Human dendritic cells transfected with allergen-DNA stimulate specific immunoglobulin G4 but not specific immunoglobulin E production of autologous B cells from atopic individuals in vitro

Abstract

Atopic/allergic diseases are characterized by T helper 2 (Th2)-dominated immune responses resulting in immunoglobulin E (IgE) production. DNA-based immunotherapies have been shown to shift the immune response towards Th1 in animal models. In further studies we showed that human dendritic cells (DC) transfected with allergen-DNA are able to stimulate autologous CD4(+) T cells from atopic individuals to produce Th1 instead of Th2 cytokines and to activate interferon-gamma (IFN-gamma)-producing CD8(+) T cells. The aim of this study was to analyse whether DC transfected with allergen-DNA are also able to influence immunoglobulin production of B cells from atopic donors. For this purpose, human monocyte-derived DC from grass-pollen allergic donors were transfected with an adenovirus encoding the allergen Phleum pratense 1 and cocultured with B cells, autologous CD4(+) T cells, and CD40 ligand-transfected L-cells. B cells receiving help from CD4(+) T cells stimulated with allergen-transfected dendritic cells produced more allergen-specific IgG4 compared to stimulation with allergen protein pulsed DC or medium, while total IgG4 production was not affected. In contrast, specific IgE production was not enhanced by stimulation with allergen-DNA transfected DC compared to medium and inhibited compared to allergen protein-pulsed DC with similar effects on total IgE production in vitro. Allergen-DNA transfected dendritic cells are able to direct the human allergic immune response from Th2-dominance towards Th1 and Tc1 also resulting in decreased IgE and increased IgG4 production.

Figure 1

Proliferation of T cells from allergic donors. (a) CD4⁺ and (b) CD8⁺ T cells were stimulated with autologous allergen (nat Phl p1)-pulsed, allergen-DNA-transfected or control (medium) DC or DC transfected with virus without encoding allergen-DNA (Psi 5). After 5 days, cells were pulsed with 1 µCi of tritiated thymidine. Results are expressed as the mean ± SD from eight atopic donors. + Indicates statistically significant differences (P < 0·04) to control (medium) DC. * Indicates statistically significant differences (P < 0·04) to allergen-protein pulsed DC.

Figure 2

Cytokine production of T cells from allergic donors. (a) CD4⁺ and (b) CD8⁺ T cells were stimulated twice with allergen (nat Phl p1)-pulsed, allergen-DNA transfected or control (medium) DC or DC transfected with virus without encoding allergen-DNA (Psi 5) and cytokines were measured by ELISA. Results are expressed as means ± SD from eight atopic donors (which were the same donors as in Fig. 1). + Indicates statistically significant differences (P < 0·04) to control (medium) DC. * Indicates statistically significant differences (P < 0·04) to allergen-protein pulsed DC.

Figure 3

Immunoglobulin G4 production of B cells from allergic donors (a) total and (b) specific IgG4. B cells and T cells were stimulated with autologous allergen (nat Phl p1)-pulsed, allergen-DNA-transfected or control (medium and Psi 5) DC, CD40L-transfected L cells and 1000 U/ml IL-4 for 12 days and immunoglobulin production was measured by ELISA. Results are expressed as means ± SD from 14 atopic donors. + Indicates statistically significant differences (P < 0·04) to control (medium) DC. * Indicates statistically significant differences (P < 0·04) to allergen (nat Phl p1)-pulsed DC.

Figure 4

Immunoglobulin E production of B cells from allergic donors (a) total and (b) specific IgE. B cells and T cells were stimulated with autologous allergen (nat Phl p1)-pulsed, allergen-DNA-transfected or control (medium and Psi 5) DC, CD40L-transfected L cells and 1000 U/ml IL-4 for 12 days and immunoglobulin production was measured by ELISA. Results are expressed as means ± SD from 14 atopic donors. The donors were the same as in Fig. 3 and included the eight donors from Figs 1 and 2. + Indicates statistically significant differences (P < 0·04) to control (medium) DC. * Indicates statistically significant differences (P < 0·04) to allergen (nat Phl p1)-pulsed DC.