A major focus of our current research is the analysis of innate recognition pathways involved in the response to mycobacteria. This topic is of importance not only for identifying host resistance mechanisms to pathogenic species such as Mycobacterium tuberculosis (MTb) but also for understanding the potent adjuvant effects of attenuated and dead mycobacteria in promoting immune responses to proteins, unrelated pathogens and tumors. In work reported in previous years, we and others have demonstrated that host resistance to Mtb is critically dependent on both IL-1beta and IL-1 alpha and identified specific myeloid cell sources of these cytokines in the lungs of infected mice. Importantly, we demonstrated that interferons play important roles in fine-tuning IL-1 production by these cell populations in vivo with type I IFNs exhibiting potent inhibitory effects. In research completed during the report period we identified one mechanism of type I interferon mediated suppression of IL-1 that involves the induction of autocrine IL-10. We found that IFN-beta potently induced IL-10 during Mtb infection in vitro, which contributed significantly to the suppression of IL-1 by the former cytokine. In additional studies we investigated the mechanism by which IL-1 mediates bacterial control. In in vitro experiments with bone marrow-derived macrophages lacking IL-1R, we found that autocrine IL-1 signals inhibit bacterial replication as measured by colony forming units as well as by means of a novel assay utilizing fluorescent Mtb to examine cell-free vs intracellular Mtb growth. These experiments suggested that, IL-1 mediated signals are instrumental in preventing extracellular bacterial replication, either through the regulation of host cell death or the release of host factors that impact on mycobacterial growth As introduced in previous reports, we have also been studying the role of innate recognition receptors in the T cell immunostimulatory activity of the mycobacteria in Complete Freund's Adjuvant (CFA). Mice immunized with OVA in CFA develop strong OVA-specific Th17 responses and we have previously shown that this cytokine polarization is dependent on IL-1beta/IL-1R signaling through MyD88 and inflammasome dependent processing of pro-IL-1beta. We have further shown that the induction of IL-1beta occurs by a CARD9 dependent pathway and that triggering of the C type lectin receptor Mincle by mycobacterial cord factor is an important component of this response. In addition our work has implicated mycobacterial peptidoglycan as the key stimulus for inflammasome activation. To better understand the role of innate immune pathways in polarizing antigen-specific T cells after immunization with complete Freund's adjuvant (CFA), we have established a system for measuring endogenous antigen-specific CD4+ T cells using an MHC class II tetramer to enumerate lymphocytes recognizing a peptide antigen derived from T.gondii. Using this tetramer, we have been able to compare the effect of IL-1R/MyD88 and Mincle/CARD9-dependent signaling on endogenous versus adoptively transferred TCR transgenic helper T cell expansion and differentiation. We found that the TCR transgenic system minimizes the effects of these innate pathways on CD4+ T cell expansion compared to what is observed with the endogenous response, probably as a result of the higher precursor frequency of the adoptively transferred transgenic cells. Analysis of these differences in CD4+ T cell expansion helped us to resolve discrepancies in the frequencies of IFN gamma-producing cells in MyD88 KO and IL-1R KO mice between the TCR transgenic and endogenous CD4+ T cell responses after CFA immunization. We have also begun to compare CD4+ T cell responses observed after immunization with CFA to those observed after immunization with incomplete Freund's adjuvant (IFA), which is mineral oil and surfactant without the additional heat-killed mycobacteria found in CFA. We have found that IFA is able to robustly prime antigen-specific CD4+ T cells and is comparable to CFA in its ability to induce follicular helper CD4+ T celll polarization, but is unable to induce Th17 polarization despite triggering modest Th1 differentiation. Also, in agreement with previous studies, we found that IFA immunization triggers robust Ab responses, consistent with the induction of Tfh cells. Thus, although the innate immune pathways involved have not yet been identified, mineral oil must be providing important signals that drive T and B cell responses and it will be of interest to determine whether similar signals are delivered by other oil based adjuvants.

Project Start
Project End
Budget Start
Budget End
Support Year
5
Fiscal Year
2012
Total Cost
$535,054
Indirect Cost
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Riteau, Nicolas; Radtke, Andrea J; Shenderov, Kevin et al. (2016) Water-in-Oil-Only Adjuvants Selectively Promote T Follicular Helper Cell Polarization through a Type I IFN and IL-6-Dependent Pathway. J Immunol 197:3884-3893
Arbore, Giuseppina; West, Erin E; Spolski, Rosanne et al. (2016) T helper 1 immunity requires complement-driven NLRP3 inflammasome activity in CD4⁺ T cells. Science 352:aad1210
Riteau, Nicolas; Sher, Alan (2016) Chitosan: An Adjuvant with an Unanticipated STING. Immunity 44:522-4
McNab, Finlay; Mayer-Barber, Katrin; Sher, Alan et al. (2015) Type I interferons in infectious disease. Nat Rev Immunol 15:87-103
Mayer-Barber, Katrin D; Sher, Alan (2015) Cytokine and lipid mediator networks in tuberculosis. Immunol Rev 264:264-75
Cardone, Marco; Dzutsev, Amiran K; Li, Hongchuan et al. (2014) Interleukin-1 and interferon-γ orchestrate β-glucan-activated human dendritic cell programming via IκB-ζ modulation. PLoS One 9:e114516
Shenderov, Kevin; Riteau, Nicolas; Yip, Ronald et al. (2014) Cutting edge: Endoplasmic reticulum stress licenses macrophages to produce mature IL-1β in response to TLR4 stimulation through a caspase-8- and TRIF-dependent pathway. J Immunol 192:2029-33
Redford, Paul S; Mayer-Barber, Katrin D; McNab, Finlay W et al. (2014) Influenza A virus impairs control of Mycobacterium tuberculosis coinfection through a type I interferon receptor-dependent pathway. J Infect Dis 209:270-4
Mayer-Barber, Katrin D; Andrade, Bruno B; Oland, Sandra D et al. (2014) Host-directed therapy of tuberculosis based on interleukin-1 and type I interferon crosstalk. Nature 511:99-103
Shenderov, Kevin; Barber, Daniel L; Mayer-Barber, Katrin D et al. (2013) Cord factor and peptidoglycan recapitulate the Th17-promoting adjuvant activity of mycobacteria through mincle/CARD9 signaling and the inflammasome. J Immunol 190:5722-30

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