title: ZnT8-Specific CD4+ T cells display distinct cytokine expression profiles between type 1 diabetes patients and healthy adults.
abstract: Determination of antigen-specific T cell repertoires in human blood has been a challenge. Here, we show a novel integrated approach that permits determination of multiple parameters of antigen-specific T cell repertoires. The approach consists of two assays: the Direct assay and the Cytokine-driven assay. Briefly, human PBMCs are first stimulated with overlapping peptides encoding a given antigen for 48 hours to measure cytokine secretion (Direct assay). Peptide-reactive T cells are further expanded by IL-2 for 5 days; and after overnight starvation, expanded cells are stimulated with the same peptides from the initial culture to analyze cytokine secretion (Cytokine-driven assay). We first applied this integrated approach to determine the type of islet-antigen-specific T cells in healthy adults. Out of ten donors, the Direct assay identified GAD65-specific CD4(+) T cells in three adults and zinc transporter 8 (ZnT8)-specific CD4(+) T cells in five adults. The intracytoplasmic cytokine staining assay showed that these islet-antigen-specific CD4(+) T cells belonged to the CD45RO(+) memory compartment. The Cytokine-driven assay further revealed that islet-antigen-specific CD4(+) T cells in healthy adults were capable of secreting various types of cytokines including type 1 and type 2 cytokines as well as IL-10. We next applied our integrated assay to determine whether the type of ZnT8-specific CD4(+) T cells is different between Type 1 diabetes patients and age/gender/HLA-matched healthy adults. We found that ZnT8-specific CD4(+) T cells were skewed towards Th1 cells in T1D patients, while Th2 and IL-10-producing cells were prevalent in healthy adults. In conclusion, the Direct assay and the Cytokine-driven assay complement each other, and the combination of the two assays provides information of antigen-specific T cell repertoires on the breadth, type, and avidity. This strategy is applicable to determine the differences in the quality of antigen-specific T cells between health and disease.
authors: Baylor Institute for Immunology Research, Dallas, Texas, USA.
authors: Daisuke
authors: D
authors: Chujo
authors: Emile
authors: E
authors: Foucat
authors: Thien-Son
authors: TS
authors: Nguyen
authors: Damien
authors: D
authors: Chaussabel
authors: Jacques
authors: J
authors: Banchereau
authors: Hideki
authors: H
authors: Ueno
mesh_terms: D000328:Adult; D000945:Antigens, Differentiation, T-Lymphocyte / Q000235:genetics* / Q000276:immunology; D016022:Case-Control Studies; D027682:Cation Transport Proteins / Q000235:genetics* / Q000276:immunology; D049109:Cell Proliferation / Q000187:drug effects; D016207:Cytokines / Q000096:biosynthesis* / Q000276:immunology; D003922:Diabetes Mellitus, Type 1 / Q000235:genetics* / Q000276:immunology / Q000473:pathology; D005260:Female; D015870:Gene Expression / Q000187:drug effects / Q000276:immunology*; D005968:Glutamate Decarboxylase / Q000235:genetics / Q000276:immunology; D006801:Humans; D007118:Immunoassay; D007156:Immunologic Memory / Q000235:genetics / Q000276:immunology; D016130:Immunophenotyping; D007376:Interleukin-2 / Q000494:pharmacology; D008213:Lymphocyte Activation / Q000187:drug effects; D008297:Male; D008875:Middle Aged; D010455:Peptides / Q000276:immunology / Q000494:pharmacology; D018417:Th1 Cells / Q000187:drug effects / Q000276:immunology / Q000473:pathology; D058408:Th1-Th2 Balance / Q000187:drug effects; D018418:Th2 Cells / Q000187:drug effects / Q000276:immunology / Q000473:pathology; D000075703:Zinc Transporter 8
chemical_list: D000945:Antigens, Differentiation, T-Lymphocyte; D027682:Cation Transport Proteins; D016207:Cytokines; D007376:Interleukin-2; D010455:Peptides; C492522:SLC30A8 protein, human; D000075703:Zinc Transporter 8; D005968:Glutamate Decarboxylase; C401141:glutamate decarboxylase 2
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