Involvement of P2Y receptors in pyridoxal-5'-phosphate-induced cardiac preconditioning

International audienceUsing an isolated non‐working rat heart model, this study investigated the mechanisms of pharmacological pre‐conditioning (PC) induced by P2Y receptor stimulation with pyridoxal‐5′‐phosphate (PLP). After 6‐hydroxydopamine pretreatment and a 15‐min stabilization period, isolated rat hearts were perfused for 25 min then subjected to 40 min of global ischemia and 30 min of reperfusion (I/R); exposed for 15 min to 0.05 μm PLP bracketed for 25 min with broad‐spectrum P2 antagonists (suramin or PPADS) or with more specific P2Y antagonists (AMPαS or MRS2578), 1 μm each, followed by a 5‐min PLP‐free perfusion before I/R; treated during 25 min with either glybenclamide (GLY, 1 μm), 5‐hydroxydecanoic acid (5‐HD, 100 μm), U73122 (0.5 μm), H89 (1 μm), or KN93 (1 μm), with an infusion starting 5 min before PLP. The main endpoints were the rate–pressure product (RPP), creatine kinase (CK) release and area necrosis. Recovery of RPP, measured 5 min after reperfusion, was rapidly improved by PLP, blocked by the P2 antagonists, and decreased with the different inhibitors. Fifteen minutes after the end of ischemia, CK release reached maximal values in all groups. PLP provided significant protection, whereas the P2 antagonists, 5‐HD, a mitochondrial selective KATP antagonist and GLY a non‐selective KATP channel blocker, suppressed the protective effect on myocardial injury. The suppression of the cardioprotective effects of PLP by AMPαS, the PKA inhibitor (H89), and phospholipase C blocker (U73122) is in agreement with the P2Y11 receptor as a receptor for PLP‐induced PC. The suppression of the cardioprotective effects of PLP by MRS2578 and U73122 is in agreement with the P2Y6 receptor as a receptor for PLP‐induced PC. Pre‐ischemic exposure to nanomolar concentrations of PLP is protective against I/R. P2Y11 and P2Y6 represents the most likely candidate receptors for PLP‐induced cardiac PC

Ticagrelor Prevents Endothelial Cell Apoptosis through the Adenosine Signalling Pathway in the Early Stages of Hypoxia

International audienceBackground: Several studies have reported the beneficial effects of anti-platelet drugs in cardioprotection against ischaemia-reperfusion injuries. To date, no studies have focused on the indirect cytoprotective effects of ticagrelor via adenosine receptor on the endothelium. Method: By evaluating cell viability and cleaved caspase 3 expression, we validated a model of endothelial cell apoptosis induced by hypoxia. In hypoxic endothelial cells treated with ticagrelor, we quantified the extracellular concentration of adenosine, and then we studied the involvement of adenosine pathways in the cytoprotective effect of ticagrelor. Results: Our results showed that 10 µM ticagrelor induced an anti-apoptotic effect in our model associated with an increase of extracellular adenosine concentration. Similar experiments were conducted with cangrelor but did not demonstrate an anti-apoptotic effect. We also found that A2B and A3 adenosine receptors were involved in the anti-apoptotic effect of ticagrelor in endothelial cells exposed to 2 h of hypoxia stress. Conclusion: we described an endothelial cytoprotective mechanism of ticagrelor against hypoxia stress, independent of blood elements. We highlighted a mechanism triggered mainly by the increased extracellular bioavailability of adenosine, which activates A2B and A3 receptors on the endothelium

Complementary Role of P2 and Adenosine Receptors in ATP Induced-Anti-Apoptotic Effects Against Hypoxic Injury of HUVECs

Background: Vascular endothelial injury during ischemia generates apoptotic cell death and precedes apoptosis of underlying tissues. We aimed at studying the role of extracellular adenosine triphosphate (ATP) on endothelial cells protection against hypoxia injury. Methods: In a hypoxic model on endothelial cells, we quantified the extracellular concentration of ATP and adenosine. The expression of mRNA (ectonucleotidases, adenosine, and P2 receptors) was measured. Apoptosis was evaluated by the expression of cleaved caspase 3. The involvement of P2 and adenosine receptors and signaling pathways was investigated using selective inhibitors. Results: Hypoxic stress induced a significant increase in extracellular ATP and adenosine. After a 2-h hypoxic injury, an increase of cleaved caspase 3 was observed. ATP anti-apoptotic effect was prevented by suramin, pyridoxalphosphate-6-azophenyl-2′,4′-disulfonic acid (PPADS), and CGS15943, as well as by selective A2A, A2B, and A3 receptor antagonists. P2 receptor-mediated anti-apoptotic effect of ATP involved phosphoinositide 3-kinase (PI3K), extracellular signal-regulated kinases (ERK1/2), mitoKATP, and nitric oxide synthase (NOS) pathways whereas adenosine receptor-mediated anti-apoptotic effect involved ERK1/2, protein kinase A (PKA), and NOS. Conclusions: These results suggest a complementary role of P2 and adenosine receptors in ATP-induced protective effects against hypoxia injury of endothelial. This could be considered therapeutic targets to limit the development of ischemic injury of organs such as heart, brain, and kidney