P2X7 receptor activation induces reactive oxygen species formation in erythroid cells

Abstract

The presence of P2X7 on erythroid cells is well established, but its physiological role remains unclear. The current study aimed to determine if P2X7 activation induces reactive oxygen species (ROS) formation in murine erythroleukaemia (MEL) cells, a commonly used erythroid cell line. ATP induced ROS formation in a time-and concentrationdependent fashion. The most potent P2X7 agonist, 2′(3′)-O-(4-benzoylbenzoyl)ATP, but not UTP or ADP, also induced ROS formation. The P2X7 antagonist, A-438079, impaired ATP-induced ROS formation. The ROS scavenger, N-acetyl-L-cysteine, and the ROS inhibitor, diphenyleneiodonium, also impaired P2X7-induced ROS formation, but use of enzymespecific ROS inhibitors failed to identify the intracellular source of P2X7-induced ROS formation. P2X7-induced ROS formation was impaired partly by physiological concentrations of Ca 2+ and Mg 2+ and almost completely in cells in N-methyl-D-glucamine chloride medium. The p38 MAPK inhibitors SB202190 and SB203580, and the caspase inhibitor Z-VAD-FMK, but not N-acetyl-L-cysteine, impaired P2X7-induced MEL cell apoptosis. ATP also stimulated p38 MAPK and caspase activation, both of which could be impaired by A-438079. In conclusion, these findings indicate that P2X7 activation induces ROS formation in MEL cells and that this process may be involved in events downstream of P2X7 activation, other than apoptosis, in erythroid cells.

CAY10593 inhibits the human P2X7 receptor independently of phospholipase D1 stimulation

“…CAY10593 also impaired ATP-induced ethidium + uptake into P2X7-transfected HEK-293 cells, and primary human B cells, T cells and monocytes. In contrast, CAY10593 failed to affect ATP-induced ethidium + uptake into murine erythroleukemia cells (unpublished observations), which like RPMI 8226 cells also express endogenous P2X7 [12,38], indicating that CAY10593 does not act on murine P2X7.…”

“…Whether this second permeability state is attributed to intrinsic channel dilation [3], the pannexin-1 channel [4] or an alternate but unknown uptake pathway [5][6][7] remains controversial. Moreover, our understanding of this permeability state is further complicated with some [8][9][10] but not other [11][12][13] studies showing that P2X7-induced dye uptake involves the p38 mitogen-activated protein kinase. Regardless of the true identity of the P2X7 pore and the mechanism by which it opens, P2X7 activation stimulates several intracellular signalling pathways to induce various cellular events including inflammatory mediator release, reactive oxygen and nitrogen species formation, and cell proliferation or death [14,15].…”

The LIM Domain Only 4 Protein Is a Metabolic Responsive Inhibitor of Protein Tyrosine Phosphatase 1B That Controls Hypothalamic Leptin Signaling

“…Because purinergic signaling increases ROS production (Kawamura et al, 2012;Wang and Sluyter, 2013), this is likely to reduce LMO4 palmitoylation and ER retention. Of note, glial cells called tanycytes that line the third ventricle and secrete ATP in response to elevated extracellular glucose (Frayling et al, 2011).…”

The P2X7 Receptor Channel: Recent Developments and the Use of P2X7 Antagonists in Models of Disease

“…The generation of reactive oxygen species (ROS) after ATP activation of P2X7 is now well established in macrophages (Lenertz et al, 2009;Moore and MacKenzie, 2009), microglia (Apolloni et al, 2013;Bartlett et al, 2013), submandibular gland cells (Seil et al, 2008), and erythroid cells (Wang and Sluyter, 2013). Activation of NADPH oxidase 2 by P2X7 was a common feature found in all cell types studied, although there is evidence that ATP, possibly via P2X7 activation, can also enhance mitochondrial reactive oxygen species (Nakahira et al, 2011).…”