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 mu M, the phosphatidylinositol 3-kinase (PI3K) inhibitor LY-294002 abolished the LPS-induced E-5'Nu activity. However, at 10 mu M, the NF-kappa B 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 mu M 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.