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Identification of N‐iminoethyl‐ l ‐ornithine as an irreversible inhibitor of nitric oxide synthase in phagocytic cells
Author(s) -
McCall T.B.,
Feelisch M.,
Palmer R.M.J.,
Moncada S.
Publication year - 1991
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1991.tb12159.x
Subject(s) - omega n methylarginine , nitric oxide , nitric oxide synthase , nitroarginine , arginine , incubation , enzyme inhibitor , chemistry , ic50 , biochemistry , microbiology and biotechnology , enzyme , biology , in vitro , amino acid , organic chemistry
1 The synthesis of nitric oxide (NO) from l ‐arginine by rat peritoneal neutrophils (PMN) and the murine macrophage cell‐line J774 and the inhibition of this synthesis by N‐iminoethyl‐ l ‐ornithine ( l ‐NIO), N G ‐monomethyl‐ l ‐arginine ( l ‐NMMA), N G ‐nitro‐ l ‐arginine ( l ‐NNA) and its methyl ester ( l ‐NAME) were investigated. 2 l ‐NIO was the most potent inhibitor in both types of cells while l ‐NMMA was less active. l ‐NNA and l ‐NAME had no significant effect in PMN and l ‐NNA produced only approximately 40% inhibition of the generation of NO in the J774 cells at the highest concentration tested (300 μ m ). 3 The inhibitory effect of l ‐NIO was rapid in onset, requiring 10 min pre‐incubation to achieve its full inhibitory activity, while the other compounds required 20–60 min pre‐incubation to achieve their full effect. 4 The inhibitory effect of l ‐NIO (10 μ m ) on intact cells could not be reversed by l ‐arginine (300 μ m ) but could be prevented by concomitant incubation with this compound (300 μ m ), while the effect of the other inhibitors could be reversed by a 3–5 fold molar excess of l ‐arginine. 5 The NO synthase from both PMN and J774 cells was cytosolic and NADPH‐ but not Ca 2+ ‐dependent, with K m values for l ‐arginine of 3.3 ± 0.8 and 4.2 ± 1.1 μ m respectively. 6 l ‐NIO was the most potent inhibitor of the neutrophil and J774 enzymes with IC 50 values of 0.8 ± 0.1 and 3 ± 0.5 μ m respectively. Furthermore, the effect of l ‐NIO was irreversible. The other three compounds were less potent, reversible inhibitors. 7 The inhibitory effects of all these compounds were enantiomerically specific. 8 These data indicate that l ‐NIO is a novel, potent, rapid in onset and irreversible inhibitor of NO synthase in phagocytic cells. The rapid uptake of l ‐NIO compared with the other compounds indicates that phagocytic cells have different uptake mechanisms for l ‐arginine analogues.