摘要
目的 :建立人脾脏单核细胞体外培养生成大量树突状细胞的新方法。 方法 :人脾脏细胞悬液培养2h获得贴壁的单核细胞 ,加入重组人粒细胞 巨噬细胞集落刺激因子 (rhGM CSF) 10 0 μg/L或rhGM CSF 10 0 μg/L +重组人白细胞介素 4(rhIL 4) 5 0 0kU/L ,体外培养 7天 ,收集悬浮细胞 ,以流式细胞仪分析细胞表型及抗原内吞能力 ,体外混合淋巴细胞反应检测细胞抗原呈递功能。 结果 :以细胞因子培养 7天生成的悬浮细胞 ,2 0 %～ 80 %表达树突状细胞 (DC)特异性标志CD1a ,表达高水平的HLA DR、B7 1、B7 2分子 ,具有较强的抗原内吞能力 ,体外可以强烈刺激同种T细胞增生 ,其形态、表型、功能与人外周血单核细胞培养获得的DC相似。 结论 :从人脾脏获得的贴壁的单核细胞 ,体外以rhGM CSF +rhIL 4培养 ,可以获得极其大量的DC(>10 9细胞 /个脾脏 ) ,为进一步研究和应用打下了基础。
Objective:To establish a new method for culture of human spleen dendritic cells(DC) for being used in vitro study. Methodology:Human spleen cells were cultured for 2 hours at 37℃,non adherent cells were removed.The adherent monocytes were then further cultured in medium supplemented with rhGM CSF(100 μg/L)and rhIL 4(500 kU/L)for 7 days.The phenotype and the endocytosis activity of the adherent cells cultured were analyzed with flow cytometry.T lymphocyte stimulating activity was determined by allo MLR. Results:Expression of human DC specific marker CDla was found in 20%～80% of the cultured adherent cells.High levels of HLA DR,B7 1 and B7 2 expression were also showed on the surface of these cells.These cells could acitvely endocytose protein particles and stimulate allogeneic T lymphocytes proliferation in vitro.Similar morphology,phenotype and functions could be observed in these human spleen DCs and the peripheral monotyes derived DCs. Conclusion:Large quantities of DC can be harvested by culturing the adherent human spleen monocytes with rhGM CSF and rhIL 4 in vitro.
出处
《肾脏病与透析肾移植杂志》
CAS
CSCD
2001年第2期133-137,共5页
Chinese Journal of Nephrology,Dialysis & Transplantation
