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目的 体外诱导高效特异的抗肿瘤免疫反应 ,以期制备新型特异性肿瘤免疫治疗效应细胞 .方法 分离健康人外周血单个核细胞 (PBMC) ,将其中非贴壁细胞 (淋巴细胞 )加高聚金葡素 (HAS)培养 (称 HASL) ;贴壁细胞经 IL- 4和 GM- CSF联合刺激诱导树突状细胞 (DC)分化 .DC用人肺癌细胞系 GL C- 82的可溶性抗原致敏 ,并用 TNF-α和 PGE2促进 DC的抗原递呈能力 .培养第 10日 ,将 DC与经或未经 HAS刺激的淋巴细胞混合培养 4h,分别称混合培养细胞为 DC- HASL 和 DC- L.将 DC- HASL,HASL及 DC- L作为效应细胞 ,肿瘤细胞作为靶细胞 ,用 MTT法检测细胞毒活性 .细胞表型用流式细胞仪分析 .结果 DC-HASL 对 GL C- 82细胞具有高效而特异性的杀伤作用 (杀伤率 10 0 % ) ;对另一肺癌细胞系 A5 49细胞的杀伤率也达 94.2 % ;对大肠癌细胞 moser和乳腺癌细胞 MCF- 7的杀伤率分别为 73.5 %和 71.2 % .而 HASL和 DC- L 对 A5 49,GL C- 82 ,moser和 MCF- 7细胞的杀伤率则分别为 48% ,6 3% ,6 0 % ,45 %和 42 .1% ,46 .4% ,39% ,2 5 .6 % .淋巴细胞经 HAS刺激后 ,CD3+ CD5 6 + 细胞的比例大幅度增加 .结论 HAS在体外刺激淋巴细胞增殖产生的 HASL 对肿瘤细胞有一定的杀伤作用 ,无明显特异性 .而 DC-HASL则可高效特异性地杀伤 G
Objective To induce highly specific anti-tumor immune responses in vitro in order to prepare novel tumor-specific immunotherapy effector cells. Methods: To isolate healthy human peripheral blood mononuclear cells (PBMCs) and to attach non-adherent cells (lymphocytes) to hyper-aggregated gold HAS culture (called HASL); adherent cells stimulate dendritic cell (DC) differentiation by IL-4 and GM-CSF co-stimulation. DC is sensitized with soluble antigen of human lung cancer cell line GL C-82, and TNF is used. -α and PGE2 promote antigen-presenting ability of DCs. On the 10th day of culture, DCs were incubated with or without HAS-stimulated lymphocytes for 4 h, and mixed culture cells were called DC-HASL and DC-L. DC- HASL, HASL and DC-L were used as effector cells and tumor cells as target cells. The cytotoxic activity was detected by MTT assay. The cell phenotype was analyzed by flow cytometry. Results DC-HASL was highly efficient and specific for GL C-82 cells. The killing effect (killing rate was 100%); the killing rate of another lung cancer cell line A5 49 cell was also 94.2%; the killing rate of colorectal cancer cell moser and breast cancer cell MCF-7 was 73.5 % and 71.2 % respectively. . And HASL and DC-L on A5 49,G The killing rates of L C-82, moser, and MCF-7 cells were 48%, 63%, 60%, 45%, 42.1%, 46.4%, 39%, and 25.6%, respectively. After stimulated by HAS, the proportion of CD3+ CD5 6 + cells increased significantly. Conclusion HAS stimulated lymphocyte proliferation in vitro HAS produced a certain killing effect on tumor cells, no obvious specificity, while DC-HASL can be highly effective. Specific killing G