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[目的]研究肝癌3种不同全细胞抗原负载方式(肿瘤细胞裂解物、融合以及肿瘤细胞总RNA)制备DC瘤苗的体外抗肿瘤活性,分析其体外诱导T细胞反应的异同,以期选择更好的DC瘤苗。[方法]分离肝癌患者外周血单核细胞,体外诱导DC;以上述3种抗原负载方式制备DC瘤苗(Lys-DC、Fus-DC、RNA-DC),刺激肝癌患者淋巴细胞作为效应细胞,肝癌细胞株HepG2为靶细胞,MTT法测定效应细胞对各肿瘤细胞的杀伤作用;分离CD8+和CD4+T细胞,瘤苗刺激后,应用IFN-γELISA检测试剂盒检测CD8+T细胞IFN-γ分泌,MTT法检测各种瘤苗刺激自体CD4+T细胞增殖的能力。[结果]三种瘤苗刺激的淋巴细胞均显示对HepG2高效特异的杀伤活性,但Fus-DC-L、RNA-DC-L对HepG2的杀伤率明显高于Lys-DC-L组;Lys-DC、Fus-DC、RNA-DC刺激的CD8+T细胞组IFN-γ分泌显著高于DC刺激的CD8+T细胞组和单纯CD8+T细胞组;但Fus-DC、RNA-DC刺激组IFN-γ分泌显著高于Lys-DC刺激组;Lys-DC、Fus-DC、RNA-DC组刺激自体CD4+T细胞增殖功能显著高于DC组;但Lys-DC、Fus-DC、RNA-DC刺激自体CD4+T细胞增殖功能无明显差异。[结论]肿瘤总RNA转染的DC以及融合瘤苗能更有效地诱导肿瘤特异性细胞毒性T淋巴细胞反应,是更为有效的DC免疫治疗方式。
[Objective] To study the antitumor activity of DCs in vitro prepared by three kinds of whole-cell antigen loading methods (tumor cell lysate, fusion and total RNA of tumor cells), and to analyze the similarities and differences of T cell responses induced in vitro in order to select better DC vaccine. [Methods] Peripheral blood mononuclear cells were isolated from patients with hepatocellular carcinoma to induce DCs in vitro. DCs (Lys-DC, Fus-DC, RNA-DC) were prepared by loading the above three antigens, stimulating lymphocytes of HCC patients as effector cells, CD8 + and CD4 + T cells were isolated from HepG2 cells by MTT assay. The IFN-γ secretion of CD8 + T cells was detected by IFN-γ ELISA kit after stimulation with the vaccine The ability of various tumor-stimulating cells to proliferate autologous CD4 + T cells was detected by MTT assay. [Results] All the three kinds of tumor-stimulated lymphocytes showed highly specific and specific killing activity against HepG2, but the killing rate of HepG2 by Fus-DC-L and RNA-DC-L was significantly higher than that of Lys- IFN-γ secretion in CD8 + T cells stimulated by DC, Fus-DC and RNA-DC was significantly higher than that of CD8 + T cells and CD8 + T cells stimulated by DCs DCs, Fus-DCs and RNA-DCs significantly enhanced the proliferation of autologous CD4 + T cells compared with DCs. However, Lys-DC, Fus-DC and RNA- Stimulation of autologous CD4 + T cell proliferation was no significant difference. [Conclusion] DC transfected with total RNA of tumor and fusion tumor vaccine can induce tumor specific cytotoxic T lymphocyte reaction more effectively, which is a more effective method of DC immunotherapy.