Dendritic cells (DC) shape immune responses by producing cytokines such as Interleukin‐12 (IL‐12). IL‐12 is important in the clearance of the intracellular bacterial pathogen Listeria monocytogenes (Lm) and activates CD8+ T cells (CTL), which are required for sterilizing immunity. Our initial studies on the role of IL‐12 on CTL function determined that IL‐12p40 signals were transduced primarily during the first 48 hours of priming. However, these studies did not address the individual contributions of IL‐12 or IL‐23 nor how they affect the physical interactions of priming. To address these questions wild‐type, IL‐12p35−/−, or IL‐12p40−/− DC were infected with Lm and then used to prime CTL. IL‐12p40−/− mice immunized with p35−/− DC or p40−/− exhibited a reduced ability to clear Listeria compared to mice immunized with wild‐type DC. We found that CTL primed in vitro by p35−/− or p40−/− DC produced less IFN‐γ with the greatest reduction seen with the p40−/− DC, whereas proliferation was not affected. CTL primed in the absence of IL‐12 engaged in shorter duration interactions with DC which correlated with the absence of the chemokines CCL1 and CCL17. Neutralization of these chemokines resulted in decreased IFN‐γ production by CTL. Thus, CTL priming by DC is influenced more by the activity of IL‐12 than IL‐23 and in part through CCL1 and CCL17 whose production requires IL‐12. 1RO1AI057770‐01A1 and 1F31AI073245‐01A1

Wild-type (wt) vesicular stomatitis virus (VSV) strains stimulate plasmacytoid dendritic cells (pDC) through

Type I IFN is key to the immune response to viral pathogens, however its role in bacterial infections is less well understood. Mice lacking the type I IFN receptor (IFNAR−/−) demonstrate enhanced resistance to infection with Listeria monocytogenes. We have now determined that following infection with Listeria, the composition of innate cells recruited to the peritoneal cavity of IFNAR−/− mice reflects an increase in the frequency of neutrophils and a decrease in monocyte frequency compared to WT controls. These differences in inflammatory infiltrates could not be attributed to distinct bone marrow composition prior to infection or to level of apoptosis. We also observed no differences in neutrophil oxidative burst. However, blocking CXCR2 prevented enhanced neutrophil influx and hampered bacterial clearance. Taken together, these studies highlight a novel mechanism by which type I interferon signaling regulates the immune response to Listeria, through negative regulation of chemokines driving neutrophil recruitment.

To determine the relative contributions of DC subsets in the development of protective immunity to Listeria monocytogenes we examined the relationship between maturation, bacterial burden, and T cell priming capacity of four well characterized subsets of splenic DC following infection with Lm. CD8α + , CD4 + , and CD8α − CD4 − DC and the B220 + plasmacytoid DC (pDC) were compared for abundance and costimulatory molecule expression at 24, 48, and 72h post i.v. infection. We further determined the bacterial burden associated with each DC subset and their relative capacities to prime CD8+ T cells at 24hpi. The CD8α + DC displayed the highest level of maturation, association with live bacteria, and T cell activation potential. Second, the CD4+ DC were also mature, yet were associated with fewer bacteria, and stimulated T cell proliferation, but not IFN-γ production. The CD8α − CD4 − DC showed a modest maturation response and were associated with a high number of bacteria, but failed to induce T cell proliferation ex vivo. pDC displayed a strong maturation response, but were not associated with detectable bacteria and also failed to stimulate T cell activation. Finally, we measured the cytokine responses in these subsets and determined that IL-12 was produced predominantly by the CD8 + DC, correlating with the ability of this subset DC to induce IFN-γ production in T cells. We conclude that Listeria-specific CD8 + T cell activation in the spleen is most effectively achieved by infection-induced maturation of the CD8α + DC subset.

BackgroundAtherosclerosis is a chronic inflammatory disorder whose development is inversely correlated with high‐density lipoprotein concentration. Current therapies involve pharmaceuticals that significantly elevate plasma high‐density lipoprotein cholesterol concentrations. Our studies were conducted to investigate the effects of low‐dose lipid‐free apolipoprotein A‐I (apoA‐I) on chronic inflammation. The aims of these studies were to determine how subcutaneously injected lipid‐free apoA‐I reduces accumulation of lipid and immune cells within the aortic root of hypercholesterolemic mice without sustained elevations in plasma high‐density lipoprotein cholesterol concentrations.Methods and Results Ldlr −/− and Ldlr −/− apoA‐I −/− mice were fed a Western diet for a total of 12 weeks. After 6 weeks, a subset of mice from each group received subcutaneous injections of 200 μg of lipid‐free human apoA‐I 3 times a week, while the other subset received 200 μg of albumin, as a control. Mice treated with lipid‐free apoA‐I showed a decrease in cholesterol deposition and immune cell retention in the aortic root compared with albumin‐treated mice, regardless of genotype. This reduction in atherosclerosis appeared to be directly related to a decrease in the number of CD131 expressing cells and the esterified cholesterol to total cholesterol content in several immune cell compartments. In addition, apoA‐I treatment altered microdomain cholesterol composition that shifted CD131, the common β subunit of the interleukin 3 receptor, from lipid raft to nonraft fractions of the plasma membrane.ConclusionsApoA‐I treatment reduced lipid and immune cell accumulation within the aortic root by systemically reducing microdomain cholesterol content in immune cells. These data suggest that lipid‐free apoA‐I mediates beneficial effects through attenuation of immune cell lipid raft cholesterol content, which affects numerous types of signal transduction pathways that rely on microdomain integrity for assembly and activation.