[A new synthetic tripeptide inhibits L-arginine transport in macrophages].
OBJECTIVE
To observe the effect of a new synthetic tripeptide [Arg(NO(3))- Lys(OCH(3))- Arg(NO(3))] on L-arginine/NO pathway in the macrophage cell strain RAW246.7.


METHODS
The cultured macrophages exposed to lipopolysaccharide (LPS, 1 microg/L) treatment were randomly divided into 3 groups (n=6) and treated with distilled water, 1x10(-4) mol/L tripeptide and 1x10(-4) mol/L L-arginine, NG-monomethyl-L-arginine (L-NMMA) for 24 h, respectively. The macrophages were incubated for 24 h with LPS (1 microg/L) and in the presence of different concentrations of L-arginine (0 to 2 mmol/L), or in normal culture medium (containing 0.5 mmol/L L-arginine) for 24 h with LPS (1 microg/L) and in the presence of tripeptide of 0 to 10x10(-4) mol/L. The changes of [(3)H]-L-arginine transport and NO production from the macrophages were measured.


RESULTS
NO release from macrophages incubated in the LPS-containing culture medium was 50 folds, and [(3)H]-L-arginine uptake 2.7 folds that in cells in normal culture medium. Tripeptide (1x10(-4) mol/L) inhibited [(3)H]-L-arginine transport and NO production by 67% and 71% respectively. The effect of tripeptide was stronger than L-NMMA (P<0.05). Extracellular L-arginine caused a concentration-dependent increase in nitrite production, which reached the maximum at concentrations above 0.5 mmol/L Km for nitrite production of 0.162+/-0.015 mmol/L and Vmax of 91.2+/-2.3 micromol/(24h.10(6) cells). L-arginine transport and NO production were inhibited by tripeptide, but their dose-dependent pattern of changes was different with EC50 of 0.21x10(-4) mol/L and 1.27x10(-4) mol/L, respectively.


CONCLUSIONS
Activation of macrophages with LPS induces nitrite accumulation in the culture medium, which is dependent on the presence of extracellular L-arginine. The tripeptide induces dysfunction of L-arginine/NO pathway in macrophages, potently inhibits L-arginine transport and competitively combine the active sites of nitric oxide synthase.
Wang
Hui-qing
HQ
Department of Cardiology, Nanfang Hospital, Southern Medical University, Guangzhou 510515, China. whqlll@126.com
Wu
Sai-zhu
SZ
Ruan
Yun-jun
YJ
Cheng
Yu-ning
YN
Sun
Fei
F
Rong
Zhi-yi
ZY
Yin
Meng-zhuo
MZ
D001120:Arginine / Q000378:metabolism*; D001692:Biological Transport / Q000187:drug effects; D002478:Cells, Cultured; D006801:Humans; D008070:Lipopolysaccharides; D008264:Macrophages / Q000166:cytology / Q000378:metabolism*; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000037:antagonists & inhibitors*; D009842:Oligopeptides / Q000494:pharmacology*
D008070:Lipopolysaccharides; D009842:Oligopeptides; D009569:Nitric Oxide; D001120:Arginine; D019001:Nitric Oxide Synthase