Essential role for Ca2+in regulation of IL-1β secretion by P2X7nucleotide receptor in monocytes, macrophages, and HEK-293 cells

Abstract

Interleukin (IL)-1β is a proinflammatory cytokine that elicits the majority of its biological activity extracellularly, but the lack of a secretory signal sequence prevents its export via classic secretory pathways. Efficient externalization of IL-1β in macrophages and monocytes can occur via stimulation of P2X7nucleotide receptors with extracellular ATP. However, the exact mechanisms by which the activation of these nonselective cation channels facilitates secretion of IL-1β remain unclear. Here we demonstrate a pivotal role for a sustained increase in cytosolic Ca2+to potentiate secretion of IL-1β via the P2X7receptors. Using HEK-293 cells engineered to coexpress P2X7receptors with mature IL-1β (mIL-1β), we show that activation of P2X7receptors results in a rapid secretion of mIL-1β by a process(es) that is dependent on influx of extracellular Ca2+and a sustained rise in cytosolic Ca2+. Moreover, reduction in extracellular Ca2+attenuates ∼90% of P2X7receptor-mediated IL-1β secretion but has no effect on enzymatic processing of precursor IL-1β (proIL-1β) to mIL-1β by caspase-1. Similar experiments with THP-1 human monocytes and Bac1.2F5 murine macrophages confirm the unique role of Ca2+in P2X7receptor-mediated secretion of IL-1β. In addition, we report that cell surface expression of P2X7receptors in the absence of external stimulation also results in enhanced release of IL-1β and that this can be repressed by inhibitors of P2X7receptors. We clarify an essential role for Ca2+in ATP-induced IL-1β secretion and indicate an additional role of P2X7receptors as enhancers of the secretory apparatus by which IL-1β is released.