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目的:探讨体外髓系培养体系中外周血单核细胞来源的CD123+髓系树突状细胞(mDC)的生物学特性。方法:分离健康人外周血单核细胞,用重组的粒/单核细胞集落刺激因子(GM-CSF)和白细胞介素4(IL-4)将其诱导为树突状细胞(DC)。用流式细胞术(FCM)检测DC表面共刺激分子、CD304、CD123和CD11C的表达,并用间接免疫磁珠法将其中CD123+DC加以分离纯化;激光共聚焦显微镜、扫描电镜观察CD123+DC形态;ELISA法检测CD123+DC的IL-12分泌量;葡聚糖吞噬试验和3H-TdR渗入法分别检测CD123+DC的吞噬功能和对同种异体T细胞的刺激能力。结果:外周血单核细胞经GM-CSF和IL-4诱导7d后,细胞表面高度表达CD86和CD11C,中等量表达CD1a和CD123,低表达CD83,丧失CD14的表达,其中,CD123和CD11C均匀分布于DC表面。免疫磁珠纯化后的CD123+DC呈现典型的不成熟DC形态,除细胞体积较小外,其表面突起类似于CD123-DC。CD123+DC仅微量分泌IL-12,其吞噬能力强于CD123-DC(P<0.05),但抗原刺激功能低于后者(P<0.05)。结论:GM-CSF和IL-4培养体系中的CD123+DC可能是DC分化发育过程中更早期的未成熟mDC,具有独特的生物学特性。
Objective: To investigate the biological characteristics of peripheral blood mononuclear cell derived CD123 + myeloid dendritic cells (mDC) in myeloid culture system in vitro. Methods: Peripheral blood mononuclear cells were isolated from healthy volunteers and induced to DC by recombinant GM-CSF and IL-4. The expression of costimulatory molecules CD304, CD123 and CD11C on DCs were detected by flow cytometry (FCM), and the CD123 + DCs were isolated and purified by indirect immunomagnetic beads method. The morphology of CD123 + DCs was observed by laser confocal microscopy and scanning electron microscopy ; IL-12 secretion of CD123 + DC was detected by ELISA; phagocytosis of CD123 + DC and stimulation of allogeneic T cells were detected by dextran phagocytosis and 3H-TdR infiltration. RESULTS: Peripheral blood mononuclear cells were highly expressed on CD86 and CD11C after induced by GM-CSF and IL-4 for 7 days. CD1a and CD123 were moderately expressed on the surface of cells, while CD83 was down-regulated and CD14 was lost. CD123 and CD11C were evenly distributed On the DC surface. Immunomagnetic beads purified CD123 + DC showed a typical immature DC morphology, with the exception of the smaller cell size, the surface prominence similar to CD123-DC. CD123 + DC secrete only a small amount of IL-12, phagocytosis than CD123-DC (P <0.05), but antigen-stimulating function was lower than the latter (P <0.05). CONCLUSION: CD123 + DC in GM-CSF and IL-4 culture system may be the early immature mDC in the process of DC differentiation development with unique biological characteristics.