摘要
目的:探讨胚胎干细胞(ESC)诱导分化为树突状细胞(DC)的实验方法,实现体外大规模扩增高纯度DC用于临床免疫治疗。方法:E14小鼠胚胎干细胞系在GM-CSF与IL-3联合作用下经胚胎体(EB)诱导分化为DC。不成熟DC(im-DC)流式细胞仪检测CD11 c、CD80、CD86、MHC II-DR表达。用磷酸脂多糖(LPS)促进DC的成熟,收获细胞后观察细胞形态及摄片并做扫描电镜检查,分析细胞表型并与不成熟DC细胞表型做对比,异体淋巴细胞增殖实验行功能检测。结果:ES细胞来源DC呈典型DC形态学表现。im-DC CD11 c、CD80、CD86、MHC-II表达低,m-DC CD11 c、CD80、CD86、MHC-II的表达均较前明显升高。功能检测发现,ES细胞来源的DC具有强烈的激发同种异体淋巴细胞增殖的作用,证实ES细胞来源的DC具备正常的免疫学功能。结论:使用GM-CSF联合IL-3能成功诱导E14胚胎干细胞发育为DC。故ES细胞可作为DC来源的一条新途径,对于体外大规模制备DC用于免疫过继治疗提供了一个较好的方法。
AIM. To investigate the method of directed differentiation dendritic cells (DC) from embryonic stem cells, and to amplify high purity dendritic cells in vitro for immunolgical therapy. METHODS: El4 embryonic stem cell line was generated ES - DC in complete medium supplemented with GM - CSF and IL - 3. Flow cytometry was used to determine CD11c, CD80, CD86, MHC- Ⅱ cell- surface phenotype in immatured ES- DC. Lipopolysaccharide (LPS) was added to induce the ES - DC maturation. The matured ES - DC was harvested 24 hours later to identifying with morphology and transmission electron microscopy. The phenotype of matured ES - DC was analyzed by flow eytometry and compared with the iramatured ES - DC. The antigen presenting was evaluated by mixed lymphocyte responses (MLR). RESULTS : The ES - DC had obviously dendritic processes under scanning electron microscope. The immature DCs expressed low level of CDIIc (4. 33±0.23)%, CD80 (7.62 ±0. 19) %, CD86 (4.77 ±1.22.) % and MHC-Ⅱ (9.68 ±0. 15) %. The mature DCs expressed higher level of CDIIc (47.36 ±2. 68)% , CD80 (74. 40 ± 1.47) %, CD86 (29.77 ±2.00) % and MHC - Ⅱ(87.56 ±2.75) %. MLR showed that ES-DCs effectively stimulated lymphocyte proliferation. CONCLUSIONS: These results provide evidence that dendritic cells can be generated from E14 embryonic cells with the stimulations of GM - CSF and IL- 3. The differentiated cells expresse high level of CD11c, CD80, CD86, MHC -Ⅱ and effectively stimulate lymphocytes to proliferate. ES cells may become new origin of DC for immunotherapy.
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2006年第8期1597-1601,共5页
Chinese Journal of Pathophysiology
基金
贵州省优秀青年科技人才专项基金资助项目
广东省自然科学基金资助项目(021839)
广东省卫生厅基金资助项目(B2002034)
关键词
树突细胞
胚胎干细胞
Dendritic cells
Embryonic stem cells