Frontline Science: Anthrax lethal toxin-induced, NLRP1-mediated IL-1β release is a neutrophil and PAD4-dependent event

Abstract

Anthrax lethal toxin (LT) is a protease that activates the NLRP1b inflammasome sensor in certain rodent strains. Unlike better‐studied sensors, relatively little is known about the priming requirements for NLRP1b. In this study, we investigate the rapid and striking priming‐independent LT‐induced release of IL‐1β in mice within hours of toxin challenge. We find IL‐1β release to be a NLRP1b‐ and caspase‐1‐dependent, NLRP3 and caspase‐11‐independent event that requires both neutrophils and peptidyl arginine deiminiase‐4 (PAD4) activity. The simultaneous LT‐induced IL‐18 response is neutrophil‐independent. Bone marrow reconstitution experiments in mice show toxin‐induced IL‐1β originates from hematopoietic cells. LT treatment of neutrophils in vitro did not induce IL‐1β, neutrophil extracellular traps (NETs), or pyroptosis. Although platelets interact closely with neutrophils and are also a potential source of IL‐1β, they were unable to bind or endocytose LT and did not secrete IL‐1β in response to the toxin. LT‐treated mice had higher levels of cell‐free DNA and HMGB1 in circulation than PBS‐treated controls, and treatment of mice with recombinant DNase reduced the neutrophil‐ and NLRP1‐dependent IL‐1β release. DNA sensor AIM2 deficiency, however, did not impact IL‐1β release. These data, in combination with the findings on PAD4, suggest a possible role for in vivo NETs or cell‐free DNA in cytokine induction in response to LT challenge. Our findings suggest a complex interaction of events and/or mediators in LT‐treated mice with the neutrophil as a central player in induction of a profound and rapid inflammatory response to toxin.

Neutro“feels” lethal toxin

“…In this issue, Greaney et al. demonstrate that neutrophils and peptidyl arginine deiminase 4 (PAD4)‐dependent event are critical for LT‐induced IL‐1β production in vivo 7 . In addition, Greaney et al.…”

“…In addition, Greaney et al. also showed that cell‐free DNA are required for LT‐induced IL‐1β production in vivo 7 . Interestingly, LT treatment could not induce IL‐1β production by neutrophil in vitro, and DNA sensing absent in melanoma 2 (AIM2) sensors were dispensable for LT‐induced IL‐1β in vivo, highlighting the complexity of LT‐induced immune responses in vivo.…”

“…Using three different chimera systems (129 vs. C57BL/6; C57BL/6 – LT‐sensitive (S) vs. resistant (R); and Nlrp1b knockout (KO) vs. wild‐type (WT) on 129 background), Greaney et al. showed that NLRP1b in the hematopoietic compartment drives LT‐induced IL‐1β and IL‐18 production 7 . LT‐induced IL‐1β production was abrogated in caspase‐1/‐11 DKO >> WT chimera, suggesting the role for hematopoietic caspase‐1 in this process.…”

“…Greaney et al. showed that neutrophils express a functional PA receptor and that LF can access neutrophil cytoplasm (shown by cleaved MEK1 substrate), demonstrating that the inability of LT to activate NLRP1b inflammasome in neutrophils is not due to LF's inability to access neutrophil cytoplasm 7 . Given these data, the authors hypothesized that whereas neutrophils are necessary for IL‐1β production in vivo, they may not be a direct cellular source of IL‐1β.…”

“…Because LT failed to induce IL‐1β production by neutrophils in vitro, Greaney et al. investigated whether neutrophils provide alarm signals for hematopoietic cells to support IL‐1β production 7 . Following LT injection of NE and PAD4 KO mice, PAD4 KO, but not NE KO mice failed to produce IL‐1β.…”