Browsing by Author "Arancibia Garavilla, Y"

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    Nitric oxide synthesis requires activity of the cationic and neutral amino acid transport system y(+)L in human umbilical vein endothelium
    (WILEY, 2003) Arancibia Garavilla, Y; Toledo, F; Casanello, P; Sobrevia, L
    L-Arginine transport is mediated by the cationic/neutral amino acid transport system y(+)L and cationic amino acid transporters y(+)/CATS in human umbilical vein endothelial cells (HUVECs). System y(+)/CATS activity may be rate-limiting for nitric oxide (NO) synthesis, but no reports have demonstrated system y(+)L involvement in NO synthesis in endothelium. We investigated the role of system y(+)L in NO synthesis in HUVECs. Transport of 1.5 muM L-arginine was inhibited (P < 0.05) by L-lysine (K-i, 1.4 muM), L-leucine (K-i,1.8 mum) and L-phenylalanine (K-i, 4.1 muM), but was unaltered (P > 0.05) by L-alanine or L-cysteine. The system y(+)/CATS inhibitor, N-ethylmaleimide (NEM), did not alter 1.5 muM L-arginine transport, but inhibited (92 +/- 3 %) 100 muM L-arginine transport. L-Arginine transport in the presence of NEM was saturable (V-max, 0.37 +/- 0.02 pmol (mug protein)(-1) min(-1); K-m, 1.5 +/- 0.3 muM) and competitively inhibited by L-leucine in the presence of Na+ (V-max, 0.49 +/- 0.06 pmol (mug protein)(-1) min(-1); K-m, 6.5 +/- 0.9 muM). HUVECs express SLC3A2/4F2hc, SLC7A7/4F2-lc2 and SLC7A6/4F2-lc3 genes encoding for the high-affinity transport system y(+)L. N-G-Nitro-L-arginine methyl ester and L-leucine, but not NEM, inhibited NO synthesis in medium containing 1.5 muM L-arginine. Cells exposed to 25 MM D-glucose (24 h) exhibited reduced system y(+)L activity (V-max, 0.15 +/- 0.008 pmol (mug protein)(-1) min(-1); K-m, 1.4 +/- 0.3 muM) and NO synthesis. However, 25 HIM D-glucose increased NO synthesis and L-arginine transport via system y(+). Thus, L-arginine transport through system y(+)L plays a role in NO synthesis, which could be a determining factor in pathological conditions where the endothelial L-arginine-NO pathway is altered, such as in diabetes mellitus.