CD38: A NAADP degrading enzyme

AbstractThe role of the multifunctional enzyme CD38 in formation of the Ca2+-mobilizing second messenger nicotinic acid adenine dinucleotide phosphate (NAADP) was investigated. Gene silencing of CD38 did neither inhibit NAADP synthesis in intact Jurkat T cells nor in thymus or spleen obtained from CD38 knock out mice. In vitro, both NAADP formation by base-exchange and degradation to 2-phospho adenosine diphosphoribose were efficiently decreased. Thus in vivo CD38 appears to be a NAADP degrading rather than a NAADP forming enzyme, perhaps avoiding desensitizing NAADP levels in intact cells

Regional IFNγ expression is insufficient for efficacious control of food-borne bacterial pathogens at the gut epithelial barrier

IFNγ is critical for host defence against various food-borne pathogens including Salmonella enterica and Listeria monocytogenes, the causative agents of salmonellosis and listeriosis, respectively. We investigated the impact of regional IFNγ expression at the intestinal epithelial barrier on host invasion by salmonellae and listeriae following oral challenge. Transgenic mice (IFNγ-gut), generated on an IFNγ knock-out (KO) background, selectively expressed IFNγ in the gut driven by the modified liver fatty acid-binding protein (Fabpl4× at −132) promoter. Infections with attenuated S. enterica Typhimurium or with L. monocytogenes did not differ significantly in IFNγ-KO, IFNγ-gut and wild-type mice. Further, Listeria-specific CD4+ and CD8+ T cells were not altered in IFNγ-gut mice. Thus, this model indicates that local IFNγ expression by non-immunological cells in the distal part of the small intestine, caecum and colon is insufficient for prevention of gut penetration by S. enterica Typhimurium and L. monocytogene

Role of 4-1BB Receptor in the Control Played by CD8+ T Cells on IFN-γ Production by Mycobacterium tuberculosis Antigen-Specific CD4+ T Cells

BACKGROUND: Antigen-specific IFN-gamma producing CD4(+) T cells are the main mediators of protection against Mycobacterium tuberculosis infection both under natural conditions and following vaccination. However these cells are responsible for lung damage and poor vaccine efficacy when not tightly controlled. Discovering new tools to control nonprotective antigen-specific IFN-gamma production without affecting protective IFN-gamma is a challenge in tuberculosis research. METHODS AND FINDINGS: Immunization with DNA encoding Ag85B, a candidate vaccine antigen of Mycobacterium tuberculosis, elicited in mice a low but protective CD4(+) T cell-mediated IFN-gamma response, while in mice primed with DNA and boosted with Ag85B protein a massive increase in IFN-gamma response was associated with loss of protection. Both protective and non-protective Ag85B-immunization generated antigen-specific CD8(+) T cells which suppressed IFN-gamma-secreting CD4(+) T cells. However, ex vivo ligation of 4-1BB, a member of TNF-receptor super-family, reduced the massive, non-protective IFN-gamma responses by CD4(+) T cells in protein-boosted mice without affecting the low protective IFN-gamma-secretion in mice immunized with DNA. This selective inhibition was due to the induction of 4-1BB exclusively on CD8(+) T cells of DNA-primed and protein-boosted mice following Ag85B protein stimulation. The 4-1BB-mediated IFN-gamma inhibition did not require soluble IL-10, TGF-beta, XCL-1 and MIP-1beta. In vivo Ag85B stimulation induced 4-1BB expression on CD8(+) T cells and in vivo 4-1BB ligation reduced the activation, IFN-gamma production and expansion of Ag85B-specific CD4(+) T cells of DNA-primed and protein-boosted mice. CONCLUSION/SIGNIFICANCE: Antigen-specific suppressor CD8(+) T cells are elicited through immunization with the mycobacterial antigen Ag85B. Ligation of 4-1BB receptor further enhanced their suppressive activity on IFN-gamma-secreting CD4(+) T cells. The selective expression of 4-1BB only on CD8(+) T cells in mice developing a massive, non-protective IFN-gamma response opens novel strategies for intervention in tuberculosis pathology and vaccination through T-cell co-stimulatory-based molecular targeting

