The activation of P2X7 receptor induces cathepsin D-dependent production of a 20-kDa form of IL-1β under acidic extracellular pH in LPS-primed microglial cells

Abstract

J. Neurochem. (2011) 117, 712–723. Abstract The potent pro‐inflammatory cytokine, interleukin‐1β (IL‐1β), is synthesized as an inactive 33‐kDa precursor (pro‐IL‐1β) and is processed by caspase 1 into the bioactive 17‐kDa mature form. The P2X7 receptor, an ATP‐gated cation channel, plays an essential role in caspase 1 activation, production and release of mature bioactive 17‐kDa form. We recently reported ATP induces the release of an unconventional 20‐kDa form of IL‐1β (p20‐IL‐1β) from lipopolysaccharide‐primed microglial cells. Emerging evidence suggests physiological relevance for p20‐IL‐1β; however, the underlying mechanisms for its production and release remain unknown. Here, we investigated the pathways involved in the ATP‐induced production of p20‐IL‐1β using lipopolysaccharide‐primed mouse microglial cells. The activation of P2X7 receptor by ATP triggered p20‐IL‐1β production under acidic extracellular conditions. ATP‐induced p20‐IL‐1β production was blocked by pepstatin A, a potent inhibitor of the lysosomal protease, cathepsin D. The removal of extracellular Ca2+ inhibited the p20‐IL‐1β production as well as ATP‐induced cathepsin D release via lysosome exocytosis. The acidic extracellular pH also facilitated the dilatation of membrane pore after ATP stimulation. Since facilitation of pore dilatation results in cytolysis accompanied with cytoplasmic pro‐IL‐1β leakage, our data suggest the leaked pro‐IL‐1β is processed into p20‐IL‐1β by cathepsin D released after ATP stimulation under acidic extracellular conditions.

Inflammasome activation by danger signals: extracellular ATP and pH

“…We found that P2X7R activation by ATP triggered the caspase-1-dependent production and secretion of mature IL-1β at pH 6.2-6.7 in LPS-primed mouse microglial cells [29]. This finding is supported by another study in which it was found that extracellular ATP triggered mature IL-1β secretion at pH 6.2 in LPS-primed mouse mixed glia [30].…”

“…This finding is supported by another study in which it was found that extracellular ATP triggered mature IL-1β secretion at pH 6.2 in LPS-primed mouse mixed glia [30]. In these experimental systems, acidic pH alone failed to induce the production and secretion of mature IL-1β [29,30]. Considering that acidic extracellular pH perturbs the functions of P2X7R channels, acidic pH-induced reductions in intracellular pH might be involved in the facilitation of P2X7R-mediated inflammasome assembly.…”

P2X7 receptors in cerebral ischemia

“…It is also known as a selective CAM-kinase-II inhibitor, a selective antagonist of the human but not the rat P2X 7 R. KN-62 attenuates the elevation of neuronal nitric oxide synthase serine phosphorylation induced by ischemia in MCAO animals, and plays a neuroprotective role early in ischemia [63] . cell culture [52] , indicating the efficacy of this drug in the brain. It is reported that this compound is essentially devoid of activity on other P2 receptors [64] .…”

“…Numerous studies indicate that P2X 7 Rs also participate in the regulation of neuroinflammation and neuroimmune reactions as an inflammatory mediator in the CNS [49][50][51][52][53] .…”

“…Three models were recently proposed to explain the non-classical secretion of mature IL-1β following P2X 7 R activation [52] . The oldest and simplest model suggests that P2X 7 R-mediated cytolysis directly triggers rapid caspase-1 activation followed by the release of mature IL-1β [10,53] depending on the K + and Ca 2+ efflux [54] .…”