We determined whether human NK cells could contribute to immune defenses against Mycobacterium tuberculosis through production of IL-22. CD3−CD56+ NK cells produced IL-22 when exposed to autologous monocytes and γ-irradiated M. tuberculosis, and this depended on the presence of IL-15 and IL-23, but not IL-12 or IL-18. IL-15-stimulated NK cells expressed 10.6 times more DAP10 mRNA compared with control NK cells, and DAP10 siRNA inhibited IL-15-mediated IL-22 production by NK cells. Soluble factors produced by IL-15-activated NK cells inhibited growth of M. tuberculosis in macrophages, and this effect was reversed by anti-IL-22. Addition of rIL-22 to infected macrophages enhanced phagolysosomal fusion and reduced growth of M. tuberculosis. We conclude that NK cells can contribute to immune defenses against M. tuberculosis through production of IL-22, which inhibits intracellular mycobacterial growth by enhancing phagolysosomal fusion. IL-15 and DAP-10 elicit IL-22 production by NK cells in response to M. tuberculosis.

Inflammasomes are important for maintaining intestinal homeostasis, and dysbiosis contributes to the pathology of inflammatory bowel disease (IBD) and increases the risk for colorectal cancer. Inflammasome defects contribute to chronic intestinal inflammation and increase the susceptibility to colitis in mice. However, the inflammasome sensor absent in melanoma 2 (AIM2) protects against colorectal cancer in an inflammasome-independent manner through DNA-dependent protein kinase and Akt pathways. Yet, the roles of the AIM2 inflammasome in IBD and the early phases of colorectal cancer remain ill-defined. Here we show that the AIM2 inflammasome has a protective role in the intestine. During steady state, Aim2 deletion results in the loss of IL-18 secretion, suppression of the IL-22 binding protein (IL-22BP) in intestinal epithelial cells and consequent loss of the STAT3-dependent antimicrobial peptides (AMPs) Reg3β and Reg3γ, which promotes dysbiosis-linked colitis. During dextran sulfate sodium-induced colitis, a dysfunctional IL-18/IL-22BP pathway in Aim2−/− mice promotes excessive IL-22 production and elevated STAT3 activation. Aim2−/− mice further exhibit sustained STAT3 and Akt activation during the resolution of colitis fueled by enhanced Reg3b and Reg3g expression. This self-perpetuating mechanism promotes proliferation of intestinal crypt cells and likely contributes to the recently described increase in susceptibility of Aim2−/− mice to colorectal cancer. Collectively, our results demonstrate a central role for the AIM2 inflammasome in preventing dysbiosis and intestinal inflammation through regulation of the IL-18/IL-22BP/IL-22 and STAT3 pathway and expression of select AMPs.

Summary Previous studies have suggested that neutrophils are required for resistance during infection with multiple pathogenic microorganisms. However, the depleting antibody used in those studies binds to both Ly6G and Ly6C (anti-Gr-1; clone RB6-8C5). This antibody has been shown to not only deplete neutrophils, but also monocytes, and a subset of CD8 T cells. Recently, an antibody against Ly6G has been characterized which specifically depletes neutrophils. In the present study, neutrophils are depleted using the antibody against Ly6G during infection with the intracellular bacterium, Listeria monocytogenes (LM). Our data show that neutrophil depleted mice are much less susceptible to infection than mice depleted with anti-Gr-1. Although neutrophils are required for clearance of LM, their importance is more pronounced in the liver and during a high-dose bacterial challenge. Furthermore, we demonstrate that protection mediated by neutrophils is due to production of TNF-α, but not IFN-γ. Additionally, neutrophils are not required for the recruitment of monocytes or the generation of adaptive T cell responses during LM infection. These studies highlight the importance of neutrophils during LM infection, and also indicate that depletion of neutrophils is less detrimental to the host than depletion of all Gr-1 expressing cell populations.

Lipopolysaccharide (LPS) of Gram-negative bacteria can elicit a strong immune response. Although extracellular LPS is sensed by TLR4 at the cell surface and triggers a transcriptional response, cytosolic LPS binds and activates non-canonical inflammasome caspases, resulting in pyroptotic cell death, as well as canonical NLRP3 inflammasome-dependent cytokine release. Contrary to the highly regulated multiprotein platform required for caspase-1 activation in the canonical inflammasomes, the non-canonical mouse caspase-11 and the orthologous human caspase-4 function simultaneously as innate sensors and effectors, and their regulation is unclear. Here we show that the oxidized phospholipid 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine (oxPAPC) inhibits the non-canonical inflammasome in macrophages, but not in dendritic cells. Aside from a TLR4 antagonistic role, oxPAPC binds directly to caspase-4 and caspase-11, competes with LPS binding, and consequently inhibits LPS-induced pyroptosis, IL-1β release and septic shock. Therefore, oxPAPC and its derivatives might provide a basis for therapies that target non-canonical inflammasomes during Gram-negative bacterial sepsis.

