中文摘要：

目的：将体外扩增的黏蛋白1（MUC1）mRNA转染入成熟的树突状细胞（DCs）,观察其体外诱导的特异性抗肿瘤效应。方法：将分离提纯的单核细胞培养诱导为DC并用流式细胞术鉴定。构建pcDNA3.1（＋）-MUC1质粒,体外转录为mRNA,电穿孔法转染DCs。定量PCR检测转染的DCs中MUC1的表达;MTT法检测T细胞增殖情况;流式细胞术检测CD8^＋在T细胞的表达;LDH释放法测定细胞毒性,ELISA检测IFN-γ分泌水平。结果：流式细胞术结果表明成熟DCs标志表型的表达明显高于对照组。定量PCR结果说明转染后的DCs MUC1 mRNA相对表达量增高。转染组DCs与T细胞按1∶10共培养时,刺激增殖能力明显高于未转染组,且CD8^＋T细胞表达率高于未转染组,诱导产生特异性的细胞毒性T细胞杀伤表达MUC1蛋白的靶细胞,而未转染组的杀伤作用较弱。转染组DCs与T细胞共培养的上清中IFN-γ的分泌水平高于未转染组。结论：电穿孔法可以将MUC1 mRNA成功转染至DCs,产生特异性杀伤效应,为以MUC1为靶点的非小细胞肺癌的免疫治疗提供实验和理论依据。

英文摘要：

AIM： To investigate the specific anti-tumor effects of mature dendritic cells（ DCs） transfected with amplified mucin 1（ MUC1） mRNA in vitro. METHODS： DCs separated and purified from the peripheral blood mononuclear cells were induced in vitro and then identified by flow cytometry. pcDNA3. 1（ ＋）-MUC1 plasmid was constructed and was able to transcribe MUC1 mRNA in vitro. The MUC1 mRNA was transfected into DCs by electroporation. MUC1-transfected DCs were used to induce T cells to be cytotoxic T-lymphocytes. Quantitative real-time PCR was performed to assess MUC1 mRNA expression in transfected DCs. The proliferation of T cells was examined by MTT assay. The proportion of CD8^＋cells in the T cells was determined by flow cytometry and the specific cytotoxicity was measured by LDH assay. The secretion of IFN-γ was detected by ELISA. RESULTS： The marker gene expression in the DCs transfected with MUC1 mRNA was significantly increased compared with control group,peaking at 24 h. The transfection group showed the higher capacity to stimulate the proliferation of T cells compared with control group when the ratio of DCs to T cells was 1∶ 10. The proportion of CD8^＋cells in transfection group was higher than that in control group. The lethal effect of special cytotoxic Tlymphocytes on target cells in transfection group was stronger than that in control group. The level of IFN-γ in the cell supernatant of transfection group was higher than that in control group. CONCLUSION： DCs plus MUC1 mRNA by electrical transfection induces specific anti-tumor effects,which provides an experiment evidence of using MUC1 as a target for immunotherapeutic strategy against non-small cell lung cancer.