Exogenous NG‐hydroxy‐l‐arginine causes nitrite production in vascular smooth muscle cells in the absence of nitric oxide synthase activity

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Abstract
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 μM) induced a dose‐dependent nitrite production with a half‐maximal effective concentration (EC50) of 18.0 ± 1.5 μM (n = 4–7). This nitrite accumulation was not inhibited by the NO‐synthase inhibitor N G‐nitro‐l‐arginine methyl ester, l‐NAME (300 μM). In contrast, it was abolished by miconazole (100 μM), an inhibitor of cytochrome P450. Incubation of vascular smooth muscle cells with LPS (10 ) 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 [3H]‐l‐arginine uptake m rat aortic smooth muscle cells, with inhibition constants of 195 ± 23 μM(n = 12), 260 ± 40 μM(n= 5) and 330 ± 10 μM(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 byproduct.

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