Stimulation of ecto-5'-nucleotidase in human umbilical vein endothelial cells by lipopolysaccharide.
The involvement of ecto-5'-nucleotidase (E-5'Nu) in the elevation of extracellular adenosine during inflammation is unclear. In the present study, the effect of lipopolysaccharide (LPS), an inflammation inducer, was investigated on E-5'Nu in human umbilical vein endothelial cells (HUVECs). E-5'Nu activity was enhanced after a 24 h exposure to LPS. This effect was dose dependent, with an EC50 of 1.66 ng/ml. At 10 microM, the phosphatidylinositol 3-kinase (PI3K) inhibitor LY-294002 abolished the LPS-induced E-5'Nu activity. However, at 10 microM, the NF-kappaB inhibitor ammonium pyrrolidine dithiocarbamate had no effect. LPS upregulated the protein expression but not the messenger RNA expression of E-5'Nu. The inhibition of E-5'Nu by 100 microM alpha,beta-methylene adenosine-5'-diphosphate increased the LPS-induced inflammation, suggesting that E-5'Nu plays a significant role in reducing inflammation, probably through the generation of adenosine. In conclusion, the experiments indicate that LPS upregulates E-5'Nu activity in HUVECs through a PI3K-dependent increase in the abundance of E-5'Nu on cell membranes. Since adenosine is an anti-inflammatory molecule, E-5'Nu upregulation may be crucial in protecting endothelial cells against inflammatory damage.
Li
R W S
RW
Department of Pharmacology, The University of Hong Kong, Pokfulam, Hong Kong.
Man
Ricky Y K
RY
Vanhoutte
Paul M
PM
Leung
George P H
GP
D015720:5'-Nucleotidase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D058917:Adenosine A2 Receptor Antagonists; D015153:Blotting, Western; D002478:Cells, Cultured; D002867:Chromones / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D016209:Interleukin-8 / Q000096:biosynthesis; D008070:Lipopolysaccharides / Q000494:pharmacology*; D009025:Morpholines / Q000494:pharmacology; D016328:NF-kappa B / Q000037:antagonists & inhibitors; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011247:Pregnancy; D012313:RNA / Q000096:biosynthesis / Q000302:isolation & purification; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013268:Stimulation, Chemical; D014471:Umbilical Veins / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D019010:Vascular Cell Adhesion Molecule-1 / Q000096:biosynthesis
D058917:Adenosine A2 Receptor Antagonists; D002867:Chromones; D004791:Enzyme Inhibitors; D016209:Interleukin-8; D008070:Lipopolysaccharides; D009025:Morpholines; D016328:NF-kappa B; D019010:Vascular Cell Adhesion Molecule-1; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D012313:RNA; D019869:Phosphatidylinositol 3-Kinases; D015720:5'-Nucleotidase