Induction of uptake of n g monomethyl-l-arginine (l-nmma) by human
endothelial cells and murine macrophages: a possible mechanism for
differential inhibition of nitric oxide biosynthesis.
Bogle, Richard G., Raymond J. Macallister, Guy St. J. Whitley, and
Patrick Vallance.
Departments of Pharmacology and Clinical Pharmacology, and Cellular
and Molecular Sciences, St. George's Hospital Medical School, Cranmer
Terrace, London SW17 0RE, U.K., Telephone: 081-725 5611, Fax:081-682
-0487
APStracts 2:0152C, 1995.
The properties, selectivity and regulation of NGmonomethyl-L-arginine
(L-NMMA) uptake were examined in a cultured human vascular
endothelial cell line (SGHEC-7) and murine macrophage J774 cells. In
both cell types the uptake of L-[14C]NMMA was time-and temperature
-dependent. In endothelial cells L-NMMA uptake occurred via a single
-saturable carrier mediated system with an apparent Kt of 77 +/- 2
[mu]M. In murine macrophage cells a saturable component with an
apparent Kt of 51 +/- 6 [mu]M and a non-saturable component of L-NMMA
uptake were identified. In both cell types uptake of L-[14C]NMMA (10
[mu]M) was significantly inhibited in the presence of 100-fold excess
of either L-NMMA, asymmetrical dimethyl-L-arginine (ADMA),
symmetrical dimethyl-L-arginine (SDMA), L-canavanine, L-arginine and
to a lesser extent D-arginine. Uptake of L-NMMA was inhibited weakly
( nearly equal to 30%) by NGnitro-L-arginine (L-NOARG), NGnitro-L
-arginine methylester (L-NAME), aminoguanidine and L-citrulline.
Incubation of macrophage J774 cells with lipopolysaccharide (1 or 10
[mu]g/ml) resulted in the induction of nitric oxide synthase activity
as determined by the accumulation of nitrite in the culture medium.
In these cells an enhanced uptake of L-NMMA uptake was also observed
which was prevented by pretreatment with cycloheximide (1 [mu]M) but
not dexamethasone (1 [mu]M). In endothelial cells stimulation for 6 h
with either lipopolysaccharide (1 [mu]g/ml) or tumour necrosis factor
(5 ng/ml) and interleukin-1[beta] (10 ng/ml) resulted in an enhanced
uptake of L-NMMA which was protein synthesis dependent but not
sensitive to inhibition by glucocorticoids. These results show that
endothelial cells and J774 macrophages transport L-NMMA by a carrier
-mediated transporter which is shared by L-arginine and other
endogenous dimethylated arginine analogues (e.g. ADMA and SDMA).
Increases in L-NMMA uptake following cytokine or lipopolysaccharide
treatment may allow a higher intracellular concentration of this
nitric oxide synthase inhibitor to be reached in such cells and
therefore may represent a mechanism for cell selective inhibition of
nitric oxide synthase.
Received 17 November 1994; accepted in final form 14 February
1995.
APS Manuscript Number C672-4.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 4 April 1995.