人外周血单核细胞体外诱导树突状细胞的形态学观察与功能研究

The morphological and functional study of dendritic cells induced from human peripheral blood monocytes

目的:探讨从人外周血单核细胞体外培养扩增的树突状细胞(dendriticcell,DC)形态学特征及其活性。方法:人外周血常规分离单个核细胞,粘附6h后去除悬浮细胞,以细胞因子粒细胞-巨噬细胞集落刺激因子(GM-CSF)、白细胞介素-4(IL-4)进行诱导培养7d,并进行形态学特征,细胞表型和淋巴细胞刺激能力鉴定。结果:培养1周即可得到大量DC,形态学观察可见细胞形态不规则,细胞表面大量突起,为典型的DC特征,免疫组化和间接免疫荧光显示CD1a阳性表达率达80%～95%,并能刺激同种淋巴细胞增殖反应。结论:人外周血单核细胞可经GM-CSF、IL-4诱导培养成DC,具有典型的树突状细胞的形态学特征及抗原呈递能力。

Objective To study the morphology and function of dendritic cells (DCs) from human peripheral blood monocytes(PBMC). Methods PBMC isolated from healthy human peripheral blood were incubated at 37℃,humidified 5%CO2. After 6 hours, nonadherent cells were gently removed, and the adherent cells were cultured in medium with Granulocyte/macrophage colony stimulating factor (GM CSF) and interleukin 4(IL 4) at 37℃, humidified 5%CO2 for more than 7 days. DCs were then collected and identified by morphological features and underwent tests of surface antigen expression and lymphocytes stimulatory ability. Results A large amount of DCs could be obtained after 7 days, DCs displayed typical morphology with many bulbous protrusions and irregular in shape when observed by phase contrast microscopy and electron microscope, the level of expression CD1a was 80%～95%with immunohistochemical staining and indirect immunofluorescent assay (IFA) and potently stimulated the proliferation of allogeneic lymphocytes. Conclusion A large amount of DCs could be induced from peripheral blood monocytes in presence of GM CSF and IL 4, the obtained DCs have typical morphology and antigen presenting ability.