Regulation of Ecto-5´-Nucleotidase by Docosahexaenoic Acid in Human Endothelial Cells

Abstract

Background/Aims: Modulation of extracellular adenine nucleotide and adenosine concentrations is one potential mechanism by which docosahexaenoic acid (DHA) may exert beneficial effects in critically ill patients. This study assessed DHA effects on extracellular adenine purines. Methods: Experiments used human pulmonary endothelial cells (HPMEC) and umbilical vein endothelial cells (HUVEC) treated with DHA (48 h). mRNA level (real-time PCR), expression (western blot, flow cytometry) and activities (hydrolysis of etheno(ε)-purines and fluorescence HPLC) of CD73 (ecto-5´-nucleotidase) and CD39 (ecto-NTPDase-1) were quantified. Results: DHA elevated total CD73 membrane protein expression concentration-dependently but CD73 mRNA level did not change. Increased expression was paralleled by increased enzyme activity. Effects observed on membrane level were reversed in intact cells, in which ε-AMP hydrolysis decreased after DHA. In intact endothelial cells ATP release was enhanced and CD39 activity blunted following DHA treatment. Hence, extracellular ATP and ADP concentrations increased and this inhibited ε-AMP hydrolysis. Conclusion: In human endothelial cells DHA caused 1) up-regulation of CD73 protein content and increased AMP hydrolysis at the cell membrane level, 2) increased cellular ATP release, and 3) decreased extracellular ATP/ADP hydrolysis. Thus, reorganization of the extracellular adenine-nucleotide-adenosine axis in response to DHA resulted in an increased extracellular ATP/adenosine ratio.

Docosahexaenoic acid reduces adenosine triphosphate-induced calcium influx via inhibition of store-operated calcium channels and enhances baseline endothelial nitric oxide synthase phosphorylation in human endothelial cells

“…Thus, it appears that eNOS activity in presence of DHA is rather augmented via enzyme phosphorylation than by calcium stimulation. In conjunction with a previous study in which we reported enhanced ATP and ADP release from endothelial cells by DHA treatment [12], multiple effects of this omega-3-fatty acid may interact stimulating endothelial cell ATP release, which via autocrine P2Y receptor activation stimulates membrane calcium influx and simultaneously changes eNOS phosphorylation in a manner predictive of enhancing nitric oxide production. Here, a separate effect of DHA also resulting in ser1177 phosphorylation may augment ATP dependent stimulation.…”

“…Once released, extracellular ATP enhances in an autocrine manner the endothelial cytosolic calcium and may subsequently increase endothelial cell nitric oxide production. As shown before, release and extracellular metabolism of ATP in endothelial cells are modulated by DHA resulting in an enhanced extracellular ATP concentration [12]. Thus, hypothetically a causal link may exist between the modulation of membrane lipid composition by DHA, extracellular ATP, endothelial cell calcium regulation and the generation of nitric oxide.…”

The Roles of CD73 in Cancer

“…Finally, some other studies have also reported that Wnt pathway [68], polyunsaturated fatty acid (PUFA) [69], and intracellular cAMP are also involved in regulation of CD73 expression [70]. Taken together, these results demonstrate that expression levels of CD73 are regulated by various factors via different mechanisms.…”

Docosahexaenoic acid (DHA) impairs hypoxia-induced cellular and exosomal overexpression of immune-checkpoints and immunomodulatory molecules in different subtypes of breast cancer cells

“…In this study, we noticed that DHA down-regulated the expression of CD39 and CD73 in two BC cells and their derived exosomes under hypoxic and normoxic conditions. Thom et al reported that CD39 activity was weaken after treatment with DHA in endothelial cells [ 32 ]. Their observations about expression of CD73 was increased in protein level but in mRNA level didn’t change after treatment with DHA [ 32 ].…”