目的： 观察IL-12与IL-18联合刺激对树突状细胞(dendritic cell, DC)分泌的exosome （DC derived exosome,Dex）活性的影响，为探索高效的exosome肿瘤疫苗奠定基础。 方法: 取正常健康人外周血单个核细胞诱导培养DCs，分别以IL-12、IL-18 或IL-12+IL-18联合刺激DC，并设空白对照组、T细胞对照组，分别提取各组的Dex， Western blotting检测Dex中HLA-DR、CD83的表达,流式细胞仪检测CD54、CD80及CD86的表达；MTT法检测Dex刺激T细胞增殖的作用， ELISA法测定T细胞IFN-γ的分泌量。 结果: IL-12、IL-18、联合组、空白组的Dex中均有HLA-DR、CD83蛋白表达；联合组Dex的CD54(323.67±44.06 vs 246.17±31.91、236.33±33.87、167.67±28.73, P<0.05)、CD80（406.37±39.18 vs 331.67±36.15、33567±41.38、 260.00± 35.58, P<005)及CD86（390.50±38.06 vs 314.33±36.64、319.00±33.10、246.83±30.55, P<005)表达均高于IL-12组、IL-18组及空白组；联合组刺激T细胞增殖高于IL-12组、IL-18组及空白组、T细胞组（1.98±0.31 vs 1.55±0.23、157±021、1.10±0.18、0.53±0.09， P<0.05)；联合组刺激T细胞分泌IFN-γ水平高于IL-12组、IL-18组及空白组、T细胞组（43667±61.80 vs 29504±40.25、358.18±55.77、225.00±3644、139.50±17.63, P<0.05)。 结论: IL-12与IL-18联合刺激能增加Dex表达CD54、CD80及CD86，促进Dex刺激的T细胞的增殖及其IFN-γ的分泌。

Objective : To observe the effect of stimulation with combined IL-12 and IL-18 on exosomes secreted by dendritic cells （DCs）, and so as to facilitate exploring high-efficiency DC derived exosome(Dex) cancer vaccines. Methods: DCs induced from human peripheral blood mononuclear cells (PBMCs) were stimulated with IL-12+IL-18，IL-12 or IL-18 for the combined group，IL-12 group and IL-18 group respectively; and no stimulant for the control group. Dex was extracted from DCs. The expressions of HLA-DR and CD83 in Dex were detected by Western blotting. CD54, CD80 and CD86 expressions were determined by flow cytometry. The T cell proliferation stimulated by Dex was detected with MTT method. IFN-γ in the T cell supernatant was detected by ELISA. Results: HLA-DR and CD83 proteins were expressed in Dex of all the four groups. The molecule expressions for combined group were CD54( 32367± 44.06 vs 246.17±31.91, 236.33±33.87, 167.67±28.73, P<0.05), CD80（406.37±39.18 vs 331.67±3615, 335.67±41.38, 260.00±35.58, P<0.05), and CD86（390.50±38.06 vs 314.33±36.64, 319.00± 33.10, 246.83± 30.55, P<0.05), which were higher than those in the IL-12 group, IL-18 group and control group. The T cell proliferation in the combined group （1.98±0.31 vs 1.55±0.23, 1.57±0.21, 1.10±0.18, P<005) was higher than that in the IL-12 group, IL-18 group and control group. IFN-γ secreted by T cells in the combined group （436.67±61.8 vs 295.04±40.25, 358.18±55.77, 225.00±36.44, P<0.05) was higher than that in the IL-12 group, IL-18 group and control group. Conclusion: Stimulation of DCs with combined IL-12 and IL-18 can enhance Dex express CD54, CD80 and CD86, promote Dex-induced T cell proliferation and increase IFN-γ secretion by T cells.

山东省自然科学基金资助项目（No.Y2007C111）