目的：体外诱导培养缓解期及未缓解期急性白血病患者来源的树突状细胞（dendritic cell, DC)联合细胞因子诱导的杀伤(cytokine-induced killer, CIK)细胞，比较两者的增殖能力、免疫表型及对白血病细胞的杀伤活性。方法: 提取2013年4月25日至2014年1月17日河北医科大学第二医院血液科收治的复发难治未缓解期白血病患者（7例）和缓解期患者（7例）的外周血单个核细胞，分别按常规方法诱导产生DC-CIK细胞，在培养过程中检测细胞增殖倍数，流式细胞术检测CD3+CD8+、CD3+CD56+细胞比例，细胞涂片法和流式细胞术检测培养前后CD34+白血病细胞比例，CCK-8法检测两组DC-CIK细胞对K562细胞及患者单个核细胞的杀伤活性。结果: 缓解组和未缓解组来源的DC-CIK细胞均以相似的速度快速增殖，CD3+CD8+和CD3+CD56+细胞比例均随培养时间延长显著升高（ P <0.05），但两组间差异没有统计学意义（ P >0.05）。培养第15天时，非缓解组DC-CIK细胞涂片检测未见CD34+原始白血病细胞，流式细胞术检测显示CD34+细胞比例较培养前显著降低\[（0.1±0.05）% vs (8.3±3.1)%， P <0.05\]。效靶比在5 ∶1~20 ∶1范围内时，缓解组与未缓解组DC-CIK细胞对K562细胞的杀伤率差异没有统计学意义（ P >0.05），但未缓解组DC-CIK对患者单个核细胞的杀伤率显著高于缓解组（ P <0.05），并且显著高于未缓解组对K562细胞的杀伤率（ P <0.05）。结论:未缓解期白血病患者来源的DC-CIK细胞在增殖活性、CD3+CD8+和CD3+CD56+表达水平和对K562细胞的非特异性杀伤活性方面与缓解期患者来源的DC-CIK细胞均相似，但对患者单个核细胞的杀伤具有更强的特异性。

Objective:To evaluate the cytotoxicity of dendritic cells-cytokine-induced killer cells (DC-CIK cells) derived from patients with remission leukemia or a non-remission disease against leukemia cells. Methods: Blood was sampled from 7 seven patients with refractory non-remission acute leukemia and 7 patients with remission leukemia who were admitted to the Department of Hematology, Hebei Medical University-Affiliated Second Hospital between April 25, 2013 and June 17, 2014. Peripheral blood mononuclear cells (PBMCs) were isolated and induced into DC-CIK cells by conventional methods. The proportions of CD3+CD8+ and CD3+CD56+ cells in the induced DC-CIK cell population were assessed by flow cytometry. Their proliferation rate was calculated and their killing activity against K562 cells and mononuclear cells were determined by CCK-8 assay. Results: DC-CIK cells derived from both patients with a remission disease and patients with a non-remission disease proliferated at similarly rapid rates ( P >0.05). Proportions of CD3+CD8+ and CD3+CD56+ cells increased dramatically with time in both groups of DC-CIK cells ( P <0.05) but there was no significant difference between the two groups ( P >0.05). After 15 days of induction, no leukemia cells were found and the proportion of CD34+ cells was significantly reduced in DC-CIK cell preparations as compared with untreated PBMCs (\[0.1±0.05\]% vs \[8.3±3.1\]%, P <0.05). At effector to target cell ratios from 5 ∶1 to 20 ∶1, DC-CIKs derived from both groups of patients displayed a similar cytotoxic activity against K562 cells ( P >0.05), but DC-CIKs derived from patients with a non-remission disease had a significantly higher cytotoxic activity against mononuclear cells ( P <0.05). Conclusion: DC-CIK cells derived from PBMCs of both patients with non-remission leukemia and patients with remission leukemia have similar proliferative activities, similar profiles of CD3+CD8+ and CD3+CD56+ expression and similar cytotoxic activities against K562 cells. However, DC-CIK cells from patients with non-remission leukemia display a higher sensitivity against autologous mononuclear cells.

国家高等学校博士学科点专项科研基金资助项目（No.200800890011）