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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±0.31vs 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 combined stimulation of IL-12 and IL-18 on the activity of exosomes (DCs) excreted by dendritic cells (DCs), so as to lay the foundation for the exploration of efficient exosome tumor vaccines. Methods: DCs were induced from normal healthy human peripheral blood mononuclear cells. DCs were stimulated with IL-12, IL-18 or IL-12 + IL-18 respectively, and blank control group and T cell control group were established. Dex and Western blotting were used to detect the expression of HLA-DR and CD83 in Dex and the expression of CD54, CD80 and CD86 by flow cytometry. The proliferation of T cells was detected by MTT assay. Secretion. Results: The expression of HLA-DR and CD83 in Dex of IL-12, IL-18, combination group and blank group were significantly higher than those in Dex group (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) IL-18 group and blank group were higher than those in IL-12 group, IL-18 group and blank group (P <0.01) 1.57 ± 0.21,1.10 ± 0.18,0.53 ± 0.09, P <0.05). The level of IFN-γ secreted by T cells in combination group was higher than that in IL-12 group, IL-18 group and blank group (436.67 ± 61.80 vs 295.04 ± 40.25, 358.18 ± 55.77, 225.00 ± 36.44, 139.50 ± 17.63, P <0.05). Conclusion: The combined stimulation of IL-12 and IL-18 can increase Dex expression of CD54, CD80 and CD86, and promote Dex-stimulated T cell proliferation and IFN-γ secretion.