P2X4 and P2X7 receptors in the expression and release of interleukin-1β

Abstract

Pro-inflammatory cytokines of innate immunity play a pivotal role in host defence against infection by modulating the gene expression of numerous target genes, including cytokines, cell surface receptors, and their own expression in neighbouring cells. Their expression can be induced by pathogen-associated molecular patterns (PAMPs) and damage-associated molecular patterns (DAMPs), such as lipopolysaccharide (LPS) and extracellular adenosine triphosphate (ATP). ATP can be released in large quantities by damaged cells, activating the P2RX7 in innate immune cells. The pro-inflammatory cytokine IL-1β is produced in its inactive form by mononuclear phagocytes upon ‘priming’ with LPS, while subsequent ATP-mediated P2RX7 activation leads to the maturation and release of the bioactive cytokine. The present study aimed to investigate the role of the P2RX4, another ATP-sensitive receptor, in the secretion of IL-1β by human mononuclear phagocytes. Moreover, the function of the P2RX4 and the P2RX7 in the priming of mononuclear phagocytes was examined. The human monocytic THP 1 cell line, THP 1 cell-derived M1 like macrophages as well as primary human peripheral blood mononuclear cells and human peritoneal macrophages were primed with LPS and stimulated with the P2RX agonists BzATP or ATP or with nigericin in the absence or presence of different P2RX4 and P2RX7 antagonists, and the release of IL-1β was quantified. The results demonstrated that the P2RX4 is also involved in the ATP-dependent secretion of IL-1β by mononuclear phagocytes. Furthermore, cells were primed with LPS in the presence of the antagonists, and the release of IL-1β and the pro-inflammatory IL-6, as well as the mRNA levels of IL1B, IL6, IL10 and other cytokines, were analysed by ELISA or by real-time RT-PCR, respectively. In monocytic cells, treatment with the P2RX4 antagonist 5 B before priming resulted in increased IL1B mRNA expression and in increased ATP independent IL-1β release. Additionally, there was a slight increase in IL6 mRNA and IL-6 secretion, as wells as a clear-cut increase in IL10 mRNA. Treatment with 5 B also resulted in the activation of caspase 3/ 7 in monocytic THP 1 cells, which suggests that 5 B may induce apoptosis. During major surgery or accidental trauma, the extensive release of ATP leads to massive cytokine release, which can cause life-threatening systemic hyperinflammation. While 5 B enhances the LPS-induced inflammation, which essentially contributes to host defence against infection, 5 B dampens IL-1β maturation and release in response to extracellular ATP, a potent trauma-associated DAMP. Presumably, the effects of 5 B are mediated via P2RX4. These findings may pave the way towards novel therapeutic strategies that protect surgical and traumatized patients against infection, while efficiently preventing sterile hyperinflammation.