Walachowski, S. | Search bioRxiv

bioRxiv2022

Immunomodulation of Neutrophil Granulocyte Functions by Bacterial Polyphosphates

Polyphosphates are highly conserved, linear polymers of monophosphates that reside in all living cells. Bacteria produce long chains containing hundreds to thousands of phosphate units, which can interfere with host defense to infection. Here, we report that intratracheal long-chain polyphosphate administration to C57BL/6J mice resulted in the release of proinflammatory cytokines and influx of Ly6G+ polymorphonuclear neutrophils in the bronchoalveolar lavage fluid causing a disruption of the physiologic endothelial-epithelial small airway barrier and histologic signs of lung injury. Polyphosphate-induced effects were attenuated after neutrophil depletion in mice. In isolated murine neutrophils, long-chain polyphosphates modulated cytokine release induced by lipopolysaccharides (LPS) from gram-negative bacteria or lipoteichoic acid from gram-positive bacteria. In addition, long-chain polyphosphates induced immune evasive effects in human neutrophils. In detail, long-chain polyphosphates down-regulated CD11b and curtailed the phagocytosis of E. coli particles by neutrophils. Polyphosphates modulated the migration capacity by inducing CD62L shedding resulting in CD62Llow and CD11blow neutrophils. The release of IL-8 induced by LPS was also significantly reduced. Pharmacologic blockade of PI3K with wortmannin antagonized long-chain polyphosphate-induced effects on LPS-induced IL-8 release. In conclusion, polyphosphates govern immunomodulation in murine and human neutrophils suggesting polyphosphates as a therapeutic target for bacterial infections to restore innate immune defense.

immunology↗

bioRxiv2022

Bacterial Polyphosphates Induce CXCL4 and Synergize with Complement Anaphylatoxin C5a in Lung Injury

Polyphosphates are linear polymers of inorganic phosphates that exist in all living cells and serve pleiotropic functions. Bacteria produce long-chain polyphosphates, which can interfere with host defense to infection. In contrast, short-chain polyphosphates are released from platelet dense granules and bind to the chemokine, CXCL4. Here, we report that long-chain polyphosphates induced the release of CXCL4 from mouse bone marrow-derived macrophages and peritoneal macrophages in a dose-/time-dependent fashion resulting from an induction of CXCL4 mRNA. This polyphosphate effect was lost after pre-incubation with recombinant exopolyphosphatase (PPX) Fc fusion protein, demonstrating the potency of long chains over monophosphates and ambient cations. In detail, polyphosphate chains longer than 70 inorganic phosphate residues were typically required to mediate robust CXCL4 release. Polyphosphates acted independently of the purinergic P2Y1 receptor and the MyD88 and TRIF adaptors of Toll-like receptors. On the other hand, polyphosphates augmented LPS/MyD88-induced CXCL4 release, which was explained by intracellular signaling convergence on PI3K/Akt. Polyphosphates alone induced phosphorylation of Akt at threonine-308. Pharmacologic blockade of PI3K (wortmannin, LY294002) antagonized polyphosphate-induced CXCL4 release from macrophages. Intra-tracheal polyphosphate administration to C57BL/6J mice caused histologic signs of lung injury, disruption of the endothelial-epithelial barrier, influx of Ly6G+ polymorphonuclear neutrophils, depletion of CD11c+SiglecF+ alveolar macrophages, and release of CXCL4. Long-chain polyphosphates synergized with the complement anaphylatoxin, C5a, which was partly explained by upregulation of the receptor, C5aR1, on myeloid cells. C5aR1-/- mice were protected from polyphosphate-induced lung injury. In conclusion, we demonstrate that polyphosphates govern immunomodulation in macrophages and are capable of inducing lung injury.

immunology↗