摘要
目的:观察黄芪多糖(Astragalus polysaccharides,APS)体外对脐血单核细胞定向分化为树突状细胞(DCs)及其对T细胞的刺激增殖作用。方法:无菌条件下采集脐血,密度梯度离心法获得脐血单个核细胞分为3组,实验组:在含有APS(浓度为100mg/L)的RPMI1640完全培养液中培养;阴性对照组:在无药物的RPMI1640完全培养液中培养;阳性对照组:在含有细胞因子(IL-4、GM-CSF、TNF-α)的RPMI1640完全培养液中培养;培养过程中用倒置光学显微镜和扫描电镜观察细胞形态;收集部分培养第12天的细胞利用流式细胞仪检测各组细胞表面CD1a、CD80、CD83和CD86的表达;收集实验组培养第12天的细胞(DCs),作为刺激细胞;分离制备健康志愿者异基因外周血单核细胞(MNC),作为反应细胞混合培养,MTT比色法检测DCs对同种异体T细胞的刺激增殖作用。结果:在培养的第72小时后实验组和阳性对照组细胞形态开始变化,随着培养时间的延长,树突状结构更加明显,第12天细胞呈典型的树突状细胞形态;阴性对照组细胞生长缓慢,培养至第12天细胞呈梭形巨噬细胞形态。培养至10天的实验组细胞扫描电镜下呈典型的树突状细胞形态。培养12天实验组、阳性对照组细胞分别高表达DCs特异性抗原CD1a、CD80、CD83和CD86,与阴性对照组对应比较差异均有显著性(P<0.01);实验组与阳性对照组之间比较无统计学意义(P>0.05)。混合淋巴细胞反应显示经黄芪多糖诱生的DCs对同种异体T淋巴细胞具有明显刺激增殖作用。结论:黄芪多糖及细胞因子体外均可诱导脐血单核细胞(DCs前体细胞)定向分化为功能性(成熟)DCs;黄芪多糖诱生的DCs具有明显刺激同种异体T淋巴细胞增殖的能力,且随DCs细胞数量的增加而作用增强。
Objective:To observe the effect of Astragalus polysaccharides (APS) on inducing the cord blood monocytes into mature dendritic cells(DCs) in vitro and to investigate their morphous, cellar immunological characteristics,and contribution to T cell proliferation. Methods:(1)The cord blood monocytes were isolated by lymphocyte isolating solution under axenic condition, and three groups were divided. (2)Cells cultured with APS in concentration of 100 mg/L as the experiment group, that with the cytokines of GMCSF/IL-4/TNF-α as the positive control, and another without either GM-CSF/IL-4/TNF-α or APS as the negative control, respectively. The morphology of DCs was identified by inverted optical microscope or scanning electron microscope. The phenotype of 12 days cultures of DCs( CD1a, CD80, CD86 and CD83) were identified by flowcytometry. The DCs preparations from the experiment group were treated with mitomycin for 45 min to remove their proliferative activity as incentive cells in the mixed cultures with allogenic peripheral blood mononuclear cells from healthy volunteers as responders. T cell proliferation induced with the DCs preparations was detected by MTr chromatometry. Results: After cultured for 72 hours, the cell of both the experiment group and the positive control grew dusteringly and began to change from round to irregularin shapes. The longer the cells were cultured, the more obvious the dendritic structure is. The cells of experiment group and the positive control group when cultured for 12 days had the most typical dendritic structure. The negative control group cells had no dendritic structure and became macrophages when cultured for 12 days. The experiment group cells cultured for 10 days showed typical dendritic morphology by SEM. The experiment group cells and the positive group cells cultured for 12 days significantly expressed high level of the phenotypes of DCs( CD1a, CD80, CD86 and CD83) by flowcytometry. And the difference exhibitied statistioal significance when
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2007年第6期539-544,550,共7页
Chinese Journal of Immunology
基金
浙江省中医药管理局科研基金资助项目(No.2005C009)
杭州市卫生局科研基金资助项目(No.2004A008)
关键词
脐血
树突状细胞
黄芪多糖/药理学
外周血单个核细胞
T细胞增殖
Cord blood
Dendritic cells(DCs)
Astragalus polysaccharides(APS)/pharmacology
Peripheral blood mononuclear cell
T cell proliferation