小鼠骨髓源性树突状细胞的体外诱导扩增和鉴定  

InvitroInduction,Expansion and Identification of Mouse Bone Marrow-derived Dendritic Cells

作　　者：马东霞[1] 赵越[1] 向莹[1] 刘斌[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院器官移植研究所,教育部/卫生部器官移植重点实验室,武汉430030

出　　处：《华中科技大学学报（医学版）》2013年第4期386-390,共5页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金资助项目(No.30972793)

摘　　要：目的体外诱导扩增获取大量高纯度的小鼠骨髓来源树突状细胞(DC),并研究不同生长状态的DC成熟情况及进行生物学特性鉴定。方法以重组小鼠粒细胞巨噬细胞集落刺激因子(rmGM-CSF)和重组小鼠白细胞介素-4(rmIL-4),体外诱导小鼠骨髓细胞分化为DC,脂多糖(LPS)刺激24h,倒置显微镜动态观察细胞形态学变化,并分别收集悬浮细胞、贴壁细胞,流式细胞术(FACS)分析DC细胞表面分子CD11c、MHCⅡ类分子、CD80和CD86的表达水平,并应用混合淋巴细胞反应(MLR)检测其刺激异基因淋巴细胞增殖的能力。结果经体外诱导培养第3天即可见大量细胞集落形成;培养至第9天,细胞形态不规则,具有典型DC形态。同组悬浮DC表面MHCⅡ类分子和共刺激分子CD80、CD86的表达水平高于贴壁DC。LPS刺激24h后,DC表面MHCⅡ类分子、CD80、CD86的表达水平显著升高,同时可以显著刺激同种异基因淋巴细胞增殖。结论体外简易诱导培养可以获得高纯度小鼠骨髓来源的各具特性的DC,为开展DC的相关实验研究提供大量稳定的细胞来源。Objective To obtain a large number of mouse bone marrow-derived dendritic cells（DC）of high purity in vitro in an attempt to examine the maturity of DCs under different growth conditions and identify their biological characteristics.Methods Bone marrow cells of mice were isolated and induced to differentiate into DCs in vitro by using recombinant mouse granulocyte-macrophage colony-stimulating factor（rmGM-CSF）and recombinant mouse interleukin-4（rmIL-4）.After 24hLPS stimulation,the morphology of DCs was observed under the light inverted microscope.The suspended and attached cells were harvested for detection of DC surface markers by flow cytometry.The mixed lymphocyte reaction was used to assess the ability of cultured cells to stimulate the proliferation of allogeneic T lymphocytes.Results On day 3of in vitro culture,a large number of colonies were found to form.On day 9,the cells assumed irregular appearance and displayed typical morphological feature of DCs.Mature DCs showed high level expression of major histocompatibility complex（MHC）classⅡ,CD80and CD86,and could significantly stimulate allogeneic T cells proliferation.Conclusion A simple protocol for in vitroinduction and expansion of DCs from mouse bone marrow is established.It may help provide sufficient naive cell sources for the experimental study.

关 键 词：树突状细胞 诱导 鉴定 

分 类 号：R329.2[医药卫生—人体解剖和组织胚胎学]