Stimulation of ecto-5′-nucleotidase in human umbilical vein endothelial cells by lipopolysaccharide

Abstract

Li RW, Man RY, Vanhoutte PM, Leung GP. Stimulation of ecto-5Ј-nucleotidase in human umbilical vein endothelial cells by lipopolysaccharide. Am J Physiol Heart Circ Physiol 295: H1177-H1181, 2008. First published July 18, 2008 doi:10.1152/ajpheart.91513.2007.-The involvement of ecto-5Ј-nucleotidase (E-5ЈNu) 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-5ЈNu in human umbilical vein endothelial cells (HUVECs). E-5ЈNu activity was enhanced after a 24 h exposure to LPS. This effect was dose dependent, with an EC 50 of 1.66 ng/ml. At 10 M, the phosphatidylinositol 3-kinase (PI3K) inhibitor LY-294002 abolished the LPS-induced E-5ЈNu activity. However, at 10 M, the NF-B inhibitor ammonium pyrrolidine dithiocarbamate had no effect. LPS upregulated the protein expression but not the messenger RNA expression of E-5ЈNu. The inhibition of E-5ЈNu by 100 M ␣,␤-methylene adenosine-5Ј-diphosphate increased the LPS-induced inflammation, suggesting that E-5ЈNu plays a significant role in reducing inflammation, probably through the generation of adenosine. In conclusion, the experiments indicate that LPS upregulates E-5ЈNu activity in HUVECs through a PI3K-dependent increase in the abundance of E-5ЈNu on cell membranes. Since adenosine is an anti-inflammatory molecule, E-5ЈNu upregulation may be crucial in protecting endothelial cells against inflammatory damage.adenosine; cytokines; inflammation ECTO-5Ј-NUCLEOTIDASE (E-5ЈNu), also known as CD73, is a 70-kDa membrane-bound glycoprotein that is abundant in the endothelium (18). E-5ЈNu extracellularly catalyzes the conversion of purine and pyrimidine ribo-and deoxyribonucleoside monophosphates to their corresponding ribo-and deoxynucleosides. For example, adenosine 5Ј-monophosphate (AMP) is one of the substrates of E-5ЈNu, and adenosine is the corresponding end product (9, 21).Adenosine exerts its effects by interacting with four receptors known as A 1 , A 2A , A 2B , and A 3 receptors (25,27). In addition to its well-known vasodilator and cardioprotective effects (20,23,25), adenosine can also serve as an immunosuppressive agent to minimize tissue damage during inflammation (13,22,25). The stimulation of A 2A receptors downregulates neutrophil functions (25), reduces the recruitment of neutrophil and macrophages to the endothelium (15), and inhibits the production of tumor necrosis factor (TNF)-␣ in peripheral mononuclear white blood cells (4).Endogenous adenosine is increased during inflammation. Since part of the adenosine is derived from the catalytic action of E-5ЈNu, it is plausible that E-5ЈNu activity is subject to change by inflammation and may play an important role in the regulation of the extracellular adenosine level during inflammation. TNF-␣, which is a proinflammatory cytokine, decreases the expression and activity of E-5ЈNu (9), suggesting that less adenosine may be produced to relieve inflammation if E-5ЈNu is downregulated. By ...

Estradiol Regulation of Nucleotidases in Female Reproductive Tract Epithelial Cells and Fibroblasts

“…Just why gene expression and biological activity were so low in our studies of endothelial cells is unclear. Endothelial cells are known to express NT5E that are stimulated by LPS [57] and inhibited by TNF-α [58]. It may be that under our culture conditions, endothelial cells are either not stimulated or that nucleotidase gene expression is negatively regulated and therefore express low levels of nucleotidases.…”

Differential Macrophage Activation Alters the Expression Profile of NTPDase and Ecto-5′-Nucleotidase

“…4B). A probable explanation for the decreased ecto-5′-NT activity in macrophages stimulated by LPS (M1) is the activation of the transcription factor NF-κB, which may suppress the transcription of the gene encoding ecto-5′-NT [56]. This notion is supported by studies that have demonstrated that anti-inflammatory effects of methotrexate results from the activation of ecto-5′-NT through suppression of NF-κB [57].…”

Lipopolysaccharide Inhibits the Channel Activity of the P2X7 Receptor

“…These putative targets include the activation of the inflammasome (in codependence of P2Xs and TLRs activation [36]) and could be related to the P2X7R-pannexin-1 complex, caspase activation, and production of inflammatory mediators [37, 38]. Ectonucleotidase inhibition by LPS [39–41] does not seem to be involved under our experimental conditions; it has been demonstrated that ATP degradation appears to be lower and not significant [14]. The possibility cannot be excluded that LPS could interact directly with ATP, decreasing its free concentration.…”