EXOGENOUS N-G-HYDROXY-L-ARGININE CAUSES NITRITE PRODUCTION IN VASCULAR SMOOTH-MUSCLE CELLS IN THE ABSENCE OF NITRIC-OXIDE SYNTHASE ACTIVITY

Nitric oxide (NO) production from exogenous N(G)-hydroxy-L-arginine (OH-L-Arg) was investigated in rat aortic smooth muscle cells in culture by measuring nitrite accumulation in the culture medium. As well. the interaction between OH-L-Arg and L-arginine uptake via the y+ cationic amino acid transporter was studied. In cells without NO-synthase activity, OH-L-Arg (1-1000 muM) induced a dose-dependent nitrite production with a half-maximal effective concentration (EC50) of 18.0 +/- 1.5 muM (n = 4-7). This nitrite accumulation was not inhibited by the NO-synthase inhibitor N(G)-nitro-L-arginine methyl ester, L-NAME (300 muM). In contrast, it was abolished by miconazole (100 muM), an inhibitor of cytochrome P450. incubation of vascular smooth muscle cells with LPS (10 mug/ml) induced an L-NAME inhibited nitrite accumulation, but did not enhance the OH-L-Arg induced nitrite production. OH-L-Arg and other cationic amino acids, L-lysine and L-ornithine, competitively inhibited [H-3]-L-arginine uptake in rat aortic smooth muscle cells, with inhibition constants of 195 +/- 23 muM (n = 12), 260 +/- 40 muM (n = 5) and 330 +/- 10 muM (n = 5), respectively. These results show that OH-L-Arg is recognized by the cationic L-amino acid carrier present in vascular smooth muscle cells and can be oxidized to NO and nitrite in these cells in the absence of NO-synthase, probably by cytochrome P450 or by a reaction involving a cytochrome P450 by-product.