Unknown,Transcriptomics,Genomics,Proteomics

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CD8+ T cell gene expression after different antigen dose stimulations

ABSTRACT: This study aimed to identify the human T cell response toward different antigen doses. Fresh isolated CD8+ T cells from peripheral blood mononuclear cells (PBMCs) were stimulated with influenza M1 peptide-loaded autologous monocyte-derived dendritic cells for 2 weeks. A high M1 peptide antigen dose (10uM on moDC, HD1-5) and an optimal antigen dose (10nM on moDC OD1-5) were used. M1-specific CD8 T cells were tetramer sorted at the end of the 2 weeks. RNA was extracted and analyzed for gene expression using Agilent 8X60K gene expression microarrays. 5 independent samples from 3 different HLA A2+ blood donors.

ORGANISM(S): Homo sapiens

SUBMITTER: Yen-Ling Chiu

PROVIDER: E-GEOD-39761 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Introduction: Expansion of antigen (Ag)-specific natural occurring regulatory T cells (nTregs) is required to obtain sufficient numbers of cells for cellular immunotherapy. In this study, different allogeneic stimuli were studied for their capacity to generate functional alloAg-specific nTregs. Methods: A highly enriched nTreg-fraction (CD4+CD25brightCD127- T cells) was alloAg-specific expanded using HLA-mismatched immature, mature monocyte-derived dendritic cells (moDC) or peripheral blood mononuclear cells (PBMC). The allogeneic mature moDC-expanded nTregs were fully characterized by analysis of the demethylation status within the TSDR of the FOXP3 gene and the expression of both protein and mRNA of FOXP3, HELIOS, CTLA4 and cytokines. In addition, the antigen-specific suppressive capacity of these expanded nTregs was tested. Results: Allogeneic mature moDC and skin-derived DC were superior in inducing nTreg-expansion compared to immature moDC or PBMC in an HLA-DR and CD80/CD86-dependent way. Remarkably, the presence of exogenous IL-15 without IL-2, could facilitate optimal mature moDC-induced nTreg-expansion. Allogeneic mature moDC-expanded nTregs were at low ratios (<1:320), potent suppressors of alloAg-induced proliferation without significant suppression of completely HLA-mismatched-Ag-induced proliferation. Mature moDC-expanded nTregs were highly demethylated at the TSDR within the FOXP3 gene and highly expressed of FOXP3, HELIOS and CTLA4. A minority of the expanded nTregs produced IL-10, IL-2, IFN-g and TNF-a but very few IL-17 producing nTregs were found. Next generation sequencing of mRNA of moDC-expanded nTregs revealed a strong induction of Treg-associated mRNAs. Conclusions: Human allogeneic mature moDC are highly efficient stimulator cells, in presence of exogenous IL-15, for expansion of stable alloAg-specific nTregs with superior suppressive function. Four different batches of highly pure regulatory T cells (all from the same donor) were expanded in two different ways, and compared to non-expanded samples.

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CD8+ T cell gene expression after different antigen dose stimulations

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