IFN-γ激活的树突状细胞对患者急性髓性白血病细胞的杀伤作用

为检测干扰素γ(IFNγ)激活的外周血树突状细胞(DC)对患者急性髓细胞白血病(AML)细胞的直接杀伤活性并探讨其杀伤机制,分离健康供者外周血单核细胞,用重组粒巨噬细胞集落刺激因子和白细胞介素4诱导生成DC。于诱导第7天在合成培养液中加入IFNγ继续培养12小时,将其激活。从6例初发的急性髓细胞白血病患者外周血分离单个核细胞作为靶细胞,以不同效靶比与DC共同培养18小时,采用51Cr释放试验检测DC激活前后抗肿瘤活性的差异。同时,用流式细胞术检测DC表面共刺激分子,细胞膜及细胞内Fas配体(FasL)、TNFα及肿瘤坏死因子相关凋亡诱导配体(TRAIL)的改变,以明确IFNγ激活的DC对肿瘤细胞直接杀伤活性的作用机制。结果表明:IFNγ可增强DC对AML细胞的杀伤活性,在效靶比为20∶1时杀伤率为(33.8±1.6)%,与未加刺激因子前相比有显著差异(P<0.05);IFNγ可上调DC表面共刺激分子CD86和CD83的表达;IFNγ刺激后,DC细胞内TRAIL表达明显增强,但其在细胞表面仍呈现低表达,而细胞内及细胞表面FasL及TNFα在IFNγ刺激前后均无明显变化。结论:IFNγ激活的DC可有效杀伤患者AML细胞,其作用机制与DC的成熟及TRAIL凋亡途径部分相关,而与FasL及TNFα凋亡途径无关。

Cytotoxicity of IFN-γ-Activated Dendritic Cells to Freshly Isolated Acute Myeloid Leukemia Cells

To investigate the tumoricidal activity of dendritic cell (DC) stimulated by interferon-gamma (IFN-gamma) against freshly isolated myeloid leukemia cells and its mechanism, the peripheral blood monocytes collected from healthy donors were cocultured with interleukin-4 and granulocyte-macrophage colony-stimulating factor in medium to induce DC for 7 days. After 12 hour culture in the absence or presence of IFN-gamma, the changes of costimulatory molecules were analyzed with flow cytometry. To assay the cytotoxicity of DC against freshly isolated acute myeloid cells, they were cocultured at various effector-to-target ratio for 18 hours, then the percentage of tumoricidal activity was measured with (51)Cr release assay. To explore the mechanism of DC-mediated cytotoxicity, the change of DC surface or intracellular protein expression of Fas ligand (Fas L), TNF-alpha and TNF related apoptosis-inducing ligand (TRAIL) were analyzed with flow cytometry. The results showed that IFN-gamma enhanced cytotoxicity of DC against AML cells was (33.8 +/- 1.6)% at E:T as 20:1, compared with unstimulated DC (P < 0.05); IFN-gamma up-regulated expression of costimulatory molecules of DC surface such as CD86 and CD83; after stimulation with IFN-gamma, expression of intracellular TRAIL of DC was significantly enhanced, but expression of TRAIL on cell surface of DC was low; while the significant changes of Fas L and TNF-alpha expression neither on cell surface or in cells were not observed before or after stimulation with IFN-gamma. It is concluded that DC stimulated by IFN-gamma exhibit tumoricidal activity against AML cells. The cytotoxicity is partially related to maturation of DC and TRAIL inducing apoptosis, but not associated with death domain-independent mechanism of Fas L and TNF-alpha.