Polarized Cytokine Release Triggered by P2X7 Receptor from Retinal Pigmented Epithelial Cells Dependent on Calcium Influx.

Xiaolei Shao,Sonia Guha,Keith E Campagno,Wenli Yang,Claire H Mitchell,Jason A Mills,Wennan Lu,Jonathan M Beckel

doi:10.3390/cells9122537

Abstract

Cytokine release from non-inflammatory cells is a key step in innate immunity, and agonists triggering cytokine release are central in coordinating responses. P2X7 receptor (P2X7R) stimulation by extracellular ATP is best known to active the NLRP3 inflammasome and release IL-1β, but stimulation also leads to release of other cytokines. As cytokine signaling by retinal pigmented epithelial (RPE) cells is implicated in retinal neurodegeneration, the role of P2X7R in release of cytokine IL-6 from RPE cells was investigated. P2X7R stimulation triggered IL-6 release from primary mouse RPE, human iPS-RPE and human ARPE-19 cells. IL-6 release was polarized, with predominant rise across apical membranes. IL-6 release was inhibited by P2X7R antagonists A438079, A839977, and AZ10606120, but not the NRTI lamivudine (3TC), P2X1R antagonist NF279, or P2Y1R antagonist MRS2179. P2X7R-mediated IL-6 release required extracellular Ca2+ and was blocked by Ca2+ chelator BAPTA. IL-6 release and Ca2+ elevation occurred rapidly, consistent with vesicular IL-6 staining in unstimulated cells. P2X7R stimulation did not trigger IL-1β release in these unprimed cells. P2X7R-mediated IL-6 release was enhanced in RPE cells from the ABCA4−/− mouse model of retinal degeneration. In summary, P2X7R stimulation triggers rapid Ca2+-dependent IL-6 release across the apical membrane of RPE cells.

Highlights

Release of cytokines from non-immune cells is a key step in innate immune responses, and the receptors that activate this cytokine release represent critical targets in controlling the response [1,2].Stimulation of the P2X7 receptor for extracellular ATP is associated with the assembly of the NOD, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome and the resulting release of cytokines interleukin (IL)-1β and IL-18 [3,4]
The findings suggest that the P2X7 receptor can trigger a rapid release of IL-6 from retinal pigmented epithelial (RPE) cells that is dependent on increased intracellular Ca2+ and not reliant on IL-1β
IL-6 Release Triggered by P2X7 Receptor Activation in Mouse, Induced Pluripotent Stem (iPS)-RPE and ARPE-19 Cells

Summary

Introduction

Release of cytokines from non-immune cells is a key step in innate immune responses, and the receptors that activate this cytokine release represent critical targets in controlling the response [1,2].Stimulation of the P2X7 receptor for extracellular ATP is associated with the assembly of the NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome and the resulting release of cytokines interleukin (IL)-1β and IL-18 [3,4]. As IL-1β is considered a “master cytokine”, capable of increasing expression and release of other cytokines [5], stimulation of the P2X7 receptor can be important in coordinating components of the innate immune response. Whether the P2X7 receptor can trigger cytokine upregulation and release independent of IL-1β is less well studied. This is relevant as the NLRP3 inflammasome requires “priming” to increase expression of key components, such as NLRP3 itself, before the activation stage can occur [6], and priming of inflammasome components may be rate-limiting in chronic age-dependent diseases [7]. The involvement of the P2X7 receptor in release of cytokines other than IL-1β may have implications for earlier stages in chronic inflammatory diseases

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