【摘要】 目的:研究肿瘤细胞分泌Sema3A对小鼠树突状细胞(dendritic cells,DCs)免疫学功能的影响。方法:以Sema3A特异性小干扰RNA片段(Si-Sema)和相应突变片段(Si-mut)分别转染肺腺癌细胞A549,以real-time PCR和Western-blot法检测干扰效果。取未转染和Si-Sema、Si-mut转染细胞的浓缩上清分别作用于DCs,并以流式细胞术分析各组细胞表面MHCⅡ分子及共刺激分子CD40、CD80的表达;酶联免疫吸附法(enzyme linked immunosorbent assay,ELISA)检测DCs表达白细胞介素(interleukin,IL)-12水平。同时,测定DCs刺激OVA-特异性CD4+T(DO11.10T)细胞表达γ干扰素(interferon-γ,IFN-γ)和IL-2水平。结果:与Si-mut转染组相比,Si-Sema转染的A549细胞表达和分泌Sema3A水平明显下降(93.4±7.3%vs 27.8±5.3%,P<0.01);Sema3A缺失引起DCs表达MHCⅡ分子和共刺激分子水平上升,分泌IL-12 p70量明显提高(449.3±29.3 pg/mlvs675.7±60.2 pg/ml,P<0.05),刺激抗原特异性T细胞表达IFN-γ(639.6±41.9 pg/mlvs905.6±68.0 pg/ml,P<0.05)和IL-2(1011.7 pg/ml±81.4 pg/mlvs1459.0 pg/ml±96.8 pg/ml,P<0.05)显著增加。结论:肺腺癌细胞A549通过分泌Sema3A,抑制DCs的成熟和免疫学功能,这可能是目前尚未完全了解的肿瘤免疫逃逸机制之一。更多还原

【Abstract】 Objective: To investigate the effects of tumor cell-derived Sema3A on the immunological functions of murine dendritic cells(DCs).Methods: Lung adenocarcinoma A549 cells were transfected with small interference RNA,Si-Sema and Si-mut,and the interference efficiency was determined by real-time PCR and Western-blot.The concentrated supernatants from cultured tumor cells,Si-Sema and Si-mut-infected tumor cells were subjected to DCs respectively.The immunophenotypes of DCs were analyzed by flow cytometry,the production of IL-12P70 and the ability of DCs to stimulate DO11.10 T cells secreting IFN-γ and IL-2 were detected by enzyme linked immunosorbent assay(ELISA).Results: Knockdown with Si-Sema3A significantly decreased the secretion of Sema3A by A549 cells in comparison with the Si-mut cells.DCs exposed to supernatants from Si-Sema cells showed elevated levels of MHC,CD40 and CD80,more production of IL-12P70,and enhanced capability of activating antigen-specific T cells,as evidenced by the remarkably increased levels of IFN-γ and IL-2.Conclusion: A549 cells secrete Sema3A to inhibit the maturation and functions of DCs,which might be associated with the unidentified mechanism of immune evasion by tumor cells.更多还原