High Dose Allergen Stimulation of T Cells from House Dust Mite-Allergic Subjects Induces Expansion of IFN-γ+ T Cells, Apoptosis of CD4+IL-4+ T Cells and T Cell Anergy

Abstract

Background: During clinically effective allergen-specific immunotherapy a shift in cytokine dominance from IL-4, IL-5 predominant to IFN-γ predominant has been observed. As antigen concentration influences Th cell priming, this study aimed to determine the effect of different allergen concentrations on human house dust mite (HDM)-specific T cell production of IL-4 and IFN-γ, proliferation and apoptosis. Methods: HDM-allergic donor PBMC were cultured for 14 days with different concentrations of HDM extract (1, 10 and 100 µg/ml). T cell intracellular IL-4 and IFN-γ, division (CFSE labelling) and apoptosis (active caspase-3 staining) were analysed by flow cytometry. Proliferation was assessed by 3H-thymidine incorporation. Results: Increased CD4+IFN-γ+ and CD8+IFN-γ+ T cell numbers were observed in high allergen concentration cultures compared with low concentration cultures whereas there were no differences in CD4+IL-4+ and CD8+IL-4+ T cell numbers. CFSE cell labelling revealed that high allergen concentration favours the expansion of IFN-γ-producing CD4+ T cells. The proportion of apoptotic cells increased with allergen concentration and there was preferential apoptosis of CD4+IL-4+ T cells. HDM-induced proliferation was decreased in high allergen concentration cultures; this was reversible by IL-2 consistent with anergy. Conclusion: These results show that T cell division and apoptosis contribute to the cytokine skewing to predominant IFN-γ production by T cells observed at high allergen concentration. Thus the use of hypoallergenic T cell reactive preparations which can be given safely at higher doses than natural extracts may enhance efficacy of allergen-specific immunotherapy.