Inflammasomes are protein platforms linking recognition of microbe, pathogen-associated and damage-associated molecular patterns by cytosolic sensory proteins to caspase-1 activation. Caspase-1 promotes pyroptotic cell death and the maturation and secretion of interleukin (IL)-1β and IL-18, which trigger inflammatory responses to clear infections and initiate wound-healing; however, excessive responses cause inflammatory disease. Inflammasome assembly requires the PYRIN domain (PYD)-containing adaptor ASC, and depends on PYD–PYD interactions. Here we show that the PYD-only protein POP2 inhibits inflammasome assembly by binding to ASC and interfering with the recruitment of ASC to upstream sensors, which prevents caspase-1 activation and cytokine release. POP2 also impairs macrophage priming by inhibiting the activation of non-canonical IκB kinase ɛ and IκBα, and consequently protects from excessive inflammation and acute shock in vivo. Our findings advance our understanding of the complex regulatory mechanisms that maintain a balanced inflammatory response and highlight important differences between individual POP members.

Sensing and responding to pathogens and tissue damage is a core mechanism of innate immune host defense, and inflammasomes represent a central cytosolic pattern recognition receptor pathway leading to the generation of the pro-inflammatory cytokines interleukin-1β and interleukin-18 and pyroptotic cell death that causes the subsequent release of danger signals to propagate and perpetuate inflammatory responses. While inflammasome activation is essential for host defense, deregulated inflammasome responses and excessive release of inflammatory cytokines and danger signals are linked to an increasing spectrum of inflammatory diseases. In this review, we will discuss recent developments in elucidating the role of PYRIN domain-only proteins (POPs) and the related CARD-only proteins (COPs) in regulating inflammasome responses and their impact on inflammatory disease.

Reactive oxygen and nitrogen species (ROS/RNS) play important roles during immune responses to bacterial pathogens. Extracellular superoxide dismutase (ecSOD) regulates extracellular concentrations of ROS/RNS and contributes to tissue protection during inflammatory insults. The participation of ecSOD in immune responses seems therefore intuitive, yet is poorly understood. In the present study, we utilized mice with varying levels of ecSOD activity to investigate the involvement of this enzyme in immune responses against Listeria monocytogenes. Surprisingly, our data demonstrate that, despite enhanced neutrophil recruitment to the liver, ecSOD activity negatively impacted host survival and bacterial clearance. Increased ecSOD activity was accompanied by decreased co-localization of neutrophils with bacteria, as well as increased neutrophil apoptosis, which reduced overall and neutrophil-specific TNF-α production. Liver leukocytes from mice lacking ecSOD produced equivalent nitric oxide (NO·) when compared to mice expressing ecSOD. However, during infection, there were higher levels of peroxynitrite (NO3·−) in livers from mice lacking ecSOD compared to mice expressing ecSOD. Neutrophil depletion studies revealed that high levels of ecSOD activity resulted in neutrophils with limited protective capacity, whereas neutrophils from mice lacking ecSOD provided superior protection compared to neutrophils from wild-type mice. Taken together, our data demonstrate that ecSOD activity reduces innate immune responses during bacterial infection and provides a potential target for therapeutic intervention.

Listeria monocytogenes (LM) is a gram-positive bacterium that is a common contaminant of processed meats and dairy products. In humans, ingestion of LM can result in intracellular infection of the spleen and liver, which can ultimately lead to septicemia, meningitis, and spontaneous abortion. Interleukin (IL)-23 is a cytokine that regulates innate and adaptive immune responses by inducing the production of IL-17A, IL-17F, and IL-22. We have recently demonstrated that the IL-23/IL-17 axis is required for optimal recruitment of neutrophils to the liver, but not the spleen, during LM infection. Furthermore, these cytokines are required for the clearance of LM during systemic infection. In other infectious models, IL-22 induces the secretion of anti-microbial peptides and protects tissues from damage by preventing apoptosis. However, the role of IL-22 has not been thoroughly investigated during LM infection. In the present study, we show that LM induces the production of IL-22 in vivo. Interestingly, IL-23 is required for the production of IL-22 during primary, but not secondary, LM infection. Our findings suggest that IL-22 is not required for clearance of LM during primary or secondary infection, using both systemic and mucosal models of infection. IL-22 is also not required for the protection of LM infected spleens and livers from organ damage. Collectively, these data indicate that IL-22 produced during LM infection must play a role other than clearance of LM or protection of tissues from pathogen- or immune-mediated damage.