Allergen-based peptide immunotherapy is associated with functional modifications in allergen-specific T-cell subpopulations

Abstract

Rationale To characterize changes in T-cell responses to allergen following peptide-based immunotherapy (PIT) in cat-allergic subjects. Methods PBMC (0.5×10 6 per 200 μl/well), obtained from cat-allergic volunteers before and after PIT, were labelled with 0.5 μM carboxyfluorescein diacetate-succinimidyl ester (CFSE) and stimulated with or without whole cat dander-allergen extract (30 μg/ml), or immobilised α-CD3/α-CD28 (5 μg/ml each). Culture supernatants were removed at 48 hours and analyzed by cytometric bead array (CBA) for Th1/Th2 cytokines. After 6 days, PBMC were stained with α-CD45RO or α-CD4, and T-cell proliferation analyzed by flow-cytometry. Results Proliferation of memory T cells (CD45RO high; 9 subjects assayed) and also non-CD4 + T cells in the lymphocyte gate (primarily CD8 + T cells; 6 subjects assayed) in response to cat-allergen was markedly reduced after PIT (p=0.004 and p=0.031 respectively). Changes in CD4 + T-cell proliferation (n=6) did not achieve statistical significance. Cytokine secretion profiles showed several distinct patterns: 8/11 subjects assayed showed an increased production of IL-10, in combination with either a decrease (5/8) or an increase (3/8) in IFN-γ. Two subjects showed a decrease in all cytokines detected, while 1 individual showed an increase in IFN-γ and a decrease in IL-10. Conclusion PIT was associated with a significant reduction in the proliferation of allergen-specific memory T cells. Changes in cytokine profiles were compatible with both the induction of regulatory T cells (increased IL-10, reduced proliferation), and a shift towards Th1 immunity (increased IFN-γ). Small-scale assays are useful for simultaneously determining cytokine production, and proliferation of specific cell populations, as defined by cell-surface marker expression.