Macrophage P2X4 receptors augment bacterial killing and protect against sepsis
Balázs Csóka, … , Pál Pacher, György Haskó
Balázs Csóka, … , Pál Pacher, György Haskó
Published June 7, 2018
Citation Information: JCI Insight. 2018;3(11):e99431. https://doi.org/10.1172/jci.insight.99431.
View: Text | PDF
Research Article Cell biology

Macrophage P2X4 receptors augment bacterial killing and protect against sepsis

Abstract

The macrophage is a major phagocytic cell type, and its impaired function is a primary cause of immune paralysis, organ injury, and death in sepsis. An incomplete understanding of the endogenous molecules that regulate macrophage bactericidal activity is a major barrier for developing effective therapies for sepsis. Using an in vitro killing assay, we report here that the endogenous purine ATP augments the killing of sepsis-causing bacteria by macrophages through P2X4 receptors (P2X4Rs). Using newly developed transgenic mice expressing a bioluminescent ATP probe on the cell surface, we found that extracellular ATP levels increase during sepsis, indicating that ATP may contribute to bacterial killing in vivo. Studies with P2X4R-deficient mice subjected to sepsis confirm the role of extracellular ATP acting on P2X4Rs in killing bacteria and protecting against organ injury and death. Results with adoptive transfer of macrophages, myeloid-specific P2X4R-deficient mice, and P2rx4 tdTomato reporter mice indicate that macrophages are essential for the antibacterial, antiinflammatory, and organ protective effects of P2X4Rs in sepsis. Pharmacological targeting of P2X4Rs with the allosteric activator ivermectin protects against bacterial dissemination and mortality in sepsis. We propose that P2X4Rs represent a promising target for drug development to control bacterial growth in sepsis and other infections.

Authors

Balázs Csóka, Zoltán H. Németh, Ildikó Szabó, Daryl L. Davies, Zoltán V. Varga, János Pálóczi, Simonetta Falzoni, Francesco Di Virgilio, Rieko Muramatsu, Toshihide Yamashita, Pál Pacher, György Haskó

×

Figure 4

P2X4Rs decrease mortality, bacterial load, inflammatory cytokines and chemokines, and organ injury in sepsis.

Options: View larger image (or click on image) Download as PowerPoint
P2X4Rs decrease mortality, bacterial load, inflammatory cytokines and ch...
(A) WT mice have improved survival compared with P2X4R–/– mice. Male WT and P2X4R–/– mice were subjected to CLP, and survival was monitored for 7 days. *P = 0.019 (WT and P2X4R–/–; n = 24 and 25, respectively). (B and C) Bacterial burden was determined by counting the number of CFUs on blood agar plates after serial dilution of blood and peritoneal lavage samples. Blood and lavage fluid were collected at 16 hour after CLP. *P < 0.05; **P < 0.01 vs. WT (WT and P2X4R–/–; n = 10 and 14, respectively, for blood; n = 10 and 14, respectively, for lavage). (D–N) WT and P2X4R–/– mice were subjected to sham or CLP operation and (D and I) IL-1β, (E and J) IL-6, (F and K) IL-10, (G and L) TNF-α, and (H and M) MIP-2 levels were determined with ELISA from blood and peritoneal lavage fluid collected at 16 hours after the operation. (N) Blood urea nitrogen (BUN) was measured in plasma of sham- or CLP-subjected WT and P2X4R–/– mice 16 hours after CLP. *P < 0.05, **P < 0.01, and ***P < 0.001 vs. WT sham- and CLP-operated WT and P2X4R–/–; n = 3, 4 for sham WT and P2X4R–/–; n = 6 and 10 for CLP WT. Data are expressed as mean ± SEM. All results are representatives of 3 experiments. Mortality curves were analyzed using Kaplan-Meier curve and log rank test and in the rest of the experiments one-way ANOVA followed by Mann Whitney test or two-tailed Student’s t test was used