论文部分内容阅读
目的利用人外周血单核细胞体外诱导培养朗格罕氏细胞(Mo-LC)和炎症性树突状表皮细胞(Mo-IDEC)并观察其表型特点。方法联合应用LymphoPrepTM梯度离心试剂和CD14+磁珠分离和筛选外周血CD14+单核细胞;在重组人粒细胞-巨噬细胞集落刺激因子(GM-CSF)和重组人白介素4(IL-4)的基础上分别于第0、2、4天加入人天然TGF-β1或β-巯基乙醇(β-ME)。培养第6天时收集细胞,通过相差显微镜观察其形态,并利用单克隆抗体和流式细胞术对诱导细胞亚群检测细胞表面分子的表达水平。结果 CD14+单核细胞体外诱导培养6 d后可形成具有树突状外观的Mo-LC和Mo-IDEC,2种细胞均不同程度表达CD1a、FcεRI、FcεRII、TLR1、TLR2;Mo-LC表达CD207,Mo-IDEC表达CD206。特应性皮炎患者来源的Mo-IDEC表面CD1a表达高于Mo-LC,且CD23、TLR2、FcεRI的表达水平显著高于健康对照组。结论利用外周血单核细胞可在体外成功诱导Mo-LC和Mo-IDEC,这2种细胞表型特点与体内分离的细胞在形态和表型上类似,为体外进一步研究这2类细胞的功能打下基础。
OBJECTIVE: To induce the expression of Mo-LC and Mo-IDEC in vitro by human peripheral blood mononuclear cells (PBMCs) and observe its phenotypic characteristics. Methods LymphhoPrepTM gradient centrifugation reagent and CD14 + magnetic beads were used to separate and screen peripheral blood CD14 + monocytes. On the basis of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) and recombinant human interleukin 4 (IL-4) Human natural TGF-β1 or β-mercaptoethanol (β-ME) was added on days 0, 2 and 4, respectively. The cells were harvested on day 6, their morphology was observed by phase-contrast microscopy, and the expression of cell-surface molecules on the induced cell subpopulations was detected using monoclonal antibodies and flow cytometry. Results Mo-LC and Mo-IDEC with dendritic appearance could be formed after cultured for 6 days in vitro. Both of them expressed CD1a, FcεRI, FcεRII, TLR1 and TLR2 to different extents; Mo-LC expressed CD207, Mo-IDEC expresses CD206. The expression of CD1a on Mo-IDEC surface in patients with atopic dermatitis was higher than that of Mo-LC, and the expression levels of CD23, TLR2 and FcεRI were significantly higher than those in healthy controls. Conclusion The use of peripheral blood mononuclear cells can be successfully induced in vitro Mo-LC and Mo-IDEC, these two kinds of cell phenotypic characteristics and in vivo separation of cells in morphology and phenotype similar to further study in vitro function of these two types of cells lay the foundation.