Inhibition of cytokine-mediated JNK signalling by purinergic P2Y11 receptors, a novel protective mechanism in endothelial cells

Abstract

Previous research from our laboratory has demonstrated a novel phenomenon whereby GPCRs play a role in inhibiting cytokine-mediated c-Jun N-terminal kinase (JNK) signalling. So far this novel phenomenon seems to have been vastly overlooked, with little research in the area. Therefore, in this study we explored this further; by assessing the potential of P2YRs to mediate inhibition of cytokine-mediated JNK signalling and related functional outcomes in human endothelial cells. We utilised primary endothelial cells, and employed the use of endogenous activators of P2YRs and well characterised pharmacological inhibitors, to assess signalling parameters mediated by P2YRs, Interleukin-1β (IL-1β), TNFα and JNK. Activation of P2YRs with adenosine tri-phosphate (ATP) resulted in a time- and concentration-dependent inhibition of IL-1β-mediated phosphorylation of JNK and associated kinase activity. The effect was specific for cytokine-mediated JNK signalling, as ATP was without effect on JNK induced by other non-specific activators (e.g. sorbitol, anisomycin), nor effective against other MAPK pathways such as p38 and the canonical NFκB cascade. Pharmacological studies demonstrated a role for the P2Y receptor in mediating this effect, but not the P2Y nor the adenosine receptors (A1, A2A, A2B & A3). The novel Gα inhibitor YM254890 and a protein kinase A (PKA) inhibitor H89 both partially reversed ATP-mediated inhibition of IL-1β-stimulated JNK indicating involvement of both Gα and Gα mediated pathways. ATP also partially reversed IL-1β-mediated induction of cyclo‑oxygenase-2 (COX-2) and E-selectin. Collectively, these studies indicate the potential for activation of purinergic receptors to protect the endothelium from inflammatory driven JNK activation and may be a new target for inflammatory disease therapy.

P2Y Purinergic Receptors, Endothelial Dysfunction, and Cardiovascular Diseases

“…NF157 decreases IL-6 and TNFα cytokine production and the activation of p38 MAPK in ECs and the consequent binding of monocytes to EC due to up-regulation of E-selectin and VCAM-1 [ 71 ]. Other reports show the anti-inflammatory effect of P2Y11 in EC, where the receptor can block the signaling activated by IL-1, TNFα, and other cytokines, particularly the activation of JNK [ 72 ]. Thus, agents activating P2Y11 could have protective therapeutic value to combat the atherosclerosis and inflammation that drives the disease.…”

Inflammatory stress-mediated chromatin changes underlie dysfunction in endothelial cells

“…For example, in rheumatoid arthritis patients, the non-immune cells responsible for removing debris or synoviocytes, but not macrophages, demonstrated prolonged expression of genes with a single TNF-alpha pulse accompanied by an increase in chromatin accessibility 10 . Endothelial responses to inflammatory stress largely rely on the mitogen-activated kinase (MAPK) signal transduction, specifically, p38MAP kinase (p38MAPK) and c-Jun N-terminal kinase (JNK), both well-known stress-activated protein kinases, due to their essential roles in orchestrated stress responses, including cytokines 11,12 . Despite the significant role of inflammation in vascular disorders, the consequences of inflammatory stimuli on the chromatin organization of vascular endothelial cells and the contribution of febrile-like fever to endothelial dysfunction are poorly understood.…”