Unknown,Transcriptomics,Genomics,Proteomics

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Molecular and functional characterization of alloantigen-specific anergic T-cell suitable for cell therapy

ABSTRACT: IL-10 is an immunomodulant cytokine that plays a central role in controlling inflammation, down-regulating immune responses, and inducing immunological tolerance. IL-10 inhibits the expression of costimulatory molecules and the secretion of inflammatory cytokines by antigen-presenting cells (APC), directly suppresses T-cell proliferation, and promotes T-cell anergy. In this study we demonstrate that IL-10 inhibits primary allogeneic proliferation in total PBMC and induces antigen(Ag)-specific T-cell hyporesponsiveness. IL-10-induced anergy is TGF-beta independent, is associated with a decrease of Ag-specific CTLp frequency, and can be obtained among cells with different degree of HLA disparity. The use of tolerogenic dendritic cells (DC) differentiated in the presence of IL-10 (DC-10) allows a more consistent anergy induction, even in the context of HLA matched donors. Importantly, alloAg-IL-10/DC-10-anergized T cells preserve their ability to respond to nominal and third party Ags. Microarray experiments were conducted on different mixed lymphocyte cultures aiming at the identification of gene expression profiles characteristic for the various T cell populations. Independently from the APC used to anergized T cells, the resulting T cell populations exhibit a specific gene signature. Based on the results of this study, we developed a method suitable for GMP scale up and for the generation of a population of anergic Ag-specific T cells for cellular therapy. The dataset comprises 12 samples divided into four sample groups each representing a certain kind of allogeneic mixed lymphocyte culture, i.e. peripheral blood mononuclear cells (PBMC) mixed with CD3-depleted cells (MEC, monocyte-enriched cells) with or without IL-10 treatment, and PBMC mixed with mature or IL-10 treated dendritic cells (DCs). Each group includes three biological replicates with cells collected from different donors.

ORGANISM(S): Homo sapiens

SUBMITTER: Stefan Tomiuk

PROVIDER: E-GEOD-17493 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Lactobacilli are probiotics that, among other health promoting effects, have been ascribed immunostimulating and virus preventive properties. Certain lactobacilli species have been shown to possess strong IL-12 inducing properties. As IL-12 production depends on the up-regulation of type I interferons, we hypothesized that the strong IL-12 inducing capacity of L. acidophilus NCFM in murine bone marrow derived DC is caused by an up-regulation of IFN-Î², which subsequently stimulates the induction of IL-12 and the dsRNA binding toll like receptor (TLR)-3. The expression of the genes encoding IFN-Î², IL-12, IL-10 and TLR-3 in DC upon stimulation with L. acidophilus NCFM was measured. L. acidophilus NCFM induced a much stronger expression of ifn-Î², il-12 and il-10 compared to the synthetic dsRNA ligand Poly I:C, whereas the levels of expressed tlr-3 were similar. By the use of whole genome microarray gene expression, we investigated whether other genes related to the viral defence were up-regulated in DC upon stimulation with L. acidophilus NCFM and found that various virus defence related genes, both early and late, were among the strongest up-regulated genes. The IFN-Î² stimulating capability was also detected in another L. acidophilus strain, but was not a property of other probiotic bacteria tested (B. bifidum and E. coli nissle).The IFN-Î² inducing capacity was markedly reduced in TLR-2 -/- DCs, dependent on endocytosis and the major cause of the induction of il-12 and tlr-3 in L. acidophilus NCFM stimulated cells. Collectively, our results reveal that certain lactobacilli trigger the expression of viral defence genes in DC in a TLR-2 manner through induction of IFN- Î². Experiment Overall Design: In the experiment Lactobacillus NCFM were added to murine dendritic cells and stimulated for 4, 10 or 18 hours. These were compared to control experiment at the same timepoints. Experiments were run in triplicates except for control 10h and control 18h which were only in duplicate, giving a total of 16 arrays.

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Molecular and functional characterization of alloantigen-specific anergic T-cell suitable for cell therapy

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