树突状细胞体外高效诱导白血病特异性细胞毒性T淋巴细胞生成  

Generation of Leukemia-specific Cytotoxic T Lymphocyte Induced by Dendritic Cells from Bone Marrow Monocytes in Vitro

作　　者：李湘新[1] 陈学良[1] 马道新[1] 何晓鹏[1] 李颢[1] 

机构地区：[1]山东大学齐鲁医院,山东济南250012

出　　处：《肿瘤学杂志》2004年第6期378-381,共4页Journal of Chinese Oncology

基　　金：国家自然科学基金资助项目(30070321)

摘　　要：[目的]研究体外诱导产生的细胞毒性T淋巴细胞(CTL)及其抗白血病反应。[方法]应用mGM鄄CSF及mIL鄄4细胞因子从小鼠骨髓细胞扩增出成熟树突状细胞(DC),使其负载冻融法制备的白血病细胞相关抗原(TAA),通过观察DC诱导的白血病特异性CTL的免疫表型,MTT分析其对于L7212细胞的抑制率,利用ELISA评价IL鄄2和IL鄄4水平。[结果]骨髓单个核细胞经mGM鄄CSF、mIL鄄4的联合作用7天后光镜及扫描电镜下观察到大量成熟DC生成。经负载TAA的DC活化后T细胞中CD3+、CD8+、CD25+细胞明显增多。FCM显示CD3+、CD8+、CD25+T细胞显著增多,CD8+细胞多于CD4+细胞。活化后T细胞对L7212细胞有特异性杀伤活性,在效靶比为50∶1培养72h后杀伤率达90.1%±2.7%。DC与T细胞共培养上清中IL鄄2的分泌水平为4.656±0.62pg/ml,明显高于普通T细胞培养组的1.436±0.11pg/ml(P=0.011),IL鄄4水平则无明显变化(P>0.05)。[结论]mGM鄄CSF及mIL鄄4配伍诱导生成的DCs经L7212冻融抗原负载后可在体外高效诱导白血病特异性CTL生成。To explore the generation of leukemia-specific cytotoxic T lymphocyte (CTL) induced by dendritic cells (DCs) from bone marrow monocytes and its anti-leukemia immune response. DCs were proliferated from bone marrow monocytes stimulated by mouse granulocyte-macrophage colony stimulating factor (mGM-CSF) and interleukin-4(mIL-4), and then they were pulsed with L7212 tumor cytolysis antigen. After coculturing of mature DCs pulsed with L7212 tumor cytolysis antigen and T cells for 3 days, flow cytometry(FCM) and Methyl thiazolyl tetrazolium(MTT)assay were used to detect the immune phenotype of CTL and the inhibition rate of CTL on LT212 cells. Enzyme-linked immunoadsordent assay(ELISA)were employed to evaluate the level of cytokine IL-2 and IL-4. Bone marrow monocytes could be induced into mature DCs stimulated by mGM-CSF and mIL-4 after 7 days. Mature DCs with tumor cytolysis antigen could active na?觙ve T cells to become leukemia-specialized CTL. FCM analysis showed significant increase of CD3+, CD8+, CD25+ T cells. The proportion of CD8+ cells was notably higher than that of CD4+ cells. This CTL had significant cytotoxicity on L7212 cells, while had no cytotoxicity on NIH3T3 cells. DCs and inactivated T lymphocyte also had no function on L7212 cells. The level of IL-2 in DCs and T cells coculture medium was 4.656±0.62pg/ml, which significantly higher than that in inactived T cell medium(1.436±0.11pg/ml, P=0.011),while the level of IL-4 showed little change(P>0.05). [Conclusion] DCs that induced by mGM-CSF and mIL-4 and pulsed with L7212 tumor cytolysis antigen can effectively activate nave T lymphocyte to leukemia-specific CTL in vitro.

关 键 词：树突状细胞 细胞毒性T淋巴细胞 白血病 免疫疗法 

分 类 号：R733.7[医药卫生—肿瘤]