Supplementary Material for: Differential T-Helper Cell Polarization after Allergen-Specific Stimulation of Autologous Dendritic Cells in Polysensitized Allergic Patients

<b><i>Background:</i></b> Dendritic cells (DCs) play an important role in the induction and regulation of adaptive immune responses by polarizing T-helper (Th) cells. In allergic disease this response is dominated by Th2 cells. It is still unclear whether the activation of Th cells by DCs in atopic individuals is allergen specific. <b><i>Methods:</i></b> Monocyte-derived DCs (MoDCs) obtained from polysensitized patients were stimulated with purified Bet v 1, Phl p 5 and Act d 10, and the surface marker expression was analysed. Proliferation and cytokine profiles of autologous naïve CD4+ T cells co-cultured with allergen-pulsed MoDCs were assessed. <b><i>Results:</i></b> The addition of either Bet v 1 or Phl p 5 did not further increase the expression of surface markers from matured MoDCs in all study groups. In co-cultures, autologous naïve CD4+ T cells proliferated when DCs obtained from individuals allergic to birch and grass pollen were stimulated with Bet v 1 and Phl p 5, respectively. In the co-culture supernatants, significantly increased levels of IL-5 and IL-13 were detected. This effect correlated with the sensitization background and was absent when applying an unspecific allergen, Act d 10. The levels of IL-10 in supernatants of MoDCs and the levels of IL-10 and IFN-γ in supernatants of T cells remained unchanged upon stimulation with allergens. <b><i>Conclusions:</i></b> In this study we observed that allergen-specific stimulation of MoDCs induces T-cell proliferation and upregulation of Th2-type cytokines. Interestingly, this Th2 polarization was only observed in cells stimulated with the allergen to which the patients were sensitized.

**背景：** 树突状细胞（Dendritic cells, DCs）通过极化辅助性T（T-helper, Th）细胞，在适应性免疫应答的诱导与调控中发挥关键作用。在变应性疾病中，此类免疫应答以Th2细胞为主导。目前尚不明确特应性个体体内DCs对Th细胞的激活是否具有变应原特异性。**方法：** 本研究从多致敏患者体内分离得到单核细胞衍生树突状细胞（Monocyte-derived DCs, MoDCs），分别以纯化的Bet v 1、Phl p 5及Act d 10进行刺激，并对其表面标志物表达情况进行分析。同时评估经变应原脉冲处理的MoDCs与自体初始CD4+ T细胞共培养时，后者的增殖情况与细胞因子谱。**结果：** 在所有研究组中，单独添加Bet v 1或Phl p 5均未进一步提升成熟MoDCs的表面标志物表达水平。共培养实验中，当分别用Bet v 1和Phl p 5刺激取自桦树花粉过敏者与青草花粉过敏者的DCs时，自体初始CD4+ T细胞出现增殖。共培养上清液中可检测到IL-5与IL-13水平显著升高。该效应与患者的致敏背景相关，而使用非特异性变应原Act d 10时则无此效应。经变应原刺激后，MoDCs上清液中的IL-10水平以及T细胞上清液中的IL-10与IFN-γ水平均未发生明显变化。**结论：** 本研究发现，对MoDCs进行变应原特异性刺激可诱导T细胞增殖并上调Th2型细胞因子表达。值得注意的是，这种Th2极化效应仅在使用患者致敏的变应原刺激的细胞中得以观测到。