耐受性树突状细胞体外培养的基本特征  

Basic characteristics of tolerogenic dendritic cells cultured in vitro

作　　者：钟志宏[1] 陈国安[2] 袁利亚[3] 鄢俊[4] 戎吉平[3] 俞火[3] 

机构地区：[1]赣南医学院教务处,江西省赣州市341000 [2]南昌大学第一附属医院血液科,江西省南昌市330006 [3]江西省医学科学研究所,江西省南昌市330006 [4]赣南医学院第一附属医院肿瘤科,江西省赣州市341000

出　　处：《中国组织工程研究与临床康复》2009年第31期6075-6079,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：国家自然科学基金资助项目(30260039);江西省自然科学基金资助项目(2007GZY2599)~~

摘　　要：背景:树突状细胞是体内最重要的抗原提呈细胞,在免疫调节中扮演着免疫应答和免疫耐受的双重角色,对维持机体的免疫平衡起重要作用。目的:探讨耐受性树突状细胞的体外培养方法,观察不同培养条件下耐受性树突状细胞的基本特征。设计、时间及地点:以细胞为观察对象,随机分组设计,于2006-09/2008-02在南昌大学第一附属医院中心实验室完成。材料:8周龄的Balb/c纯系雄性小鼠,体质量20～25g,用于分离和制备骨髓单个核细胞。方法:将小鼠骨髓单个核细胞分为5组在不同的培养体系中进行体外诱导培养:空白对照组(不加任何因子)、树突状细胞组(粒-巨噬细胞集落刺激因子+白细胞介素4)、血管活性肠肽组(粒-巨噬细胞集落刺激因子+血管活性肠肽)、地塞米松组(粒-巨噬细胞集落刺激因子+地塞米松)、血管活性肠肽+地塞米松组(粒-巨噬细胞集落刺激因子+血管活性肠肽+地塞米松)。主要观察指标:光镜下动态观察细胞形态,流式细胞仪检测细胞CD80、CD86、CD40、CD11c表达,四甲基偶氮唑盐法检测树突状细胞刺激淋巴细胞增殖的活性,特异性夹心酶联免疫分析测定树突状细胞上清液中白细胞介素10、白细胞介素12的水平。结果:①利用粒-巨噬细胞集落刺激因子、白细胞介素4、血管活性肠肽、地塞米松、脂多糖等因子体外培养能生成具有典型树枝状突起的树突状细胞。②树突状细胞组、血管活性肠肽组、地塞米松组、血管活性肠肽+地塞米松组CD11c的表达高于空白对照组(P<0.05)。血管活性肠肽组、地塞米松组、血管活性肠肽+地塞米松组CD40、CD80和CD86表达,淋巴细胞增殖活性及培养上清液白细胞介素12水平均低于树突状细胞组(P<0.05),血管活性肠肽组以质量浓度为40μg/L、地塞米松组以10μg/L减低明显,其中血管活性肠肽+地塞米松组在血管活性肠肽质量浓度为40μg/L、地塞BACKGROUND： Dentritic cells are the most important antigen presenting cells in vivo and play a double role of immune response and immune tolerance; in addition, the cells are significance for maintaining immune balance. OBJECTIVE： To study the culture method of tolerogenic dendritic cells in vitro, and observe the basic characters. DESIGN, TIME AND SETTING： A randomized grouping study based on cytology level was performed at the Central Laboratory of the First Affiliated Hospital of Nanchang University from September 2006 to February 2008. MATERIALS： Male Balb/c mice aging 8 weeks and weighing 20 25 g were used for separate and establish mononuclear cells of bone marrow. METHODS： Mononuclear cells derived from bone marrow were randomly assigned into 5 groups： blank control group （without any factors）, dendritic cells group [granulocyte-macrophage colony stimulating factor （GM-CSF） ＋ interleukin-4 （IL-4）], vasoactive intestinal peptide （VIP） group （GM-CSF ＋ VIP）, dexamethasone （DXM） group （GM-CSF ＋ DXM）, and VIP ＋ DXM group （GM-CSF ＋ VIP ＋ DXM）. MAIN OUTCOME MEASURES： Cell morphology was observed under optic microscope; CD80, CD86, CD40, and CD1 lc expressions were detected using flow cytometry; the capability to stimulate the proliferation of lymphocytes by dendritic cells was examined with MFI- assay; IL-10 and IL-12 levels in dendritic cell supernatant were measured with enzyme linked immunosorbent assay （ELISA）. RESULTS： （1） The dendritic cells which could generate typical dendritic processes were cultured with GM-CSF, IL-4, VIP, DXM, and lipopolysaccharide in vivo. （2） CD1 lc expression in the dendritic cells, VIP, DXM, and VIP ＋ DXM groups was significantly higher than blank control group （P 〈 0.05）. CD40, CDSO, and CD86 expressions, proliferative activity of lymphocytes, and IL-12 level in the VlP, DXM, and VlP ＋ DXM groups were significantly lower than dendritic cells group （P 〈 0.05）. Density of 40 μg/L in the VlP gro

关 键 词：树突状细胞 免疫耐受 小鼠 细胞因子 

分 类 号：R318[医药卫生—生物医学工程]