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.

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Listeria monocytogenes (LM) is a Gram-positive, intracellular bacterium that can induce spontaneous abortion, septicemia, and meningitis. Although it is known that neutrophils are required for elimination of the bacteria and for survival of the host, the mechanisms governing the recruitment of neutrophils to LM-infected tissues are not fully understood. We demonstrate here that IL-23 and the IL-17 receptor A (IL-17RA), which mediates both IL-17A and IL-17F signaling, are necessary for resistance against systemic LM infection. LM-infected IL-23p19 knockout (KO) mice have decreased production of IL-17A and IL-17F, while IFN-γ production is not altered by the lack of IL-23. LM induces the production of IL-17A from γδ T cells, but not CD4, CD8, or NK cells. Furthermore, a lack of efficient neutrophil recruitment to the liver is evident in both IL-23p19 KO and IL-17RA KO mice during LM infection. Immunocytochemical analysis of infected livers revealed that neutrophils were able to localize with LM in IL-23p19 KO and IL-17RA KO mice, indicating that IL-23 and IL-17RA do not regulate the precise localization of neutrophils with LM. The importance of IL-23-induced IL-17A was demonstrated by injecting IL-23p19 KO mice with recombinant IL-17A. These mice had reduced LM bacterial burdens compared with IL-23p19 KO mice that did not receive IL-17A. These results indicate that during LM infection, IL-23 regulates the production of IL-17A and IL-17F from γδ T cells, resulting in optimal liver neutrophil recruitment and enhanced bacterial clearance.

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Brief-moderate shock (3, 0.75 s, 1.0 mA) has opposite effects on different measures of pain, inducing antinociception on the tail-flick test while lowering vocalization thresholds to shock and heat (hyperalgesia) and enhancing fear conditioned by a gridshock unconditioned stimulus (US). This study examined the generality of shock-induced hyperalgesia under a range of conditions and explored parallels to sensitized startle. Reduced vocalization thresholds to shock and antinociception emerged at a similar shock intensity. Severe shocks (3, 25 s, 1.0 mA or 3, 2 s, 3.0 mA) lowered vocalization threshold to shock but increased vocalization and motor thresholds to heat and undermined fear conditioned by a gridshock or a startling tone US. All shock schedules facilitated startle, but only brief-moderate shock inflated fear conditioning. The findings suggest that brief-moderate shock enhances the affective impact of aversive stimuli, whereas severe shocks attenuate pain.

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.

Most studies investigating the function of IL-23 have concluded that it promotes IL-17-secreting T cells. Although some reports have also characterized IL-23 as having redundant pro-inflammatory effects with IL-12, we have instead found that IL-23 antagonizes IL-12-induced secretion of IFN-c. When splenocytes or purified populations of T cells were cultured with IL-23, IFN-c secretion in response to IL-12 was dramatically reduced. The impact of IL-23 was most prominent in CD8 1 T cells, but was also observed in NK and CD81 T cells in IL-23p19-deficient mice as compared with WT mice. This increase in IFN-c production coincided with increased LM clearance at days 2 and 3 post-infection. Our data suggest that IL-23 may be a key factor in determining the responsiveness of lymphocytes to IL-12 and their subsequent secretion of IFN-c.Key words: CD8 1 T cells . Cytokines . Innate immunity . Listeria monocytogenes Introduction IL-23 is an IL-12 family cytokine which is composed of the IL-12p40 subunit and a novel cytokine subunit, p19. The heterodimeric structure of IL-23 binds to a receptor complex containing the IL-12Rb1 and a novel receptor termed IL-23R [1]. Similar to IL-12, IL-23 is secreted by activated macrophages and dendritic cells [2] in response to gram-positive and -negative bacterial, viral, and fungal infections, as well as multiple other stimuli [3][4][5][6][7][8][9][10][11]. The majority of the research investigating the actions of IL-23 has focused on its role in maintaining populations of IL-17-secreting cells as well as the possibility of directly inducing IL-17 and IL-22 secretion from a variety of cell types (for reviews see [2,12]). Initially, however, IL-23 was found to have some overlapping functions with IL-12. In a previous report, stimulation of human CD4 1 T cells with anti-CD3 and anti-CD28 resulted in IFN-g secretion, which could be significantly enhanced by IL-12 and slightly enhanced by . Contrary to this, using the murine system, several studies have suggested that IL-23 does not induce production of IFN-g in CD4 1 T cells [13,14]. In this article, we also provide evidence that murine CD4 1 and CD8 1T cells do not respond to IL-23 by secreting IFN-g. IL-23 has been shown to play a role in infectious diseases, autoimmunity and cancer. Mice deficient in IL-23 show increased susceptibility to Mycobacterium tuberculosis, Klebsiella pneumoniae, Citrobacter rodentium, Toxoplasma gondii, Salmonella enterica, and Cryptococcus neoformans [15][16][17][18][19][20][21][22][23]. Increased susceptibility to these infections was linked to the ability of IL-23 to regulate the production of IL-17 and IL-22, although during certain infections, IL-23 does not regulate both cytokines. Figure 1. IL-12, but not IL-23, induces the differentiation of IFN-g secreting T cells. Purified CD8 1 (A, B, D, and E) or CD4 1 (C and F) T cells from naïve WT B6 mice were cultured 6 days on plates coated with anti-CD3 and soluble anti-CD28 in IL-2-supplemented medium with 5 ng/mL IL-12, 10 ng/ mL IL-23, or no additional ...

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