulline through a two-step oxidation of L-arginine by three different NO synthase isoforms. The endothelial and neuronal isoforms are constitutively expressed, and their activities are regulated by calcium and calmodulin (Ca 2ϩ /CaM) (1, 2). The third NOS isoform is induced in response to cytokines or lipopolysaccharide, and its activity is independent of Ca 2ϩ /CaM (3). Despite the modest sequence homology and different regulation among the three NOS isoforms, they share a similar cofactor composition and possess a bidomain structure (4 -8). The C-terminal reductase domain is homologous to NADPH-cytochrome P450 reductase and has binding regions for NADPH, FAD, and FMN. The N-terminal oxygenase domain containing heme, BH 4 , and L-arginine binding sites is a P450-type hemoprotein, but does not show sequence homology to other known P450s. A CaM binding module exists near the center of the NOS sequences (9), and binding of Ca 2ϩ /CaM facilitates electron transfer from the reductase to the oxygenase domain (10, 11).Because of the obvious impact of NO on human health, a detailed understanding of the NOS reaction mechanism is essential for selective pharmacological intervention against individual isoforms. The oxygenase active site domain, including the proximal heme thiolate ligand, the distal heme pocket, and the substrate binding region is the center of NOS catalysis. Defining the amino acid residues making up the heme binding region and substrate oxidation site is a key s...

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“…In the first set of experiments, conditioned medium from unstimulated cells and cells treated with LPS (1000 ng/ml) and IFN-␥ (100 U/ml) was collected. Duplicate preparations of cells were treated with either the NO donor S-nitroso-Nacetyl-D,L-penicillamine (SNAP; Alexis) or L-N 5 -(1-iminoethyl)-ornithine (L-NIO; 80 M; Alexis), a selective iNOS inhibitor to block production of NO by these cells (21). Thus, exogenous NO was added to cells from non-OVA-treated mice through application of SNAP to approximate the NO production of cells from OVA-treated mice.…”

Section: Bal Cell Culturementioning

confidence: 99%

Alterations in Nitric Oxide and Cytokine Production with Airway Inflammation in the Absence of IL-10

Ameredes

Zamora

Sethi

et al. 2005

The Journal of Immunology

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IL-10 is an anti-inflammatory cytokine that suppresses NO synthase (NOS) and production of NO; its lack may promote NO production and alterations in cytokines modulated by NO with allergic airway inflammation (AI), such as IL-18 and IL-4. Therefore, we induced AI in IL-10 knockout (−/−) and IL-10-sufficient C57BL/6 (C57) mice with inhaled OVA and measured airway NO production, as exhaled NO (ENO) and bronchoalveolar lavage fluid nitrite levels. ENO and nitrite levels were elevated significantly in naive IL-10−/− mice as compared with C57 mice. With AI, ENO and nitrite levels increased in C57 mice and decreased in IL-10−/− mice. IL-18 production fell with both AI and addition of S-nitroso-N-acetyl-d,l-penicillamine (a NO donor) but was not significantly increased by chemical NOS inhibition by l-N5-(1-iminoethyl)-ornithine. IL-4 AI was increased significantly (up to 10-fold greater) in the absence of IL-10 but was reduced significantly with chemical inhibition of NOS. Airway responsiveness was lower in IL-10−/− mice and was associated with alteration in production of NO and IL-4. Thus, IL-4 production was increased, and likely decreased NO production, in a way not predicted by the absence of IL-10. Inhibition of IL-4 production, with inhibition of NOS in the absence of IL-10, demonstrated the importance of a NO and IL-4 feedback mechanism regulating this interaction.

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Identification of N‐iminoethyl‐l‐ornithine as an irreversible inhibitor of nitric oxide synthase in phagocytic cells

Cited by 246 publications

References 17 publications

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

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

Mutation of Glu-361 in Human Endothelial Nitric-oxide Synthase Selectively Abolishes L-Arginine Binding without Perturbing the Behavior of Heme and Other Redox Centers

Alterations in Nitric Oxide and Cytokine Production with Airway Inflammation in the Absence of IL-10

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