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背景与目的:白血病细胞能在体外分化为树突细胞(dendriticcells,DCs),从而有希望用于白血病的免疫治疗。本研究旨在探讨CD14高表达的单核细胞系白血病(M4、M5)细胞分化而来的DCs体外诱导抗白血病T细胞应答的能力。方法:取5例初诊CD14高表达的M4或M5型白血病患者的骨髓标本,分离单个核细胞(bonemarrowmononuclearcells,BMMNCs),将白血病细胞分为3组:贴壁白血病细胞组、非贴壁白血病细胞组及总白血病细胞组。流式细胞术(flowcytometry,FCM)比较3组细胞的CD14表达。用含GM-CSF、IL-4和TNF-α或不含细胞因子的培养液培养细胞7～10天后,通过细胞形态学观察及FCM检测细胞表型,鉴定单核白血病细胞来源的DCs(monocyticleukemiacell-deriveddendriticcells,Mo-LDCs);采用异基因混合淋巴细胞反应(allogeneicmixedlymphocytereaction,Allo-MLR)以及细胞毒性T淋巴细胞(cytotoxicTlymphocytes,CTL)抗白血病细胞毒分析检测Mo-LDCs的免疫功能,染色体核型分析结合异常表面抗原确定Mo-LDCs的白血病来源。结果:3组中贴壁白血病细胞的CD14含量最高,在细胞因子联合诱导下,可分化为大量CD83+成熟DCs。在同一病例的3组细胞以及不同病例的总单核细胞组间,培养前CD14的表达率与诱导后CD83+DCs的产率成正相关(r=0.967,P=0.007)。Mo-LDCs具有典型的成熟DCs的形态及表型特征,在Allo-MLR中能刺激同种T细胞明显增殖,并能刺激扩增特异性抗白血病CTL。同时,Mo-LDCs持续存在所起源白血病的核型异常和异常表达的髓系抗原。结论:在细胞因子组合诱导下,M4、M5亚型AML的CD14+细胞可分化为具有免疫功能的Mo-LDCs,单核系白血病细胞的CD14表达高低可能预示其DCs分化能力。Mo-LDCs具有经典的DCs的表型及功能,还具有白血病的克隆异常,可用于M4、M5患者的免疫治疗。
BACKGROUND & OBJECTIVE: Leukemic cells can differentiate into dendritic cells (DCs) in vitro, which may be promising for immunotherapy of leukemia. This study was designed to investigate the ability of DCs differentiated from CD14-overexpressed monocytic leukemia (M4, M5) cells to induce leukemic T cell responses in vitro. Methods: Bone marrow specimens from 5 newly diagnosed patients with M4 or M5 leukemia with CD14 overexpression were isolated and bone marrow mononuclear cells (BMMNCs) were isolated. Leukemic cells were divided into 3 groups: adherent leukemia cell group, non-adherent leukemia cell group And total leukemia cell group. Flow cytometry (FCM) was used to compare CD14 expression in three groups of cells. After cultured for 7-10 days in culture medium containing GM-CSF, IL-4 and TNF-α or without cytokines, monocytic leukemia cells (DCs) were identified by morphological observation and FCM. -derived dendritic cells, Mo-LDCs). The immune function of Mo-LDCs was detected by allogeneic mixed lymphocyte reaction (Allo-MLR) and cytotoxic T lymphocytes (CTL) Combined with abnormal surface antigens to determine the source of leukemia Mo-LDCs. Results: Adherent leukemia cells in the three groups had the highest levels of CD14. Under the induction of cytokines, CD14 could differentiate into a large number of CD83 + mature DCs. The positive rate of CD14 before culturing was positively correlated with the yield of CD83 + DCs after induction (r = 0.967, P = 0.007) among the 3 groups of cells in the same case and the total mononuclear cells in different cases. Mo-LDCs have the morphological and phenotypic characteristics of typical mature DCs. Allo-MLR stimulates the proliferation of allogeneic T cells and stimulates the proliferation of specific anti-leukemia CTLs. At the same time, Mo-LDCs persist in the presence of myeloid abnormalities and abnormally expressed myeloid antigens of the origin of leukemia. CONCLUSION: CD14 + cells of M4 and M5 subtypes AML can differentiate into immune-competent Mo-LDCs induced by cytokines. The expression of CD14 in mononuclear leukemia cells may predict the differentiation ability of DCs. Mo-LDCs have the phenotype and function of classical DCs, and also have the clonal abnormalities of leukemia, which can be used in the immunotherapy of patients with M4 and M5.