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目的观察趋化因子巨噬细胞炎症蛋白-1α(MIP-1α)体外动员的树突状细胞(DC)经基因修饰后在荷瘤小鼠体内的抗胃癌效应。方法615小鼠通过尾静脉注射MIP-1α,流式细胞仪分选出B220~- CD11c~+细胞,加入鼠粒-巨噬细胞集落刺激因子(mGM-CSF)、白细胞介素- 4(IL-4)和鼠肿瘤坏死因子-α(mTNF-α)贯续培养进行诱导分化。通过细胞表型和混合淋巴细胞反应,检测细胞因子培养前后B220~- CD11c~+细胞的异同。收集培养后的B220~- CD11c~+细胞,加入编码黑色素瘤抗原基因-3(MAGE-3)的重组腺病毒进行转染,制备表达肿瘤抗原的DC疫苗。制备小鼠前胃癌(MFC)实体瘤模型,DC疫苗于MFC细胞接种后经皮下注射,观察小鼠瘤体生长和存活情况,研究DC疫苗的免疫治疗作用。结果MIP-1α注射8h后外周血中B220~- CD11c~+细胞数量即升高,48h达到高峰,占外周血单个核细胞(MNCs)(13.68±0.95)%。新鲜分离的B220~- CD11c~+细胞不具有成熟DC的特征,而经体外细胞因子诱导分化后则具有典型的DC表型,并具有极强的刺激T细胞增殖的能力。小鼠皮下接种MFC细胞后注射基因修饰的DC疫苗,小鼠瘤体生长缓慢,存活时间明显延长,与对照组之间差异有统计学意义(P<0.01)。结论注射趋化因子MIP-1α可快速动员B220~- CD11c~+细胞进入小鼠外周血,并经细胞因子可诱导分化为成熟DC。MIP-1α动员的DC经基因转染制备的DC疫苗,在体内对荷瘤小鼠有明显的免疫治疗作用,且较荷载全肿瘤细胞抗原的DC疫苗作用明显增强。
Objective To observe the anti-gastric cancer effect of genetically modified dendritic cells (DCs) stimulated by chemokine MIP-1α in tumor-bearing mice. Methods 615 Mice were injected with MIP-1α via the tail vein and the B220 ~ - CD11c ~ + cells were sorted by flow cytometry. The mGM-CSF, IL- -4) and murine tumor necrosis factor-α (mTNF-α) were cultured continuously. The similarities and differences of B220 ~ - CD11c ~ + cells before and after cytokine culture were detected by cell phenotype and mixed lymphocyte reaction. The cultured B220 ~ - CD11c ~ + cells were collected and transfected with the recombinant adenovirus encoding the melanoma antigen gene-3 (MAGE-3) to prepare a DC vaccine expressing tumor antigens. Preparation of mouse solid gastric cancer (MFC) model, DC vaccine was injected subcutaneously in MFC cells after inoculation to observe the growth and survival of tumor in mice to study the immunotherapy of DC vaccine. Results The number of B220 ~ - CD11c ~ + cells in peripheral blood increased after 8h injection of MIP-1α and peaked at 48h, accounting for 13.68 ± 0.95% of peripheral blood mononuclear cells (MNCs). Freshly isolated B220 ~ - CD11c ~ + cells do not have the characteristics of mature DCs, but have typical DC phenotype after differentiation induced by cytokines in vitro, and have strong ability of stimulating T cell proliferation. Mice were inoculated with MFC cells subcutaneously after injection of genetically modified DC vaccine, the growth of tumor in mice slow, survival time was significantly longer, with the difference between the control group was statistically significant (P <0.01). Conclusion Injection of chemokine MIP-1α can rapidly mobilize B220 ~ - CD11c ~ + cells into peripheral blood of mice and can differentiate into mature DCs by cytokines. DC vaccines prepared by MIP-1α-mobilized DCs have obvious immunotherapy effect on tumor-bearing mice in vivo, and the effect of DC vaccines is better than that of DC vaccine loaded with whole tumor cell antigens.