摘要
目的 研究小鼠骨髓树突状细胞体外诱导分化、成熟及对T细胞增殖的影响 ,为进一步研究树突状细胞的功能及临床肿瘤治疗的应用提供技术方法。方法 应用IL 4、GM CSF培养小鼠骨髓细胞 5～ 7d ,镜下观察细胞形态学变化 ;培养至第 5～ 6d ,加入黑色素瘤 (B16 )冻融抗原继续培养 1～ 2d ,流式细胞仪检测细胞表面分子CD83、CD86 ,并与同种异体T细胞混合培养 72h ,终止培养前 16h加入 3H TdR(0 .5 μCi 孔 ) ,γ 液体闪烁仪测定cpm值。 结果 IL 4、GM CSF培养 3d可见细胞形态发生改变 ,细胞形状不规则 ,培养 5～ 6d时 ,有刺状突起、拉长 ,为典型树突状细胞形态学特征 ;抗原装载后流式细胞仪检测CD83、CD86表达 ,IL 4 +GM CSF +TNF α组 (6 1.6 8% ,71.2 5 % )及IL - 4 +GM -CSF +冻融抗原组 (6 3.11% ,76 .88% )明显高于对照组 (2 .4 1% ,3.88% )及单纯IL - 4 +GM -CSF培养组 (2 1.86 % ,2 8.6 9% ) (P <0 .0 0 1) ,IL - 4 +GM -CSF +TNF -α组与IL - 4 +GM -CSF +冻融抗原组比较CD83、CD86表达没有显著性差异 ;液闪仪检测结果显示 ,IL - 4 +GM -CSF +冻融抗原组刺激T细胞增殖的能力明显强于其他组 ,且有显著性差异(P <0 .0 0 1,P <0 .0 5 )。
Objective To study function of dendritic cells and provide techinology methods for clinical tumor-therapy, the differentiation induced in vitro of dendritic cells and T cells proliferation stimulated by mature DCs were observed.Methods with IL-4 and GM-CSF, the cells from mouse bone marrow were cultured. Morphology was observed on 3 and 5 day of culture; immature DCs on 5 or 6 day of culture were loaded with tumor-antigen for 1～2 day ; CD83 and CD86 were tested by flow-cytometric analysis; T-cell-stimulatory activity in DCs derived from mouse bone marrow and loaded with tumor-antigen was analysed by γ- scintillomete. Results alteration of cell form and dendritic protrusion appear on 3 day of culture; dendritic protrusion prolong on 5 day of culture; flow-cytometric analysis show that expression of CD83 and CD86 of IL-4+GM-CSF+TNF-αgroup and IL-4+GM-CSF+tumor-antigen group have significant difference comparison with control group and IL-4+GM-CSF group( P <0.001)and IL-4+GM-CSF+tumor-antigen group have also significant difference comparison with IL-4+GM-CSF+TNF-αgroup( P <0.01); proliferation of T cells are evaluated that IL-4+GM-CSF+tumor-antigen group strongly enhance proliferating activities of T cells comparison with other( P <0.001, P <0.05). Conclusion sufficient DCs generated from bone marrow cells are induced in vitro and strongly stimulate proliferation of T cells after tumor antigen are loaded.
出处
《中国实验诊断学》
2003年第6期484-486,共3页
Chinese Journal of Laboratory Diagnosis
