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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
Author(s) -
ArancibiaGaravilla Yerko,
Toledo Fernando,
Casanello Paola,
Sobrevia Luis
Publication year - 2003
Publication title -
experimental physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.925
H-Index - 101
eISSN - 1469-445X
pISSN - 0958-0670
DOI - 10.1113/eph8802647
Subject(s) - nitric oxide , arginine , leucine , umbilical vein , amino acid , chemistry , human umbilical vein endothelial cell , alanine , biochemistry , endothelium , biology , endocrinology , in vitro , organic chemistry
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 μM L‐arginine was inhibited (P < 0.05) by L‐lysine (K i , 1.4 μM), L‐leucine (K i , 1.8 μM) and L‐phenylalanine (K i , 4.1 μM), 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 μM L‐arginine transport, but inhibited (92 ± 3%) 100 μM L‐arginine transport. L‐Arginine transport in the presence of NEM was saturable (V max , 0.37 ± 0.02 pmol (μg protein) −1 min −1 ; K m , 1.5 ± 0.3 μM) and competitively inhibited by L‐leucine in the presence of Na + (V max , 0.49 ± 0.06 pmol (μg protein) −1 min −1 ; K m , 6.5 ± 0.9 μM). 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 μM L‐arginine. Cells exposed to 25 mM D‐glucose (24 h) exhibited reduced system y + L activity (V max , 0.15 ± 0.008 pmol (μg protein) −1 min −1 ; K m , 1.4 ± 0.3 μM) and NO synthesis. However, 25 mM 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.