The mechanism of the inhibitory effect of polyamines on the induction of nitric oxide synthase: role of aldehyde metabolites

Abstract

1 We have recently found that in the presence, but not in the absence, of foetal calf serum, spermine inhibits the production of nitric oxide (NO) in cultured J774.2 macrophages stimulated with bacterial endotoxin (lipopolysaccharide; LPS) or with '-interferon (IFN), showing that polyamines may act as suppressants of NO-mediated immune functions. Here, we have studied the mechanisms and the specificity of this inhibitory action. 2 Other polyamines, as well as spermine, inhibit the formation of NO in cultured J774.2 macrophages, with the order of potency being spermine > spermidine >> putrescine = cadaverine. This inhibition of NO formation is not due to any cytotoxic effect of these agents for they neither reduced mitochondrial respiration nor increased the release of lactate dehydrogenase into the supernatant. 3 Spermine is not a direct inhibitor of the activity of iNOS in induced J774.2 cells as measured by its lack of effect on the conversion of L-arginine to L-citrulline in homogenates. Neither spermine, nor its metabolites, interfere with the production of nitrite from NO or act as scavengers of NO. Thus, spermine is an inhibitor of the induction of iNOS. 4 Spermine inhibits nitrite formation in the presence of foetal, newborn or adult bovine serum, but not rat or human serum. 5 The effect of sper mine on nitrite production can be prevented by isoniazid, hydrazine or hydroxylamine, inhibitors of spermine oxidase, as well as by phenylhydrazine, an aldehyde inhibitor. We have, therefore, tested the effects of spermine dialdehyde or malon dialdehyde on the induction of iNOS. Spermine dialdehyde (SDA, 10-M) inhibits nitrite formation by IFN-activated J774.2 cells in the absence of serum when given as a pretreatment but not when given 6 h after stimulation. In contrast, malon dialdehyde was ineffective. Thus, aldehyde metabolites of spermine, such as SDA, account for the inhibitory effect of polyamines on the induction of NOS in vitro. 6 The inhibitory effect of polyamines on iNOS induction appears to be fairly specific to iNOS, for spermine does not inhibit LPS-induced production of prostaglandin F2g or tumour necrosis factor.

Spermine Causes Loss of Innate Immune Response to Helicobacter pylori by Inhibition of Inducible Nitric-oxide Synthase Translation

“…Arginase converts L-arginine to L-ornithine, which is metabolized by ODC to produce the polyamine putrescine that is converted to the polyamines spermidine and spermine. Spermine has been shown to inhibit the immune effector function of monocytes and macrophages in response to LPS (11,12). We now report that spermine inhibits H. pylori-stimulated NO production in macrophages by a post-transcriptional effect on iNOS translation.…”

“…Spermine has been reported to inhibit NO production in LPS-stimulated J774 macrophages (12,16). However, in these studies, the effect on iNOS itself was not studied.…”

The Effect of e-, i-, and n-Nitric Oxide Synthase Inhibition on Colonic Motility in Normal and Muscular Dystrophy (Mdx) Mice

“…Both L-NIO and 1,3-PBIT inhibit e-, i-, and n-NOS at different concentrations [20][21][22]. Spermine, an end-product of polyamine biosynthesis from L-arginine, is a competitive inhibitor of n-and i-NOS [23][24][25]. DPI is an i-NOS inhibitor [26,27].…”

Spontaneous rearrangement of aminoalkylisothioureas into mercaptoalkylguanidines, a novel class of nitric oxide synthase inhibitors with selectivity towards the inducible isoform

“…Nitrite production, an indicator of NO synthesis, was measured in the supernatant of J774.2 macrophages as previously described (Szabo et al, 1994b (Szabo et al, 1994b). Cells in 96-well plates were incubated with MTT (0.2 mg ml-1) for 60 min at 370C.…”

“…To induce iNOS, fresh culture medium containing E. coli LPS (10 Mg ml-') and murine y-interferon (IFN, 50 u ml-') was added. Nitrite accumulation in the cell culture medium (in the absence or presence of various inhibitors) was measured after 24 h. Unless stated otherwise, NOS inhibitors were added 6 h after LPS/IFN in order to prevent interference with the process of iNOS induction (see: Szabo et al, 1994b). In some experiments, agents were applied together with excess L-arginine (in total 1.4, 10 or 30 mM in the culture medium, as opposed to 0.4 mm in normal DMEM).…”