Impact of Ectoenzymes on P2 and P1 Receptor Signaling

Purinergic signaling regulates a diverse and biologically relevant group of processes in the liver. However, progress of research into functions regulated by purinergic signals in the liver has been hampered by the complexity of systems probed. Specifically, there are multiple liver cell subpopulations relevant to hepatic functions, and many of these have been effectively modeled in human cell lines. Furthermore, there are more than 20 genes relevant to purinergic signaling, each of which has distinct functions. Hence, we felt the need to categorize genes relevant to purinergic signaling in the best characterized human cell line models of liver cell subpopulations. Therefore, we investigated the expression of adenosine receptor, P2X receptor, P2Y receptor, and ecto-nucleotidase genes via RT-PCR in the following cell lines: LX-2, hTERT, FH11, HepG2, Huh7, H69, and MzChA-1. We believe that our findings will provide an excellent resource to investigators seeking to define functions of purinergic signals in liver physiology and liver disease pathogenesis.

Section: Discussionmentioning

confidence: 99%

Expression of mediators of purinergic signaling in human liver cell lines

Goree

Lavoie

Fausther

et al. 2014

“…Extracellular nucleotide levels are determined by factors that control both cellular release and extracellular metabolism/degradation of nucleotides. Nucleotides are degraded by membrane adenosine triphosphate (ATP)-metabolizing proteins and by specific ectonucleotidases, including NTPDase1, 2, 3, and 8 and CD73 [16,17]. Knockout of the NTPDase1 (CD39) in mice causes an elevation in blood nucleotides and a metabolic syndrome phenotype (inflammation, hypertriglyceridemia, and insulin resistance) [18].…”

Section: Introductionmentioning

confidence: 99%

P2X receptors regulate adenosine diphosphate release from hepatic cells

Chatterjee

Sparks

2014

Extracellular nucleotides act as paracrine regulators of cellular signaling and metabolic pathways. Adenosine polyphosphate (adenosine triphosphate (ATP) and adenosine diphosphate (ADP)) release and metabolism by human hepatic carcinoma cells was therefore evaluated. Hepatic cells maintain static nanomolar concentrations of extracellular ATP and ADP levels until stress or nutrient deprivation stimulates a rapid burst of nucleotide release. Reducing the levels of media serum or glucose has no effect on ATP levels, but stimulates ADP release by up to 10-fold. Extracellular ADP is then metabolized or degraded and media ADP levels fall to basal levels within 2-4 h. Nucleotide release from hepatic cells is stimulated by the Ca 2+ ionophore, ionomycin, and by the P2 receptor agonist, 2′3′-O-(4-benzoyl-benzoyl)-adenosine 5′-triphosphate (BzATP). Ionomycin (10 μM) has a minimal effect on ATP release, but doubles media ADP levels at 5 min. In contrast, BzATP (10-100 μM) increases both ATP and ADP levels by over 100-fold at 5 min. Ion channel purinergic receptor P2X7 and P2X4 gene silencing with small interference RNA (siRNA) and treatment with the P2X inhibitor, A438079 (100 μM), decrease ADP release from hepatic cells, but have no effect on ATP. P2X inhibitors and siRNA have no effect on BzATP-stimulated nucleotide release. ADP release from human hepatic carcinoma cells is therefore regulated by P2X receptors and intracellular Ca 2+ levels. Extracellular ADP levels increase as a consequence of a cellular stress response resulting from serum or glucose deprivation.

“…The number of phosphate residues removed and the kinetic properties and substrate preferences vary from enzyme to enzyme [34,35]. Alkaline phosphatase, which is highly expressed in neutrophils, can convert ATP directly to adenosine, while CD73 converts AMP to adenosine [33]. In order to determine whether the expression of ectonucleotidases in breast cancer cells could be involved in their unique motility patterns, we used qPCR analysis to compare ectonucleotidase transcripts in HMEC and MDA-MB-231 cells.…”

Section: Breast Cancer Cells Express Ectonucleotidases That Promote Amentioning

confidence: 99%

Adenosine arrests breast cancer cell motility by A3 receptor stimulation

Ledderose

Hefti

Chen

et al. 2016

In neutrophils, adenosine triphosphate (ATP) release and autocrine purinergic signaling regulate coordinated cell motility during chemotaxis. Here, we studied whether similar mechanisms regulate the motility of breast cancer cells. While neutrophils and benign human mammary epithelial cells (HMEC) form a single leading edge, MDA-MB-231 breast cancer cells possess multiple leading edges enriched with A3 adenosine receptors. Compared to HMEC, MDA-MB-231 cells overexpress the ectonucleotidases ENPP1 and CD73, which convert extracellular ATP released by the cells to adenosine that stimulates A3 receptors and promotes cell migration with frequent directional changes. However, exogenous adenosine added to breast cancer cells or the A3 receptor agonist IB-MECA dose-dependently arrested cell motility by simultaneous stimulation of multiple leading edges, doubling cell surface areas and significantly reducing migration velocity by up to 75 %. We conclude that MDA-MB-231 cells, HMEC, and neutrophils differ in the purinergic signaling mechanisms that regulate their motility patterns and that the subcellular distribution of A3 adenosine receptors in MDA-MB-231 breast cancer cells contributes to dysfunctional cell motility. These findings imply that purinergic signaling mechanisms may be potential therapeutic targets to interfere with the motility of breast cancer cells in order to reduce the spread of cancer cells and the risk of metastasis.