Flow cytometric analysis of cytokine expression in short-term allergen-stimulated T cells mirrors the phenotype of proliferating T cells in long-term cultures
| Autor: | Martijn C. Nawijn, J. N. G. Oude Elberink, A. J. M. Van Oosterhout, D. Van Hemelen, J. Heimweg, Barbara Bohle |
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| Přispěvatelé: | Groningen Research Institute of Pharmacy, Groningen Research Institute for Asthma and COPD (GRIAC) |
| Rok vydání: | 2011 |
| Předmět: |
Adult
Male BLOOD medicine.medical_treatment T cell Immunology CD40 Ligand Antigen-specific T(H) cells Cell Culture Techniques ANTIGENS CFSE Immunophenotyping Interleukin 21 Antigen medicine otorhinolaryngologic diseases Immunology and Allergy Cytotoxic T cell Humans CD154 HEALTHY Cell Proliferation GRASS-POLLEN Immunoassay CD40 II PEPTIDE TETRAMERS biology food and beverages Rhinitis Allergic Seasonal Grass-pollen allergy T-Lymphocytes Helper-Inducer Allergens Flow Cytometry INDIVIDUALS Cytokine medicine.anatomical_structure Phenotype BALANCE biology.protein Cytokines Pollen Female RESPONSES |
| Zdroj: | Journal of Immunological Methods, 371(1-2), 114-121. ELSEVIER SCIENCE BV |
| ISSN: | 1872-7905 0022-1759 |
| Popis: | Background: Allergen-specific T(H) cells play an important role in IgE-mediated disorders as allergies. Since this T(H) cell-population only accounts for a small percentage of Tv, cells, they are difficult to phenotype without prior selection or expansion.Methods: Grass-pollen-specific T(H) cell profiles were evaluated in 5 allergic and 4 non-allergic individuals using three different approaches: CD154 expression on ex vivo grass-pollen-activated PBMCs (i): CFSE-dilution in grass-pollen-restimulated PBMCs (ii) and T cell lines enriched for allergen-specific T cells (iii).Results: Relatively low numbers of allergen-specific T(H) cells were detected using CD154 expression, limiting the power to detect phenotypic differences between allergic and non-allergic individuals. In contrast, higher frequencies of proliferating T(H) cells were detected by loss-of-CFSE intensity in PBMCs and TCLs after grass-pollen-stimulation, resulting in the detection of significantly more IL-4 producing T(H) cells in allergic vs non-allergic individuals. In addition, higher numbers of IFN gamma producing T(H) cells were detected in long-term cultures compared to the CD154 expressing T(H) cells.Conclusion: To detect allergen-specific T(H) cells for a common allergen as grass-pollen, expansion is not absolutely necessary, although within 8-day grass-pollen cultures, higher numbers of proliferating T(H) cells resulted in increased statistical power to detect phenotypic differences. However, this approach also detects more bystander activated T(H) cells. TCLs resulted in comparable percentages of cytokine expressing T cells as 8-day cultures. Therefore enrichment can be necessary for detection of T(H) cells specific for a single allergen or allergen-derived peptides, but is dispensable for the detection and phenotyping of allergen-specific T(H) cells using crude extracts. (C) 2011 Elsevier B.V. All rights reserved. |
| Databáze: | OpenAIRE |
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