Induction of Bet-specific iTreg from birch-pollen allergic patients with associated hazelnut allergy by autologous IL-10 DC
As previously demonstrated by us, human IL-10 DC induce iTreg with a high capacity to suppress T cell responses in an antigen-specific manner.35,38,42,43 In this study, we wanted to investigate whether these tolerogenic IL-10 DC are able to induce an allergen-specific as well as cross-reactive tolerance in patients suffering from a pollen (birch, Bet v 1) and associated food (hazelnut, Cor a 1) allergy (for patients´ details see Supplementary Table 1). For this purpose, we used our previously established in vitro model with human DC to analyze allergen-specific and cross-reactive T cell responses.12
IL-10 DC obtained from birch pollen allergic patients with associated hazelnut allergy were loaded with Bet or left unloaded (IL-10 DCBet/IL-10 DC0) and cocultured with autologous CD4+ T cells to induce Bet-specific or control iTreg, respectively (iTregBet/iTreg0). In addition, mDC (mDCBet/mDC0) were used to generate Bet-specific and non-specific Teff (TeffBet/Teff0) as controls (see Methods, Supplementary Methods and Supplementary Figure 1).
After primary culture, TeffBet exhibited a significantly increased proliferative capacity compared to Teff0, demonstrating an allergen (Bet)-specific T cell response (Figure 1A). In contrast, stimulation with IL-10 DC0 or IL-10DCBet resulted in a significantly reduced T cell proliferation of both iTreg0 and iTregBet.
In line with the data of allergen-specific T cell proliferation, analysis of cytokine production after primary culture revealed significantly increased levels of TH2 cytokines (IL-5, IL-9 and IL-13) and of IL-2 (T cell activation) in supernatants of mDCBet activated TeffBet compared to control Teff0, demonstrating highly stimulated Bet-specific Teff (Figure 1B-E). In contrast, supernatants of iTreg primed by IL-10 DC0 or IL-10 DCBet, respectively, exhibited reduced amounts of TH2 cytokines in comparison to Bet-specific control Teff, confirming the diminished activity of iTreg after primary culture. However, we observed very high IL-10 concentrations produced by iTregBet compared to control TeffBet/Teff0 as well as to iTreg0, suggesting the immunosuppressive cytokine IL-10 as mediator of allergen-specific iTreg suppressor function (Figure 1F).