Human Neutrophils Generate Extracellular Vesicles That Modulate Their Functional Responses.

Neutrophils influence innate and adaptive immunity by releasing various cytokines and chemokines, by generating neutrophil extracellular traps (NETs), and by modulating their own survival. Neutrophils also produce extracellular vesicles (EVs) termed ectosomes, which influence the function of other immune cells. Here, we studied neutrophil-derived ectosomes (NDEs) and whether they can modulate autologous neutrophil responses. We first characterized EV production by neutrophils, following MISEV 2018 guidelines to facilitate comparisons with other studies. We found that such EVs are principally NDEs, that they are rapidly released in response to several (but not all) physiological stimuli, and that a number of signaling pathways are involved in the induction of this response. When co-incubated with autologous neutrophils, NDE constituents were rapidly incorporated into recipient cells and this triggered and/or modulated neutrophil responses. The pro-survival effect of GM-CSF, G-CSF, IFNγ, and dexamethasone was reversed; CXCL8 and NET formation were induced in otherwise unstimulated neutrophils; the induction of inflammatory chemokines by TNFα was modulated depending on the activation state of the NDEs' parent cells; and inducible NET generation was attenuated. Our data show that NDE generation modulates neutrophil responses in an autocrine and paracrine manner, and indicate that this probably represents an important aspect of how neutrophils shape their environment and cellular interactions.

Regulation of Discrete Functional Responses by Syk and Src Family Tyrosine Kinases in Human Neutrophils.

Neutrophils play a critical role in innate immunity and also influence adaptive immune responses. This occurs in good part through their production of inflammatory and immunomodulatory cytokines, in conjunction with their prolonged survival at inflamed foci. While a picture of the signaling machinery underlying these neutrophil responses is now emerging, much remains to be uncovered. In this study, we report that neutrophils constitutively express various Src family isoforms (STKs), as well as Syk, and that inhibition of these protein tyrosine kinases selectively hinders inflammatory cytokine generation by acting posttranscriptionally. Accordingly, STK or Syk inhibition decreases the phosphorylation of signaling intermediates (e.g., eIF-4E, S6K, and MNK1) involved in translational control. By contrast, delayed apoptosis appears to be independent of either STKs or Syk. Our data therefore significantly extend our understanding of which neutrophil responses are governed by STKs and Syk and pinpoint some signaling intermediates that are likely involved. In view of the foremost role of neutrophils in several chronic inflammatory conditions, our findings identify potential molecular targets that could be exploited for future therapeutic intervention.