Macrophages mediate flagellin induced inflammasome activation and host defense in zebrafish

Abstract

Summary The inflammasome is an innate immune complex whose rapid inflammatory outputs play a critical role in controlling infection, however the host cells that mediate inflammasome responses in vivo are not well defined. Using zebrafish larvae, we examined the cellular immune responses to inflammasome activation during infection. We compared the host responses to two Listeria monocytogenes strains: wild type and Lm-pyro, a strain engineered to activate the inflammasome via ectopic expression of flagellin. Infection with Lm-pyro led to activation of the inflammasome, macrophage pyroptosis, and ultimately attenuation of virulence. Depletion of caspase A, the zebrafish caspase-1 homolog, restored Lm-pyro virulence. Inflammasome activation specifically recruited macrophages to infection sites, whereas neutrophils were equally recruited to WT and Lm-pyro infections. Similar to caspase A depletion, macrophage deficiency rescued Lm-pyro virulence to wild type levels, while defective neutrophils had no specific effect. Neutrophils were however important for general clearance of L. monocytogenes, as both wild type and Lm-pyro were more virulent in larvae with defective neutrophils. This study characterizes a novel model for inflammasome studies in an intact host, establishes the importance of macrophages during inflammasome responses, and adds importance to the role of neutrophils in controlling L. monocytogenes infections.

Listeria monocytogenes and the Inflammasome: From Cytosolic Bacteriolysis to Tumor Immunotherapy

“…Similar early innate immune cell infiltrates consisting of predominantly macrophages were observed in zebrafish infected with inflammasome-activating L. monocytogenes ultimately resulting in attenuation (Fig. 7.2) (Vincent et al 2015). Previous work with Salmonella enterica subsp Typhimurium similarly engineered to robustly activate the inflammasome suggests that pyroptosis expels bacteria for uptake by bactericidal neutrophils ultimately resulting in phagocyte oxidase dependent attenuation (Miao et al 2010a).…”

“…Both Sauer et al (2011) and Tsuchiya et al (2014) observed no increased susceptibility to L. monocytogenes in Caspase-1/11 −/− mice at 2 days post infection. Similarly, there was no increased susceptibility to L. monocytogenes in zebrafish lacking Caspase-1 (Vincent et al 2015). These studies are consistent with the tight levels of regulation observed with potential inflammasome agonists as well as the critical nature of host cell survival that has previously been reported to promote L. monocytogenes virulence.…”

“…Because L. monocytogenes largely avoids activating the inflammasome, much of the work examining the host response to pyroptosis has used an engineered strain of L. monocytogenes that ectopically expresses flagellin resulting in hyper-activation of the inflammasome and attenuation (Sauer et al 2011a; Warren et al 2011; Williams et al 2013; Vincent et al 2015). Infection with inflammasome-activating L. monocytogenes resulted in an earlier MMC infiltrate that penetrates into the deep T-cell zones of the spleen; however, this infiltrate is less robust and clears earlier than with wild-type infection, corresponding with a drop in burden (Williams et al 2013).…”

“…Previous work with Salmonella enterica subsp Typhimurium similarly engineered to robustly activate the inflammasome suggests that pyroptosis expels bacteria for uptake by bactericidal neutrophils ultimately resulting in phagocyte oxidase dependent attenuation (Miao et al 2010a). However, neutrophil depletion does not specifically rescue virulence defects following activation of pyroptosis by L. monocytogenes (Sauer et al 2011a; Vincent et al 2015). Instead, depletion of macrophages rescues the virulence defect of inflammasome-activating L. monocytogenes (Vincent et al 2015).…”

zWEDGI: Wounding and Entrapment Device for Imaging Live Zebrafish Larvae

“…Statistical analyses for body and regenerate length were performed in R (www.r-project.org) comparing between experimental conditions using analysis of variance with Tukey method for multiple comparisons. 25 Graphs and survival analysis (logrank test) were performed using Prism 6 software (GraphPad). For all tests, p-value <0.05 was considered statistically significant.…”