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有关树状突细胞(Dendritic Cell,DC)在肿瘤免疫中作用的研究,多限于观察肿瘤局部DC的变化,而对DC抗肿瘤作用的探讨尚属起步.本研究采用多因素不同水平的全面试验设计,进行DC的体外抗肿瘤实验.首先应用新的三步分离法分离、纯化,获得纯度为60～70%的DC;又在用重组IL-2诱导淋巴因子激活的杀伤细胞(LAK 细胞)的基础上,以中性红摄入比色法检测细胞毒活性,观察了体外DC对LAK细胞杀伤H_7402肿瘤细胞活性的影响,也观察了形态学变化.结果:活细胞观察发现DC联合LAK作用组残存的活肿瘤细胞比单LAK作用组明显减少,HE染色见肿瘤细胞呈不同程度的坏死状态.免疫细胞化学染色显示S-100蛋白阳性的DC与LAK细胞形成花环,DC、LAK和肿瘤细胞三者形成细胞簇.扫描电镜下可见DC借助突起与肿瘤细胞相连,也与LAK细胞接触,三者形成细胞簇.细胞毒活性检测所得各孔光密度OD值经方差分析和参数估计,表明LAK细胞体外杀伤H_(7402)肿瘤细胞的活性随效靶比增加而增强,DC能协同LAK细胞的活性,上调其杀伤肿瘤的作用(P<0.01),并以中等剂量的DC上调速度最明显,加入IL-2后,DC的调节作用更有上升.提示人外周血DC在细胞免疫抗肿瘤过程中起重要使用.
Studies on the role of dendritic cells (DCs) in tumor immunity are mostly limited to the observation of local DC changes in tumors, but the investigation of the anti-tumor effects of DCs is still in its infancy. This study uses multiple tests at different levels and different levels. Design and conduct DC in vitro anti-tumor experiments. First use a new three-step isolation method to separate and purify to obtain DCs with a purity of 60-70%; In addition, lymphokine activated killer cells (LAK cells) are induced with recombinant IL-2. Based on the assay, cytotoxicity was measured by neutral red colorimetric assay, and the effect of DC on killing of H_7402 tumor cells by LAK cells in vitro was observed. Morphological changes were also observed. Results: Live cell observations revealed the role of DC in combination with LAK Liver tumor cells in the group survived significantly less than those in the single LAK group. HE staining showed necrosis of the tumor cells in different degrees. Immunocytochemical staining showed that DCs, LAKs, and tumor cells formed by S-100 protein positive DCs and LAK cells. The three formed cell clusters. Under scanning electron microscopy, DCs were connected to tumor cells via projections, and were also in contact with LAK cells. The three cells formed clusters of cells. The optical density OD of each well was measured by variance analysis. The parameter estimation showed that the activity of LAK cells killing H 7402 tumor cells in vitro increased with the increase of effector-to-target ratio, and DCs could synergize the activity of LAK cells, up-regulate their tumor-killing effects (P<0.01), and treated with medium-dose DCs. The most obvious up-regulation rate was that after the addition of IL-2, the regulatory effect of DC increased. This suggests that human peripheral blood DC plays an important role in the cellular anti-tumor process.