Development of Functional and Molecular Correlates of Vaccine-Induced Protection for a Model Intracellular Pathogen, F. tularensis LVS

In contrast with common human infections for which vaccine efficacy can be evaluated directly in field studies, alternative strategies are needed to evaluate efficacy for slowly developing or sporadic diseases like tularemia. For diseases such as these caused by intracellular bacteria, serological measures of antibodies are generally not predictive. Here, we used vaccines varying in efficacy to explore development of clinically useful correlates of protection for intracellular bacteria, using Francisella tularensis as an experimental model. F. tularensis is an intracellular bacterium classified as Category A bioterrorism agent which causes tularemia. The primary vaccine candidate in the U.S., called Live Vaccine Strain (LVS), has been the subject of ongoing clinical studies; however, safety and efficacy are not well established, and LVS is not licensed by the U.S. FDA. Using a mouse model, we compared the in vivo efficacy of a panel of qualitatively different Francisella vaccine candidates, the in vitro functional activity of immune lymphocytes derived from vaccinated mice, and relative gene expression in immune lymphocytes. Integrated analyses showed that the hierarchy of protection in vivo engendered by qualitatively different vaccines was reflected by the degree of lymphocytes' in vitro activity in controlling the intramacrophage growth of Francisella. Thus, this assay may be a functional correlate. Further, the strength of protection was significantly related to the degree of up-regulation of expression of a panel of genes in cells recovered from the assay. These included IFN-γ, IL-6, IL-12Rβ2, T-bet, SOCS-1, and IL-18bp. Taken together, the results indicate that an in vitro assay that detects control of bacterial growth, and/or a selected panel of mediators, may ultimately be developed to predict the outcome of vaccine efficacy and to complement clinical trials. The overall approach may be applicable to intracellular pathogens in general

Analysis of eight genes modulating interferon gamma and human genetic susceptibility to tuberculosis: a case-control association study

<p>Abstract</p> <p>Background</p> <p>Interferon gamma is a major macrophage-activating cytokine during infection with <it>Mycobacterium tuberculosis</it>, the causative pathogen of tuberculosis, and its role has been well established in animal models and in humans. This cytokine is produced by activated T helper 1 cells, which can best deal with intracellular pathogens such as <it>M. tuberculosis</it>. Based on the hypothesis that genes which regulate interferon gamma may influence tuberculosis susceptibility, we investigated polymorphisms in eight candidate genes.</p> <p>Methods</p> <p>Fifty-four polymorphisms in eight candidate genes were genotyped in over 800 tuberculosis cases and healthy controls in a population-based case-control association study in a South African population. Genotyping methods used included the SNPlex Genotyping System™, capillary electrophoresis of fluorescently labelled PCR products, TaqMan^®SNP genotyping assays or the amplification mutation refraction system. Single polymorphisms as well as haplotypes of the variants were tested for association with TB using statistical analyses.</p> <p>Results</p> <p>A haplotype in interleukin 12B was nominally associated with tuberculosis (p = 0.02), but after permutation testing, done to assess the significance for the entire analysis, this was not globally significant. In addition a novel allele was found for the interleukin 12B D5S2941 microsatellite.</p> <p>Conclusions</p> <p>This study highlights the importance of using larger sample sizes when attempting validation of previously reported genetic associations. Initial studies may be false positives or may propose a stronger genetic effect than subsequently found to be the case.</p

T cell fate and clonality inference from single-cell transcriptomes.

We developed TraCeR, a computational method to reconstruct full-length, paired T cell receptor (TCR) sequences from T lymphocyte single-cell RNA sequence data. TraCeR links T cell specificity with functional response by revealing clonal relationships between cells alongside their transcriptional profiles. We found that T cell clonotypes in a mouse Salmonella infection model span early activated CD4(+) T cells as well as mature effector and memory cells