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Abstract：
Eimeria maxima possesses integral families of immunogenic constituents that promote differentiation of immune cells during host-parasite interactions. Dendritic cells (DCs) have an irreplaceable role in the modulation of the host immunity. However, the selection of superlative antigen with immune stimulatory efficacies on host DCs is lacking. In this study, 5 recombinant proteins of E. maxima (Em), including Em14-3-3, rhomboid family domain containing proteins (ROM) EmROM1 and EmROM2, microneme protein 2 (EmMIC2), and Em8 were identified to stimulate chicken splenic derived DCs in vitro. The cultured populations were incubated with recombinant proteins, and typical morphologies of stimulated DCs were obtained. DC-associated markers major histocompatibility complex class II, CD86, CD11c, and CD1.1, showed upregulatory expressions by flow cytometry assay. Immunofluorescence assay revealed that recombinant proteins could bind with the surface of chicken splenic derived DCs. Moreover, quantitative real-time PCR results showed that distinct gene expressions of Toll-like receptors and Wnt signaling pathway were upregulated after the coincubation of recombinant proteins with DCs. The ELISA results indicated that the DCs produced a significant higher level of interleukin (IL)-12 and interferon-γ secretions after incubation with recombinant proteins. While transforming growth factor-β was significantly increased with rEmROM1, rEmROM2, and rEmMIC2 as compared to control groups, and IL-10 did not show significant alteration. Taken together, these results concluded that among 5 potential recombinant antigens, rEm14-3-3 could promote immunogenic functions of chicken splenic derived DCs more efficiently, which might represent an effective molecule for inducing the host Th1-mediated immune response against Eimeria infection.
摘要：
最大艾美球虫具有免疫原性成分的完整家族，可在宿主-寄生虫相互作用期间促进免疫细胞的分化。树突状细胞(DC)在调节宿主免疫中具有不可替代的作用。然而,缺乏对宿主DC具有免疫刺激作用的最高级抗原的选择。在这项研究中,5个E.maxima（Em）的重组蛋白，包括Em14-3-3，含有菱形家族域的蛋白质（ROM）EmROM1和EmROM2，微丝蛋白2（EmMIC2），和Em8被鉴定为体外刺激鸡脾来源的DC。培养的群体与重组蛋白一起孵育，并获得了经刺激的DCs的典型形态。DC相关标志物主要组织相容性复合体II类，CD86,CD11c,和CD1.1，通过流式细胞术检测显示表达上调。免疫荧光分析表明，重组蛋白可以与鸡脾来源的DC表面结合。此外，实时定量PCR结果显示,重组蛋白与DCs共孵育后,Toll样受体和Wnt信号通路的明显基因表达上调。ELISA结果表明，与重组蛋白孵育后，DC产生的白细胞介素（IL）-12和干扰素-γ分泌水平显着升高。与对照组相比，rEmROM1，rEmROM2和rEmMIC2的转化生长因子-β显着增加，IL-10无明显改变。一起来看,这些结果得出结论，在5种潜在的重组抗原中，rEm14-3-3能更有效地促进鸡脾来源DC的免疫原性功能,这可能是诱导宿主Th1介导的针对艾美球虫感染的免疫应答的有效分子。
