A23187诱导HL-60细胞分化为树突状细胞的研究

目的 探讨用钙离子载体A23187通过钙动员机制使人急性粒细胞性白血病细胞系HL-60细胞迅速分化为树突状细胞(DCs)的方法,为今后临床用DCs进行抗白血病免疫治疗打下基础.方法 将生长良好的HL-60细胞加入不同浓度A23187(45～720 ng/mL)或联用重组人γ-干扰素(rhIFN-γ,1000 U/mL)培养20～96 h,倒置显微镜和扫描电镜观察其形态学特征,流式细胞术检测其免疫表型,混合淋巴细胞反应检测其刺激T淋巴细胞增殖的能力.结果 HL-60细胞加入适量A23187(180 ng/mL)或联合rhIFN-γ(1000 U/mL)诱导20 h后,即可出现细胞表面树状突起,且树突状细胞表面特异性成熟标志抗原CD83、共刺激分子CD80、CD86表达升高;培养48 h,CD83分子表达开始降低;培养72 h,可获得DC的典型形态,CD80、CD86分子表达持续升高.同种异体混合淋巴细胞反应检测经A23187或联用rhIFN-γ诱生的DCs可明显刺激同种异体T淋巴细胞增殖.结论 钙离子载体可诱导HL-60细胞分化为高表达共刺激分子及具有刺激同种异体T淋巴细胞增殖的成熟DCs,钙动员有望成为一种快速获取DCs的有效方法.

Research on A23187 Inducing HL-60 Cells to Differentiate into Dendritic Cells

OBJECTIVE: To investigate the method for calcium mobilization inducing rapidly to produce the dendritic cells (DC) from myeloid leukemia cells. METHODS: HL-60 cells were cultured with calcium ionophore A23187 (45-720 ng/mL) or plus rhIFN-gamma (1000 U/mL) for 20-96 hours. The morphologic features of cells were observed under inverted microscope and scanning electron microscope, when the cell phenotypes of HL-60 treated with A23187 were determined by flow cytometry. The proliferation of allogeneic human T cells was tested by mixed lymphocyte reaction (Allo-MLR). RESULTS: After treated with A23187 (180 ng/mL) or plus rhIFN-gamma (1000 U/mL) for 20 hours, some of HL-60 cells were found to have the dendritic appearance on cell surface; and the CD83 as a characteristic marker of mature DCs, co-stimulating molecules CD80 and CD86 got an up-regulated expression. When HL-60 cells were cultured with A23187 for 48 hours, the expression of CD83 began going down. A large number of cells with typical dendritic appearance were observed after cultured with A23187 for 72 hours, and the expressions of CD80 and CD86 were up-regulated continuously. Allo-MLR revealed that DCs derived from HL-60 cells treated with A23187 or plus rhIFN-gamma could stimulate the proliferation of allogeneic human T cells. CONCLUSION: The calcium ionophore A23187 can induce the HL-60 cells into mature DCs. This study suggests that calcium mobilization for cell differentiation may be an alternative way to rapidly acquire DCs.